Erna Sziksz

Fibrosis Related Inflammatory Mediators: Role of the IL-10 Cytokine Family

Importance of chronic fibroproliferative diseases (FDs) including pulmonary fibrosis, chronic kidney diseases, inflammatory bowel disease, and cardiovascular or liver fibrosis is rapidly increasing and they have become a major public health problem. According to some estimates about 45% of all deaths are attributed to FDs in the developed world. Independently of their etiology the common hallmark of FDs is chronic inflammation. Infiltrating immune cells, endothelial, epithelial, and other resident cells of the injured organ release an orchestra of inflammatory mediators, which stimulate the proliferation and excessive extracellular matrix (ECM) production of myofibroblasts, the effector cells of organ fibrosis. Abnormal amount of ECM disturbs the original organ architecture leading to the decline of function. Although our knowledge is rapidly expanding, we still have neither a diagnostic tool to detect nor a drug to specifically target fibrosis. Therefore, there is an urgent need for the more comprehensive understanding of the pathomechanism of fibrosis and development of novel diagnostic and therapeutic strategies. In the present review we provide an overview of the common key mediators of organ fibrosis highlighting the role of interleukin-10 (IL-10) cytokine family members (IL-10, IL-19, IL-20, IL-22, IL-24, and IL-26), which recently came into focus as tissue remodeling-related inflammatory cytokines.

Increased expression of hypoxia-inducible factor 1alpha in coeliac disease.

Previously, it has been suggested that hypoxia-inducible factor (HIF) 1 signaling may play determinative role in the maintenance of the barrier function of the intestinal epithelium in inflammatory bowel disease. Our aim was to depict the alteration of HIF-1alpha and related genes in celiac disease (CD) where the importance of the barrier function is well known. Duodenal biopsy specimens were collected from 16 children with untreated CD, 9 children with treated CD and 10 controls. HIF-1alpha, trefoil factor 1 (TFF1), ecto-5-prime nucleotidase (CD73), and multi drug resistance gene 1 (MDR1) mRNA and HIF-1alpha protein expression were determined by real-time PCR and Western blot, respectively. Localization of HIF-1alpha was determined by immunofluorescent staining. We found increased HIF-1alpha and TFF1 mRNA and HIF-1alpha protein expression in the duodenal mucosa of children with untreated CD compared with controls or children with treated CD (p < 0.05). In untreated CD children, HIF-1alpha staining was present in cytoplasmic and nuclear region of the villous enterocytes. In treated CD mRNA expression of CD73 and MDR1 were increased compared with controls (p < 0.01 and 0.05, respectively). Our results of increased mucosal HIF-1alpha expression in CD children suggest the contribution of this signaling pathway in the pathomechanism of CD.

Role of Altered Expression of miR-146a, miR-155, and miR-122 in Pediatric Patients with Inflammatory Bowel Disease.

Background:Evidence suggests the central role of tumor necrosis factor (TNF)-&agr; in the pathomechanism of inflammatory bowel disease (IBD); however, its effect on epigenetic factors, including small non-coding microRNAs (miRs), is less known. Our present aim was the comparative investigation of the expression of TNF-&agr; and immune response–related miRs in children with Crohns disease (CD) and ulcerative colitis (UC). Methods:Fresh-frozen (FF) and formalin-fixed, paraffin-embedded (FFPE) biopsies were used to analyze the expression of miR-146a, -155, -122, and TNF-&agr; by real-time reverse transcription polymerase chain reaction in macroscopically inflamed (CD: 12 FFPE and 24 FF; UC: 10 FF) and intact (CD: 12 FFPE; 14 FF) colonic biopsies of children with IBD and controls (16 FFPE; 23 FF). The expression of miR-146a, -155, and -122 was also determined in TNF-&agr;–treated HT-29 colonic epithelial cells. Results:Increased expression of TNF-&agr; was observed in the colonic mucosa of children with CD and UC in comparison with controls. Expression of miR-146a and -155 was higher in the inflamed mucosa of children with CD and UC than in the intact mucosa. Expression of miR-122 elevated in the macroscopically intact colonic regions of CD compared with controls and patients with UC. In HT-29 cells, TNF-&agr; treatment increased the expression of miR-146a and -155, but not that of miR-122. Conclusions:Our results showed altered expression of miR-146a, -155, and -122 in the colonic mucosa of children with IBD and in TNF-&agr;–treated colonic epithelial cells. Our data suggest the TNF-&agr;–related involvement of these miRs in the pathogenesis of IBD.

Increased Heat Shock Protein 72 Expression in Celiac Disease

Background and Objectives: Heat shock protein (HSP) 72, a known chaperone, has potential epithelial barrier protecting, antiapoptotic, and immune system regulatory effects; therefore, our aim was to study its involvement in the pathology of celiac disease (CD). Patients and Methods: Duodenal biopsy specimens were collected from children with untreated and treated CD and from controls. mRNA expression, protein level, and localization of HSP72 were determined. Results: Elevated HSP72 mRNA expression and higher protein levels were found in the duodenal mucosa of children with untreated CD as well as in children with treated CD compared with those in controls. In the duodenal mucosa of children with treated CD, HSP72 mRNA expression was decreased and HSP72 protein levels were lower than those in children with untreated CD. We detected intensive HSP72 staining in the villous enterocytes and immune cells of the lamina propria in the duodenal villi of children with untreated CD compared with that in controls. Conclusions: The increased expression and altered localization of HSP72 in CD indicate that HSP72 should have a role in protection against gliadin-induced cytotoxicity. HSP72 may exert antiapoptotic effect and contribute to preservation of intestinal epithelial barrier integrity. Moreover, HSP72 as a ligand of TLR2 and TLR4 may promote innate immune responses and warn the cells of the potential injury.

Increased Expression of Hypoxia-Inducible Factor 1α in Coeliac Disease

Previously, it has been suggested that hypoxia-inducible factor (HIF) 1 signaling may play determinative role in the maintenance of the barrier function of the intestinal epithelium in inflammatory bowel disease. Our aim was to depict the alteration of HIF-1α and related genes in celiac disease (CD) where the importance of the barrier function is well known. Duodenal biopsy specimens were collected from 16 children with untreated CD, 9 children with treated CD and 10 controls. HIF-1α, trefoil factor 1 (TFF1), ecto-5-prime nucleotidase (CD73), and multi drug resistance gene 1 (MDR1) mRNA and HIF-1α protein expression were determined by real-time PCR and Western blot, respectively. Localization of HIF-1α was determined by immunofluorescent staining. We found increased HIF-1α and TFF1 mRNA and HIF-1α protein expression in the duodenal mucosa of children with untreated CD compared with controls or children with treated CD (p < 0.05). In untreated CD children, HIF-1α staining was present in cytoplasmic and nuclear region of the villous enterocytes. In treated CD mRNA expression of CD73 and MDR1 were increased compared with controls (p < 0.01 and 0.05, respectively). Our results of increased mucosal HIF-1α expression in CD children suggest the contribution of this signaling pathway in the pathomechanism of CD.

Th17 cells in rheumatoid arthritis

Th17 cells are the newly described subset of the CD4(+) T lymphocytes. Activated Th17 cells are characterized by their ability to produce IL-17A and other pro-inflammatory cytokines. IL-17A regulates immune function through its cell-surface receptor expressed on epithelial-and endothelial cells, fibroblasts and leukocytes by promoting neutrophil recruitment and releasing further pro-inflammatory mediators. Failures of the susceptible balance of the immunoregulation may lead to unchecked immune response and autoimmune diseases. The central role of Th17 cells and cytokines produced by Th17 cells were confirmed in a wide variety of human autoimmune diseases, including rheumatoid arthritis. Recently Th17 cells and its cytokines come into the focus of immunological research as potential therapeutic targets.A T helper 17 (Th17) populáció a CD4+ T-lymphocyták újonnan felfedezett csoportja. Az aktivált Th17 sejtek az őket leginkább jellemző interleukin (IL) -17A mellett számos más proinfl ammatorikus citokint is termelnek. Az IL-17A epithelés endothelsejteken, fi broblastokon és a leukocytákon található sejtfelszíni receptorán keresztül további gyulladásos mediátorok felszabadításával és neutrophil granulocyták aktiválásával vesz részt az immunválasz szabályozásában. Az immunreguláció érzékeny egyensúlyának megbomlása gyulladásos és autoimmun betegségekhez vezet. A Th17 sejtek és az általuk termelt citokinek kiemelt szerepét számos humán autoimmun kórképben, köztük a rheumatoid arthritisben is igazolták. Terápiás célpontként való felhasználásuk napjaink immunológiai kutatásainak egyik ígéretes területe.

Dehydroepiandrosterone Pretreatment Alters the Ischaemia/Reperfusion-Induced VEGF, IL-1 and IL-6 Gene Expression in Acute Renal Failure

Background: Beneficial effects of dehydroepiandrosterone (DHEA) pretreatment were reported in ischaemia/reperfusion (I/R)-induced kidney damage. Methods: To investigate the mechanism of DHEA pretreatment during renal I/R injury, the left renal pedicles of DHEA- [G_DHEA; 4.0 mg/kg/day DHEA dissolved in propylene glycol (PG)] and PG-pretreated male Wistar rats (G_PG) were clamped for 55 min followed by 2 (T₂) and 24 h (T₂₄) of reperfusion. Sham-operated, non-clamped animals (T₀) served as controls in both groups. Renal function, kidney morphology and interleukin 1β (IL-1β), interleukin 6 (IL-6) and vascular endothelial growth factor (VEGF) expression were determined in the kidneys of both groups. Results: Renal functional parameters and kidney structure did not differ in G_DHEA versus G_PG at any time point. Renal mRNA expression of IL-1β was lower at T₀, while IL-6 at T₂ was lower in G_DHEA than in G_PG.While renal VEGF mRNA expression remained unchanged, protein levels were increased at T₂ and T₂₄compared to T₀ kidneys in both groups. VEGF protein levels were lower at T₂ and T₂₄in G_DHEA than in G_PG. Conclusion: We found that DHEA pretreatment alters renal IL-1β, IL-6 and VEGF synthesis. Moreover, contrary changes in VEGF mRNA and protein levels suggest that VEGF synthesis – distinct from other organs – might be primarily posttranscriptionally regulated in postischaemic rat kidneys.

Altered mucosal expression of microRNAs in pediatric patients with inflammatory bowel disease

INTRODUCTION MicroRNAs (miRs) came recently into focus as promising novel research targets offering new insights into the pathogenesis of inflammatory bowel diseases (IBD). AIMS The aim of our study was to identify a pediatric IBD (pIBD) characteristic miR profile serving as potential Crohns disease (CD) and ulcerative colitis (UC) specific diagnostic pattern and to further analyze the related target genes. METHODS Small RNA sequencing was performed on inflamed and intact colonic biopsies of CD, and control patients. Selected miRs were further investigated by RT-PCR, complemented with an UC group, in order to address the differential diagnostic potential of miRs in the two IBD subtypes. To analyze network connection of differentially expressed miRs and their target genes MiRTarBase database and previous transcriptome sequencing data from pediatric patient groups were used. RESULTS Sequencing analysis identified 170 miRs with altered expression. RT-PCR analysis revealed altered expression of miR-31, -125a, -142-3p, and -146a discriminating between the inflamed mucosa of CD and UC. In the intact mucosa of CD patients the expression of miR-18a, -20a, -21, -31, -99a, -99b, -100, -125a, -126, -142-5p, -146a, -185, -204, -221, and -223 was elevated compared to the controls. The expression of miR-20a, -204 and -221 was elevated exclusively in the intact region of CD patients compared to the controls. Enrichment analysis identified main IBD-related functional groups. CONCLUSIONS We demonstrated a characteristic colonic miR pattern in pIBD that could facilitate deeper understanding of the pathomechanism of IBD and may serve as a diagnostic tool.

Involvement of heat shock proteins in gluten-sensitive enteropathy.

Gluten-sensitive enteropathy, also known as coeliac disease (CD), is an autoimmune disorder occurring in genetically susceptible individuals that damages the small intestine and interferes with the absorption of other nutrients. As it is triggered by dietary gluten and related prolamins present in wheat, rye and barley, the accepted treatment for CD is a strict gluten-free diet. However, a complete exclusion of gluten-containing cereals from the diet is often difficult, and new therapeutic strategies are urgently needed. A class of proteins that have already emerged as drug targets for other autoimmune diseases are the heat shock proteins (HSPs), which are highly conserved stress-induced chaperones that protect cells against harmful extracellular factors. HSPs are expressed in several tissues, including the gastrointestinal tract, and their levels are significantly increased under stress circumstances. HSPs exert immunomodulatory effects, and also play a crucial role in the maintenance of epithelial cell structure and function, as they are responsible for adequate protein folding, influence the degradation of proteins and cell repair processes after damage, and modulate cell signalling, cell proliferation and apoptosis. The present review discusses the involvement of HSPs in the pathophysiology of CD. Furthermore, HSPs may represent a useful therapeutic target for the treatment of CD due to the cytoprotective, immunomodulatory, and anti-apoptotic effects in the intestinal mucosal barrier.

Microarray Analysis Reveals Increased Expression of Matrix Metalloproteases and Cytokines of Interleukin-20 Subfamily in the Kidneys of Neonate Rats Underwent Unilateral Ureteral Obstruction: A Potential Role of IL-24 in the Regulation of Inflammation and Tissue Remodeling

Background/Aims: Congenital obstructive nephropathy (CON) is the main cause of pediatric chronic kidney diseases leading to renal fibrosis. High morbidity and limited treatment opportunities of CON urge the better understanding of the underlying molecular mechanisms. Methods: To identify the differentially expressed genes, microarray analysis was performed on the kidney samples of neonatal rats underwent unilateral ureteral obstruction (UUO). Microarray results were then validated by real-time RT-PCR and bioinformatics analysis was carried out to identify the relevant genes, functional groups and pathways involved in the pathomechanism of CON. Renal expression of matrix metalloproteinase (MMP)-12 and interleukin (IL)-24 were evaluated by real-time RT-PCR, flow cytometry and immunohistochemical analysis. Effect of the main profibrotic factors on the expression of MMP-12 and IL-24 was investigated on HK-2 and HEK-293 cell lines. Finally, the effect of IL-24 treatment on the expression of pro-inflammatory cytokines and MMPs were tested in vitro. Results: Microarray analysis revealed 880 transcripts showing >2.0-fold change following UUO, enriched mainly in immune response related processes. The most up-regulated genes were MMPs and members of IL-20 cytokine subfamily, including MMP-3, MMP-7, MMP-12, IL-19 and IL-24. We found that while TGF-β treatment inhibits the expression of MMP-12 and IL-24, H2O2 or PDGF-B treatment induce the epithelial expression of MMP-12. We demonstrated that IL-24 treatment decreases the expression of IL-6 and MMP-3 in the renal epithelial cells. Conclusions: This study provides an extensive view of UUO induced changes in the gene expression profile of the developing kidney and describes novel molecules, which may play significant role in the pathomechanism of CON.