Étude comparative de l'angiotensine II et de l'angiotensine IV in vitro au niveau des cellules musculaires lisses vasculaires humaines (caractérisation fonctionnelle des effets de l'angiotensine IV sur le transport de glucose insulino-dépendant)

LYON1-BU.Sciences (692662101) / SudocSudocFranceF

Extracellular matrix remodelling in rat heart in an unpredictable chronic mild stress model associated with diabetes

630-639The coexistence of depression and diabetes is a serious challenge in cardiovascular disease. However, extracellular matrix (ECM) under stress model associated or not with diabetes remains unknown. This study aims to investigate the involvement of diabetes and unpredictable chronic mild stress (UCMS) on ECM remodelling. Rats were exposed to UCMS, diabetes or combined treatment. mRNA expression of matrix metalloproteinases (MMP-2 and MMP-9), plasminogen activator (t-PA) and inhibitor (PAI-1) was examined by Q-RT-PCR. ECM was determined by ELISA. MMP-2 and MMP-9 mRNA was lower in diabetes (P <0.05). UCMS increased MMP-2 and MMP-9 compared to control and diabetic group. Compared to the control and diabetes groups, MMP-2 and MMP-9 mRNA was significantly increased in the combined treatment group. UCMS increased MMP-2 expression in the diabetic group (P <0.01). Compared to control, gelatinase activity was higher in diabetes and UCMS (P <0.05). Combined diabetes and UCMS significantly increased gelatinase activity compared to T2D and UCMS groups. Collagen, hydroxyproline and fibronectin content were significantly increased in diabetes and combined groups. UCMS combined with diabetes exacerbated diabetes-induced MMPs and fibronectin deposition. In conclusion, comorbidity between diabetes and UCMS may exacerbate ECM remodelling. These findings will be useful in understanding diabetes-induced cardiovascular disease

Combined diabetes and chronic stress exacerbates cytokine production and oxidative stress in rat liver and kidney

AbstractInflammatory response and oxidative stress state have been largely described in diabetes and depression separately but not in combination. We aimed to explore the involvement of diabetes and unpredictable chronic mild stress (UCMS) separately or in combination on inflammatory response and oxidative stress. Diabetes, UCMS or combined rat models were used. Proinflammatory cytokines, Interleukin 6 (IL-6) and Tumor necrosis factor alpha (TNF-α) were assessed by real time quantitative polymerase chain reaction (q-PCR) and Enzyme-Linked Immunosorbent Assay (ELISA). Superoxide dismutase (SOD) and catalase (CAT) were determined by colorimetric assays. In the diabetes group, IL-6 mRNA expression increased by 55% and 34% (p < 0.05), respectively, in liver and kidney. UCMS alone or in combination with diabetes increased the mRNA of IL-6, respectively, by 85% and 78% in the liver and by 28% and 63% in the kidney (p < 0.01 for all). ELISA showed that diabetes and UCMS act in synergy on TNF-α and IL-6 expression. Diabetes and UCMS separately or in combination inhibited significantly (p < 0.01) the activities of the two anti-oxidant enzymes when compared to controls. The malondialdehyde (MDA) level was significantly enhanced in the group with diabetes combined with UCMS compared to UCMS alone in both organs. UCMS enhances the proinflammatory cytokines release and induces oxidative stress imbalance, and diabetes comorbidity with depression aggravates the inflammatory response and lipid peroxidation. These observations can be useful to better understand depression-induced organ damage

mTORC1 Directly Phosphorylates and Regulates Human MAF1▿

mTORC1 is a central regulator of growth in response to nutrient availability, but few direct targets have been identified. RNA polymerase (pol) III produces a number of essential RNA molecules involved in protein synthesis, RNA maturation, and other processes. Its activity is highly regulated, and deregulation can lead to cell transformation. The human phosphoprotein MAF1 becomes dephosphorylated and represses pol III transcription after various stresses, but neither the significance of the phosphorylations nor the kinase involved is known. We find that human MAF1 is absolutely required for pol III repression in response to serum starvation or TORC1 inhibition by rapamycin or Torin1. The protein is phosphorylated mainly on residues S60, S68, and S75, and this inhibits its pol III repression function. The responsible kinase is mTORC1, which phosphorylates MAF1 directly. Our results describe molecular mechanisms by which mTORC1 controls human MAF1, a key repressor of RNA polymerase III transcription, and add a new branch to the signal transduction cascade immediately downstream of TORC1

Cellular and Molecular Effect of MEHP Involving LXRa in Human Fetal Testis and Ovary

International audienceBackground: Phthalates have been shown to have reprotoxic effects in rodents and human during fetal life. Previous studies indicate that some members of the nuclear receptor (NR) superfamilly potentially mediate phthalate effects. This study aimed to assess if expression of these nuclear receptors are modulated in the response to MEHP exposure on the human fetal gonads in vitro. Methodology/Principal Findings: Testes and ovaries from 7 to 12 gestational weeks human fetuses were exposed to 1024M MEHP for 72 h in vitro. Transcriptional level of NRs and of downstream genes was then investigated using TLDA (TaqMan Low Density Array) and qPCR approaches. To determine whether somatic or germ cells of the testis are involved in the response to MEHP exposure, we developed a highly efficient cytometric germ cell sorting approach. In vitro exposure of fetal testes and ovaries to MEHP up-regulated the expression of LXRa, SREBP members and of downstream genes involved in the lipid and cholesterol synthesis in the whole gonad. In sorted testicular cells, this effect is only observable in somatic cells but not in the gonocytes. Moreover, the germ cell loss induced by MEHP exposure, that we previously described, is restricted to the male gonad as oogonia density is not affected in vitro. Conclusions/Significance: We evidenced for the first time that phthalate increases the levels of mRNA for LXRa, and SREBP members potentially deregulating lipids/cholesterol synthesis in human fetal gonads. Interestingly, this novel effect is observable in both male and female whereas the germ cell apoptosis is restricted to the male gonad. Furthermore, we presented here a novel and potentially very useful flow cytometric cell sorting method to analyse molecular changes in germ cells versus somatic cells

<i>In vitro</i> exposure to MEHP does not affect the number or apoptosis rate of germ cells in human fetal ovaries.

<p>Ovaries from 7 to 12 week/old human embryos were cultured with or without 10⁻⁴ M MEHP for three days. At the end of the culture period, they were fixed with Bouin’s solution and stained with hematoxylin and eosin. Oogonia density (number of germ cells per mm² tissue) was measured based on the morphological analysis (A) and germ cell apoptosis rate (B) based on the expression of cleaved Caspase-3 (brown) (C). Histograms represent the mean ± SEM of four different ovaries from different fetuses (as indicated in the columns). Arrowheads indicate oogonia and arrows cleaved Caspase- 3 positive oogonia. Bar, 15 µm.</p

Isolation of M2A-positive and –negative cells from human fetal testes by flow cytometric cell sorting.

<p>Histological immunofluorescent staining for M2A (green) and AMH (red) in human fetal testis (A). M2A staining in living dissociated testicular cells (B). DNA was stained with DAPI (blue color in (A) and (B)). Cell sorting profiles of M2A fluorescence in fetal testicular cells incubated with isotype control (C) or anti-M2A antibody (D). The green square (D) represents the cell fraction sorted as M2A-positive. mRNA expression of different cell type markers in M2A-positive and -negative cell fractions (E). <i>AMH</i> and <i>StAR</i> are markers of Sertoli and Leydig cells, respectively, while <i>VASA</i> and <i>M2A</i> characterize the germ cell population. <i>Actin β</i> was used as control. White arrow indicates M2A positive cell, white arrowhead indicates AMH positive cell.</p