Identifikasi dan Purifikasi Vitelogenin Arwana Asia: Banjar, Pinoh (Scleropages macrocephalus), Papua (Scleropages jardinii) dan Super Red (Scleropages legendrei)

Arowana fish (Scleropages sp.) are monomorphic species, those animals that physically could not be distinguished between male and female. The research was aimed to identify and purify Vitellogenin of four variants of Asian Arowana: Banjar, Papua, Pinoh, and Super red. 12 fishes, 3 from each variant were given estradiol stimulation through toad for vitellogenin production purpose. The SDS-PAGE results expressed that there were two types of Asian Arowana vitellogenin characters, single Vtg for Arowana Banjar, Pinoh (Scleropages macrocephalus) and Super Red (Scleropages legendrei) with a molecular weight of 180 kDa and double Vtg in Papuan Arowana (Scleropages jardinii) with molecular weight 180 and 110 kDa. Pure vitellogenin has been collected from 3 varieties, Banjar, Papua, and Super red with a concentration range between 0.1 - 0.67 mg / mL. The actual concentration is believed to be greater than the measured concentration.

Quantitative Proteomics of Epigenetic Histone Modifications in MCF-7 cells under Estradiol Stimulation

Estrogen exposure has already been considered to be associated with tumorigenesis and breast cancer progression. To study the epigenetic regulation mechanism in MCF-7 cells under estrogen exposure, which normally results...

SUN-551 Impaired Vascular Relaxation and Altered eNOS Regulation in Boys with Hypospadias

Background: Sex hormones influence vascular function. Whether boys with hypospadias who have insufficient androgen exposure during the masculinisation programming window have altered vascular function is unknown. Objective: To investigate whether vascular function is impaired in boys with hypospadias and to explore the putative role of eNOS. Methods: Peripheral arteries from excess foreskin tissue obtained from boys undergoing hypospadias repair (cases) or circumcision (controls) were used. Vascular function was assessed by myography. mRNA expression was measured by qPCR in vascular smooth muscle cells (VSMCs). Nitric oxide (NO) was measured by DAF fluorescence assay and peroxynitrite levels measured via ELISA. Results: 23 boys with hypospadias and 34 age-matched controls were studied. There were 18 (52%) cases of distal, 7 (22%) of midshaft and 9 (26%) of proximal hypospadias and none of them had biochemical evidence of hypogonadism or a variant in AR. Clinical cardiometabolic parameters were similar between groups. Endothelium-dependent relaxation to acetylcholine (ACh) and endothelium-independent relaxation to sodium nitroprusside (SNP) were reduced in arteries from cases vs controls (Emax %U46619: 72.4 vs 1.2, p<0.0001 and Emax %U46619: (42.7 vs 11.8, p<0.01 respectively). Incubation with the NO synthase inhibitor, L-NAME (1x10-5 M) worsened endothelial-dependent relaxation in controls (Emax % U46619: 76.8 vs 1.2, p<0.0001) but had no effect in cases (Emax % U46619:60.6 vs 72.4, p=0.3). Testosterone (1x10-7 M) ameliorated vascular relaxation (p<0.05), whereas17[[Unsupported Character - Symbol Font 𝝱]];-estradiol stimulation (1x10-9 M) did not. In cultured VSMCs, mRNA expression of eNOS and iNOS was reduced whereas that of nNOS was increased in cases versus controls. Nitric oxide production was reduced in cases (5 fold, p<0.01), as was peroxynitrite production (0.5 fold, p<0.05). Testosterone increased expression of eNOS in VSMCs. There was no difference in mRNA expression of the AR and GPRC6A but cases had increased expression of ESR1 (2.71 fold), ESR2 (2.63 fold) and GPR30 (2.86 fold) (p<0.05). Conclusion: Arteries in eugonadal boys with hypospadias show vascular dysfunction which involves impaired NOS/NO regulation effects that are ameliorated with testosterone but not oestrogen. These processes may predispose to long-term cardiovascular disease.

Estrogen Regulation of the Glucuronidation Enzyme UGT2B15 in Estrogen Receptor-Positive Breast Cancer Cells

Estrogens and androgens influence many properties of breast cancer cells; hence, regulation of local estrogen and androgen levels by enzymes involved in steroid hormone biosynthesis and metabolism would impact signaling by these hormones in breast cancer cells. In this study, we show that the UDP-glucuronosyltransferase (UGT) enzyme UGT2B15, a member of the UGT family of phase II enzymes involved in the glucuronidation of steroids and xenobiotics, is a novel, estrogen-regulated gene in estrogen receptor (ER)-positive human breast cancer cells (MCF-7, BT474, T47D, and ZR-75). UGT2B15 is the only UGT2B enzyme up-regulated by estrogen, and marked estradiol stimulation of UGT2B15 mRNA levels is observed, in a time- and dose-dependent manner. UGT2B15 stimulation by estradiol is blocked by the antiestrogen ICI182,780, but not by the translational inhibitor cycloheximide, indicating that UGT2B15 is likely a primary transcriptional response mediated through the ER. UGT2B15 up-regulation is also evoked by other estrogens (propylpyrazoletriol, genistein) and by the androgen 5α-dihydrotestosterone working through the ER, but not by other steroid hormone receptor ligands. Western blot and immunocytochemical analyses with several UGT2B-specific antibodies we have designed and steroid glucuronidation assays indicate a large increase in both cellular UGT2B15 protein and enzyme activity after estrogen treatment. Due to the important role of UGT enzymes in forming conjugates between steroids and glucuronic acid, thereby inactivating them and targeting them for removal, the estrogen-induced up-regulation of UGT2B15 might have a significant moderating effect on estrogen and androgen concentrations, thereby reducing their signaling in breast cancer cells.