Prolonged Treatment with Inhaled Corticosteroids does not Normalize High Activity of Matrix Metalloproteinase-9 in Exhaled Breath Condensates of Children with Asthma
Katarzyna Grzela • Wioletta Zagorska • Alicja Krejner • Malgorzata Litwiniuk • Anna Zawadzka-Krajewska • Aleksandra Banaszkiewicz • Marek Kulus • Tomasz Grzela
Abstract The airway remodeling in asthma is associated with increased amount of matrix metalloproteinase (MMP)-9. High levels of MMP-9 were found in mucosal biopsies, sputum and in exhaled breath condensates (EBC) of asthma patients. However, there are no data concerning real in vivo activity. Inhaled corticosteroids are effective in asthma control, but it is unclear, whether they only attenuate inflammation, or also protect against progressive remodeling of respiratory tract. Therefore, the aim of the study was to assess the amount and activity of MMP-9 in context of pro-inflammatory cytokines (IL-6, IL-8 and tumor necrosis factor, TNF), measured in EBC of asthmasuffering children, treated with inhaled steroids. The study involved 27 children with asthma, continuously treated with inhaled fluticasone propionate, and 22 healthy controls. In addition to routine clinical screening, the selected cytokines in EBC were analyzed using Ultrasensitive ELISA, whereas activity of MMP-9 was assessed using a novel immunozymography method. Despite chronic treatment with inhaled steroids mean MMP-9/EBC activity in asthma group was significantly higher than in healthy controls. Moreover, high MMP-9/EBC in asthma-suffering children significantly correlated with IgE serum levels. The IL-6 and IL-8 concentration was below the detection limit in all EBC samples. TNF/EBC levels were similar in both, asthma and healthy children. We hypothesize that MMP-9 hyperactivity in asthma may be closely related to high IgE serum levels. Our results suggest that inhaled steroids may be ineffective to prevent asthma-associated airway remodeling. Finally, we emphasize the necessity of further research focused on MMP-9 inhibition in asthma treatment.
Keywords Asthma Exhaled breath condensate Inhaled steroids MMP-9 Remodeling
Mucosal Immune Development in Early Life: Setting the Stage
Sylvia Brugman • Olaf Perdijk • R. J. Joost van Neerven • Huub F. J. Savelkoul
Abstract Our environment poses a constant threat to our health. To survive, all organisms must be able to discriminate between good (food ingredients and microbes hat help digest our food) and bad (pathogenic microbes, viruses and toxins). In vertebrates, discrimination between beneficial and harmful antigens mainly occurs at the mucosal surfaces of the respiratory, digestive, urinary and genital tract. Here, an extensive network of cells and organs form the basis of what we have come to know as the mucosal immune system. The mucosal immune system is composed of a single epithelial cell layer protected by a mucus layer. Different immune cells monitor the basolateral side of the epithelial cells and dispersed secondary lymphoid organs, such as Peyer’s patches and isolated lymphoid follicles are equipped with immune cells able to mount appropriate and specific responses. This review will focus on the current knowledge on host, dietary and bacterial-derived factors that shape the mucosal immune system before and after birth. We will discuss current knowledge on fetal immunity (both responsiveness and lymphoid organ development) as well as the impact of diet and microbial colonization on neonatal immunity and disease susceptibility. Lastly, inflammatory bowel disease will be discussed as an example of how the composition of the microbiota might predispose to disease later in life. A fundamental understanding of the mechanisms involved in mucosal immune development and tolerance will aid nutritional intervention strategies to improve health in neonatal and adult life.
Keywords Mucosal immunity Development Airways Fetal Neonatal feeding Inflammatory bowel disease
CD133-Positive Cells from Non-Small Cell Lung Cancer Show
Distinct Sensitivity to Cisplatin and Afatinib
Angela Alama • Rosaria Gangemi • Silvano Ferrini •
Gaia Barisione • Anna Maria Orengo • Mauro Truini •
Maria Giovanna Dal Bello • Francesco Grossi
Received: 15 May 2014 / Accepted: 7 October 2014 / Published online: 13 February 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract The standard of care for advanced non-small
cell lung cancer (NSCLC) consists in cisplatin-combina-
tion chemotherapy. In patients bearing tumors with
activating mutations of the epidermal growth factor re-
ceptor (EGFR), the inhibition of the EGFR intracellular
tyrosine kinase can induce up to 80 % response rates.
However, both therapeutic strategies will eventually lead to
recurrent disease due to the development of drug resis-
tance. The identification of rare cancer stem-like cells able
to repopulate the tumor, after failure to standard treatment
modalities, has led to characterize these cells as potential
therapeutic targets. This article will address the role of the
CD133/EpCAM stem cell-related markers and explore cell
sensitivity to cisplatin and to the EGFR-tyrosine kinase
inhibitor, afatinib. Three human NSCLC cell lines, one
wild-type (A549) and two harboring EGFR mutations
(H1650 and H1975), as well as 20 NSCLC primary cul-
tures, were grown in non-differentiating culture conditions
for stem cell enrichment. Flow-cytometry analyses of
CD133 and EpCAM and cell sensitivity to cisplatin and
afatinib were performed. Moreover, the expression of ac-
tivated EGFR was assessed by Western blot. The cell lines
and primary cultures grown in non-differentiating culture
conditions were enriched with CD133/EpCAM-positive
cells and were significantly more resistant to cisplatin and
more sensitive to afatinib as compared to the differentiated
counterpart. In addition, increased EGFR-phosphorylation
in non-differentiated cultures was observed. The present
findings suggest that afatinib might be beneficial for pa-
tients bearing tumors with constitutively activated EGFR,
to target chemo-resistant CD133/EpCAM-positive cancer
stem cells.
Keywords NSCLC CD133 EpCAM Cisplatin
Afatinib Drug-resistance
From Mysterious Supernatant Entity to miRNA-150 in Antigen-
Specific Exosomes: a History of Hapten-Specific T Suppressor
Factor
Włodzimierz Ptak • Katarzyna Nazimek •
Philip W. Askenase • Krzysztof Bryniarski
Received: 18 November 2014 / Accepted: 26 January 2015 / Published online: 18 February 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract Soon after the discovery of T suppressor cells
by Gershon in 1970, it was demonstrated that one sub-
population of these lymphocytes induced by i.v. hapten
injection suppresses contact sensitivity response mediated
by effector CD4? or CD8? T cells in mice through the
release of soluble T suppressor factor (TsF) that acts
antigen specifically. Our experiments showed that
biologically active TsF is a complex entity consisting of
two subfactors, one antigen specific and other non-specific,
produced by differently induced populations of cells. In
following years, we found that the antigen-specific sub-
factor is a light chain of IgM antibody that is produced by
B1a lymphocytes. However, the exact nature of non-
specific part remained a mystery for about 30 years. Our
current studies characterized TsF as regulatory miRNA-
150 carried by T suppressor cell-derived exosomes that are
antigen specific due to a surface coat of IgM antibody light
chains produced by B1a cells. The present communication
briefly summarizes our studies on TsF that led to discovery
of regulating miRNA that acts antigen specifically to sup-
press immune response.
Keywords T suppressor factor T suppressor cells
miRNA Exosomes Immune suppression
Immune tolerance Tolerosomes
Human Microbiome: When a Friend Becomes an Enemy
Magdalena Muszer • Magdalena Noszczyn´ ska •
Katarzyna Kasperkiewicz • Mikael Skurnik
Received: 10 July 2014 / Accepted: 12 December 2014 / Published online: 15 February 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract The microorganisms that inhabit humans are
very diverse on different body sites and tracts. Each
specific niche contains a unique composition of the mi-
croorganisms that are important for a balanced human
physiology. Microbial cells outnumber human cells by
tenfold and they function as an invisible organ that is called
the microbiome. Excessive use of antibiotics and unhealthy
diets pose a serious danger to the composition of the mi-
crobiome. An imbalance in the microbial community may
cause pathological conditions of the digestive system such
as obesity, cancer and inflammatory bowel disease; of the
skin such as atopic dermatitis, psoriasis and acne and of the
cardiovascular system such as atherosclerosis. An unbal-
anced microbiome has also been associated with
neurodevelopmental disorders such as autism and multiple
sclerosis. While the microbiome has a strong impact on the
development of the host immune system, it is suspected
that it can also be the cause of certain autoimmune dis-
eases, including diabetes or rheumatoid arthritis. Despite
the enormous progress in the field, the interactions between
the human body and its microbiome still remain largely
unknown. A better characterization of the interactions may
allow for a deeper understanding of human disease states
and help to elucidate a possible association between the
composition of the microbiome and certain pathologies.
This review focuses on general findings that are related to
the area and provides no detailed information about the
case of study. The aim is to give some initial insight on the
studies of the microbiome and its connection with human
health.
Keywords Microbiome Bacteria Homeostasis
Health
A Targeted Multiple Antigenic Peptide Vaccine Augments
the Immune Response to Self TGF-b1 and Suppresses Ongoing
Hepatic Fibrosis
Yaping Li • Wenjun Wang • Xiaoli Jia • Song Zhai •
Xiufang Wang • Yuan Wang • Shuangsuo Dang
Received: 18 July 2014 / Accepted: 5 January 2015 / Published online: 5 March 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Transforming growth factor (TGF)-b1 expres-
sion is induced upon liver injury, and plays a critical role in
hepatic fibrosis. Antibodies against TGF-b1 represent a
novel approach in the treatment of hepatic fibrosis. However,
TGF-b1 is not a suitable antigen due to immunological
tolerance. In the current study, we synthesized a multiple
antigenic peptide (MAP) vaccine against the dominant
B-cell epitope of TGF-b1. The immunogenicity and poten-
tial therapeutic effects of this vaccine were examined using a
rat model of hepatic fibrosis. Dominant B-cell epitopes of
TGF-b1 were identified using bioinformatic program. An
MAP vaccine corresponding to the 90–98 amino acid do-
main of TGF-b1 and containing four dendritic arms was
synthesized using a 9-fluorenylmethoxycarbonyl solid phase
method. Hepatic fibrosis which was induced in male Spra-
gue-Dawley rats received a high-fat diet and ethanol (1.8 g/
kg). Starting from the third week, rats were exposed to 40 %
carbon tetrachloride (CCl4; 150 ll/100 g body weight twice
weekly, initially 200 ll/100 g) treatment for a duration of
8 weeks. Rats received the MAP vaccine (100 lg) or Fre-
und’s adjuvant at weeks 1, 3, 5. A group of rats receiving the
fibrosis-inducing regimen alone and a group of healthy rats
(receiving an olive oil vehicle alone) were included as
controls. At the conclusion of the experiment, serum titre of
TGF-b1 antibody was measured using ELISA and a standard
liver functional test panel was examined. The extent of
hepatic fibrosis was determined by measuring hydroxypro-
line content in the liver as well as hematoxylin–eosin (HE)
and van Gieson (VG) staining. The expression of TGF-b1
and alpha-smooth muscle actin (a-SMA) was examined
using immunohistochemistry, and presented as positive
staining cells. The MAP purity was [90 % upon reverse
phase high-performance liquid chromatography, with ap-
parent molecular weight at 4.77 kDa. Serum TGF-b1
antibody titre was 1:256. The fibrosis-inducing treatment
produced significant liver damage, as reflected by increases
in liver functional test, HE and VG staining. The MAP
vaccine attenuated such damage, as reflected by decreased
alanine aminotransferase, aspartate aminotransferase, total
bilirubin, and hepatic hydroxyproline (116.78 ± 23.76 vs.
282.71 ± 136.94 IU/L; 319.78 ± 82.48 vs. 495.29 ±
137.13 IU/L; 2.02 ± 0.27 vs. 4.01 ± 0.52 lmol/L;
263.67 ± 41.18 vs. 439.14 ± 43.29 lg/g vs. in model rats,
respectively; p \ 0.01), as well as fibrosis extent by HE and
VG staining. The MAP vaccine reduced TGF-b1 and a–
SMA expression in rats (0.325 ± 0.059 vs. 0.507 ± 0.044
IOD/area; 0.318 ± 0.058 vs. 0.489 ± 0.029 IOD/area vs.
model rats, respectively; p \ 0.05). The TGF-b1 MAP
vaccine could generate sufficient antibody that suppresses
the development of hepatic fibrosis.
Keywords Hepatic fibrosis Transforming growth
factor-b1 Multiple antigenic peptides Vaccine
9a,11b-PGF2, a Prostaglandin D2 Metabolite, as a Marker of Mast
Cell Activation in Bee Venom-Allergic Patients
Marita Nittner-Marszalska • Ewa Cichocka-Jarosz •
Marek Sanak • Magdalena Wujczyk • Anna Dor-Wojnarowska •
Grzegorz Lis • Jerzy Liebhart
Received: 23 July 2014 / Accepted: 27 February 2015 / Published online: 13 March 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract Mast cell (MC) mediators, among them
prostaglandin D2 (PGD2) and 9a,11b-PGF2, PGD2’s
metabolite, play a key role in allergic reactions, including
bee venom anaphylaxis (BVA). Assessment of these me-
diators has never been performed in BVA. The aim of the
study was to assess the activation of MC during in vivo
provocation with bee venom (BV) and to measure PGD2
and 9a,11b-PGF2 in the course of an allergen challenge.
The second aim was to determine if assessment of these
mediators could be useful for predicting adverse events
during venom immunotherapy (VIT). In 16 BV-VIT pa-
tients and 12 healthy subjects, levels of PGD2 and 9a,11b–
PGF2 were assessed during BV provocation by means of
the skin chamber method. Chamber fluids, collected at 5
and 15 min, were analyzed for both mediators by gas
chromatography mass spectrometry negative ion chemical
ionization. BVA in comparison to non-allergic patients had
a significantly higher ratio of 9a,11b-PGF2 in allergen-
challenged chambers to 9a,11b-PGF2 in allergen-free
chambers after 15 min of provocation (p = 0.039). Aller-
gen challenge resulted in a significant increase of 9a,11b–
PGF2 levels between 5 and 15 min after provocation only
in BVA patients (p \ 0.05). Analysis of log-transformed
PGD2 levels showed significant difference between chan-
ges in PGD2 concentration between BVA and healthy
subjects. No study patient developed adverse reactions
during. 9a,11b-PGF2 is actively generated during the early
allergic response to BV. Skin chamber seems to be a
promising, non-invasive and safe model of in vivo allergen
provocation in BV-allergic patients. High or low levels of
both mediators do not predict occurrence of adverse events
during VIT.
Keywords PGD2 9a,11b-PGF2 Skin chamber method
Bee venom provocation
Macrophage Phenotype in the Ocular Surface of Experimental
Murine Dry Eye Disease
In-Cheon You • Terry G. Coursey • Fang Bian • Flavia L. Barbosa •
Cintia S. de Paiva • Stephen C. Pflugfelder
Received: 16 December 2014 / Accepted: 27 February 2015 / Published online: 13 March 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract To evaluate the phenotype of macrophages in
the cornea and conjunctiva of C57BL/6 mice with induced
experimental dry eye. C57BL/6 mice exposed to desic-
cating stress (DS) were evaluated at 1, 5, and 10 days and
C57BL/6 mice maintained in non-stressed environment
were used as controls. Whole eyes and adnexa were ex-
cised for histology or used for gene expression analysis.
Location and phenotype of macrophages infiltrating the
cornea and conjunctiva was evaluated by immunofluores-
cence analysis. Quantitative polymerase chain reaction
evaluated macrophage markers and T cell-related and in-
flammatory cytokine expression in cornea and conjunctiva.
Immunofluorescence staining demonstrated that macro-
phages reside in the conjunctiva of control and dry eye
mice and their number did not change with DS. Real-time
RT-PCR demonstrated that the level of M1 macrophage
marker, iNOS, increased prominently in the conjunctiva at
DS 10 days. In contrast, there was a non-significant de-
crease of the M2 marker Arg1 with DS. The levels of
inflammatory cytokine, IL-12a mRNA transcript in the
conjunctiva increased significantly at DS1 and decreased at
DS5, while levels of IL-18 were significantly increased at
DS 10. Macrophages reside in the ocular surface tissues of
C57BL/6 mice. Although the number of macrophages in
the conjunctiva does not change, evidence of inflammatory
M1 activation after desiccating stress was observed. Better
understanding of phagocyte diversity and activation in dry
eye disease provide a basis for the development of
phagocyte-targeted therapeutic strategies.
Keywords Dry eye disease Macrophage phenotype
Desiccating stress Ocular surface
Reestablishing T Cell Tolerance by Antibody-Based Therapy
in Type 1 Diabetes
Y. Maurice Morillon1 • Aaron Martin1 • Gregory Gojanovich1 • Bo Wang1 •
Roland Tisch1,2
Received: 11 November 2014 / Accepted: 2 February 2015 / Published online: 20 March 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Type 1 diabetes (T1D) is an autoimmune dis-
ease in which the insulin-producing b cells are selectively
destroyed. b cell-specific T cells are considered to be the
major mediators of pathology. Accordingly, most im-
munotherapies tested in the clinic to date have focused on
reestablishing self-tolerance within the T cell compart-
ment. Monoclonal antibodies (Ab) targeting a variety of
lymphocyte surface proteins have demonstrated benefits in
preclinical and clinical settings. Indeed, the use of Ab to
target T cells directly or indirectly has proven to be an
effective strategy to rapidly suppress b cell autoimmunity
and establish tissue-specific, long-term tolerance in rodent
T1D models. In this review, we describe a number of these
Ab-based immunotherapies, discuss associated immune
regulatory mechanisms, and highlight results obtained in
T1D clinical trials.
Keywords Autoimmunity Immunotherapy
Immunoregulation T cells
Combined Tumor Cell-Based Vaccination and Interleukin-12
Gene Therapy Polarizes the Tumor Microenvironment in Mice
Magdalena Jarosz-Biej1 • Ryszard Smolarczyk1 • Tomasz Cichon´ 1 •
Natalia Kułach1,2 • Justyna Czapla1 • Sybilla Matuszczak1 • Stanisław Szala1
Received: 2 December 2014 / Accepted: 9 February 2015 / Published online: 24 March 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract Tumor progression depends on tumor milieu,
which influences neovasculature formation and immuno-
suppression. Combining immunotherapy with antiangiogenic/
antivascular therapy might be an effective therapeutic ap-
proach. The aim of our study was to elaborate an anticancer
therapeutic strategy based on the induction of immune re-
sponse which leads to polarization of tumor milieu. To
achieve this, we developed a tumor cell-based vaccine.
CAMEL peptide was used as a B16-F10 cell death-inducing
agent. The lysates were used as a vaccine to immunize mice
bearing B16-F10 melanoma tumors. To further improve the
therapeutic effect of the vaccine, we combined it with in-
terleukin (IL)-12 gene therapy. IL-12, a cytokine with
antiangiogenic properties, activates nonspecific and specific
immune responses. We observed that combined therapy is
significantly more effective (as compared with mono-
therapies) in inhibiting tumor growth. Furthermore, the
tested combination polarizes the tumor microenvironment,
which results in a switch from a proangiogenic/immuno-
suppressive to an antiangiogenic/immunostimulatory one.
The switch manifests itself as a decreased number of tumor
blood vessels, increased levels of tumor-infiltrating CD4?,
CD8? and NK cells, as well as lower level of suppressor
lymphocytes (Treg). Our results suggest that polarizing
tumor milieu by such combined therapy does inhibit tumor
growth and seems to be a promising therapeutic strategy.
Keywords Tumor cell-based vaccine CAMEL
IL-12 Combined anti-tumor therapy
Polarization of tumor microenvironment
Assessment of the Effect of Methotrexate Therapy on Bone
Metabolism in Patients with Rheumatoid Arthritis
Jerzy S´wierkot1 • Katarzyna Gruszecka1 • Agnieszka Matuszewska2 •
Piotr Wiland1
Received: 22 April 2014 / Accepted: 2 February 2015 / Published online: 3 April 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract Proinflammatory cytokines and growth factors,
which regulate mutual interactions between immune sys-
tem cells and bone tissue cells, play a major role in the
formation of bone changes in rheumatoid arthritis (RA).
The aim of the work was to assess serum concentration of
osteoprotegerin (OPG), RANKL, Dkk-1 and sclerostin in
RA patients compared to a control group and to analyze
changes of these concentrations during methotrexate
(MTX) therapy. Patients enrolled in the study were 30
women of Caucasian origin aged 30–74 years with RA.
Patients with active form of the disease were administered
recommended doses of MTX for at least 6 months. The
study group was divided into subgroup I—patients with
improvement; and subgroup II—patients with no im-
provement. The control group consisted of 12 healthy
women in the age of 41–73. Before MTX therapy, RA
patients had higher levels of RANKL (644.97 ± 477.13 vs.
255.19 ± 130.26 pmol/l), lower values of OPG/RANKL
(0.01 ± 0.0101 vs. 0.02 ± 0.0078) and higher levels of
Dkk-1 protein (1821.32 ± 1060.28 vs. 548.52 ± 36.35 pg/
ml) compared to the control group. In the analyzed group
of patients (all patients receiving MTX regardless of re-
sponder non responder status) after 6 months of therapy, a
statistically significant increase in the ratio of OPG/
RANKL was found (0.0118 ± 0.0102 vs. 0.0141 ± 0.0118;
p = 0.02). The index value of OPG/RANKL differed sig-
nificantly depending on the resultant effect of treatment
(0.01702 ± 0.01274 in the subgroup of improvement vs.
0.00675 ± 0.00289 in the subgroup without improvement).
The difference in the mean concentrations of Dkk-1 before
and after treatment with MTX between subgroups I and II
was statistically significant (p = 0.002). In subgroup I,
mean concentration of Dkk-1 decreased after 6 months of
treatment with MTX (2054.72 ± 1004.74 vs. 1831.70 ±
851.70 pg/ml); while in subgroup II, the mean concentra-
tion of Dkk-1 increased (1214.48 ± 738.32 vs. 2275.01 ±
1385.23 pg/ml). There were no statistically significant
changes in the mean concentrations of sclerostin before and
after treatment with MTX (in whole group treatment with
MTX, in subgroup I, and in subgroup II). The results
confirm the presence of disorders of bone metabolism in
patients with RA. Treatment with MTX affects the value of
the ratio of OPG/RANKL and concentration of Dkk-1.
Keywords Dkk-1 Methotrexate Osteoprotegerin
RANKL Rheumatoid arthritis
Central Nervous System and Peripheral Expression of CCL19,
CCL21 and Their Receptor CCR7 in Experimental Model
of Multiple Sclerosis
Bartosz Bielecki1,4 • Izabela Jatczak-Pawlik2 • Pawel Wolinski2 • Andrzej Bednarek3 •
Andrzej Glabinski1,2
Received: 13 November 2014 / Accepted: 16 April 2015 / Published online: 10 May 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract It is well documented that inflammatory
chemokines play a significant role in the development of
multiple sclerosis (MS) and its model, experimental au-
toimmune encephalomyelitis (EAE). Recently, the
involvement of homeostatic (or lymphoid) chemokines in
the pathogenesis of autoimmune diseases has become an
object of intensive study. In this work, quantitative analysis
of CCL19, CCL21 and CCR7 expression in the central
nervous system (CNS), as well as in inflammatory
mononuclear cells isolated from several organs during the
first attack, remission and the second attack of chronic-
relapsing EAE (ChREAE), was performed. Using real-time
PCR, RNAse Protection Assay and immunohistochemistry,
the expression of both chemokines, as well as of their
common receptor CCR7, was analyzed in the brain, spleen,
lymph nodes and peripheral blood mononuclear cells. In-
creased expression of CCL19 and CCL21 was observed
mostly in mononuclear inflammatory cells isolated from
the CNS during active ChREAE. At the same time the
expression of CCR7 in blood mononuclear leukocytes was
reduced. This observation extends our current knowledge
about the possible role of chemokines CCL19, CCL21 and
their receptor CCR7 in the pathogenesis of ChREAE and,
by extension, MS.
Keywords Experimental autoimmune
encephalomyelitis Chemokine Chemokine receptor
Neuroinflammation
Transdifferentiation of Bone Marrow Mesenchymal Stem Cells
into the Islet-Like Cells: the Role of Extracellular Matrix Proteins
Marta Pokrywczynska1 • Marzena Anna Lewandowska2,3 • Sandra Krzyzanowska1 •
Arkadiusz Jundzill1 • Marta Rasmus1 • Karolina Warda1 • Maciej Gagat4 •
Aleksander Deptula5 • Anna Helmin-Basa6 • Marcin Holysz7 • Maciej Nowacki1 •
Lukasz Buchholz1 • Magdalena Bodnar8 • Andrzej Marszalek8,9 • Alina Grzanka4 •
Wojciech Jozwicki10 • Jacek Michalkiewicz6 • Tomasz Drewa1,11
Received: 12 August 2014 / Accepted: 16 April 2015 / Published online: 10 May 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Pancreatic islet implantation has been recently
shown to be an efficient method of treatment for type 1
diabetes. However, limited availability of donor islets re-
duces its use. Bone morrow would provide potentially
unlimited source of stem cells for generation of insulin-
producing cells. This study was performed to evaluate the
influence of extracellular matrix proteins like collagen,
laminin, and vitronectin on bone marrow mesenchymal
stem cells (BM-MSCs) transdifferentiation into islet-like
cells (ILCs) in vitro. To our knowledge, this is the first
report evaluating the importance of vitronectin in transdif-
ferentiation of BM-MSCs into ILCs. Rat BM-MSCs were
induced to ILCs using four-step protocol on plates coated
with collagen type IV, laminin type I and vitronectin type I.
Quantitative real-time PCR was performed to detect gene
expression related to pancreatic b cell development. The
induced cells expressed islet-related genes including:
neurogenin 3, neurogenic differentiation 1, paired box 4,
NK homeobox factor 6.1, glucagon, insulin 1 and insulin 2.
Laminin but not collagen type IV or vitronectin enhanced
expression of insulin and promoted formation of islet-like
structures in monolayer culture. Laminin triggered
transdifferentiation of BM-MSCs into ILCs.
Keywords Mesenchymal stem cells Islet-like cells
Collagen Laminin Vitronectin Transdifferentiation
Epstein–Barr Virus-Positive Diffuse Large B cell Lymphoma
in the Experience of a Tertiary Medical Center in Poland
Mateusz Ziarkiewicz1 • Dominika Wołosz2 • Tomasz Dziecia˛tkowski3,4 •
Ewa Wilczek2 • Jadwiga Dwilewicz-Trojaczek1 • Wiesław Wiktor Je˛drzejczak1 •
Beata Gierej2 • Bogna Ziarkiewicz-Wro´blewska2
Received: 9 October 2014 / Accepted: 15 May 2015 / Published online: 18 June 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract The role of Epstein–Barr virus (EBV) in the
biology and clinical characteristics of diffuse large B cell
lymphoma (DLBCL) is still poorly defined. A new provi-
sional entity EBV-positive DLBCL of the elderly has been
described in Asian population. Its incidence and prognosis
remains unknown in middle European patients. Clinical data
and tissue samples were collected from 74 Caucasian
patients with DLBCL, aged between 23 and 86 years,
treated at a single institution. Lymphoma morphology was
reassessed, laboratory procedures included in situ
hybridization specific for EBV-encoded small RNAs
(EBER), immunohistochemical staining for latent mem-
brane protein and serological testing for EBV-specific
antibodies. EBER staining revealed 12.2 % of EBV-positive
cases, whereas 9.5 % were diagnosed as EBV-positive
DLBCL of the elderly. Serologic EBV markers did not
correlate with the presence of EBV in tissue samples
(P [ 0.10). Elderly EBV-positive cases had lower BCL-6
(P = 0.038) and higher CD30 (P = 0.049) expression and
were characterized by higher progression risk (median time-
to-progression 12.5 months vs not reached; P = 0.029) and
a trend towards worse overall survival (median overall
survival 24.5 months vs not reached; P = 0.059). EBV-
positive DLBCL of the elderly occurs relatively frequently
in Polish population and may be associated with inferior
prognosis in comparison with DLBCL, not otherwise
specified.
Keywords Diffuse large B cell lymphoma DLBCL
Epstein–Barr virus EBV EBV-positive
DLBCL of the elderly EBER EBV serology
Insights into Myeloid-Derived Suppressor Cells in Inflammatory
Diseases
Yewon Kwak1 • Hye-Eun Kim1 • Sung Gyoo Park1
Received: 5 December 2014 / Accepted: 14 May 2015 / Published online: 20 May 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Myeloid-derived suppressor cells (MDSCs) are
a heterogeneous population of cells involved in immune
regulation. This population subdivides into granulocytic
MDSCs and monocytic MDSCs, which regulate immune
responses via the production of various molecules includ-
ing reactive oxygen species, nitric oxide, arginase-1,
interleukin-10, and transforming growth factor-b. Most
studies of MDSCs focused on their role in tumors. MDSCs
protect tumor cells from immune responses, and thus the
frequency of MDSCs associates with poor prognosis. Many
recent studies reported an important role for MDSCs in
inflammatory diseases via the regulation of immune cells.
In addition, the utilization of MDSCs by infectious
pathogens suggests an immune evasion mechanism. Thus,
MDSCs are important immune regulators in inflammatory
diseases, as well as in tumors. This review focuses on the
role of MDSCs in the regulation of inflammation in non-
tumor settings.
Keywords Myeloid-derived suppressor cells
Inflammation Autoimmunity Infection
Transplantation
Clinical and In Vitro Studies on Impact of High-Dose Etoposide
Pharmacokinetics Prior Allogeneic Hematopoietic Stem Cell
Transplantation for Childhood Acute Lymphoblastic Leukemia
on the Risk of Post-Transplant Leukemia Relapse
Joanna Sobiak1 • Urszula Kazimierczak2 • Dariusz W. Kowalczyk2,3 •
Maria Chrzanowska1 • Jan Styczyn´ ski4 • Mariusz Wysocki4 • Dawid Szpecht5 •
Jacek Wachowiak5
Received: 9 October 2014 / Accepted: 26 May 2015 / Published online: 4 June 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract The impact of etoposide (VP-16) plasma con-
centrations on the day of allogeneic hematopoietic stem
cell transplantation (allo-HSCT) on leukemia-free survival
in children with acute lymphoblastic leukemia (ALL) was
studied. In addition, the in vitro effects of VP-16 on the
lymphocytes proliferation, cytotoxic activity and on Th1/
Th2 cytokine responses were assessed. In 31 children
undergoing allo-HSCT, VP-16 plasma concentrations were
determined up to 120 h after the infusion using the HPLC–
UV method. For mentioned in vitro studies, VP-16 plasma
concentrations observed on allo-HSCT day were used. In
84 % of children, VP-16 plasma concentrations
(0.1–1.5 lg/mL) were quantifiable 72 h after the end of the
drug infusion, i.e. when allo-HSCT should be performed.
In 20 (65 %) children allo-HSCT was performed 4 days
after the end of the drug infusion, and VP-16 was still
detectable (0.1–0.9 lg/mL) in plasma of 12 (39 %) of
them. Post-transplant ALL relapse occurred in four
children, in all of them VP-16 was detectable in plasma
(0.1–0.8 lg/mL) on allo-HSCT day, while there was no
relapse in children with undetectable VP-16. In in vitro
studies, VP-16 demonstrated impact on the proliferation
activity of stimulated lymphocytes depending on its con-
centration and exposition time. The presence of VP-16 in
plasma on allo-HSCT day may demonstrate an adverse
effect on graft-versus-leukemia (GvL) reaction and
increase the risk of post-transplant ALL relapse. Therefore,
if 72 h after VP-16 administration its plasma concentration
is still above 0.1 lg/mL then the postponement of trans-
plantation for next 24 h should be considered to protect
GvL effector cells from transplant material.
Keywords Etoposide Conditioning
Hematopoietic stem cell transplantation
Graft versus leukemia
Pediatric acute lymphoblastic leukemia
The Role of IL-17 and Th17 Lymphocytes in Autoimmune
Diseases
Jacek Tabarkiewicz1 • Katarzyna Pogoda1 • Agnieszka Karczmarczyk2 •
Piotr Pozarowski3 • Krzysztof Giannopoulos2
Received: 10 October 2014 / Accepted: 26 May 2015 / Published online: 11 June 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract The end of twentieth century has introduced
some changes into T helper (Th) cells division. The iden-
tification of the new subpopulation of T helper cells
producing IL-17 modified model of Th1–Th2 paradigm and
it was named Th17. High abilities to stimulate acute and
chronic inflammation made these cells ideal candidate for
crucial player in development of autoimmune disorders.
Numerous publications based on animal and human models
confirmed their pivotal role in pathogenesis of human
systemic and organ-specific autoimmune diseases. These
findings made Th17 cells and pathways regulating their
development and function a good target for therapy.
Therapies based on inhibition of Th17-dependent pathways
are associated with clinical benefits, but on the other hand
are frequently inducing adverse effects. In this review, we
attempt to summarize researches focused on the impor-
tance of Th17 cells in development of human autoimmune
diseases as well as effectiveness of targeting IL-17 and its
pathways in pre-clinical and clinical studies.
Keywords Th17 IL-17 Autoimmune diseases
Therapy
Airway Remodeling in Chronic Obstructive Pulmonary Disease
and Asthma: the Role of Matrix Metalloproteinase-9
Katarzyna Grzela1 • Malgorzata Litwiniuk2,3 • Wioletta Zagorska1 •
Tomasz Grzela2
Received: 19 July 2014 / Accepted: 23 February 2015 / Published online: 28 June 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract Chronic obstructive pulmonary disease (COPD)
and asthma are both associated with airflow restriction and
progressive remodeling, which affect the respiratory tract.
Among various biological factors involved in the path-
omechanisms of both diseases, proteolytic enzymes—
matrix metalloproteinases (MMPs)—play an important
role, especially MMP-9. In this review, the authors discuss
the current topics of research concerning the possible role of
MMP-9 in both mentioned diseases. They include the
analysis of protein levels, nucleotide polymorphisms of
MMP-9 gene and their possible correlation with asthma and
COPD. Finally, the authors refer to the studies on MMP-9
inhibition as a new perspective for increasing the effec-
tiveness of treatment in asthma and COPD.
Keywords Asthma Chronic obstructive pulmonary
disease COPD MMP-9 Remodeling
Activation and Inhibition of ATM by Phytochemicals: Awakening
and Sleeping the Guardian Angel Naturally
Ammad Ahmad Farooqi1 • Shyh-Jong Wu2 • Yung-Ting Chang3 • Jen-Yang Tang4,5,6 •
Kun-Tzu Li7 • Muhammad Ismail8 • Chih-Chuang Liaw2,9 • Ruei-Nian Li7 •
Hsueh-Wei Chang7,10,11,12
Received: 3 December 2014 / Accepted: 17 February 2015 / Published online: 19 June 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Double-stranded breaks (DSBs) are cytotoxic
DNA lesions caused by oxygen radicals, ionizing radiation,
and radiomimetic chemicals. Increasing understanding of
DNA damage signaling has provided an ever-expanding
list of modulators reported to orchestrate DNA damage
repair and ataxia telangiectasia mutated (ATM) is the
master regulator and main transducer of the DSB response.
Increasingly, it is being realized that DNA damage
response is a synchronized and branched network that
functionalizes different molecular cascades to activate
special checkpoints, thus temporarily arresting progression
of the cell cycle while damage is being assessed and pro-
cessed. It is noteworthy that both nutrigenetics and
nutrigenomics have revolutionized the field of molecular
biology and rapidly accumulating experimental evidence
has started to shed light on biological activities of a wide
range of phytochemicals reported to modulate cell cycle,
DNA repair, cell growth, differentiation and apoptosis as
evidenced by cell-based studies. In this review, we have
attempted to provide an overview of DNA damage sig-
naling, how ATM signaling regulates tumor necrosis
factors-related apoptosis inducing ligand (TRAIL)-induced
intracellular network. We also illuminate on how resvera-
trol, epigallocatechin gallate, curcumin, jaceosidin,
cucurbitacin, apigenin, genistein, and others trigger acti-
vation of ATM in different cancer cells as well as agents
for ATM inactivation. Understanding the interplay of
TRAIL-induced intracellular signaling and ATM modula-
tion of downstream effectors is very important. This holds
particularly for a reconceptualization of the apparently
paradoxical roles and therapeutically targetable for
enhancing the response to DNA damage-inducing therapy
Keywords ATM inducers ATM inhibitors
Phytochemicals p53
Mesenchymal Stromal Cell Therapy in MDR/XDR Tuberculosis:
A Concise Review
Lavanya Joshi1 • Lakshmi Kiran Chelluri2 • Sumanlatha Gaddam1,3
Received: 12 November 2014 / Accepted: 14 April 2015 / Published online: 23 June 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Multi-drug-resistant (MDR) tuberculosis is a
major public health problem worldwide. Drug resistance
arises due to non-compliance of antibiotic therapy. Herein,
we explored the therapeutic options available ranging from
conservative treatment approaches to alternate adjunct
therapies such as mesenchymal stromal cell (MSC) therapy
interventions. It is attractive to understand the scientific
rationale of using cells as drugs, in particular mesenchymal
stem/stromal cells. The review dwells and attempts to
analyze the mechanistic approaches of the current treat-
ment modalities to modern therapies. MSCs have
demonstrated profound capacity to regenerate and repair.
They appear to modulate that the activities of dendritic
cells regulate T cells, both in vivo and in vitro. While there
seems to be some benefit of such therapies, its use warrants
further research. The merits and de-merits of autologous
therapy/allogeneic therapy are ill understood. The chal-
lenges of requirement of large number of cells for infusion,
the route of administration, choice of timing are complex
issues that need to be addressed. Furthermore, the host
immune responses, environmental factors and epigenetic
mechanisms compound the problem. Although, clinical
studies are being performed using autologous MSCs in
different inflammatory models, it is important that such an
intervention should be based on sound scientific rationale.
The current review examines the immunomodulatory
properties of MSCs, its interactions with other cell types, in
assessing the basis for autologous/allogeneic cell-based
therapies in the treatment of XDR/MDR tuberculosis.
Keywords Mesenchymal stem cell therapy
Immunomodulation Clinical trials Adjunct therapy
Endotoxin Elimination in Patients with Septic Shock:
An Observation Study
Barbara Adamik1 • Stanislaw Zielinski1 • Jakub Smiechowicz1 • Andrzej Ku¨ bler1
Received: 17 November 2014 / Accepted: 14 April 2015 / Published online: 21 June 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract To evaluate the effectiveness of endotoxin
elimination with an adsorption column in patients with
septic shock and endotoxemia. The elimination therapy
was guided by a new bedside method of measuring endo-
toxin activity (EA). Intensive care unit (ICU) patients with
septic shock and suspected Gram-negative infection were
consecutively added to the study group within the first
24 h. Endotoxin elimination was performed using hemop-
erfusion with the Alteco LPS Adsorber. The primary
endpoint was improvement in organ function within the
first 24 h of treatment. A secondary objective was to assess
the usefulness of a new method of measuring EA to help
guide endotoxin elimination therapy. Out of 64 patients 18
had a high baseline EA [0.70 EA units (0.66–0.77)]. Those
patients had endotoxin elimination treatment in addition to
conventional medical therapy. At 24 h after endotoxin
elimination, the EA had decreased to 0.56 EA units
(0.43–0.77), (p = 0.005); MAP increased from 69 (62–80)
to 80 mm Hg (68–88), (p = 0.002), and noradrenaline use
decreased from 0.28 (0.15–0.80) to 0.1 lg/kg/min
(0.00–0.70) at the same time (p = 0.04). The SOFA score
had decreased from 11 (9–15) to 9 (7–14) points 24 h after
endotoxin elimination (p = 0.01) with a median delta
SOFA –2 points. Endotoxin elimination did not have a
significant effect on the ICU length of stay or ICU mor-
tality. Effective endotoxin elimination resulted in a
significant improvement in hemodynamic parameters and
of organ function. The application of the EA assay was
useful for the bedside monitoring of endotoxemia in criti-
cally ill ICU patients.
Keywords Septic shock Endotoxin elimination
Gram-negative infection LPS measurement
Lymphangiogenesis and Inflammation—Looking for the ‘‘Missing
Pieces’’ of the Puzzle
Anca Maria Cimpean1 • Marius Raica1
Received: 8 February 2015 / Accepted: 27 April 2015 / Published online: 14 July 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Several papers about lymphangiogenesis and
inflammation focused on the detailed and complicated
descriptions of the molecular pathways accompanying both
non-tumor and tumor inflammatory-induced lymphatic
vessel development. Many authors are tempted to present
inflammatory-induced lymphangiogenesis in pathologic
conditions neglecting the role of inflammatory cells during
embryonic lymphatic vessel development. Some of the
inflammatory cells are largely characterized in inflamma-
tory-induced lymphangiogenesis, while others as mast
cells, eosinophils, or plasma cells are less studied. No
phenotypic characterization of inflammation-activated
lymphatic endothelial cell is available in this moment.
Another paradox is related to the existence of few papers
regarding lymphangiogenesis inside lymphoid organs and
for their related pathology. There are still several ‘‘missing
pieces of such a big puzzle’’ of lymphangiogenesis and
inflammation, with a direct impact on the ineffectiveness of
the anti-inflammatory therapy as lymphangiogenesis inhi-
bitors. The present paper will focus on the controversial
issues of lymphangiogenesis and inflammation.
Keywords Lymphangiogenesis Inflammation
Lymphatic vessels Inflammatory cells Endothelial cells
Genetic (KIR, HLA-C) and Some Clinical Parameters Influencing
the Level of Liver Enzymes and Early Virologic Response
in Patients with Chronic Hepatitis C
Iwona Mozer-Lisewska1 • Katarzyna Zwolin´ska2 • Arleta El_zbieta Kowala-Piaskowska1 •
Maciej Bura1 • Bła_zej Rozpłochowski1 • Anna Pauli1 • Jan _Zeromski3 •
Egbert Piasecki2 • Piotr Kus´nierczyk4
Received: 22 January 2015 / Accepted: 14 April 2015 / Published online: 24 July 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract Natural killer cells play an important role as
effectors of innate immunity and regulators of adaptive
immunity. They are important elements of the innate
response to viral infections, which they detect using human
leukocyte antigen (HLA) class I-binding receptors. Most
polymorphic of these are killer cell immunoglobulin-like
receptors (KIRs) which exist as two basic isotypes, acti-
vating or inhibitory receptors and are encoded by genes
distributed differently in unrelated individuals. We sear-
ched for links between selected clinical data (including
HCV viremia, liver enzymes level and liver histology
parameters) and the presence of genes encoding these
receptors and their ligands in hepatitis C virus-infected
individuals subjected to pegylated interferon-a and rib-
avirin therapy. Genomic DNA samples from two hundred
and ninety-two chronically infected patients were typed by
polymerase chain reaction for the presence or absence of
genes for KIRs and their ligands, class I HLA molecules,
and clinical data of the patients were collected. Our results
suggest an importance of clinical parameters and the con-
tribution of KIR and HLA genes to the course of hepatitis
C virus infection and the response to therapy. The study
revealed that levels of liver enzymes before therapy were
about 30 % higher in patients who possessed a variant
KIR2DS4 gene with 22-base pair deletion. Decrease of
ALT activity after treatment was higher in HLA–C C2–
positive than negative individuals. Beside it, patients
demonstrated early virologic response to the therapy if the
time lag before treatment was short, particularly in women.
Keywords KIR2DS4 HLA-C C2 ALT AST
HCV chronic infection Early virologic response
Inflammation in Alzheimer’s Disease and Molecular Genetics:
Recent Update
Zhi-Gang Zhang1 • Yan Li2 • Cheung Toa Ng1 • You-Qiang Song1,3
Received: 29 October 2014 / Accepted: 3 March 2015 / Published online: 1 August 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Alzheimer’s disease (AD) is a complex age-
related neurodegenerative disorder of the central nervous
system. Since the first description of AD in 1907, many
hypotheses have been established to explain its causes. The
inflammation theory is one of them. Pathological and
biochemical studies of brains from AD individuals have
provided solid evidence of the activation of inflammatory
pathways. Furthermore, people with long-term medication
of anti-inflammatory drugs have shown a reduced risk to
develop the disease. After three decades of genetic study in
AD, dozens of loci harboring genetic variants influencing
inflammatory pathways in AD patients has been identified
through genome-wide association studies (GWAS). The
most well-known GWAS risk factor that is responsible for
immune response and inflammation in AD development
should be APOE e4 allele. However, a growing number of
other GWAS risk AD candidate genes in inflammation
have recently been discovered. In the present study, we try
to review the inflammation in AD and immunity-associated
GWAS risk genes like HLA–DRB5/DRB1, INPP5D,
MEF2C, CR1, CLU and TREM2.
Keywords Inflammation Alzheimer’s disease
Genetics GWAS TREM2
Cannabis Allergy: What do We Know Anno 2015
Ine Decuyper1 • Hanne Ryckebosch1 • Athina L. Van Gasse1 • Vito Sabato1 •
Margaretha Faber1 • Chris H. Bridts1 • Didier G. Ebo1
Received: 10 January 2015 / Accepted: 27 February 2015 / Published online: 16 July 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract For about a decade, IgE-mediated cannabis
(marihuana) allergy seems to be on the rise. Both active
and passive exposure to cannabis allergens may lead to a
cannabis sensitization and/or allergy. The clinical mani-
festations of a cannabis allergy can vary from mild to life-
threatening reactions, often depending on the route of
exposure. In addition, sensitization to cannabis allergens
can trigger various secondary cross-allergies, mostly for
plant-derived food. This clinical entity, which we have
designated as the ‘‘cannabis-fruit/vegetable syndrome’’
might also imply cross-reactivity with tobacco, latex and
plant-food derived alcoholic beverages. These secondary
cross-allergies are mainly described in Europe and appear
to result from cross-reactivity between non-specific lipid
transfer proteins or thaumatin-like proteins present in
Cannabis sativa and their homologues that are ubiquitously
distributed throughout plant kingdom. At present, diagnosis
of cannabis-related allergies rests upon a thorough history
completed with skin testing using native extracts from buds
and leaves. However, quantification of specific IgE anti-
bodies and basophil activation tests can also be helpful to
establish correct diagnosis. In the absence of a cure,
treatment comprises absolute avoidance measures includ-
ing a stop of any further cannabis (ab)use.
Keywords Allergy Cannabis Prevalence Can s 3
Cross-reactivity Diagnosis IgE CD63
Basophil activation
The Q705K and F359L Single-Nucleotide Polymorphisms
of NOD-Like Receptor Signaling Pathway: Association
with Chronic Pancreatitis, Pancreatic Cancer, and Periodontitis
Andrzej Miskiewicz1 • Grzegorz Szparecki2 • Marek Durlik3 • Gra_zyna Rydzewska4 •
Ireneusz Ziobrowski3 • Renata Go´rska1
Received: 26 December 2014 / Accepted: 3 July 2015 / Published online: 8 August 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract The aim of this study was to establish the
correlation between the occurrence of Q705K and F359L
polymorphisms in patients diagnosed with pancreatic dis-
eases and periodontal conditions of various degrees of
severity. The above-mentioned genetic markers were
assessed in patients with pancreatic cancer (n = 18) and
chronic pancreatitis (n = 39) as well as in a healthy control
group (n = 115). The established inclusion criteria were
the following: Caucasian descent, non-smoking, and age
range 20–80, with different levels of periodontitis activity
according to S. Offenbacher’s scale. The genotyping
reactions were performed by means of an RT-PCR with the
use of TaqManÒ genotyping assay. Results of the study
revealed that the state of periodontium was significantly
worse in patients with chronic pancreatitis. The Q705K and
F359L polymorphisms were associated with more
advanced cases of periodontitis measured by clinical
attachment level, whereas the Q705K was associated with
intensified bleeding index. Furthermore, the F359L single-
nucleotide polymorphism was significantly higher in the
group with chronic pancreatitis (p \ 0.0001;
OR = 6.8571). Whereas, the prevalence of Q705K
polymorphism was higher in the group of pancreatic cancer
(p = 0.107; OR = 3.3939). This study suggests that the
exaggerated inflammatory response provoked by Q705K
and F359L might be the common denominator for peri-
odontitis, pancreatic cancer, and chronic pancreatitis.
These findings might constitute the basis for a new diag-
nostic and therapeutic approach.
Keywords Inflammasomes Innate immunity
Inflammation mediators Pancreatic cancer
Chronic pancreatitis Periodontitis
The Role of CXC Chemokines in Pulmonary Fibrosis of Systemic
Lupus Erythematosus Patients
Agnieszka Nielepkowicz-Goz´dzin´ ska1 • Wojciech Fendler2 • Ewa Robak3 •
Lilianna Kulczycka-Siennicka3 • Paweł Go´rski4 • Tadeusz Pietras4 •
Ewa Brzezian´ ska5 • Małgorzata Pietrusin´ ska4 • Adam Antczak1
Received: 27 October 2014 / Accepted: 5 May 2015 / Published online: 15 August 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract The inflammatory process in systemic lupus
erythematosus (SLE) affects many organs including the
lungs. CXC chemokines are suggested to play an important
role in the pathogenesis of SLE and pulmonary fibrosis. To
estimate the concentrations of CXCL9, CXCL10, CXCL11
in bronchoalveolar lavage fluid (BALF) of patients with
and without pulmonary involvements of SLE to evaluate
CXC chemokines role in the pathogenesis of pulmonary
fibrosis in SLE. Twenty-six SLE patients and 31 healthy
controls were evaluated using high-resolution computed
tomography (HRCT), pulmonary function tests, the SLE
Disease Activity Index (SLEDAI), assessing CXCL9,
CXCL11, CXCL10 level in BALF (an enzyme-im-
munosorbent assay kit). The mean CXCL9 and CXCL11
concentrations in BALF were higher in SLE patients
compared to healthy controls (34.09 ± 102.34 vs
10.98 ± 14.65 pg/mL, p \ 0.001; 72.65 ± 112.89 vs
16.12 ± 83.75 pg/mL, p = 0.012, respectively). The dis-
ease activity scored by SLEDAI and the concentration of
CXCL10 in BALF were significantly higher in the SLE
patients with pulmonary fibrosis when compared with
patients with normal HRCT (8.23 ± 3.19 vs 5.01 ± 2.41;
73.45 ± 34.12 vs 40.76 ± 41.65, respectively, in both
p \ 0.05). In SLE patients positive correlations were found
between SLEDAI and the percentage of lymphocytes in
BALF (r = 0.51, p \ 0.05); CXCL9 and CXCL10 con-
centrations in BALF (r = 0.65, p \ 0.001); CXCL9 and
CXCL11 concentrations in BALF (r = 0.69, p \ 0.001).
In lupus patients with pulmonary manifestations positive
correlations were found between CXCL11 concentration in
BALF and SLEDAI (r = 0.55, p \ 0.05), CXCL11 con-
centration and the percentage of neutrophils in BALF
(r = 0.69, p \ 0.05), CXCL10 concentration and the per-
centage of neutrophils in BALF (r = 0.57, p \ 0.05). Our
observations indicate that CXCL9 and CXCL11 play an
important role in the pathogenesis of SLE but it needs
further studies. These results suggest that CXCL10 and
CXCL11 are associated with neutrophils accumulation in
the alveolar space of SLE patients with pulmonary fibrosis
and should be considered as potential factor of interstitial
fibrosis.
Keywords Bronchoalveolar lavage fluid CXCL9
CXCL10 CXCL11 Pulmonary fibrosis
Systemic lupus erythematosus
Inflammatory Markers Change with Age, but do not Fall Beyond
Reported Normal Ranges
Aleksandra Wyczalkowska-Tomasik1 • Bozena Czarkowska-Paczek2 •
Magdalena Zielenkiewicz3 • Leszek Paczek1
Received: 16 March 2015 / Accepted: 25 May 2015 / Published online: 18 August 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract We examined the serum levels of IL-6, IL-8,
TNF, IL-6R, TNF-R1, and CRP and the dynamics of
changes in these levels according to age. The study
included healthy individuals of 20–90 years of age. Par-
ticipants were divided into subgroups based on their decade
of life, and into subgroups of C65 or \65 years. Serum
cytokine levels were assayed by ELISA, and CRP using an
immunoturbidimetric method. Serum CRP levels were
within the normal range for all subgroups. The 60- to
70-year age group showed higher CRP than the 20- to 30-
(p = 0.003), 30- to 40- (p = 0.009), and 40- to 50-
(p = 0.030) year age groups. Serum cytokine levels were
low. It was greater in the 60- to 70-year age group than in
the 20- to 30- (p = 0.008) and 30- to 40- (p = 0.040) year
groups, and was greater in the 70- to 90-year group than the
20- to 30-year group (p = 0.043). Serum TNF-R1 level in
the 70- to 90-year group was greater than in all other age
groups (p = 0.000 for all comparisons). Other measured
parameters did not differ between groups. Serum levels of
IL-6, CRP, and TNF-R1 were greater in participants C65
than \65 years of age. Healthy older people showed low
serum levels of CRP and pro-inflammatory cytokines, but
higher than in younger population. Therefore, the adjust-
ment of normal ranges in the elderly should be considered.
Serum levels of pro-inflammatory cytokines elevated
beyond normal ranges indicate particular diseases.
Keywords CRP Cytokines Healthy aging
Inflammation
Modulatory Effects of Oral Bovine Lactoferrin on the IgA
Response at Inductor and Effector Sites of Distal Small Intestine
from BALB/c Mice
Ivonne Maciel Arciniega-Martı´nez1 • Rafael Campos-Rodrı´guez1 •
Marı´a Elisa Drago-Serrano2 • Luvia Enid Sa´nchez-Torres3 •
Teresita Rocı´o Cruz-Herna´ndez1 • Aldo Arturo Rese´ndiz-Albor1
Received: 11 February 2015 / Accepted: 29 May 2015 / Published online: 30 August 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Bovine lactoferrin (bLf) up-modulates intestinal
IgA that is essential for homeostasis and which might confer
protection to the distal small intestine that is vulnerable to
inflammation. This study analyzed the effects of bLf
administered orally on the IgA response at inductive (Pey-
er’s patches) and effector (lamina propria) sites of the distal
small intestine in mice. Groups of five healthy male BALB/c
mice were orally treated with 5 mg of bLf for 7, 14, 21, or
28 days. Then, mice were killed and the distal small intes-
tine was dissected. Intestinal fluid samples were analyzed to
determine IgA and IgM levels by enzyme-immuno assay.
Peyer’s patches and lamina propria were analyzed for IgA?
or IgM? plasma cells, B, CD4? T and CD8? T cells as well
as CD4? T cells positive for either pro-inflammatory
cytokines [tumor necrosis factor (TNF)-a, interferon-c and
interleukin (IL)-12] or for IgA-producing ILs (IL-4, -5, -10
and -6) by cytofluorometry. Antibodies, antibody-secreting
cells, and B and T responses in both Peyer’s patches and
lamina propria were higher in bLf-treated than bLf-un-
treated mice. The generation of IL-10 and IL-6 CD4? T cells
in Peyer’s patches or TNF-a and IL-12 CD4? T cells in
lamina propria showed similar response patterns. On days 14
and 28, cytokine/IL CD4? T cell responses were increased
in Peyer’s patches or decreased in lamina propria. The effect
of bLf on the elicitation of IgA indicates a potential appli-
cation of bLf as a nutraceutical to control inflammation in
the distal small intestine.
Keywords Bovine lactoferrin IgA
Intestinal modulation Ileum Peyer’s patches
Intestinal lamina propria
The Role of Cathelicidin LL-37 in Cancer Development
Ewelina Piktel1 • Katarzyna Niemirowicz1 • Urszula Wnorowska1 •
Marzena Wa˛tek2 • Tomasz Wollny2 • Katarzyna Głuszek2 • Stanisław Go´z´dz´3 •
Ilya Levental4 • Robert Bucki1,5
Received: 1 April 2015 / Accepted: 29 May 2015 / Published online: 22 September 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract LL-37 is a C-terminal peptide proteolytically
released from 18 kDa human cathelicidin protein
(hCAP18). Chronic infections, inflammation, tissue injury
and tissue regeneration are all linked with neoplastic
growth, and involve LL-37 antibacterial and
immunomodulatory functions. Such a link points to the
possible involvement of LL-37 peptide in carcinogenesis.
An increasing amount of evidence suggests that LL-37 can
have two different and contradictory effects—promotion or
inhibition of tumor growth. The mechanisms are tissue-
specific, complex, and depend mostly on the ability of LL-
37 to act as a ligand for different membrane receptors
whose expression varies on different cancer cells. Over-
expression of LL-37 was found to promote development
and progression of ovarian, lung and breast cancers, and to
suppress tumorigenesis in colon and gastric cancer. This
review explores and summarizes the current views on how
LL-37 contributes to immunity, pathophysiology and cell
signaling involved in malignant tumor growth.
Keywords Cancer Immune system Carcinogenesis
Cathelicidin LL-37
Polymorphisms of the Toll-Like Receptor-3 Gene in Autoimmune
Adrenal Failure and Type 1 Diabetes in Polish Patients
Marta Fichna1,2,3 • Magdalena _Zurawek1 • Piotr Fichna4 •
Danuta Januszkiewicz-Lewandowska1 • Marek Ruchała2 • Jerzy Nowak1
Received: 26 January 2015 / Accepted: 8 June 2015 / Published online: 30 August 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract Infectious agents are plausible environmental
triggers for autoimmunity in genetically susceptible indi-
viduals. Polymorphic variants of genes implicated in innate
immunity may affect immune responses and hence pro-
mote auto-aggressive reactions. Genes such as Toll-like
receptor-3 (TLR3), which participate in recognizing con-
served foreign molecules and mounting the first line of
defence against viral infections, are promising functional
candidates in autoimmune conditions. We investigated the
association of the TLR3 variants, rs13126816 and
rs3775291, with the autoimmune endocrine disorders,
Addison’s disease (AD) and type 1 diabetes (T1D) in the
Polish population. The study comprised 168 AD patients,
524 individuals with T1D and 592 healthy controls.
Genotyping was performed by real-time PCR. Distribution
of the TLR3 genotypes and alleles did not reveal significant
differences between patients and controls (p [ 0.05). No
effect on age at disease onset was found in affected
cohorts. This analysis does not support an association
between TLR3 variants and the risk for autoimmune
destruction of the adrenal cortex and beta cells. However,
innate immunity merits further studies in autoimmune
endocrine conditions.
Keywords Addison’s disease Polymorphism
TLR3 Type 1 diabetes
Immunopathogenic Background of Pars Planitis
Joanna Przez´dziecka-Dołyk1 • Agnieszka We˛grzyn2 • Anna Turno-Kre˛cicka1 •
Marta Misiuk-Hojło1
Received: 27 October 2014 / Accepted: 29 April 2015 / Published online: 5 October 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract Pars planitis is defined as an intermediate
uveitis of unknown background of systemic disease with
characteristic formations such as vitreous snowballs,
snowbanks and changes in peripheral retina. The incidence
of pars planitis varies 2.4–15.4 % of the uveitis patients.
The pathogenesis of the disease is to be determined in
future. Clinical and histopathological findings suggest an
autoimmune etiology, most likely as a reaction to
endogenous antigen of unknown source, with T cells pre-
dominant in both vitreous and pars plana infiltrations. T
cells subsets play an important role as a memory-effector
peripheral cell. Snowbanks are formed as an effect of post
inflammatory glial proliferation of fibrous astrocytes. There
is also a genetic predisposition for pars planitis by human
leukocyte antigen and several other genes. A coexistence of
multiple sclerosis and optic neuritis has been described in
numerous studies. Epiretinal membrane, cataract, cystoid
macular edema, retinal detachment, retinal vasculitis,
neovascularization, vitreous peripheral traction, peripheral
hole formation, vitreous hemorrhage, disc edema are
common complications observed in pars planitis. There is a
need to expand the knowledge of the pathogenic and
immunologic background of the pars planitis to create an
accurate pharmacological treatment.
Keywords Pars planitis
Idiopathic uveitis intermedialis Iris Ciliary body
Immunology Eye
Targeting Th17 Effector Cytokines for the Treatment
of Autoimmune Diseases
Tetsuya Yamagata1 • Jill Skepner1 • Jianfei Yang1
Received: 19 March 2015 / Accepted: 28 July 2015 / Published online: 10 September 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Interleukin (IL)-17-producing T cells, espe-
cially T helper (Th)17 cells, play a critical role in the
pathogenesis of a variety of autoimmune inflammatory
diseases. The pathogenic function of Th17 cells results
from their production of Th17 effector cytokines, namely
IL-17 (or IL-17A), IL-17F, IL-22 and IL-26. The impor-
tance of IL-17 has been demonstrated by antibody
neutralization studies in both animal models of autoim-
mune diseases as well as in human clinical trials. This
review highlights the current knowledge of the clinical
aspects of the Th17 cytokines as well as therapeutic anti-
bodies against IL-17, IL-17F, IL-17 receptor, IL-22, IL-26
and granulocyte macrophage colony-stimulating factor for
the future treatment of autoimmune inflammatory diseases.
Keywords Th17 IL-17 IL-22 IL-26
Autoimmune disease
Impact of Whole Body Cryotherapy at 2110 °C on Subjects
with Arterial Hypertension
M. Missmann1 • M. Himsl2 • E. Mur3 • H. Ulmer4 • P. Marschang3
Received: 10 June 2014 / Accepted: 12 May 2015 / Published online: 25 September 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Whole body cryotherapy (WBC) in a cryo-
chamber as a medical treatment was first established in
Japan in the 1980s, later in Central Europe, and is now
becoming more popular also in the United States. The
exposure to extreme, non-physiological environmental
conditions in a cryo-chamber at –110 °C may exceed the
normal adaption capacity. The aim of this study was to
investigate the effects of WBC on blood pressure (BP)
readings in adult subjects with rheumatic disorders and
normal or moderately elevated BP. A sample of 23 subjects
(8 female, 15 male) which were recruited according to their
pathology between the age of 35 and 69 years undergoing
21 WBC applications was divided into three groups: a
group of subjects with anti-hypertensive therapy, a group
of subjects with mild arterial hypertension without medical
treatment, and a normotensive control-group. A total of
483 BP readings were taken immediately before and after
each WBC application. The systolic and diastolic BP were
recorded, and the mean arterial pressure, and the amplitude
of BP were calculated. A statistically significant rise of BP
after WBC was found in the whole sample and in the
normotensive group. Over the course of time, no significant
change of BP behavior was observed, except for nor-
motensive subjects, who showed a wider range in their
systolic BP values. Generally accepted exclusion criteria
were applied, and in our sample group WBC was safe with
respect to unwanted BP alterations for adult subjects under
70 years—regardless of a pre-existing untreated mild or
pharmacologically treated arterial hypertension. Greater
changes of BP values might infrequently occur, so an
individual monitoring of subjects is necessary.
Keywords Whole body cryotherapy Cryo-chamber
Blood pressure Blood pressure response –110 °C
Arterial hypertension
Virus-Specific Cellular Response in Hepatitis C Virus Infection
Justyna Kaz´mierczak1 • Kamila Caraballo Cortes1 • Iwona Bukowska-Os´ko1 •
Marek Radkowski1
Received: 4 February 2015 / Accepted: 8 June 2015 / Published online: 1 October 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Studies performed on chimpanzees and humans
have revealed that strong, multispecific and sustained
CD4? and CD8? T cell immune responses is a major
determinant of hepatitis C virus (HCV) clearance. How-
ever, spontaneous elimination of the virus occurs in
minority of infected individuals and cellular response
directed against HCV antigens is not persistent in indi-
viduals with chronic infection. This review presents
characteristics of the HCV-specific T cell response in
patients with different clinical course of infection, includ-
ing acute and chronic infection, persons who spontaneously
eliminated HCV and non-infected subjects exposed to
HCV. Detection of HCV-specific response, especially in
non-infected subjects exposed to HCV, may be indicative
of HCV prevalence in population and rate of spontaneous
viral clearance. Understanding the mechanisms and role of
HCV-specific cellular immune response would contribute
to better understanding of HCV epidemiology,
immunopathogenesis and may help to design an effective
vaccine.
Keywords HCV Virus-specific cellular response
CD4? CD8? T cells
Type I, II, and III Interferons: Regulating Immunity
to Mycobacterium tuberculosis Infection
Maja Travar1,2 • Miroslav Petkovic2 • Antonija Verhaz3
Received: 20 April 2015 / Accepted: 22 July 2015 / Published online: 11 September 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Interferons (IFNs) are cytokines released by
host cells in response to the presence of pathogens or tumor
cells. The aim of this review was to present the previously
known and new findings about the role of interferons type I
and II, and recently discovered type III in Mycobacterium
tuberculosis (M. tuberculosis) infection control. Infection
of various cell types with M. tuberculosis induce both IFN-
a and IFN-b synthesis. The majority of the studies support
the findings that IFN type I actually promotes infection
with M. tuberculosis. It has been well establish that IFN-c
has protective function against M. tuberculosis and the
other mycobacteria and that the primary source of this
cytokine are CD4? and CD8? T cells. Recently, it has been
shown that also the innate lymphocytes, cd T cells, natural
killer (NK) T cells, and NK cells can also be the source of
IFN-c in response to mycobacterial infection. Several
studies have shown that CD4? T cells protect mice against
M. tuberculosis independently of IFN-c. The balance
between IFN-c and different cytokines such as IL-10 and
other Th2 cell cytokines is likely to influence disease
outcome. Type I IFN appears to be detrimental through at
least three separate, but overlapping, type I IFN-mediated
mechanisms: induction of excessive apoptosis, specific
suppression of Th1 and IFN-c responses, and dampening of
the immune response by strong IL-10 induction. Recently it
has been found that M. tuberculosis infection in A549 lung
epithelial cells stimulate up-regulation of IFN-k genes
in vitro. IFN-ks also have a role in modulation of Th1/Th2
response. IFN-ks are not essential for M. tuberculosis
infection control, but can give some contribution in
immune response to this pathogen.
Keywords Interferon Interferon lambda
Mycobacterium tuberculosis
HMGB1 Inhibition During Zymosan-Induced Inflammation:
The Potential Therapeutic Action of Riboflavin
Agnieszka Irena Mazur-Bialy1 • Ewa Pochec´2
Received: 12 February 2015 / Accepted: 31 August 2015 / Published online: 7 October 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract Sepsis, also known as systemic inflammatory
response syndrome, is a life-threatening condition caused
by a pathogenic agent and leading to multiple organ dys-
function syndrome. One of the factors responsible for the
excessive intensification of the inflammatory response in
the course of inflammation is high-mobility group protein
B1 (HMGB1). HMG-1 is a nuclear protein which, after
being released to the intercellular space, has a highly pro-
inflammatory effect and acts as a late mediator of lethal
damage. The purpose of this study was to examine whether
the anti-inflammatory action of riboflavin is accompanied
by inhibition of HMGB1 release during peritoneal
inflammation and zymosan stimulation of macrophages.
Peritonitis was induced in male BALB/c and C57BL/6J
mice via intraperitoneal injection of zymosan (40 mg/kg).
RAW 264.7 macrophages were activated with zymosan
(250 lg/ml). Riboflavin (mice, 50 mg/kg; RAW 264.7,
25 lg/ml) was administered 30 min before zymosan,
simultaneously with, or 2, 4, 6 h after zymosan. Addi-
tionally, mRNA expression of HMGB1 and its intracellular
and serum levels were evaluated. The research showed that
riboflavin significantly reduces both the expression and the
release of HMGB1; however, the effect of riboflavin was
time-dependent. The greatest efficacy was found when
riboflavin was given 30 min prior to zymosan, and also 2
and 4 h (C57BL/6J; RAW 264.7) or 4 and 6 h (BALB/c)
after zymosan. Research showed that riboflavin influences
the level of HMGB1 released in the course of inflamma-
tion; however, further study is necessary to determine its
mechanisms of action.
Keywords Macrophages Vitamin B2 HMGB1
Inflammation Sepsis Peritonitis
Studies on Immunogenicity and Antigenicity of Baculovirus-
Expressed Binding Region of Plasmodium falciparum EBA-140
Merozoite Ligand
Agata Zerka1 • Joanna Rydzak1 • Anna Lass2 • Beata Szostakowska2 •
Wacław Nahorski3 • Agnieszka Wroczyn´ ska3 • Przemyslaw Myjak2 •
Hubert Krotkiewski1 • Ewa Jaskiewicz1,4
Received: 31 March 2015 / Accepted: 31 August 2015 / Published online: 6 October 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract The erythrocyte binding ligand 140 (EBA-140)
is a member of the Plasmodium falciparum erythrocyte
binding antigens (EBA) family, which are considered as
prospective candidates for malaria vaccine development.
EBA proteins were identified as important targets for nat-
urally acquired inhibitory antibodies. Natural antibody
response against EBA-140 ligand was found in individuals
living in malaria-endemic areas. The EBA-140 ligand is a
paralogue of the well-characterized P. falciparum EBA-
175 protein. They both share homology of domain struc-
ture, including the binding region (Region II), which
consists of two homologous F1 and F2 domains and is
responsible for ligand–erythrocyte receptor interaction
during merozoite invasion. It was shown that the erythro-
cyte receptor for EBA-140 ligand is glycophorin C-a minor
human erythrocyte sialoglycoprotein. In studies on the
immunogenicity of P. falciparum EBA ligands, the
recombinant proteins are of great importance. In this
report, we have demonstrated that the recombinant bac-
ulovirus-obtained EBA-140 Region II is immunogenic and
antigenic. It can raise specific antibodies in rabbits, and it is
recognized by natural antibodies present in sera of patients
with malaria, and thus, it may be considered for inclusion
in multicomponent blood-stage vaccines.
Keywords Plasmodium falciparum
Recombinant binding region of EBA-140 ligand
Region II baculovirus expression Immunogenicity
Antigenicity Natural anti-EBA-140 IgG antibodies
Quinoline-Based Compound BPIQ Exerts Anti-Proliferative
Effects on Human Retinoblastoma Cells via Modulating
Intracellular Reactive Oxygen Species
Kai-Chun Cheng1,2,3 • Chun-Tzu Hung3,4 • Kuo-Jen Chen2 • Wen-Chuan Wu3,5 •
Jau-Ling Suen1 • Cheng-Hsien Chang1,2,3,5 • Chi-Yu Lu6 • Chih-Hua Tseng7 •
Yeh-Long Chen8 • Chien-Chih Chiu9,10,11
Received: 13 February 2015 / Accepted: 31 August 2015 / Published online: 12 November 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Retinoblastoma (Rb) is the most common pri-
mary intraocular malignant tumor of childhood. It is
important to develop the strategy for Rb treatment. We have
tested a quinolone derivative 2,9-bis[2-(pyrrolidin-1-yl)e-
thoxy]-6-{4-[2-(pyrrolidin-1-yl)ethoxy]phenyl}-11H-in-
deno[1,2-c]quinolin-11-one (BPIQ) for its anti-cancer
effects against Rb via cultured human Rb cell line Y79. Our
results showed that BPIQ significantly inhibits cell growth
of Y79. Furthermore, the flow cytometer-based assays and
Western blotting showed that BPIQ induces the apoptosis of
Y79 via increasing the level of reactive oxygen species
(ROS). Besides, the activation of cH2AX, a DNA damage
sensor in human Y79 cells was also observed, indicating the
potential of BPIQ for causing DNA damage of Rb cells. On
the contrary, BPIQ-induced apoptosis of Y79 cells was
attenuated significantly by N-acetyl-L-cysteine (NAC), an
ROS scavenger. The results of Western blot showed that
BPIQ down-regulates the levels of anti-apoptotic proteins
Bcl-2, survivin and XIAP while up-regulates the pro-
apoptotic proteins Bad, Bax and Bid. Our present study
demonstrated the anti-proliferative effect of BPIQ in human
Y79 cells. The inhibitory effect of BPIQ on the proliferation
of Y79 cells is, at least, partly mediated by the regulation of
ROS and DNA damage pathway. In conclusion, BPIQ may
provide an alternative option in the chemotherapeutics or
chemoprevention on the Rb therapy in the future.
Keywords BPIQ Quinoline Retinoblastoma
Reactive oxygen species DNA damage Apoptosis
The Molecular Influence of Graphene and Graphene Oxide
on the Immune System Under In Vitro and In Vivo Conditions
Ilona Dudek1 • Marta Skoda1 • Anna Jarosz1 • Dariusz Szukiewicz1
Received: 10 March 2015 / Accepted: 31 August 2015 / Published online: 26 October 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Graphene and graphene oxide (GO), due to
their physicochemical properties and biocompatibility, can
be used as an innovative biomedical material in biodetec-
tion, drug distribution in the body, treating neoplasms,
regenerative medicine, and in implant surgery. Research on
the biomedical use of graphene and GO that has been
carried out until now is very promising and shows that
carbon nanomaterials present high biocompatibility.
However, the intolerance of the immune system to gra-
phene nanomaterials, however low, may in consequence
make it impossible to use them in medicine. This paper
shows the specific mechanism of the molecular influence of
graphene and GO on macrophages and lymphocytes under
in vitro and in vivo conditions and their practical appli-
cation in medicine. Under in vitro conditions graphene and
GO cause an increased production of pro-inflammatory
cytokines, mainly IL-1, IL-6, IL-10 and TNF-a, as a result
of the activation of Toll-like receptors in macrophages.
Graphene activates apoptosis in macrophages through the
TGFbr/Smad/Bcl-2 pathway and also through JNK kinases
that are stimulated by an increase of ROS in the cell or
through a signal received by Smad proteins. Under in vivo
conditions, graphene nanomaterials induce the develop-
ment of the local inflammatory reaction and the
development of granulomas in parenchymal organs. How-
ever, there is a huge discrepancy between the results
obtained by different research groups, which requires a
detailed analysis. In this work we decided to collect and
analyze existing research and tried to explain the discrep-
ancies. Understanding the precise mechanism of how this
nanomaterial influences immune system cells allows esti-
mating the potential influence of grapheme and GO on the
human body.
Keywords Graphene Graphene oxide Immune cells
Inflammatory reaction
Antigen Cross-Presentation and Heat Shock Protein-Based
Vaccines
Katerina Zachova1 • Michal Krupka1 • Milan Raska1,2
Received: 28 January 2015 / Accepted: 31 August 2015 / Published online: 30 October 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Vaccines currently in the clinical use contain
adjuvants stimulating preferably Th2 type of immune
response associated with the production of specific anti-
bodies, mostly of neutralizing isotypes. This kind of
immune response is effective only against some types of
pathogens and has limited effect against tumors and many
viruses where parallel activation of antigen-specific
humoral and cell-mediated immunity is required. One of
the main objectives of the current vaccine research is the
development of approaches leading to the induction of
antigen-specific CD8? T cell response including cytotoxic
T lymphocyte (CTL). Induction of antigen-specific CD8?
T cell response to exogenously delivered antigen requires
their cross-presentation by antigen presenting cells, espe-
cially dendritic cells. The cross-presentation principles
seem to be crucial for effective activation of CTL. In this
paper, we discuss some approaches to employing heat
shock proteins for induction of antigen-specific CD8? T
cells in the context of cross-presentation and cross-priming
principles.
Keywords Cross-presentation
Heat shock protein (hsp) Dendritic cells
Fetal/Neonatal Alloimmune Thrombocytopenia: Pathogenesis,
Diagnostics and Prevention
Ewa Brojer1 • Anne Husebekk2 • Marzena De˛bska3 • Małgorzata Uhrynowska1 •
Katarzyna Guz1 • Agnieszka Orzin´ ska1 • Romuald De˛bski3 • Krystyna Mas´lanka1
Received: 12 March 2015 / Accepted: 31 August 2015 / Published online: 12 November 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract Fetal/neonatal alloimmune thrombocytopenia
(FNAIT) is a relatively rare condition (1/1000–1/2000) that
was granted orphan status by the European Medicines
Agency in 2011. Clinical consequences of FNAIT, how-
ever, may be severe. A thrombocytopenic fetus or new-
born is at risk of intracranial hemorrhage that may result in
lifelong disability or death. Preventing such bleeding is
thus vital and requires a solution. Anti-HPA1a antibodies
are the most frequent cause of FNAIT in Caucasians. Its
pathogenesis is similar to hemolytic disease of the newborn
(HDN) due to anti-RhD antibodies, but is characterized by
platelet destruction and is more often observed in the first
pregnancy. In 75 % of these women, alloimmunization by
HPA-1a antigens, however, occurs at delivery, which
enables development of antibody-mediated immune sup-
pression to prevent maternal immunization. As for HDN,
the recurrence rate of FNAIT is high. For advancing
diagnostic efforts and treatment, it is thereby crucial to
understand the pathogenesis of FNAIT, including cellular
immunity involvement. This review presents the current
knowledge on FNAIT. Also described is a program for
HPA-1a screening in identifying HPA-1a negative preg-
nant women at risk of immunization. This program is now
performed at the Institute of Hematology and Transfusion
Medicine in cooperation with the Department of Obstetrics
and Gynecology of the Medical Centre of Postgraduate
Education in Warsaw as well as the UiT The Arctic
University of Norway.
Keywords Human platelet alloantigens Pregnancy
Fetal/neonatal alloimmune thrombocytopenia
Therapeutic intervention
HLA-G and MHC Class II Protein Expression in Diffuse Large
B-Cell Lymphoma
Dorota Jesionek-Kupnicka1 • Marcin Bojo2 • Monika Prochorec-Sobieszek3 •
Anna Szumera-Ciec´kiewicz3 • Joanna Jabłon´ ska1 • Ewa Kalinka-Warzocha2 •
Radzisław Kordek1 • Wojciech Młynarski4 • Tadeusz Robak5 • Krzysztof Warzocha6 •
Ewa Lech-Maranda6,7
Received: 7 June 2015 / Accepted: 14 September 2015 / Published online: 14 December 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract The expression of human leukocyte antigen-G
(HLA-G) and HLA class II protein was studied by
immunohistochemical staining of lymph nodes from 148
patients with diffuse large B-cell lymphoma (DLBCL) and
related to the clinical course of the disease. Negative HLA-
G expression was associated with a lower probability of
achieving a complete remission (p = 0.04). Patients with
negative HLA-G expression tended towards a lower 3-year
overall survival (OS) rate compared to those with positive
expression of HLA-G (p = 0.08). When restricting the
analysis to patients receiving chemotherapy with ritux-
imab, the estimated 3-year OS rate of patients with positive
HLA-G expression was 73.3 % compared with 47.5 %
(p = 0.03) in those with negative expression. Patients with
negative HLA class II expression presented a lower 3-year
OS rate compared to subjects with positive expression
(p = 0.04). The loss of HLA class II expression (p = 0.05)
and belonging to the intermediate high/high IPI risk group
(p = 0.001) independently increased the risk of death.
HLA class II expression also retained its prognostic value
in patients receiving rituximab; the 3-year OS rate was
65.3 % in patients with positive HLA class II expression
versus 29.6 % (p = 0.04) in subjects that had loss of HLA
class II expression. To our knowledge, for the first time, the
expression of HLA-G protein in DLBCL and its association
with the clinical course of the disease was demonstrated.
Moreover, the link between losing HLA class II protein
expression and poor survival of patients treated with
immunochemotherapy was confirmed.
Keywords Human leukocyte antigen-G
Major histocompatibility class II Immunohistochemistry
Diffuse large B-cell lymphoma Outcome
Bacterial Infections and Osteoclastogenesis Regulators in Men
and Women with Cholesteatoma
Wirginia Likus1 • Krzysztof Siemianowicz2 • Jarosław Markowski3 •
Jan Wiaderkiewicz4,5 • Anna Kostrza˛b-Zdebel6 • Edyta Jura-Szołtys7 •
Włodzimierz Dziubdziela8 • Ryszard Wiaderkiewicz9 • Marek J. Łos10,11
Received: 1 May 2015 / Accepted: 12 October 2015 / Published online: 19 November 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract One of the most distinct features of middle ear
cholesteatoma is bone destruction. Aetiology of choles-
teatoma is thought to be multifactorial. Endotoxins
produced by bacteria are thought to initiate the inflamma-
tion process in the middle ear leading to cholesteatoma.
There are physiological differences in bone metabolism
between men and women. The aim of our study was the
immunohistochemical evaluation of the contents of two
key components of the OPG/RANK/RANKL triad—
RANKL and OPG in cholesteatoma, to analyse if there are
any differences between the sexes and to evaluate the
bacteria species isolated from cholesteatoma just before
surgical treatment and to evaluate their plausible influence
on the expression of OPG and RANKL in cholesteatoma.
Twenty-one adult patients with acquired cholesteatoma
who underwent surgery were analysed. There were no
statistically significant differences in the expression of both
regulators of osteoclastogenesis between the sexes. In
38.1 % patients cholesteatoma was not infected, whereas in
61.9 % patients various bacterial infections or mycosis
were found. The most frequently isolated species was
Pseudomonas aeruginosa (14.29 % infections) followed by
Staphylococcus aureus (9.52 % infections). There were no
statistically significant differences in expression of both
OPG and RANKL between uninfected and infected
cholesteatomas.
Keywords Cholesteatoma Osteoprotegerin RANKL
Bacterial infection Sexes
Erratum to: Studies on Immunogenicity and Antigenicity
of Baculovirus-Expressed Binding Region of Plasmodium
falciparum EBA-140 Merozoite Ligand
Agata Zerka1 • Joanna Rydzak1 • Anna Lass2 • Beata Szostakowska2 •
Wacław Nahorski3 • Agnieszka Wroczyn´ ska3 • Przemyslaw Myjak2 •
Hubert Krotkiewski1 • Ewa Jaskiewicz1,4
Published online: 19 November 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Erratum to: Arch Immunol Ther Exp
DOI 10.1007/s00005-015-0367-5
The author would like to amend the Acknowledgment
section of his online published article. The complete
Acknowledgment section should read as follows:
The research was supported by Grants No. N N302 281436
from the Ministry of Science and Higher Education of
Poland and Grant No. 2012/05/N/NZ6/00667 from the
National Centre of Science. The publication was supported
by Wroclaw Centre of Biotechnology, programme The
Leading National Research Centre (KNOW) for the years
2014–2018.
Matrix Metalloproteinases and Their Inhibitors in Chronic
Obstructive Pulmonary Disease
Zdenka Navratilova1 • Vitezslav Kolek2 • Martin Petrek1
Received: 19 December 2014 / Accepted: 25 September 2015 / Published online: 26 November 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Chronic obstructive pulmonary disease
(COPD) is characterised by irreversible airflow limitation
associated with chronic inflammation. Matrix metallopro-
teinases (MMPs) are proteolytic enzymes that contribute to
the inflammatory response in COPD and degrade extra-
cellular matrix components. Their enzymatic activity is
inhibited by a four-member family of tissue inhibitors of
metalloproteinases (TIMPs). In COPD, the MMP/TIMP
network, mainly MMP-9, has been repeatedly observed to
be dysregulated at both the local (lung) and systemic
levels. Here, we review the findings reported in numerous
cross-sectional studies with our primary focus on longitu-
dinal observations in human COPD studies. The data from
longitudinal prospective studies on the MMP/TIMP net-
work may lead to the introduction of novel prognostic
biomarkers into clinical management of COPD. We
address the relationship between the systemic and local
lung MMP/TIMP network in COPD patients and briefly
describe the involvement of microRNAs. Finally, the role
of the MMP/TIMP network in COPD treatment is
discussed.
Keywords Chronic obstructive pulmonary disease
Matrix metalloproteinases
Tissue inhibitors of metalloproteinases MicroRNAs
Local and systemic response Biomarker
Neurotrophic Factors and Their Potential Applications in Tissue
Regeneration
Nan Xiao1 • Quynh-Thu Le2
Received: 21 September 2015 / Accepted: 2 November 2015 / Published online: 26 November 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Neurotrophic factors are growth factors that
can nourish neurons and promote neuron survival and
regeneration. They have been studied as potential drug
candidates for treating neurodegenerative diseases. Since
their identification, there are more and more evidences to
indicate that neurotrophic factors are also expressed in non-
neuronal tissues and regulate the survival, anti-inflamma-
tion, proliferation and differentiation in these tissues. This
mini review summarizes the characteristics of the neu-
rotrophic factors and their potential clinical applications in
the regeneration of neuronal and non-neuronal tissues.
Keywords Neurotrophic factor Tissue regeneration
Inflammaging and Anti-Inflammaging: The Role of Cytokines
in Extreme Longevity
Paola Lucia Minciullo1 • Antonino Catalano2 • Giuseppe Mandraffino3 •
Marco Casciaro1 • Andrea Crucitti2 • Giuseppe Maltese4 • Nunziata Morabito2 •
Antonino Lasco2 • Sebastiano Gangemi1 • Giorgio Basile2
Received: 16 June 2015 / Accepted: 2 November 2015 / Published online: 12 December 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Longevity and aging are two sides of the same
coin, as they both derive from the interaction between
genetic and environmental factors. Aging is a complex,
dynamic biological process characterized by continuous
remodeling. One of the most recent theories on aging focuses
on immune response, and takes into consideration the acti-
vation of subclinical, chronic low-grade inflammation which
occurs with aging, named ‘‘inflammaging’’. Long-lived
people, especially centenarians, seem to cope with chronic
subclinical inflammation through an anti-inflammatory
response, called therefore ‘‘anti-inflammaging’’. In the pre-
sent review, we have focused our attention on the contrast
between inflammaging and anti-inflammaging systems, by
evaluating the role of cytokines and their impact on extreme
longevity. Cytokines are the expression of a network
involving genes, polymorphisms and environment, and are
involved both in inflammation and anti-inflammation. We
have described the role of IL-1, IL-2, IL-6, IL-12, IL-15, IL-
18, IL-22, IL-23, TNF-a, IFN-c as pro-inflammatory
cytokines, of IL-1Ra, IL-4, IL-10, TGF-b1 as anti-inflam-
matory cytokines, and of lipoxin A4 and heat shock proteins
as mediators of cytokines. We believe that if inflammaging is
a key to understand aging, anti-inflammaging may be one of
the secrets of longevity.
Keywords Inflammaging Anti-inflammaging
Cytokine Aging Longevity
Search for the Function of NWC, Third Gene Within RAG Locus:
Generation and Characterization of NWC-Deficient Mice
Monika Kasztura1 • Lukasz Sniezewski1 • Agnieszka Laszkiewicz1 •
Michal Majkowski1 • Kamil Kobak1 • Karolina Peczek1 • Sylwia Janik1 •
Violetta Kapusniak2 • Arkadiusz Miazek3 • Malgorzata Cebrat1 • Pawel Kisielow1,3
Received: 3 June 2015 / Accepted: 2 November 2015 / Published online: 24 December 2015
Ó The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract NWC is a third gene within recombination
activating gene (RAG) locus, which unlike RAG genes is
ubiquitously expressed and encodes a unique protein con-
taining three strongly evolutionarily conserved domains
not found in any other known protein. To get insight into
its function we identified several proteins co-immunopre-
cipitating with NWC protein and generated new NWC-
deficient mice. Here, we present evidence that unlike many
other ubiquitously expressed evolutionarily conserved
proteins, functional inactivation of NWC does not cause
any gross developmental, physiological or reproductive
abnormalities and that under physiological conditions
NWC may be involved in assembling and functioning of
cilia, cell surface organelles found on nearly every
eukaryotic cell.
Keywords Evolutionarily conserved genes
NWC-knockout mice NWC protein function/partners
Recombination activating gene (RAG) locus Cilia
Immunomodulatory Effects of Different Cellular Therapies
of Bone Marrow Origin on Chimerism Induction
and Maintenance Across MHC Barriers in a Face
Allotransplantation Model
Mikael Hivelin1 • Aleksandra Klimczak1,2 • Joanna Cwykiel1,3 • Erhan Sonmez1 •
Serdar Nasir1 • James Gatherwright1 • Maria Siemionow1,3
Received: 5 May 2015 / Accepted: 5 October 2015 / Published online: 26 December 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Many more patients would benefit from vas-
cularized composite allotransplantation if less toxic and
safer immunosuppressive protocols will become available.
Tolerance induction protocols with donor cells co-trans-
plantation are one of the promising pathways to reduce
maintenance immunosupressive regimens. We investigated
the role of donor bone marrow cells (BMC), mesenchymal
stromal cells (MSC) and in vivo created chimeric cells
(CC) used as supportive therapies in a fully MHC-mis-
matched rat face transplantation model. Twenty-four fully
MHC-mismatched hemiface transplantations were per-
formed between ACI (RT1a) donors and Lewis (RT1l)
recipients under combined seven-day immunosuppressive
regimen of anti-ab-T-cell receptor (TCR) monoclonal
antibody and cyclosporin A. We studied four experimental
groups—group 1: no cellular therapy; group 2: supportive
therapy with BMC; group 3: supportive therapy with MSC;
group 4: supportive therapy with CC generated in a pri-
mary chimera. We evaluated clinical and histological
rejection grades, transplanted cells migration, donor-
specific chimerism in the peripheral blood and bone mar-
row compartments, and CD4?/CD25? T-cell levels. Face
allograft rejection was observed at 26.8 ± 0.6 days post-
transplant (PT) in the absence of cellular therapy, at
34.5 ± 1.1 days for group 2, 29.3 ± 0.8 days for group 3,
and 30.3 ± 1.38 PT for group 4. The longest survival was
observed in allografts supported by co-transplantation of
BMC. All support in cellular therapies delayed face allo-
graft rejection by chimerism induction and/or
immunomodulatory properties of co-transplanted cells.
Survival time was comparable between groups, however,
further studies, with different cell dosages, delivery routes
and delivery times are required.
Keywords Vascularized composite allograft/
allotransplantation Face transplantation
Chimerism and tolerance Bone marrow
Mesenchymal stromal cells Chimeric cells
From the Deep Sea to Everywhere: Environmental Antigens
for iNKT Cells
Gerhard Wingender1
Received: 18 August 2015 / Accepted: 2 November 2015 / Published online: 24 December 2015
Ó L. Hirszfeld Institute of Immunology and Experimental Therapy, Wroclaw, Poland 2015
Abstract Invariant natural killer T (iNKT) cells are a
unique subset of innate T cells that share features with
innate NK cells and adaptive memory T cells. The first
iNKT cell antigen described was found 1993 in a marine
sponge and it took over 10 years for other, bacterial anti-
gens to be described. Given the paucity of known bacterial
iNKT cell antigens, it appeared as if iNKT cells play a very
specialist role in the protection against few, rare and unu-
sual pathogenic bacteria. However, in the last few years
several publications painted a very different picture, sug-
gesting that antigens for iNKT cells are found almost
ubiquitous in the environment. These environmental iNKT
cell antigens can shape the distribution, phenotype and
function of iNKT cells. Here, these recent findings will be
reviewed and their implications for the field will be
outlined.
Keywords Innate T cells
Invariant natural killer T (iNKT) cells
Environmental antigens Mucosal immunology
Novel Insights into the Roles of Rho Kinase in Cancer
Lei Wei1,2 • Michelle Surma1 • Stephanie Shi1 • Nathan Lambert-Cheatham1 •
Jianjian Shi1
Received: 12 July 2015 / Accepted: 24 November 2015 / Published online: 2 January 2016
Ó The Author(s) 2016. This article is published with open access at Springerlink.com
Abstract Rho-associated coiled-coil kinase (ROCK) is a
major downstream effector of the small GTPase RhoA. The
ROCK family, consisting of ROCK1 and ROCK2, plays a
central role in the organization of the actin cytoskeleton,
and is involved in a wide range of fundamental cellular
functions such as contraction, adhesion, migration, prolif-
eration, and apoptosis. Since the discovery of effective
inhibitors such as fasudil and Y27632, the biological roles
of ROCK have been extensively explored in numerous
diseases, including cancer. Accumulating evidence sup-
ports the concept that ROCK plays important roles in
tumor development and progression through regulating
many key cellular functions associated with malignancy,
including tumorigenicity, tumor growth, metastasis,
angiogenesis, tumor cell apoptosis/survival and chemore-
sistance as well. This review focuses on the new advances
of the most recent 5 years from the studies on the roles of
ROCK in cancer development and progression; the dis-
cussion is mainly focused on the potential value of ROCK
inhibitors in cancer therapy.
Keywords Rho kinase Rho kinase pan-inhibitor
ROCK isoform-specific inhibitor Cancer therapy