Extracellular Vesicles of Patients on Peritoneal Dialysis Inhibit the TGF-ß- and PDGF-B-Mediated Fibrotic Processes.

Among patients on peritoneal dialysis (PD), 50-80% will develop peritoneal fibrosis, and 0.5-4.4% will develop life-threatening encapsulating peritoneal sclerosis (EPS). Here, we investigated the role of extracellular vesicles (EVs) on the TGF-ß- and PDGF-B-driven processes of peritoneal fibrosis. EVs were isolated from the peritoneal dialysis effluent (PDE) of children receiving continuous ambulatory PD. The impact of PDE-EVs on the epithelial-mesenchymal transition (EMT) and collagen production of the peritoneal mesothelial cells and fibroblasts were investigated in vitro and in vivo in the chlorhexidine digluconate (CG)-induced mice model of peritoneal fibrosis. PDE-EVs showed spherical morphology in the 100 nm size range, and their spectral features, CD63, and annexin positivity were characteristic of EVs. PDE-EVs penetrated into the peritoneal mesothelial cells and fibroblasts and reduced their PDE- or PDGF-B-induced proliferation. Furthermore, PDE-EVs inhibited the PDE- or TGF-ß-induced EMT and collagen production of the investigated cell types. PDE-EVs contributed to the mesothelial layer integrity and decreased the submesothelial thickening of CG-treated mice. We demonstrated that PDE-EVs significantly inhibit the PDGF-B- or TGF-ß-induced fibrotic processes in vitro and in vivo, suggesting that EVs may contribute to new therapeutic strategies to treat peritoneal fibrosis and other fibroproliferative diseases.

Optimization of Sirius Red-Based Microplate Assay to Investigate Collagen Production In Vitro.

Tissue fibrosis is characterized by chronic fibroblast activation and consequently excessive accumulation of collagen-rich extracellular matrix. In vitro microplate-based assays are essential to investigate the underlying mechanism and the effect of antifibrotic drugs. In this study, in the absence of a gold-standard method, we optimized a simple, cost-effective, Sirius Red-based colorimetric measurement to determine the collagen production of fibroblasts grown on 96-well tissue culture plates. Based on our findings, the use of a serum-free medium is recommended to avoid aspecific signals, while ascorbate supplementation increases the collagen production of fibroblasts. The cell-associated collagens can be quantified by Sirius Red staining in acidic conditions followed by alkaline elution. Immature collagens can be precipitated from the culture medium by acidic Sirius Red solution, and after subsequent centrifugation and washing steps, their amount can be also measured. Increased attention has been paid to optimizing the assay procedure, including incubation time, temperature, and solution concentrations. The resulting assay shows high linearity and sensitivity and could serve as a useful tool in fibrosis-related basic research as well as in preclinical drug screening.

High Salt Promotes Inflammatory and Fibrotic Response in Peritoneal Cells.

Recent studies draw attention to how excessive salt (NaCl) intake induces fibrotic alterations in the peritoneum through sodium accumulation and osmotic events. The aim of our study was to better understand the underlying mechanisms. The effects of additional NaCl were investigated on human primary mesothelial cells (HPMC), human primary peritoneal fibroblasts (HPF), endothelial cells (HUVEC), immune cells (PBMC), as well as ex vivo on peritoneal tissue samples. Our results showed that a high-salt environment and the consequently increased osmolarity increase the production of inflammatory cytokines, profibrotic growth factors, and components of the renin-angiotensin-aldosterone system, including IL1B, IL6, MCP1, TGFB1, PDGFB, CTGF, Renin and Ace both in vitro and ex vivo. We also demonstrated that high salt induces mesenchymal transition by decreasing the expression of epithelial marker CDH1 and increasing the expression of mesenchymal marker ACTA2 and SNAIL1 in HPMCs, HUVECs and peritoneal samples. Furthermore, high salt increased extracellular matrix production in HPFs. We demonstrated that excess Na+ and the consequently increased osmolarity induce a comprehensive profibrotic response in the peritoneal cells, thereby facilitating the development of peritoneal fibrosis.

Zonulin as a Potential Therapeutic Target in Microbiota-Gut-Brain Axis Disorders: Encouraging Results and Emerging Questions.

The relationship between dysbiosis and central nervous diseases has been proved in the last 10 years. Microbial alterations cause increased intestinal permeability, and the penetration of bacterial fragment and toxins induces local and systemic inflammatory processes, affecting distant organs, including the brain. Therefore, the integrity of the intestinal epithelial barrier plays a central role in the microbiota-gut-brain axis. In this review, we discuss recent findings on zonulin, an important tight junction regulator of intestinal epithelial cells, which is assumed to play a key role in maintaining of the blood-brain barrier function. In addition to focusing on the effect of microbiome on intestinal zonulin release, we also summarize potential pharmaceutical approaches to modulate zonulin-associated pathways with larazotide acetate and other zonulin receptor agonists or antagonists. The present review also addresses the emerging issues, including the use of misleading nomenclature or the unsolved questions about the exact protein sequence of zonulin.

PARK7/DJ-1 as a Therapeutic Target in Gut-Brain Axis Diseases.

It is increasingly known that Parkinson's (PD) and Alzheimer's (AD) diseases occur more frequently in patients with inflammatory gastrointestinal diseases including inflammatory bowel (IBD) or celiac disease, indicating a pathological link between them. Although epidemiological observations suggest the existence of the gut-brain axis (GBA) involving systemic inflammatory and neural pathways, little is known about the exact molecular mechanisms. Parkinson's disease 7 (PARK7/DJ-1) is a multifunctional protein whose protective role has been widely demonstrated in neurodegenerative diseases, including PD, AD, or ischemic stroke. Recent studies also revealed the importance of PARK7/DJ-1 in the maintenance of the gut microbiome and also in the regulation of intestinal inflammation. All these findings suggest that PARK7/DJ-1 may be a link and also a potential therapeutic target in gut and brain diseases. In this review, therefore, we discuss our current knowledge about PARK7/DJ-1 in the context of GBA diseases.

Urocortin stimulates ERK1/2 phosphorylation and proliferation but reduces ATP production of MCF7 breast cancer cells.

Urocortins are members of the stress-related corticotropin-releasing factor family. Small amounts of them are present in the circulation and they are produced locally in various tissues of higher vertebrates. Aside from regulating circulation, or food uptake they also influence, via auto- and paracrine mechanisms, cell proliferation. In the present study we investigated in MCF7 human breast cancer cells the effect of urocortin onto mitogenic signaling via ERK1/2. Our results revealed that already 10 nM urocortin could stimulate the phosphorylation of these kinases and cell proliferation of MCF7 cells while ATP production was reduced when kept in the presence of the peptide up to two days. We examined the expression and contribution of the specific receptors of urocortin to the activation of ERK1/2 and to cell proliferation, the intracellular distribution of phosphorylated ERK1/2, and the involvement of additional proteins like PKA, PKB/Akt, MEK, p53, Rb and E2F-1 behind the observed phenomena.

Transient Agarose Spot (TAS) Assay: A New Method to Investigate Cell Migration.

Fibroblasts play a central role in diseases associated with excessive deposition of extracellular matrix (ECM), including idiopathic pulmonary fibrosis. Investigation of different properties of fibroblasts, such as migration, proliferation, and collagen-rich ECM production is unavoidable both in basic research and in the development of antifibrotic drugs. In the present study we developed a cost-effective, 96-well plate-based method to examine the migration of fibroblasts, as an alternative approach to the gold standard scratch assay, which has numerous limitations. This article presents a detailed description of our transient agarose spot (TAS) assay, with instructions for its routine application. Advantages of combined use of different functional assays for fibroblast activation in drug development are also discussed by examining the effect of nintedanib-an FDA approved drug against IPF-on lung fibroblasts.

High salt diet impairs dermal tissue remodeling in a mouse model of IMQ induced dermatitis.

Recent animal studies, as well as quantitative sodium MRI observations on humans demonstrated that remarkable amounts of sodium can be stored in the skin. It is also known that excess sodium in the tissues leads to inflammation in various organs, but its role in dermal pathophysiology has not been elucidated. Therefore, our aim was to study the effect of dietary salt loading on inflammatory process and related extracellular matrix (ECM) remodeling in the skin. To investigate the effect of high salt consumption on inflammation and ECM production in the skin mice were kept on normal (NSD) or high salt (HSD) diet and then dermatitis was induced with imiquimod (IMQ) treatment. The effect of high salt concentration on dermal fibroblasts (DF) and peripheral blood mononuclear cells (PBMC) was also investigated in vitro. The HSD resulted in increased sodium content in the skin of mice. Inflammatory cytokine Il17 expression was elevated in the skin of HSD mice. Expression of anti-inflammatory Il10 and Il13 decreased in the skin of HSD or HSD IMQ mice. The fibroblast marker Acta2 and ECM component Fn and Col1a1 decreased in HSD IMQ mice. Expression of ECM remodeling related Pdgfb and activation phosphorylated (p)-SMAD2/3 was lower in HSD IMQ mice. In PBMCs, production of IL10, IL13 and PDGFB was reduced due to high salt loading. In cultured DFs high salt concentration resulted in decreased cell motility and ECM production, as well. Our results demonstrate that high dietary salt intake is associated with increased dermal pro-inflammatory status. Interestingly, although inflammation induces the synthesis of ECM in most organs, the expression of ECM decreased in the inflamed skin of mice on high salt diet. Our data suggest that salt intake may alter the process of skin remodeling.

Immunomodulatory role of Parkinson's disease 7 in inflammatory bowel disease.

Recently the role of Parkinson's disease 7 (PARK7) was studied in gastrointestinal diseases, however, the complex role of PARK7 in the intestinal inflammation is still not completely clear. Expression and localization of PARK7 were determined in the colon biopsies of children with inflammatory bowel disease (IBD), in the colon of dextran sodium sulphate (DSS) treated mice and in HT-29 colonic epithelial cells treated with interleukin (IL)-17, hydrogen peroxide (H2O2), tumor necrosis factor (TNF)-α, transforming growth factor (TGF)-ß or lipopolysaccharide (LPS). Effect of PARK7 on the synthesis of IBD related cytokines was determined using PARK7 gene silenced HT-29 cells and 3,4,5-trimethoxy-N-(4-(8-methylimidazo(1,2-a)pyridine-2-yl)phenyl)benzamide (Comp23)-compound increasing PARK7 activity-treated mice with DSS-colitis. PARK7 expression was higher in the mucosa of children with Crohn's disease compared to that of controls. While H2O2 and IL-17 treatment increased, LPS, TNF-α or TGF-ß treatment decreased the PARK7 synthesis of HT-29 cells. PARK7 gene silencing influenced the synthesis of IL1B, IL6, TNFA and TGFB1 in vitro. Comp23 treatment attenuated the ex vivo permeability of colonic sacs, the clinical symptoms, and mucosal expression of Tgfb1, Il1b, Il6 and Il10 of DSS-treated mice. Our study revealed the role of PARK7 in the regulation of IBD-related inflammation in vitro and in vivo, suggesting its importance as a future therapeutic target.

Interleukin-24 regulates mucosal remodeling in inflammatory bowel diseases.

BACKGROUND: Recently, increased interleukin (IL)-24 expression has been demonstrated in the colon biopsies of adult patients with inflammatory bowel disease (IBD). However, the role of IL-24 in the pathomechanism of IBD is still largely unknown. METHODS: Presence of IL-24 was determined in the samples of children with IBD and in the colon of dextran sodium sulfate (DSS) treated mice. Effect of inflammatory factors on IL24 expression was determined in peripheral blood (PBMCs) and lamina propria mononuclear cells (LPMCs). Also, the impact of IL-24 was investigated on HT-29 epithelial cells and CCD-18Co colon fibroblasts. Expression of tissue remodeling related genes was investigated in the colon of wild type (WT) mice locally treated with IL-24 and in the colon of DSS treated WT and Il20rb knock out (KO) mice. RESULTS: Increased amount of IL-24 was demonstrated in the serum and colon samples of children with IBD and DSS treated mice compared to that of controls. IL-1ß, LPS or H2O2 treatment increased the expression of IL24 in PBMCs and LPMCs. IL-24 treatment resulted in increased amount of TGF-ß and PDGF-B in HT-29 cells and enhanced the expression of extracellular matrix (ECM)-related genes and the motility of CCD-18Co cells. Similarly, local IL-24 treatment increased the colonic Tgfb1 and Pdgfb expression of WT mice. Moreover, expression of pro-fibrotic Tgfb1 and Pdgfb were lower in the colon of DSS treated Il20rb KO compared to that of WT mice. The disease activity index of colitis was less severe in DSS treated Il20rb KO compared to WT mice. CONCLUSION: Our study suggest that IL-24 may play a significant role in the mucosal remodeling of patients with IBD by promoting pro-fibrotic processes.

PARK7 Diminishes Oxidative Stress-Induced Mucosal Damage in Celiac Disease.

Coeliac disease (CD) is a chronic, immune-mediated small intestinal enteropathy, accompanied with gluten-triggered oxidative damage of duodenal mucosa. Previously, our research group reported an increased mucosal level of the antioxidant protein Parkinson's disease 7 (PARK7) in children with CD. In the present study, we investigated the role of increased PARK7 level on the epithelial cell and mucosal integrity of the small intestine. The presence of PARK7 was investigated using immunofluorescent staining on duodenal mucosa of children with CD and on FHs74Int duodenal epithelial cells. To investigate the role of oxidative stress, FHs74Int cells were treated with H2O2 in the absence or presence of Comp23, a PARK7-binding compound. Intracellular accumulation of reactive oxygen species (ROS) was determined by DCFDA-based assay. Cell viability was measured by MTT, LDH, and Annexin V apoptosis assays. Disruption of cytoskeleton and cell adhesion was investigated by immunofluorescence staining and by real-time RT PCR. Effect of PARK7 on mucosal permeability was investigated ex vivo using intestinal sacs derived from control and Comp-23-pretreated mice. Comp23 treatment reduced the H2O2-induced intracellular accumulation of ROS, thus preserving the integrity of the cytoskeleton and also the viability of the FHs74Int cells. Accordingly, Comp23 treatment increased the expression of antioxidants (NRF2, TRX1, GCLC, HMOX1, NQO1), cell-cycle regulators (TP53, CDKN1A, PCNA, BCL2, BAX), and cell adhesion molecules (ZO1, CDH1, VCL, ITGB5) of H2O2-treated cells. Pretreatment with Comp23 considerably decreased the small intestinal permeability. In this study, we demonstrate that PARK7-binding Comp23 reduces the oxidative damage of duodenal epithelial cells, via increased expression of NRF2- and P53-regulated genes. Our results suggest that PARK7 plays a significant role in the maintenance of mucosal integrity in CD.

Characterization of IL-19, -20, and -24 in acute and chronic kidney diseases reveals a pro-fibrotic role of IL-24.

BACKGROUND: Recently, the role of IL-19, IL-20 and IL-24 has been reported in renal disorders. However, still little is known about their biological role. METHODS: Localization of IL-20RB was determined in human biopsies and in the kidneys of mice that underwent unilateral ureteral obstruction (UUO). Renal Il19, Il20 and Il24 expression was determined in ischemia/reperfusion, lipopolysaccharide, streptozotocin, or UUO induced animal models of kidney diseases. The effects of H2O2, LPS, TGF-ß1, PDGF-B and IL-1ß on IL19, IL20 and IL24 expression was determined in peripheral blood mononuclear cells (PBMCs). The extents of extracellular matrix (ECM) and α-SMA, Tgfb1, Pdgfb, and Ctgf expression were determined in the kidneys of Il20rb knockout (KO) and wild type (WT) mice following UUO. The effect of IL-24 was also examined on HK-2 tubular epithelial cells and NRK49F renal fibroblasts. RESULTS: IL-20RB was present in the renal biopsies of patients with lupus nephritis, IgA and diabetic nephropathy. Amount of IL-20RB increased in the kidneys of mice underwent UUO. The expression of Il19, Il20 and Il24 increased in the animal models of various kidney diseases. IL-1ß, H2O2 and LPS induced the IL19, IL20 and IL24 expression of PBMCs. The extent of ECM, α-SMA, fibronectin, Tgfb1, Pdgfb, and Ctgf expression was lower in the kidney of Il20rb KO compared to WT mice following UUO. IL-24 treatment induced the apoptosis and TGF-ß1, PDGF-B, CTGF expression of HK-2 cells. CONCLUSIONS: Our data confirmed the significance of IL-19, IL-20 and IL-24 in the pathomechanism of renal diseases. Furthermore, we were the first to demonstrate the pro-fibrotic effect of IL-24.

Role of IL-24 in the mucosal remodeling of children with coeliac disease.

BACKGROUND: Recently, involvement of IL-19, IL-20 and IL-24 has been reported in inflammatory diseases associated with tissue remodeling. However, their impact on the pathomechanism of coeliac disease (CD) is still completely unknown. METHODS: Expression of IL19, IL20 and IL24 was measured by real-time RT-PCR, protein amount of IL-24, α smooth muscle actin (α-SMA) and fibronectin (FN) was determined by Western-blot analysis in the duodenal biopsies of therapy naive children with CD and controls. Localization of IL-24 and IL-20RB was investigated by immunofluorescent staining in the duodenal mucosa. Effect of recombinant IL-1ß, TNF-α, TGF-ß and IL-17 treatment on the expression of IL19, IL20, IL24 and their receptors was investigated by real-time RT-PCR in small intestinal epithelial cells (FHs74Int), in primary duodenal myofibroblasts (pdMFs) and in peripheral blood mononuclear cells (PBMCs). Effect of IL-24 on H2O2 treated FHs74Int cells and on pdMFs was measured by MTT, LDH, Annexin V assays, real-time RT-PCR and by fluorescent microscopy. RESULTS: We found increased level of IL-24 (3.3×, p < 0.05), α-SMA (2.4×, p < 0.05) and FN (2.3×, p < 0.05) in the duodenal mucosa and increased expression of IL19 (3.6×, p < 0.05) and IL24 (5.2×, p < 0.05) in the PBMCs of children with CD compared to that of controls. IL-1ß was a strong inducer of IL24 expression of FHs74Int cells (9.9×, p < 0.05), pdMFs (552.9×, p < 0.05) or PBMCs (17.2×, p < 0.05), as well. IL-24 treatment reduced the number of apoptotic cells (0.5×, p < 0.05) and decreased the expression of inflammatory factors, including IL1A, IL6 and TNF of H2O2-treated FHs74Int cells. IL-24 decreased the proliferation (0.6×, p < 0.05) of PDGF-B treated pdMFs. Moreover, IL-24 treatment altered the morphology of pdMFs by influencing the size of the angles between stress fibers and the longitudinal axis of the cells (2.0×, p < 0.05) and the expression of cytoskeletal components, including ACTA2, ACTB, VIM, SNAI1 and SNAI2. CONCLUSION: Our results suggest that IL-24 plays a significant role in the maintenance of duodenal mucosal integrity in CD.

Role of microRNA-223 in the regulation of poly(ADP-ribose) polymerase in pediatric patients with Crohn's disease.

OBJECTIVES: Crohn's disease (CD) is a multifactorial disease, characterized by oxidant-induced tissue injury with a possible activation of poly(ADP-ribose) polymerase (PARP)-1. MicroRNAs (miRs) can offer a potential link between the genetic susceptibility, environmental and immunologic factors in the pathogenesis of CD. Previously, PARP-1 was identified as a direct target gene of miR-223 in an epithelial cell line. Our aim was to examine PARP activation and miR-223 expression in colonic biopsies of pediatric CD. To support our in vivo findings, the effect of lipopolysaccharide (LPS) on same parameters was examined in HT-29 colonic epithelial cell line. METHODS: Colonic biopsies were taken from patients with macroscopically inflamed and intact mucosa with CD and controls. LPS treated HT-29 cells served as our in vitro model. To analyze the PARP-1 expression real-time PCR, Western blot and immunohistochemical analyses were used. PARP-1 enzymatic activity was assessed on the basis of poly(ADP-ribosyl)ated proteins. Expression of miR-223 was examined by real-time PCR. RESULTS: PARP-1 mRNA and miR-223 expression was significantly elevated, however, the amount of PARP-1 protein and poly(ADP-ribose) was reduced in pediatric CD compared to controls. LPS incubation did not affect the expression of PARP-1 mRNA, however, decreased miR-223 expression, and enhanced PARP-1 activity. CONCLUSIONS: In our study, we showed that the expression of miR-223 is up-regulated and poly(ADP-ribosyl)ation is reduced in pediatric patients with CD. Moreover, we confirmed their opposite change in LPS treated epithelial cells, too. These data suggest that the hypofunctionality of PARP-1 may play a potential role in the pathomechanism of CD.

The role of Sigma-1 receptor in sex-specific heat shock response in an experimental rat model of renal ischaemia/reperfusion injury.

We previously showed that female rats are more protected against renal ischaemia/reperfusion (I/R) injury than males, which is partly attributed to their more pronounced heat shock response. We recently described that Sigma-1 receptor (S1R) activation improves postischaemic survival and renal function. 17ß-estradiol activates S1R, thus here we investigated the role of sex-specific S1R activation and heat shock response in severe renal I/R injury. Proximal tubular cells were treated with 17ß-estradiol, which caused direct S1R activation and subsequent induction of heat shock response. Uninephrectomized female, male and ovariectomized female (Ovx) Wistar rats were subjected to 50-min renal ischaemia followed by 2 (T2) and 24 (T24) hours of reperfusion. At T24 renal functional, impairment was less severe and structural damage was less prominent in females versus males or Ovx. Postischaemic increase in S1R, pAkt, HSF-1, HSP72 levels were detected as early as at T2, while pHSP27 was elevated later at T24. Abundance of heat shock proteins was higher in healthy female rats and remained higher at T2 and T24 (female versus male or Ovx; resp.). We propose a S1R-dependent mechanism, which contributes to the relative renoprotection of females after I/R injury by enhancing the heat shock response.

Selective measurement of α smooth muscle actin: why ß-actin can not be used as a housekeeping gene when tissue fibrosis occurs.

BACKGROUND: Prevalence of fibroproliferative diseases, including chronic kidney disease is rapidly increasing and has become a major public health problem worldwide. Fibroproliferative diseases are characterized by increased expression of α smooth muscle actin (α-SMA) that belongs to the family of the six conserved actin isoforms showing high degree homology. The aim of the present study was to develop real-time PCRs that clearly discriminate α-SMA and ß-actin from other actin isoforms. RESULTS: Real-time PCRs using self-designed mouse, human and rat specific α-SMA or ß-actin primer pairs resulted in the specific amplification of the artificial DNA templates corresponding to mouse, human or rat α-SMA or ß-actin, however ß-actin showed cross-reaction with the housekeeping γ-cyto-actin. We have shown that the use of improperly designed literary primer pairs significantly affects the results of PCRs measuring mRNA expression of α-SMA or ß-actin in the kidney of mice underwent UUO. CONCLUSION: We developed a set of carefully designed primer pairs and PCR conditions to selectively determine the expression of mouse, human or rat α-SMA and ß-actin isoforms. We demonstrated the importance of primer specificity in experiments where the results are normalized to the expression of ß-actin especially when fibrosis and thus increased expression of α-SMA is occur.

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-ß 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.

The role of sigma-1 receptor and brain-derived neurotrophic factor in the development of diabetes and comorbid depression in streptozotocin-induced diabetic rats.

RATIONALE: Depression is highly prevalent in diabetes (DM). Brain-derived neurotrophic factor (BDNF) which is mainly regulated by the endoplasmic reticulum chaperon sigma-1 receptor (S1R) plays a relevant role in the development of depression. OBJECTIVES: We studied the dose-dependent efficacy of S1R agonist fluvoxamine (FLU) in the prevention of DM-induced depression and investigated the significance of the S1R-BDNF pathway. METHODS: We used streptozotocin to induce DM in adult male rats that were treated for 2 weeks p.o. with either different doses of FLU (2 or 20 mg/bwkg) or FLU + S1R antagonist NE100 (1 mg/bwkg) or vehicle. Healthy controls were also enrolled. Metabolic, behaviour, and neuroendocrine changes were determined, and S1R and BDNF levels were measured in the different brain regions. RESULTS: In DM rats, immobility time was increased, adrenal glands were enlarged, and thymuses were involuted. FLU in 20 mg/bwkg, but not in 2 mg/bwkg dosage, ameliorated depression-like behaviour. S1R and BDNF protein levels were decreased in DM, while FLU induced SIR-BDNF production. NE100 suspended all effects of FLU. CONCLUSIONS: We suggest that disturbed S1R-BDNF signaling in the brain plays a relevant role in DM-induced depression. The activation of this cascade serves as an additional target in the prevention of DM-associated depression.

Peroxisome proliferator-activated receptor-γ and thymic stromal lymphopoietin are involved in the pathophysiology of childhood coeliac disease.

Celiac disease (CD) is a chronic autoimmune enteropathy caused by exposure to dietary gluten in genetically predisposed individuals. The transcription factor peroxisome proliferator-activated receptor gamma (PPARγ) was shown to exert protective effects in several immune-mediated disorders. Activation of PPARγ suppressed the expression of thymic stromal lymphopoietin (TSLP), an inducer of proinflammatory cytokines. Since the role of TSLP in gluten-sensitive enteropathy is completely unknown, we investigated the involvement of TSLP and its regulator PPARγ in childhood CD. We collected duodenal biopsy specimens from 19 children with newly diagnosed CD, 6 children with treated CD (gluten-free diet, GFD), and 10 controls. Expression of mRNA and protein levels of PPARγ, TSLP, and TSLP receptor were determined by real-time RT-PCR and Western blot, respectively. Duodenal localization of PPARγ and TSLP was studied by immunohistochemistry. In duodenal mucosa of children with CD, the amount of PPARγ was significantly lower and simultaneously that of TSLP significantly higher compared to controls (p < 0.05). In GFD-treated patients, the levels of PPARγ mRNA and protein were significantly higher while that of TSLP markedly lower compared to newly diagnosed CD (p < 0.05). Immunohistochemistry revealed PPARγ and TSLP expression in lamina propria immune cells and in enterocytes. Low expression of PPARγ and high expression of TSLP in the duodenal mucosa of children with newly diagnosed CD suggest that they are involved in the pathophysiology of CD. We hypothesize that PPARγ may be an inhibitory regulator of TSLP-stimulated inflammatory processes in CD.