The role of occlusion and micro-incontinence in the pathogenesis of penile lichen sclerosus: an observational study of pro-inflammatory cytokines' gene expression.

PURPOSE: To assess the expression of selected cytokines in penile lichen sclerosus (PLS) and associate them with the occurrence of micro-incontinence (MI) in different stages of PLS. METHODS: The skin biopsies from 49 PLS affected, and 13 from nonlesional foreskins (healthy control adult males undergoing circumcision due to phimosis caused by short frenulum) were obtained. All specimens were used for RNA extraction and RT-qPCR. Quantitative assessment of the gene expression of interleukin 1-A (IL-1A), interleukin 1-B (IL-1B), interleukin 1 receptor antagonist (IL-1RN), interleukin 6 (IL-6), transforming growth factor ß1 (TGF-ß1), and interferon-gamma (INF-γ) was performed. To determinate the presence of MI, the patients were asked about voiding patterns, especially leaking tiny drops of urine from the urethral meatus after urination. RESULTS: IL-1A, IL-6, and INF-γ mRNA levels were approximately 150, 16, and 59 times higher in PLS than in control samples, respectively. The highest IL-1A mRNA levels were observed in early PLS (n = 13), INF-γ in moderate PLS (n = 32), while IL-6 in severe PLS (n = 4). MI was noted in 45 PLS patients vs. 0 in control (p < 0.0001). IL-1A and IL-6 vs control ratios were concentration (ca.) 400 and 30 times higher, respectively, in MI PLS samples than in PLS without MI. CONCLUSION: Occlusion and irritating urine effect are associated with the clinical progression of penile LS with increased mRNA expression of IL-1A, INF-γ, and IL-6 pro-inflammatory cytokines in the foreskin.

The Immunoexpression of YAP1 and LATS1 Proteins in Clear Cell Renal Cell Carcinoma: Impact on Patients' Survival.

The aim of the study was to determine by immunohistochemistry cellular localization and immunoreactivity levels of YAP1 and LATS1 proteins in paired sections of tumor and unchanged renal tissues of 54 clear cell renal cell carcinoma (ccRCC) patients. Associations between clinical-pathological and overall survival (OS; median follow-up was 40.6 months) data of patients and YAP1 and LATS1 immunoreactivity were analyzed by uni- and multivariate Cox regression model and log-rank test. YAP1 immunoreactivity was found in the nuclei of tumor cells in 64.8% of ccRCC patients, whereas only 24.1% of tumors revealed cytoplasmic YAP1 expression. LATS1 immunoexpression was observed only in the cytoplasm of tumor cells in 59.3% of patients. LATS1 immunoreactivity in cancer cells negatively correlated with the size of primary tumor. The overall YAP1 immunoreactivity did not correlate with clinical-pathological data of patients. However, the subgroup of ccRCC patients who presented with cytoplasmic YAP1 immunoexpression had significantly shorter OS (median = 26.8 months) than patients without cytoplasmic YAP1 expression (median undefined). Multivariate Cox analysis revealed that increased cytoplasmic YAP1 (HR = 4.53) and decreased LATS1 immunoreactivity levels (HR = 0.90) were associated with worse prognosis, being independent prognostic factors. These results suggest that YAP1 and LATS1 can be considered as new prognostic factors in ccRCC.

Differential antitumor effects of vitamin D analogues on colorectal carcinoma in culture.

Colorectal cancer (CRC) is an emerging global problem with the rapid increase in its incidence being associated with an unhealthy lifestyle. Epidemiological studies have shown that decreased levels of vitamin D3 significantly increases the risk of CRC. Furthermore, negative effects of vitamin D3 deficiency can be compensated by appropriate supplementation. Vitamin D3 was shown to inhibit growth and induce differentiation of cancer cells, however, excessive vitamin D3 intake leads to hypercalcemia. Thus, development of efficient vitamin D3 analogues with limited impact on calcium homeostasis is an important scientific and clinically relevant task. The aims of the present study were to compare the antiproliferative potential of classic vitamin D3 metabolites (1α,25(OH)2D3 and 25(OH)D3) with selected low calcemic analogues (calcipotriol and 20(OH)D3) on CRC cell lines and to investigate the expression of vitamin D-related genes in CRC cell lines and clinical samples. Vitamin D3 analogues exerted anti-proliferative effects on all CRC cell lines tested. Calcipotriol proved to be as potent as 1α,25(OH)2D3 and had more efficacy than 20-hydroxyvitamin D3. In addition, the analogs tested effectively inhibited the formation of colonies in Matrigel. The expression of genes involved in 1α,25(OH)2D3 signaling and metabolism varied in cell lines analysed, which explains in part their different sensitivities to the various analogues. In CRC biopsies, there was decreased VDR expression in tumor samples in comparison to the surgical margin and healthy colon samples (p<0.01). The present study indicates that vitamin D3 analogues which have low calcemic activity, such as calcipotriol or 20(OH)D3, are very promising candidates for CRC therapy. Moreover, expression profiling of vitamin D-related genes is likely to be a powerful tool in the planning of anticancer therapy. Decreased levels of VDR and increased CYP24A1 expression in clinical samples underline the importance of deregulation of vitamin D pathways in the development of CRC.

TL1A as a Potential Local Inducer of IL17A Expression in Colon Mucosa of Inflammatory Bowel Disease Patients.

Interaction between TL1A and death receptor 3 (DR3) co-stimulates T cells, induces production of several pro-inflammatory cytokines and has been linked to pathogenesis of inflammatory bowel disease (IBD). This study aimed to establish a link between expression of TL1A and selected TL1A-induced pro-inflammatory cytokines involved in IBD pathogenesis (IL-4, IL-13, IL-17A and IFN-γ) and to investigate a connection between serum concentration of TL1A in patients with IBD and activation of peripheral blood T cells. Elevated levels of IL-4 (2.91-fold) and IL-13 (4.05-fold) mRNA were detected in the inflamed colon mucosa of patients with ulcerative colitis (UC), IFN-γ mRNA was upregulated (3.23-fold) in the inflamed colon mucosa of patients with Crohn's disease (CD), whereas upregulation of IL-17A and TL1A mRNA was present in the inflamed colon mucosa of patients with both CD and UC (IL-17A: 4.48-fold and 2.74-fold, TL1A: 3.19-fold and 3.22-fold, respectively) vs. control subjects. We did not detect any changes in DR3 mRNA expression in the investigated groups of patients. TL1A mRNA level in colon mucosa of patients with IBD correlated only with the level of IL-17A mRNA but no other investigated cytokines. In colon mucosa, expression of TL1A and DR3 was localized to enterocytes and lamina propria mononuclear cells. We did not find any correlation between serum concentrations of TL1A and IL-17A or changes of CD4(+) or CD8(+) lymphocytes phenotype in patients with IBD. Therefore, our data indicate that TL1A may contribute to pathogenesis of IBD via local but not systemic induction of IL-17A but not IL-4, IL-13 or IFN-γ.

Short-term calorie restriction and refeeding differently affect lipogenic enzymes in major white adipose tissue depots of young and old rats.

The metabolic effects of short-term calorie restriction (SCR) and subsequent refeeding were compared in different white adipose tissue (WAT) depots of young (5-month old) and old (24-month) male Wistar rats. The animals were subjected to a 40% calorie restricted diet (i.e. 40% lower food supply than of control rats) for 30 days, and then re-fed for 0, 2, or 4 days. WAT samples from perirenal (pWAT), epididymal (eWAT), and subcutaneous (sWAT) depots were analysed for the enzymatic activities of ATP-citrate lyase (ACL), fatty acid synthase (FAS), and glucose-6-phosphate dehydrogenase (G6PD). The total WAT mass almost doubled in old rats, however, aging did not alter the relative proportions of the major regional fat depots. Serum leptin concentration was prominently higher in old rats, in which SCR resulted in less suppression of leptin level than in young animals, whereas refeeding increased leptin concentration in young, but not old, rats. In young rats refeeding elevated leptin gene expression only in pWAT, while in old rats the expression was induced first in eWAT, and later in pWAT. A prominent age-related decrease of ACL and FAS activities, but not of G6PD activity, was found in all the studied WAT depots. In young control rats, ACL activity was highest in pWAT, FAS activity was similar in all WAT depots, and G6PD activity was lowest in eWAT. In old rats, the enzymatic activities were lower in eWAT than in the other depots. The patterns of response to SCR and refeeding varied by age and WAT location. SCR stimulated ACL activity in pWAT but not in other depots of young rats, while FAS activity in pWAT and sWAT did not change in young and decreased in the old animals. Among the studied depots, pWAT was most responsive to refeeding in both age groups. In conclusion, SCR in old rats, as compared to the young, may be accompanied by reduced 'rebound effect' upon returning to unrestricted diet.

Influence of receptor activator of nuclear factor kappa B ligand, osteoprotegerin and interleukin-33 on bone metabolism in patients with long-standing ulcerative colitis.

BACKGROUND: Ulcerative colitis (UC) is a chronic disease with periods of remission and recurrences. Dysfunction of the local immune response leads to chronic inflammation within the large intestine which triggers morphological changes in the intestinal wall as well as induces the synthesis of numerous factors that have an adverse impact on the bone metabolism. The aim of the study was to determine the expression of RANKL, OPG and IL-33 in mucosal biopsies of UC patients with long disease duration as well as serum level of these cytokines in the context of bone density and bone metabolism. MATERIALS AND METHODS: The UC group consisted of 56 patients with average disease duration of 16y. The control group comprised 37 healthy individuals. Local expression of cytokines was assessed in the biopsies of colonic mucosa by the real-time PCR and immunohistochemistry (IHC), and their serum concentration was measured by ELISA. RESULTS: The increased bone resorption observed in patients with UC was reflected by low bone density and high serum level of C-terminal telopeptide (CTX). Mucosal RANKL expression and serum concentration were similar in UC group and healthy subjects, however, UC patients had higher local expression of OPG and serum OPG concentration. Increased IL-33 gene expression was observed only in UC at the mRNA level. We propose that bone resorption in UC patients despite OPG up-regulation could be caused by IL-33-induced mucosal synthesis of a potent proinflammatory cytokine, such as TNF-α, known as a possible inducer of osteoclastogenesis in the way independent of RANKL.

Human peripheral blood CD8+ CD28- T cells of renal allograft recipients do not express FOXP3 protein.

INTRODUCTION: In recent studies, the FOXP3 molecule has been suggested to be a marker of a suppressor subset of human CD8+ CD28- T cells based on correlations between the level of its mRNA and allograft function. Because this transcriptional factor produces a protein, we suggest that these correlations should focus on the FOXP3 protein. The aim of our study was to evaluate whether FOXP3 protein was present in cells of the CD8+ CD28- population in the peripheral blood of renal allograft recipients and whether the level of CD8+ CD28- FOXP3+ cells correlated with allograft function. METHODS: The study was performed on 30 renal allograft recipients with uneventful stable courses (n=18) or biopsy-proven chronic rejection (n=12). The immunosuppression was based on cyclosporine (n=12) or rapamycin (n=9). Peripheral blood mononuclear cells isolated from recipient blood samples were labeled with anti-CD8 and anti-CD28 MAbs conjugated with fluorochromes. After incubation, washing, and labeling using a PE anti-human FOXP3 Kit, we determined the percentage of cells by flow cytometry. RESULTS: FOXP3 protein expression was not observed either in the CD8+ CD28- population, or the whole populations of CD8+ or CD28- cells among patient groups. CONCLUSIONS: The expression of FOXP3 protein in CD8+ CD28- cells seems to be of a questionable value as a diagnostic tool for allograft function, it is probably not a marker for the CD8+ CD28- T cell subset.

Overexpression of the fragile histidine triad (FHIT) gene in inflammatory bowel disease.

OBJECTIVE: FHIT gene encodes human diadenosine triphosphate hydrolase involved in the regulation of cell cycle and nucleotide metabolism and is a candidate tumor suppressor gene. AIM: To investigate expression of FHIT gene at the mRNA and protein levels in sporadic inflammatory bowel disease (IBD). MATERIALS AND METHODS: FHIT mRNA was quantified by the validated real-time PCR (QPCR) and FHIT protein was detected by immunohistochemistry (IHC) in mucosal biopsies of 139 ulcerative colitis (UC), 19 Crohn's disease (CD) and 37 control patients. RESULTS: Significant FHIT gene overexpression was found in 78% of active UC but not in CD. IHC showed comparable results to QPCR. CONCLUSION: The local up-regulation of FHIT gene and protein expression in active UC may represent an adequate response against inflammatory challenge of epithelial cell homeostasis and protect against DNA damage and cell cycle disturbances.

Fragile histidine triad (FHIT) gene is overexpressed in colorectal cancer.

OBJECTIVE: FHIT gene, mapped at FRA3B site, encodes human diadenosine triphosphate hydrolase involved in the regulation of cell cycle and nucleotide metabolism. Decreased FHIT gene expression was previously observed in various types of human cancer, however, quantification of FHIT mRNA was seldom performed. AIM: To investigate loss of heterozygosity (LOH) at FRA3B, expression of FHIT gene at the mRNA and protein levels in sporadic colorectal carcinoma (CRC) and benign colon adenoma. MATERIALS AND METHODS: FHIT mRNA was quantified by the validated realtime PCR (QPCR) in tumor samples of 84 CRC patients and mucosal biopsies of 15 adenomas, in comparison to 37 control patients, whereas subgroup of 57 CRC, 10 adenoma and 10 control cases were selected for immunohistochemical (IHC) detection of the native FHIT protein and LOH determination at FRA3B. RESULTS: Higher level of FHIT mRNA was found in 86% of CRC (P<0.001) and 60% of adenomas (P=0.016). IHC showed comparable results to QPCR (P=0.003), revealing the strongest presence of FHIT protein in Dukes' C/D stages (P<0.001) and N1/N2 lymph nodes metastasis in CRC (P=0.04). FHIT gene expression and Dukes' and G staging were positively correlated in CRC as analyzed by QPCR and IHC. Deletion analysis of the fragile FRA3B site revealed the highest LOH frequency at D3S1234 in 32.5% of CRC informative cases, however, LOH did not correspond to QPCR, IHC or clinical-pathological variables. CONCLUSION: Our data suggest that reduction or absence of the FHIT gene expression is not a prerequisite for colorectal cancer development and progression.

Decreased Toll-like receptor-5 (TLR-5) expression in the mucosa of ulcerative colitis patients.

OBJECTIVE: Although there is a convincing evidence supporting an important role for microorganisms in the pathogenesis of Inflammatory Bowel Disease (IBD) which comprises ulcerative colitis (UC) and Crohn's disease (CD), the specific mechanisms involved remain unclear. Toll-like receptors (TLR) recognize various molecules of microbiota including flagellin, the principal protein of motile comensal and pathogenic bacteria implicated in the pathogenesis of IBD. AIM: To investigate the expression of the TLR-5 receptors at the mRNA and protein levels in the mucosa of UC patients. MATERIALS AND METHODS: TLR-5 mRNA was quantified by the validated real-time PCR (QPCR) in mucosal biopsies of 99 UC patients and 34 control patients and TLR-5 protein was detected by immunohistochemistry (IHC) in 57 UC and 10 control patients. RESULTS: Significantly decreased TLR-5 gene expression at mRNA and protein level was found in the mucosa of patients with moderate and severe disease activity as compared to patients with low UC activity and control. TLR-5 immunoreactivity was found in the mucosa of UC patients and normal controls in the cytoplasm of enterocytes and at their basolateral domain. However, the intensity of the IHC reaction in specimens from UC patients was substantially lower than in control samples. CONCLUSION: The decreased expression of TLR-5 gene and protein in the mucosa of UC patients suggests that down-regulation of TLR-5 is probably caused by the increased number of ligand molecules in the proximity of epithelial cells in the inflamed tissue.

The influence of immuosuppressive therapy on the development of CD4+CD25+ T cells after renal transplantation.

A growing number of studies suggest that CD4(+)CD25(+) T regulatory (Treg) cells play a significant role to downregulate the immune response to alloantigens. In this study, we investigated the possible influence of immunosuppressive therapy, including cyclosporine (CsA) or rapamycin (sirolimus), on the level of CD4(+)CD25(+), CD4(+)CD25(+)FOXP3(+), and CD4(+)CD25(+)CTLA-4(+) T cells in the peripheral blood of renal allograft recipients. The study was performed on renal allograft recipients who displayed uneventful stable courses (RAR-S; n = 15) versus biopsy-proven chronic rejection (RAR-CH; n = 12). The patients were divided based on the immunosuppressive protocol: group 1 (prednisone+CsA+Aza) and group II (prednisone+sirolimus). The control group consisted of 10 healthy blood donors. We examined the expression of CD4, CD25, CTLA-4, and Foxp3 in peripheral blood T cells. Flow cytometry was performed with a FACSCalibur (BD Biosciences) instrument with data analyzed using Cell Quest software. The percentage of CD4(+)CD25(+)Foxp3(+) T cells in rapamycin (sirolimus) treated patients did not differ from that observed in healthy individuals, but was significantly higher compared with CsA-treated patients. CsA therapy resulted in a reduction in the percentage of CD4(+)CD25(+)CTLA-4(+) and CD4(+)CD25(+)Foxp3(+) regulatory T cells after renal transplantation in both groups (RAR-S and RAR-CH) compared with patients treated with rapamycin or to healthy donors. The type of immunosuppressive therapy (with or without calcineurin inhibitors) may have an important role in tolerance induction and graft function.

Beta 3 integrin expression on T cells from renal allograft recipients.

Recent studies emphasize the paramount significance of beta 3 integrin in cell adhesion and homing, which may be particularly relevant in cancer progression and metastasis. In contrast, the presence and potential role of beta 3 on human T cells is practically unknown. We show that T cells can express significant amounts of alpha-beta 3 integrin (CD41/CD61), and the expression of alpha(v)-beta 3 (CD51/CD61) remains very low. T-cell beta 3 integrin is probably transferred by platelet-derived microparticles.

Parametric generation of forward and phase-conjugated spin-wave bullets in magnetic films.

We show experimentally as well as by numerical simulation that interaction of a linear two-dimensional spin-wave packet with quasiuniform pulsed pumping leads to the formation of strongly self-focused nonlinear spin-wave bullets propagating in both forward and reversed directions. The focusing of the reversed, phase-conjugated wave bullet is stronger than that of the forward one, because not only the nonlinear four-wave self-focusing effect but also linear focusing due to two-dimensional phase conjugation contributes to the focusing of the reversed bullet.

New insights into the possible role of bacteriophages in transplantation.

Due to the increasing prevalence of drug-resistant bacterial infections in the "post-antibiotic era," bacteriophages (bacterial viruses, BP) may be useful to administer to transplant recipients without exposing them to an increased risk of rejection, which occurs consequent to some viral infections. Herein we present evidence that at least some coliphages (T4) do not pose such risk. Interestingly, they may produce immunosuppressive effects extending transplant survival. Our data suggest that BP may be used in clinical transplantation to treat drug-resistant bacterial infections and perhaps as an adjunct to immunosuppressive therapy.

Involvement of beta2-microglobulin in CD69 expression on T cells.

Beta2-microglobulin (beta2M) is the light chain of the class I HLA molecule. The serum level of beta2M is elevated in various diseases including lymphoma, inflammation, viral infections and chronic renal dysfunction. The present study addressed the possible influence of beta2M on T lymphocyte activation in vitro. Peripheral blood mononuclear cells from a group of 17 healthy subjects were examined. Stimulation with OKT3 and fibronectin in combination with 30 mg beta2M/dl resulted in a two-fold increase of cell proliferation. A similar effect was observed when OKT3 and collagen I were applied as well as when OKT3 and collagen IV were used as costimulation to T cells. The CD69 expression, measured by flow cytometry was significantly enhanced above the control level (1.52 +/- 1.03% vs 33.21 +/- 20.26%, p<0.01, control group and 30 mg beta2M/dl, respectively). Together, these observations suggest that beta2M may play a role in modulating lymphocyte proliferation, possibly through modification of the CD69 molecule.

[Doppler sonography assessment of renal arteries against the cardiovascular risk factors in young men--preliminary report]. / Ultrasonograficzna dopplerowska ocena tetnic nerkowych a czynniki ryzyka chorób sercowo-naczyniowych u mlodych mezczyzn--doniesienie wstepne.

The purpose of this study was to find out a relationship between cardiovascular risk factors (CVRF) and abnormal results of Duplex Doppler sonography (DD), suggesting renal artery stenosis (RAS). The group of 30 potentially healthy men (age: 35.5 +/- 7.9) randomly selected from population of 200 young men, underwent DD of renal arteries. Abnormal results of DD study, suggesting RAS were found in 19 of 30 men (63%). In this group, occurrence of 3 or more CVRF was significantly higher (15/19) than in the group with normal results of DD (5/11; p < 0.05). The significantly higher (77%; p < 0.05) occurrence of the abnormal DD and the significantly longer (p < 0.001) duration of habitual tobacco smoking were also found out in the group with RR > or = 130/85 mm Hg, in comparison to the group with RR < 130/85 mm Hg. Emphasizing preliminary character of this study and the need of confirmation on larger population, it seems reasonable to consider the assessment of renal arteries with DD in the population of men with cardiovascular risk factors.

[Biochemical risk factor for atherosclerosis in young men. Preliminary report]. / Biochemiczne czynniki ryzyka miazdzycy u mlodych mezczyzn. Doniesienie wstepne.

Atherosclerotic damage of cardiovascular system, including kidneys, is an increasing problem not only in the modern cardiology but also in nephrology and dialysotherapy. The purpose of our study was to evaluate the frequency of particular biochemical risk factors for atherosclerosis (RFfA) in young men being over-pressed with psychological stress because of decisive character of their professional job. There were 68 men in mean age 33.3 +/- 7.4 years, being employed in the state administration for mean 10.9 +/- 7.5 years. On the basis of the results of environmental questionnaire the studied group was divided into two subgroups: subgroup A "passive" (30 persons) which was physically passive (employed in the administration) and subgroup B "active" (38 persons) whose professional job was connected with physical activity. The following biochemical RFfA in the blood were examined: cholesterol and its fractions, triglycerides, urea acid, glucose, fibrinogen, C-reactive protein, homocysteine and antibodies against Chlamydia pneumoniae. The results were as follows: total cholesterol above 200 mg/dl in 63%, LDL cholesterol above 130 mg/dl in 60% and HDL cholesterol below 50 mg/dl in 31% of men. Among "new" RFfA we found "cardiological" (above 0.2 mg/dl) levels of CRP almost in 25% of the whole group. There were no statistically significant differences with respect to the studied RFfA between "active" and "passive" group. On the basis of these preliminary results we presume, that potentially healthy men over-pressed with psychological stress because of decisive character of their professional job, are characterized--independently on the proclaimed physical activity--by high frequency of risk factors for premature atherosclerosis.