The 14-Kilodalton Human Growth Hormone Fragment a Potent Inhibitor of Angiogenesis and Tumor Metastasis.

The 14-kilodalton human growth hormone (14 kDa hGH) N-terminal fragment derived from the proteolytic cleavage of its full-length counterpart has been shown to sustain antiangiogenic potentials. This study investigated the antitumoral and antimetastatic effects of 14 kDa hGH on B16-F10 murine melanoma cells. B16-F10 murine melanoma cells transfected with 14 kDa hGH expression vectors showed a significant reduction in cellular proliferation and migration associated with an increase in cell apoptosis in vitro. In vivo, 14 kDa hGH mitigated tumor growth and metastasis of B16-F10 cells and was associated with a significant reduction in tumor angiogenesis. Similarly, 14 kDa hGH expression reduced human brain microvascular endothelial (HBME) cell proliferation, migration, and tube formation abilities and triggered apoptosis in vitro. The antiangiogenic effects of 14 kDa hGH on HBME cells were abolished when we stably downregulated plasminogen activator inhibitor-1 (PAI-1) expression in vitro. In this study, we showed the potential anticancer role of 14 kDa hGH, its ability to inhibit primary tumor growth and metastasis establishment, and the possible involvement of PAI-1 in promoting its antiangiogenic effects. Therefore, these results suggest that the 14 kDa hGH fragment can be used as a therapeutic molecule to inhibit angiogenesis and cancer progression.

miRNA implication in the pathogenesis and the outcome of Tunisian endemic pemphigus foliaceus.

Pemphigus foliaceus (PF) is a bullous autoimmune skin disease diagnosed through sera and skin analyses. PF severity is associated with maintained anti-Dsg1 sera levels and its prognosis is unpredictable. MicroRNA (miRNA), dynamic regulators of immune function, have been identified as potential biomarkers for some autoimmune diseases. This study aimed to assess the miRNA expression of miR-17-5p, miR-21-5p, miR-146a-5p, miR-155-5p and miR-338-3p using quantitative real-time PCR in peripheral blood mononuclear cells (PBMC) and lesional skin samples from untreated and treated PF patients (both remittent and chronic) over 3 months. Overall, miRNA expression was significantly higher in PBMC than in biopsy samples. Blood miR-21 expression was increased in untreated patients compared to controls and had a diagnostic value with an AUC of 0.78. After 6 weeks, it decreased significantly, similar to anti-Dsg1 antibodies and the PDAI score. In addition, a positive correlation was observed between cutaneous miR-21 expression and the disease activity score. Conversely, cutaneous expressions of miR-17, miR-146a and miR-155 were significantly higher in treated chronic patients compared to remittent ones. The cutaneous level of miR-155 positively correlated with pemphigus activity, making it a potential predictive marker for patients' clinical stratification with an AUC of 0.86.These findings suggest that blood miR-21 and cutaneous miR-155 can be used as supplemental markers for PF diagnosis and activity, respectively in addition to classical parameters.

Nonstop mutation in the Kisspeptin 1 receptor (KISS1R) gene causes normosmic congenital hypogonadotropic hypogonadism.

PURPOSE: Congenital hypogonadotropic hypogonadism (CHH) is a rare genetic disorder mostly characterized by gonadotropins release and/or action deficiencies. Both isolated (idiopathic hypogonadotropic hypogonadism) and syndromic (Kallmann) forms are identified depending on the olfactory ability. Clinical and genetic heterogeneities of CHH have been widely explored, thus improving our understanding of the disease's pathophysiology. This work aims to (1) provide a detailed clinical and hormonal description of normosmic CHH patients and (2) identify the mutation linked to the studied phenotype. PARTICIPANTS AND METHODS: We investigated three affected patients with normosmic CHH, belonging to a consanguineous Tunisian family. Patients underwent an insulin-induced hypoglycemia test. We performed whole exome sequencing to identify the causal mutation. RESULTS: At first diagnosis, a total gonadotropic deficiency was identified in all patients. The insulin-induced hypoglycemia test has also revealed a reduced cortisol secretion and complete growth hormone deficiency. At 20.8 years, one female exhibited a spontaneous recovery of the hypothalamic-pituitary-adrenal axis function, unlike her affected siblings who still depend on corticosteroid replacement therapy. Herein, we identified a novel homozygous nonstop mutation (c.1195T>C) in KISS1R gene in all affected subjects. This mutation led to the substitution of the physiologic stop codon by an arginine (p.X399R). CONCLUSIONS: Our study highlights the importance of the KISS1R signaling, in gonadotropin-releasing hormone neurons, in the control of reproductive function. Additionally, our data suggests a complex central and peripheral metabolic control of puberty, through the hypothalamic KISS1R signaling. We suggest a mutual link between the hypothalamic-pituitary-gonadal, -adrenal, and -somatotropic axes.

Genotyping of Tunisian azoospermic men with Sertoli cell-only and maturation arrest.

Azoospermia factor (AZF) deletions were associated with severe oligospermia and azoospermia with testicular histologies varying from maturation arrest (MA) to Sertoli cell-only (SCO) phenotypes. Abnormal androgen receptor (AR) structure or function has also been implicated in male infertility. To assess the contribution of these genetic defects to azoospermic patients, 19 Tunisian men with SCO (n = 13) or MA (n = 6) were enrolled in this study. Using immunohistochemistry method, we evaluated the expression of AR in testicular biopsy for the two phenotypes. PCR with primers flanking the AR-(CAG)n region and direct sequencing were used to determine AR-(CAG)n length. And PCR amplification of 14 sequence-tagged sites (STSs) located at Yq was used to determine the rate and extent of Y microdeletions. We found a significant difference of the AR expression between SCO and MA cases. Hence, this expression in the testis depends on the status of spermatogenesis. However, we did not find any relationship between the (CAG) repeat and the testicular histology (neither for SCO nor MA). On the other hand, we found a high frequency of AZF deletions (46.2%) in SCOS and in MA (50%). The present results also suggest the contribution of Y chromosome microdeletions in SCO and MA pathogenesis.

SLC26A4 expression among autoimmune thyroid tissues.

CONTEXT: The PDS gene (SLC26A4) is responsible for Pendred syndrome (PS). Genetic analysis of PDS using Tunisian samples showed evidence for linkage and association with autoimmune thyroid diseases (AITD) emergence. In addition, the PDS gene product, pendrin, was recently identified as a novel autoantigen in Graves' disease (GD) or Hashimoto thyroiditis (HT) patients' sera. OBJECTIVE: The aim of this study was to quantify the PDS gene expression and to evaluate the pendrin in vivo and in vitro immunolocalisation. PATIENTS: A total of 52 thyroid gland tissue samples (22 GD, 11 HT, 5 multinodular goiter (MNG), 3 normal thyroid tissues, 8 papillary thyroid carcinoma (PTC), 1 follicular thyroid carcinoma (FTC) and 2 medullar thyroid carcinoma (MTC)) were explored. METHOD: PDS and pendrin expression levels were determined using quantitative RT-PCR and immuno-detection methods. TSH and thyroglobulin (Tg) effects on pendrin expression were investigated by immunofluorescence on primary cell culture from GD thyroid tissues. RESULTS: The relative quantification using PDS transcript level among GD thyroid tissues was increased compared to normal thyroid tissues used as calibrator (mean: 27.17-fold higher than normal thyroid tissues). However, thyroids with HT, carcinoma and MNG showed a decrease expression level (means: 92.05-, 77.68-, 14.3-fold lower than normal thyroid tissues, respectively). These results were confirmed by immunoanalysis. Immunofluorescence results showed an apical and a cytoplasmic pendrin localisation on GD thyroid tissues and a marked pendrin expression reduction on HT thyroid tissues. GD primary cell cultures under TSH and Tg stimulation showed a trafficking improvement of pendrin apical localisation. CONCLUSIONS: Our data point to the presence of a relation between SLC26A4 expression in AITD and thyroid function.

Polymorphisms in the MUC16 gene: potential implication in epithelial ovarian cancer.

MUC16 plays an important role in epithelial ovarian cancer. In this paper, we studied the association between two tags SNPs of MUC16 and the risk of epithelial ovarian cancer. We aimed also to test the association between these tags SNPs and elevated level of the protein CA125. We analyzed a collection of 117 cases. Forty-one samples of patients with epithelial ovarian cancer and 76 samples from Tunisian volunteers were genotyped for two synonymous coding tags SNPs of the MUC16 gene (rs1596797, A/C and rs2547065, C/G) using polymerase chain reaction and sequencing. For the rs1596797 SNP, there was no significant difference in genotype distribution, a rare variation observed in only one patient. For the polymorphism rs2547065, mean CA125 levels were 24 and 78 UI/ml in patients with GG and GC genotypes versus 230 UI/ml in patients with CC genotype (P = 0.36). Compared to the C/C genotype, the 'G' allele (C/G+G/G genotypes) did not significantly modified the risk of developing epithelial ovarian cancer (OR = 0.43; 95% CI). As for the polymorphism rs1596797, compared to the C/C genotype, the 'A' allele (C/A+A/A genotypes) did not significantly modified the risk of developing epithelial ovarian cancer (OR = 881.7; 95% CI). MUC16 gene polymorphisms selected in this study are neither involved in genetic predisposition to epithelial ovarian cancer nor associated with CA125 level.

Absence of anti-pendrin auto-antibodies in the sera of Tunisian patients with autoimmune thyroid diseases.

BACKGROUND: We previously demonstrated that the PDS gene is involved in the genetic susceptibility to autoimmune thyroid diseases (AITD) in Tunisia. In the same population, we now investigated the presence of anti-pendrin auto-antibodies (aAbs) in AITD patients' sera. METHODS: Thirty seven Tunisian AITD patients and 19 healthy subjects from families previously linked to the PDS gene, 75 unrelated patients and 20 healthy unrelated subjects were included in our study. The detection of anti-pendrin aAbs in patients' sera was performed by ELISA using membrane protein extracts of CHO cells expressing pendrin (CHO-hPDS) and by immunofluorescence using transient COS-7 cells expressing a GFP tagged pendrin. CHO cells transfected with human TPO in the same ELISA conditions were used as positive control. RESULTS: The majority of AITD patients' sera were positive for the presence of anti-TPO aAbs. In contrast, no reactivity was detected with CHO-hPDS membrane protein extracts. Likewise, no significant immunostaining was found on transfected COS-7cells upon exposure to patients' and controls' sera. CONCLUSIONS: Our data point to the absence of anti-pendrin aAbs in Tunisian AITD patients' sera.

Genetic investigation of FOXE1 polyalanine tract in thyroid diseases: new insight on the role of FOXE1 in thyroid carcinoma.

FOXE1 polyalanine tract (poly-Ala) has been associated with thyroid dysgenesis. Recently, the SNP (rs1867277:-238G>A) within the FOXE1 5'UTR was involved in the genetic susceptibility to thyroid cancer (TC). In the aim to assess the influence of FOXE1 poly-Ala length on the genetic susceptibility to TC and autoimmune thyroid diseases (AITD), a case-control design (including 261 Tunisian AITD, 170 Spanish TC and respectively 171 and 218 matched healthy subjects) was performed. The effect of Ala length and rs1867277 alleles on FOXE1 expression was investigated by mRNA relative real time quantification on 8 papillary thyroid carcinoma (PTC) and 10 Graves' disease (GD) genotyped thyroid biopsies. The fluorescent genotyping of poly-Ala polymorphism revealed nine alleles (from 12 to 22 repetitions). The association of poly-Ala polymorphism with AITD was rejected (Pc>0.05). However, a significant association was found with TC. In addition, the genotypic distributions revealed the predispositional effect of the 16/16 genotype (OR = 2.71; 95%CI: 1.36-5.42; p=0.001) and the protector effect of the 14/14 genotype (OR= 0.46; 95%CI: 0.29-0.72; p=0.003). The quantification studies reveal that FOXE1 transcripts were less abundant in PTC than GD samples. Moreover, FOXEI gene was 4,8 fold less expressed among PTC protected patients compared to homozygous 16/16-A/A. In conclusion, by exploring the poly-Ala polymorphism, we confirmed the involvement of {\it FOXE1} gene in the genetic susceptibility to TC and we reported its down expression among PTC tissues.