35 research outputs found
The effects of streptozotocin-induced diabetes on ghrelin expression in rat testis: biochemical and immunohistochemical study
Introduction. Ghrelin is a hormone which has effects on the secretion of growth hormone, gastrointestinal system, cardiovascular system, cell proliferation and reproductive system. The present study we focused on the relation between ghrelin and GHS-R1a gene expression and the regulation of their expression in the testes of diabetic rats. Material and methods. 40 male Wistar albino rats were divided into four groups: control, and sampled 4, 8 and 12 weeks after induction of diabetes by streptozotocin (STZ) intraperitoneal injection (40 mg/kg). The rats were decapitated under ketamine anesthesia and their testes were removed. Blood was obtained from heart and serum follicle stimulating hormone (FSH), luteinizing hormone (LH), and testosterone levels were measured by ELISA. Tissue ghrelin and GHS-R mRNA levels were determined by qRT-PCR, while ghrelin protein expression was studied by immunohistochemistry. Histopathological damage scores were also assessed. Results. Eight weeks after diabetes induction serum FSH level was increased, whereas LH and testosterone concentrations decreased. The ghrelin and GHS-R1a gene expression and ghrelin immunohistochemistry score first tended to increase after first four weeks of diabetes, and then tended to decrease. Ghrelin-immunopositive cells were detected in Leydig cells in all groups of rats, however, not in the germinal epithelium. Congestion of vessels and hemorrhage, formation of the vacuoles in spermatogonia and spermatocytes, desquamation of spermatids in the lumen and disorganization of seminiferous tubule germinal epithelium were observed in testis of all the diabetic rats. In addition, mean testicular biopsy score and mean seminiferous tubule diameter were getting lower in diabetic animals. Conclusion. Our results suggest that diabetes affects ghrelin expression in rat testis.
Chromatin Central: towards the comparative proteome by accurate mapping of the yeast proteomic environment
High resolution mapping of the proteomic environment and proteomic hyperlinks in fission and budding yeast reveals that divergent hyperlinks are due to gene duplications
The Schizosaccharomyces pombe JmjC-protein, Msc1, prevents H2A.Z localization in centromeric and subtelomeric chromatin domains
Eukaryotic genomes are repetitively packaged into chromatin by nucleosomes, however they are regulated by the differences between nucleosomes, which establish various chromatin states. Local chromatin cues direct the inheritance and propagation of chromatin status via self-reinforcing epigenetic mechanisms. Replication-independent histone exchange could potentially perturb chromatin status if histone exchange chaperones, such as Swr1C, loaded histone variants into wrong sites. Here we show that in Schizosaccharomyces pombe, like Saccharomyces cerevisiae, Swr1C is required for loading H2A.Z into specific sites, including the promoters of lowly expressed genes. However S. pombe Swr1C has an extra subunit, Msc1, which is a JumonjiC-domain protein of the Lid/Jarid1 family. Deletion of Msc1 did not disrupt the S. pombe Swr1C or its ability to bind and load H2A.Z into euchromatin, however H2A.Z was ectopically found in the inner centromere and in subtelomeric chromatin. Normally this subtelomeric region not only lacks H2A.Z but also shows uniformly lower levels of H3K4me2, H4K5, and K12 acetylation than euchromatin and disproportionately contains the most lowly expressed genes during vegetative growth, including many meiotic-specific genes. Genes within and adjacent to subtelomeric chromatin become overexpressed in the absence of either Msc1, Swr1, or paradoxically H2A.Z itself. We also show that H2A.Z is N-terminally acetylated before, and lysine acetylated after, loading into chromatin and that it physically associates with the Nap1 histone chaperone. However, we find a negative correlation between the genomic distributions of H2A.Z and Nap1/Hrp1/Hrp3, suggesting that the Nap1 chaperones remove H2A.Z from chromatin. These data describe H2A.Z action in S. pombe and identify a new mode of chromatin surveillance and maintenance based on negative regulation of histone variant misincorporation.publishedVersio
Roles of H2A.z in Fission Yeast Chromatin
Covalent histone modifications such as methylation, acetylation as well as differential incorporation of histone variants are shown to coincide with different chromatin compartments and mark active or repressed genes. Msc1 is one of the seven JmjC Domain Proteins (JDPs) in Fission Yeast. JDPs are known to function in chromatin and some act as histone demethylases. We found that Msc1 is a member of Swr1 Complex which is known to exchange histone H2A variant H2A.z in nucleosomes. We purified H2A.z as a member of Swr1 Complex and its interaction with Swr1 Complex depends on Swr1. We’ve shown that histone H4 Lysine 20 trimethylation (H4 K20 Me3) is lost in h2A.z and msc1 deletion strains and these strains are sensitive to UV. Deletion strain of h2A.z is sensitive to Camptothecin. Histones H3 and H4 are obtained in Msc1 and H2A.z purifications and we’ve shown that histone H4 from these purifications has low level of Lysine 16 acetylation (H4 K16 Ac). Deletion strains of h2A.z, swr1 and msc1 are shown to be sensitive to TSA, a histone deacetylase (HDAC) inhibitor suggesting that H2A.z cooperates with HDACs. TSA treatment of wild type cells cause an increase in H4 K16 Ac and a decrease in H4 K20 Me3. Gene expression profiles of h2A.z, swr1 and msc1 are significantly similar and upregulated genes in deletion strains localize at chromosome ends (a region of 160 kb for each end). The number of stress or meiotic inducible genes is increased in deletion strains suggesting that H2A.z has a role in regulation of inducible genes. We suggest that H2A.z, in cooperation with HDACs, functions in regulation of chromatin accessibility of inducible promoters
Priming hMSCs with a putative anti-cancer compound, myrtucommulone-a: a way to harness hMSC cytokine expression via modulating PI3K/Akt pathway?
Tumour microenvironment is a key factor for cancer growth and metastasis. Tumour surrounding tissue is known to include high number of mesenchymal stem cells which have been thought to have a role in regulating cancer cell behaviour via paracrine signalling. Therefore, modulating human mesenchymal stem cell (hMSC) secretome is highly significant for controlling and treating disease. Since common therapeutic agents are known to enhance cancer resistance, there is a strong urge to define novel agents for developing cell-based therapies. In the present study, we aimed at investigating the effect of active compounds, myrtucommulone-A (MC-A) and thymoquinone (TQ), on hMSC cytokine expression. Our data revealed that MC-A treatment have significantly altered cytokine expression in hMSCs. Upon MC-A treatment, hMSCs decreased the expression levels of various cytokines including TNF-alpha, VEGF, IL-6, IL-8 and FGF-2. hMSC conditioned medium (CM) primed with MC-A decreased the proliferation, migration ability and clonogenicity of bladder cancer cells and breast cancer cells in comparison to non-primed hMSC medium and hMSC medium primed with TQ. To the best of our knowledge, this study is the first report showing the effects of active compounds, MC-A and TQ, on hMSCs and therefore valuable for highlighting the potential use of active compounds in combination with hMSCs for cell-based targeted cancer therapy
Higher Frequency of rs4977574 (the G Allele) on Chromosome 9p21.3 in Patients with Myocardial Infarction as Revealed by PCR-RFLP Analysis
Single Nucleotide Polymorphisms (SNPs) can genetically predispose individuals for certain diseases and therefore are of clinical significance. Myocardial infarction (MI) was investigated in large genetic association studies revealing novel SNPs associated with MI. rs4977574 is a non-protein coding SNP (A>G) that is located in proximity of cyclin-dependent kinase inhibitor 2A and B genes on chromosome 9p21.3. rs4977574 has been recently found to be associated with the early-onset of MI, and rs4977574 is characterized by a guanine nucleotide (G) instead of an adenine nucleotide (A). rs4977574 has been reported to increase the risk for MI by 28%. In this study, we developed a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method for detecting rs4977574 in Turkish population that consisted of 28 controls without previous MI record and 44 patients with MI. An intergenic genomic region containing the target SNP was amplified by PCR using patient's genomic DNA. Amplified DNA fragments were digested with a restriction enzyme, Hhal that cuts the amplified sequence if only the sequence has GCGC that carries rs4977574. After digestion with Hhal, DNA fragments were visualized in order to detect genotypes. PCR-RFLP revealed that the frequency of rs4977574, the MI-associated allele (G), was 56.8% (25/44) in patients with MI and 33.9% (9.5/28) in controls; the frequency of rs4977574 in patients with MI was significantly higher compared to controls (P = 0.027). Importantly, for the first time in this study, we have developed a novel PCR-RFLP method to detect the presence of rs4977574
Molecular characterization of Acanthamoeba isolated from Kayseri well water
Aim: Potentially pathogenic free-living amoebae have a cosmopolitan distribution in soil, dust, air, and water. Generally, environmental free-living amoebae do not threaten human health. This study aimed to investigate the presence of Acanthamoeba in well waters drawn from different locations in Kayseri, Turkey, which are mainly used for drinking or irrigation by the residents in the region
Canatan H: Pronounced transcriptional regulation of apoptotic and TNF-NF-kappa-B signaling genes during the course of thymoquinone mediated apoptosis in HeLa cells. Mol Cell Biochem 2013, 383(1–2):243–251. doi:10.1186/1471-2407-14-414 Cite this article as
Abstract Thymoquinone (TQ) is the active ingredient extracted from the essential oil of Nigella sativa. A number of studies implicated TQ as an antitumor agent. In this study, cytotoxic effects of the oil of N. sativa and TQ were evaluated on human cervical cancer cell line, HeLa cells
Evaluation of two different adjuvants with immunogenic uroplakin 3A-derived peptide for their ability to evoke an immune response in mice
Rationale: Organ- or tissue-specific antigens produced by normal tissue or by cancer cells could be used in cancer immunotherapy, to target the tumor. In our previous study, we induced T-cell-mediated, bladder-specific autoimmunity by targeting the bladder-specific protein Uroplakin 3A (UPK3A). UPK3A is a well-chosen target for developing an autoimmune response against bladder cancer since the antigen is also expressed in bladder tumors. To use this peptide, which was derived from the UPK3A protein in a bladder cancer vaccine study, it is necessary to induce a strong immune response. In this study, we aimed to develop a robust immune response in BALB/c mice using the well-characterized keyhole limpet hemocyanin (KLH)-conjugated peptide antigen (UPK3A 65-84) conjugated with an immunogenic carrier protein. In combination with the peptide, we used either Freund's complete adjuvant (CFA) or CpG (cytosine-phosphate-guanine oligonucleotides) as effective adjuvants in order to overcome tumor tolerance. Objectives: The immune response evoked by UPK3A 65-84 peptide, using two different adjuvants, was compared by detection of changes in the proliferative response of immune cells, in the cytokine profile, and in the immune cell populations. Findings: We demonstrated that CpG, combined with KLH-UPK3A 65-84, promoted a more robust immune response, via induction of higher IL-2, IFN-gamma, TNF-beta, IL-17 production and activation of more immune cells (CD4(+) T cells, CD8(+) T cells, NK cells CD11b, CD45), than CFA and the KLH-UPK3A 65-84. Conclusion: CpG as an adjuvant combined with KLH-UPK3A 65-84 could be used in preclinical models of bladder cancer for the development of cancer immunotherapy strategies
Pronounced transcriptional regulation of apoptotic and TNF-NF-kappa-B signaling genes during the course of thymoquinone mediated apoptosis in HeLa cells
Thymoquinone (TQ) is the active ingredient extracted from the essential oil of Nigella sativa. A number of studies implicated TQ as an antitumor agent. In this study, cytotoxic effects of the oil of N. sativa and TQ were evaluated on human cervical cancer cell line, HeLa cells. IC50 value was similar to 0.125 mu l/ml for N. sativa oil preparations and 12.5 mu M for TQ. TQ strongly inhibited wound healing at all concentrations ranging from 12.5 to 100 mu M in a scratch wound healing assay. Additionally, induction of apoptosis by TQ was assessed by Giemsa staining and TQ was found to induce apoptosis in cancer cells especially at concentrations of 50 and 100 mu M. TQ-mediated transcriptional regulation of 84 genes involved in apoptosis was studied using a PCR array. At low dose (12.5 mu M), TQ was found to induce expression of four pro-apoptotic genes: BIK (similar to 22.7-fold), FASL (similar to 2.9-fold), BCL2L10 (similar to 2.1-fold), and CASP1 (similar to 2-fold). TQ was also found to reduce the expression of an anti-apoptotic gene implicated in NF-kappa-B signaling and cancer: RELA (similar to 8-fold). At high dose (100 mu M), TQ mediated the expression of 21 genes implicated directly in apoptosis (6 genes), TNF signaling (10 genes), and NF-kappa-B signaling (3 genes) such as BIK, BID, TNFRSF10A, TNFRSF10B, TNF, TRAF3, RELA, and RELB. In conclusion, this study implicates the role of TQ in the inhibition of cancer cell proliferation and migration. At the same time, our results strongly suggest that TQ intervenes with TNF and NF-kappa-B signaling during TQ-mediated induction of apoptosis in cancer cells. Thymoquinone (TQ) is the active ingredient extracted from the essential oil of Nigella sativa. A number of studies implicated TQ as an antitumor agent. In this study, cytotoxic effects of the oil of N. sativa and TQ were evaluated on human cervical cancer cell line, HeLa cells. IC50 value was ~0.125 μl/ml for N. sativa oil preparations and 12.5 μM for TQ. TQ strongly inhibited wound healing at all concentrations ranging from 12.5 to 100 μM in a scratch wound healing assay. Additionally, induction of apoptosis by TQ was assessed by Giemsa staining and TQ was found to induce apoptosis in cancer cells especially at concentrations of 50 and 100 μM. TQ-mediated transcriptional regulation of 84 genes involved in apoptosis was studied using a PCR array. At low dose (12.5 μM), TQ was found to induce expression of four pro-apoptotic genes: BIK (~22.7-fold), FASL (~2.9-fold), BCL2L10 (~2.1-fold), and CASP1 (~2-fold). TQ was also found to reduce the expression of an anti-apoptotic gene implicated in NF-kappa-B signaling and cancer: RELA (~8-fold). At high dose (100 μM), TQ mediated the expression of 21 genes implicated directly in apoptosis (6 genes), TNF signaling (10 genes), and NF-kappa-B signaling (3 genes) such as BIK, BID, TNFRSF10A, TNFRSF10B, TNF, TRAF3, RELA, and RELB. In conclusion, this study implicates the role of TQ in the inhibition of cancer cell proliferation and migration. At the same time, our results strongly suggest that TQ intervenes with TNF and NF-kappa-B signaling during TQ-mediated induction of apoptosis in cancer cells.</p