The Role of T Follicular Helper Cells in Clinical Remission and Relapse in Patients with Pemphigus Treated with Rituximab.

BACKGROUND: Pemphigus is a rare chronic autoimmune disease. Recent studies have found that T follicular helper (Tfh) cells may play a role in autoimmune diseases. In this study, Tfh cells frequency, BCL6 gene expression, IL-21, and IL-6 cytokines levels were examined, with the aim of understanding the effect of RTX on these cells in the onset of clinical remission or relapse in patients with pemphigus. METHODS: 20 patients with pemphigus vulgaris and 20 healthy controls without any autoimmune diseases that were admitted to the Dermatology and Venereology Clinic of the Akdeniz University Hospital were included. Peripheral blood sample was taken from all individuals and studied to analyze Tfh cell distribution, IL-21 and IL-6 distribution in CD3+CD4+CXCR5+ lymphocytes with flow cytometry, plasma IL-21 levels with ELISA, and mRNA levels that refer to BCL6 expression with PCR. RESULTS: Circulating Tfh cell distribution and IL-21 and IL-6 distribution in CD3+CD4+CXCR5+ lymphocytes and mRNA levels that refer to BCL6 expression showed no difference between patient and control groups. However, in patients who had received rituximab treatment there was a significant reduction in Tfh cells compared with other groups. Plasma IL-21 levels were significantly higher in the patient group. CONCLUSIONS: We found that plasma concentrations of the cytokine IL-21 were greatly increased in the pemphigus compared with the control group. There were no significant differences in Tfh cell percentages between the patient and control groups. Tfh cells were decreased in patients who received rituximab treatment. Our findings show that the response to RTX in pemphigus causes a reduction in circulating T follicular helper cells, but not in the plasma IL-21 level. Further studies are required to clarify the role of Tfh cells in pemphigus vulgaris.

Phenotypic and Molecular Spectrum of a Turkish Cohort with Hereditary Multiple Osteochondromas.

OBJECTIVE: Hereditary multiple osteochondromas is an autosomal dominant disorder caused by heterozygous pathogenic variants in EXT1 or EXT2. We aimed to evaluate the clinical and molecular findings of a Turkish cohort with hereditary multiple osteochondroma. MATERIALS AND METHODS: Thirty-two patients aged 1.3-49.6 years from 22 families were enrolled. Genetic analyses were made by EXT1 and/or EXT2 sequencing and chromosomal microarray analyses. RESULTS: We found 17 intragenic pathogenic variants in EXT1 (13/17) and EXT2 (4/17), 12 of which are novel. Four probands had EXT1 deletions, including 2 patients with partial EXT1 microdeletions involving exons 2-11 and 5-11, and 2 patients with whole-gene deletions. In 21 variants, the frequency of truncating and missense variants was 76.1% and 23.8%, respectively. Two families had no detectable variants in EXT1 and EXT2. All patients had multiple osteochondromas at the long bones, mainly at the tibia, forearm, femur, and humerus. Bowing deformity of the forearms (9/32) and the lower extremities (2/32), and scoliosis (6/32) were observed. The clinical severity was not different between patients with EXT1 or EXT2 variants. One patient with an EXT2 variant and another with an EXT1 microdeletion had the most severe phenotype with class III disease. Four patients with no EXT1 or EXT2 variants had milder phenotypes. Intrafamilial variability in disease severity was not observed. CONCLUSION: We report a hereditary multiple osteochondroma cohort with clinical and molecular data including 12 novel intragenic variants in EXT1 or EXT2, and 4 microdeletions involving EXT1. Taken together, our data expand the existing knowledge of the phenotype-genotype spectrum in hereditary multiple osteochondroma.

Investigation of the relationship between reproductive disorders and chromosomal abnormalities in a large-scale, single-center 10-year retrospective study.

OBJECTIVE: Chromosomal changes are an important cause of reproductive disorders. This study investigated the chromosomal changes and prevalence of pathologies in individuals admitted to our Genetic Evaluation Center over a 10-year period due to a reproductive disorder. MATERIALS & METHODS: The chromosomal findings of 4345 individuals with reproductive disorders who applied to our Genetic Evaluation Center at Akdeniz University in Antalya, Turkey between 2011 and 2021 were retrospectively evaluated. RESULTS: In this study, an abnormal karyotype was found in a total of 138 individuals (87 males and 51 females). Although the incidence of this abnormal karyotype varied among the diseases in the reproductive disorder subgroups, it was most frequently seen in azoospermia (17.0%). Of the 138 abnormalities, 75 were numerical and 54 were structural. The remaining 9 abnormalities consisted of 6 sex reversals and 3 patients with both numerical and structural anomalies. Additionally, the X chromosome was the chromosome most frequently involved in these abnormalities, being observed in 40.6% of patients. CONCLUSION: This 10-year, single-center study involved one of the largest case series in the literature to investigate the subtypes of reproductive disorders and their chromosomal relationship. Although the importance of chromosome analysis has been deemphasized, it is still recommended for use by the guidelines and, as the results of this study demonstrate, is still a highly effective method in the investigation of reproductive disorders. Furthermore, chromosome analysis of individuals diagnosed with a reproductive disorder is also very important in the practice of the increasingly utilized preimplantation genetic diagnosis (PGD).

Evaluation of exonic copy numbers of SMN1 and SMN2 genes in SMA.

SMA is a neuromuscular disease and occurs primarily through autosomal recessive inheritance. Identification of deletions in the SMN1 gene especially in the exon 7 and exon 8 regions (hot spot), are used in carrier testing. The exact copy numbers of those exons in the SMN1 and SMN2 genes in 113 patients who presented with a pre-diagnosis of SMA were determined using MLPA method. We aimed to reveal both the most common copy number profiles of different SMA types. It was found that the frequency of homozygous deletions in SMN1 was 15.9%, while heterozygous deletions was 16.9%. The most common SMN-MLPA profile was 0-0-3-3. In the cases with homozygous deletion, SMA type III diagnosis was observed most frequently (44%), and the rate of consanguineous marriage was found 33%. Two cases with the same exonic copy number profile but with different clinical subtypes were identified in a family. We also detected distinct exonic deletion and duplication MLPA profiles for the first time. We created "the SMA signature" that can be added to patient reports. Furthermore, our data are important for revealing potential local profiles of SMA and describing the disease in genetic reports in a way that is clear and comprehensive.

Alterations in plasma miR-21, miR-590, miR-192 and miR-215 in idiopathic pulmonary fibrosis and their clinical importance.

BACKGROUND: Many studies have revealed that microRNA (miRNA) molecules may take part in idiopathic pulmonary fibrosis (IPF). But, the role of miRNAs in the development of IPF is not yet clear. METHODS: We investigated the plasma levels of miR-21, miR-590, miR-192, and miR-215 in IPF (n = 88) and healthy control (n = 20) groups in this study. We compared the expression levels of target miRNAs in patients with IPF and healthy participants. We grouped the patients with IPF according to age, forced vital capacity, carbon monoxide diffusing capacity (DLCO), gender-Age-pulmonary physiology (GAP) score, the presence of honeycombing and compared the expression levels of target miRNAs in these clinical subgroups. RESULTS: 82 (93.18%) of the patients with IPF were male and the mean age was 66.6 ± 8.6 years. There was no significant difference between the gender and age distributions of IPF and the control group. The mean plasma miR-21 and miR-590 levels in IPF group were significantly higher than in the control group (p < 0.0001, p < 0.0001, respectively). There was no significant difference between the miR-192 and miR-215 expression levels of the IPF and control group. Both miR-21 and miR-590 correlated positively with age (p = 0.041, p = 0.007, respectively) while miR-192 and miR-215 displayed a negative correlation with age (p = 0.0002, p < 0.0001, respectively). The levels of miR-192 and miR-215 increased as the GAP score decreased. The levels of miR-192 in patients with honeycombing were significantly lower than in those without honeycombing (p = 0.003). CONCLUSIONS: Our study showed that both miR-21 and miR-590 were overexpressed in IPF. The miR-21 and miR-590 were associated with DLCO, while miR-192 and miR-215 were associated with the GAP score and honeycombing.

The Expression of GDF-15 in the Human Vitreous in the Presence of Retinal Pathologies with an Inflammatory Component.

PURPOSE: The presence of growth differentiation factor-15 (GDF-15), a protein implicated in the regulation of the inflammatory response, was investigated in the vitreous of patients with vitreoretinal disorders. METHODS: Vitreous and plasma samples were collected from patients with idiopathic epiretinal membrane (IERM), macular hole (MH), rhegmatogenous retinal detachment (RRD), nucleus drop (ND), or proliferative diabetic retinopathy (PDR). GDF-15 concentrations were measured using ELISA. RESULTS: The vitreous levels of GDF-15 were higher in ND (5) and PDR (14) patients (1494 ± 243 and 904 ± 138 pg/mL, respectively) than RRD (3), MH (3), and IERM (8) patients (302 ± 160, 288 ± 24, and 254 ± 91 pg/mL, respectively). The vitreous levels of GDF-15 were significantly higher in patients with inflammatory vitreoretinal disorders (p < 0.0001). CONCLUSIONS: This is the first report showing that GDF-15 appears to be expressed in the vitreous, and that its expression is significantly higher in the presence of a vitreoretinal disorder in which there is an inflammatory component.

Downregulation of VANGL1 inhibits cellular invasion rather than cell motility in hepatocellular carcinoma cells without stimulation.

AIMS: The Wnt planar cell polarity (PCP) pathway is one of the Wnt pathways which plays a critical role in cell proliferation and fate. The VANGL1 protein is one of Wnt-PCP pathway components. It is known that Wnt-PCP pathway has major roles in cell motility but its role in hepatocellular carcinoma (HCC) progression through invasion and metastasis needs to be clarified. METHODS: We silenced VANGL1 gene expression in the HepG2 HCC cell line by stable transfection with a vector containing siRNA template for VANGL1 and investigated the change in cell invasion and motility. RESULTS: Transfected cells with the siRNA template showed significantly suppressed invasive capacity when compared to controls although cellular motility was only slightly affected. CONCLUSION: Our study showed a basal role for VANGL1 with respect to the invasive capacity of HCC cells. This suggests that the Wnt-PCP pathway may play a role in progression of HCC through cellular invasion but further studies are needed to clarify its role in cell motility.

The association of Klinefelter syndrome and multiple pterygium syndrome: an unusual presentation.

Multiple pterygium syndrome is characterized by a number of phenotypic features, small stature, webbing of the neck, elbows, and/or knees, and joint contractures. In this report, we present an 11-year-old boy who had the classical findings of multiple pterygium syndrome, and his chromosomal analysis revealed a 47,XXY karyotype. Interestingly, he did not show any of the main clinical signs of Klinefelter syndrome. This patient appears to be the first reported case in the literature in which a non-mosaic 47,XXY karyotype has been found in a patient with multiple pterygium syndrome. The aim of the present report is to describe a non-classic Klinefelter syndrome associated with multiple pterygium syndrome and to emphasize the importance of karyotype analysis in patients with multiple pterygium syndrome.

The levels of hepatocyte growth factor in serum and follicular fluid and the expression of c-Met in granulosa cells in patients with polycystic ovary syndrome.

OBJECTIVE: To evaluate the levels of hepatocyte growth factor (HGF) in follicular fluid (FF) and the expression of c-Met in granulosa cells (GCs) with respect to the quality of the oocyte and embryo both in patients with polycystic ovary syndrome (PCOS) and in the normal ovary during controlled ovarian hyperstimulation cycles. DESIGN: Prospective controlled study. SETTING: University hospital. PATIENT(S): Fifty-nine women undergoing IVF treatment (of whom 21 had PCOS and 38 were in the control group). INTERVENTION(S): A total of 168 FF samples were collected at the time of oocyte retrieval. The HGF levels were measured by ELISA, and the mRNA expression of c-Met in GCs was detected by real-time polymerase chain reaction. MAIN OUTCOME MEASURE(S): The predictive values of HGF levels in serum and FF and the mRNA expression of c-Met in GCs for successful fertilization and oocyte-embryo quality. RESULT(S): The levels of HGF in serum and FF and the c-Met expression in GCs were similar between the PCOS and control groups. Granulosa cells of fertilized oocytes (2PN) had a significantly higher level of c-Met expression than that in oocytes that failed to fertilize. The mean HGF level in FF was significantly higher in the grade 1 embryos than in the grades 2-4 embryos. CONCLUSION(S): This study suggests that HGF/c-Met signaling may be a crucial determinant of fertilization success.

Filamin a mediates HGF/c-MET signaling in tumor cell migration.

Deregulated hepatocyte growth factor (HGF)/c-MET axis has been correlated with poor clinical outcome and drug resistance in many human cancers. Identification of novel regulatory mechanisms influencing HGF/c-MET signaling may therefore be necessary to develop more effective cancer therapies. In our study, we show that multiple human cancer tissues and cells express filamin A (FLNA), a large cytoskeletal actin-binding protein, and expression of c-MET is significantly reduced in human tumor cells deficient for FLNA. The FLNA-deficient tumor cells exhibited poor migrative and invasive ability in response to HGF. On the other hand, the anchorage-dependent and independent tumor cell proliferation was not altered by HGF. The FLNA-deficiency specifically attenuated the activation of the c-MET downstream signaling molecule AKT in response to HGF stimulation. Furthermore, FLNA enhanced c-MET promoter activity by its binding to SMAD2. The impact of FLNA deficiency on c-MET expression and HGF-mediated cell migration in human tumor cells was confirmed in primary mouse embryonic fibroblasts deficient for Flna. These data suggest that FLNA is one of the important regulators of c-MET signaling and HGF-induced tumor cell migration.

Canonical Wnt signaling is antagonized by noncanonical Wnt5a in hepatocellular carcinoma cells.

BACKGROUND: beta-catenin mutations that constitutively activate the canonical Wnt signaling have been observed in a subset of hepatocellular carcinomas (HCCs). These mutations are associated with chromosomal stability, low histological grade, low tumor invasion and better patient survival. We hypothesized that canonical Wnt signaling is selectively activated in well-differentiated, but repressed in poorly differentiated HCCs. To this aim, we characterized differentiation status of HCC cell lines and compared their expression status of Wnt pathway genes, and explored their activity of canonical Wnt signaling. RESULTS: We classified human HCC cell lines into "well-differentiated" and "poorly differentiated" subtypes, based on the expression of hepatocyte lineage, epithelial and mesenchymal markers. Poorly differentiated cell lines lost epithelial and hepatocyte lineage markers, and overexpressed mesenchymal markers. Also, they were highly motile and invasive. We compared the expression of 45 Wnt pathway genes between two subtypes. TCF1 and TCF4 factors, and LRP5 and LRP6 co-receptors were ubiquitously expressed. Likewise, six Frizzled receptors, and canonical Wnt3 ligand were expressed in both subtypes. In contrast, canonical ligand Wnt8b and noncanonical ligands Wnt4, Wnt5a, Wnt5b and Wnt7b were expressed selectively in well- and poorly differentiated cell lines, respectively. Canonical Wnt signaling activity, as tested by a TCF reporter assay was detected in 80% of well-differentiated, contrary to 14% of poorly differentiated cell lines. TCF activity generated by ectopic mutant beta-catenin was weak in poorly differentiated SNU449 cell line, suggesting a repressive mechanism. We tested Wnt5a as a candidate antagonist. It strongly inhibited canonical Wnt signaling that is activated by mutant beta-catenin in HCC cell lines. CONCLUSION: Differential expression of Wnt ligands in HCC cells is associated with selective activation of canonical Wnt signaling in well-differentiated, and its repression in poorly differentiated cell lines. One potential mechanism of repression involved Wnt5a, acting as an antagonist of canonical Wnt signaling. Our observations support the hypothesis that Wnt pathway is selectively activated or repressed depending on differentiation status of HCC cells. We propose that canonical and noncanonical Wnt pathways have complementary roles in HCC, where the canonical signaling contributes to tumor initiation, and noncanonical signaling to tumor progression.

Increased expression of NKX3.1 in benign prostatic hyperplasia.

OBJECTIVES: To establish the role of the NKX3.1 gene in the development of benign prostatic hyperplasia by comparing the expression of NKX3.1 in messenger ribonucleic acid (mRNA) and protein levels in young adult prostate and BPH tissues. METHODS: Normal prostate tissue samples (n = 4) were obtained from prostate biopsies of patients less than 40 years of age who underwent diagnostic cystoscopy for microscopic hematuria. Benign prostatic hyperplasia tissues (n = 12) were obtained from patients who underwent transurethral prostate resection for bladder outlet obstruction. The RNAs isolated from these tissue samples were analyzed with quantitative reverse transcriptase polymerase chain reaction; the proteins were analyzed with Western blotting and immunohistochemistry. RESULTS: The mean NKX3.1 mRNA transcript expression was 19.17 +/- 3.05 vs 1.24 +/- 1.32 in BPH and normal tissues, respectively, and NKX3.1 protein expression of BPH was approximately 2.4-fold higher than in normal prostate tissue. Reverse transcriptase polymerase chain reaction and Western blot analyses revealed that NKX3.1 gene expression in BPH patient tissues were higher compared with normal prostate tissues. Immunohistochemistry results indicated that most of the BPH tissues stained diffusely, and there was no BPH tissue that lacked NKX3.1 expression. CONCLUSIONS: NKX3.1 expression is elevated in BPH tissues when compared with normal tissues, which may be important in the development of BPH.