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Short-chain fatty acids (SCFAs) play a pivotal role in regulating the proliferation and development of bovine rumen epithelial cells (BRECs). G protein-coupled receptor 41 (GPR41) is involved in the signal transduction in BRECs as a receptor for SCFAs. Nevertheless, the impact of GPR41 on the proliferation of BRECs has not been reported. The results of this research showed that the knockdown of GPR41 (GRP41KD) decreased BRECs proliferation compared with the wild-type BRECs (WT) (p < 0.001). The RNA sequencing (RNA-seq) analysis showed that the gene expression profiles differed between WT and GPR41KD BRECs, with the major differential genes enriched in phosphatidylinositol 3-kinase (PIK3) signaling, cell cycle, and amino acid transport pathways (p < 0.05). The transcriptome data were further validated by Western blot and qRT-PCR. It was evident that the GPR41KD BRECs downregulated the level of the PIK3-Protein kinase B (AKT)-mammalian target of the rapamycin (mTOR) signaling pathway core genes, such as PIK3, AKT, eukaryotic translation initiation factor 4E binding protein 1 (4EBP1) and mTOR contrasted with the WT cells (p < 0.01). Furthermore, the GPR41KD BRECs downregulated the level of Cyclin D2 p < 0.001) and Cyclin E2 (p < 0.05) compared with the WT cells. Therefore, it was proposed that GPR41 may affect the proliferation of BRECs by mediating the PIK3-AKT-mTOR signaling pathway.

Aldose Reductase Inhibitor Fidarestat Prevents Retinal Oxidative Stress and Vascular Endothelial Growth Factor Overexpression in Streptozotocin-Diabetic Rats Irina G. Obrosova 1 , Alexander G. Minchenko 2 , Rukmini Vasupuram 1 , Lauren White 1 , Omorodola I. Abatan 1 , Arno K. Kumagai 1 , Robert N. Frank 3 and Martin J. Stevens 1 1 Division of Endocrinology and Metabolism, Department of Internal Medicine, University of Michigan Medical Center, Ann Arbor, Michigan 2 Department of Anesthesiology, Thomas Jefferson University, Philadelphia, Pennsylvania 3 Department of Ophthalmology, Kresge Eye Institute, Wayne State University School of Medicine, Detroit, Michigan Abstract The study addressed the role for aldose reductase (AR) in 1 ) retinal oxidative stress and vascular endothelial growth factor (VEGF) overexpression in early diabetes, and 2 ) high glucose-induced oxidative stress in retinal endothelial cells. In vivo experiments were performed on control rats and diabetic rats treated with or without low or high dose of the AR inhibitor (ARI) fidarestat (2 or 16 mg · kg −1 · day −1 ). In vitro studies were performed on bovine retinal endothelial cells (BREC) cultured in either 5 or 30 mmol/l glucose with or without 1 μmol/l fidarestat. Intracellular reactive oxygen species were assessed using the 5-(and-6)-chloromethyl-2′,7′-dichlorodihydrofluorescein diacetate (H 2 DCFDA) probe and flow cytometry. Both low and high doses of fidarestat (i.e., the doses that partially and completely inhibited sorbitol pathway hyperactivity) arrested diabetes-induced retinal lipid peroxidation. This was achieved due to upregulation of the key antioxidative defense enzyme activities rather than changes in reduced glutathione, oxidized glutathione, ascorbate and dehydroascorbate concentrations, and the glutathione and ascorbate redox states. Diabetes-associated 2.1-fold VEGF protein overexpression (enzyme-linked immunosorbent assay; ELISA) was dose-dependently prevented by fidarestat, whereas total VEGF mRNA and VEGF-164 mRNA (RT-PCR) abundance were not affected by either diabetes or the ARI. In BREC, fidarestat corrected hyperglycemia-induced increase in H 2 DCFDA fluorescence but not oxidative stress caused by three different pro-oxidants in normoglycemic conditions. In conclusion, increased AR activity contributes to retinal oxidative stress and VEGF protein overexpression in early diabetes. The findings justify the rationale for evaluation of fidarestat on diabetic retinopathy. Footnotes Address correspondence and reprint requests to Dr. Irina G. Obrosova, Division of Endocrinology and Metabolism, Department of Internal Medicine, University of Michigan Medical Center 1150 West Medical Center Drive, MSRB II, Rm 5570, Ann Arbor, MI 48109-0678. E-mail: iobrosso{at}umich.edu . Received for publication 6 August 2001 and accepted in revised form 12 December 2002. AA, ascorbate; AGE, glycation end products; AR, aldose reductase; ARI, AR inhibitor; BSO, l -buthionine(S,R)-sulfoximine; BREC, bovine retinal endothelial cells; DHAA, dehydroascorbate; DMEM, Dulbecco’s modified Eagle’s medium; DR, diabetic retinopathy; ELISA, enzyme-linked immunosorbent assay; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; GSH, reduced glutathione; GSHPx, glutathione peroxidase; GSSG, oxidized glutathione; GSSGRed, glutathione reductase; GSSGTrans, glutathione transferase; 4-HA, 4-hydroxyalkenals; MDA, malondialdehyde; PKC, protein kinase C; ROS, reactive oxygen species; SOD, superoxide dismutase; VEGF, vascular endothelial growth factor. DIABETES

Aim: To assess the effect of crystalline triamcinolone acetonide on retinal endothelial cell proliferation in vivo and in vitro. Methods: For in vitro analysis, a sprouting assay was employed. Bovine retinal endothelial cells were stimulated with basic fibroblast growth factor (bFGF) and incubated with different concentrations of triamcinolone acetonide (0.05 mg/ml to 8 mg/ml). For in vivo analysis, a retinopathy of prematurity (ROP) model was used. 16 C57BL/J6 mice were exposed to 75% oxygen from postnatal day 7 to day 12. On day 12, triamcinolone acetonide was intravitreally injected into one eye (“study eye”) and isotonic saline into the contralateral eye (“control eye”). On day 17, the mice were sacrificed and the eyes removed for quantitative analysis of preretinal neovascularisation. Four non-exposed mice served as negative control. Results: The sprouting assay demonstrated a dose dependent inhibition of bovine retinal endothelial cell proliferation from 0.05 mg triamcinolone acetonide/ml (no inhibition) to 3 mg triamcinolone acetonide/ml (complete inhibition). Dosages of more than 2 mg/ml resulted in cytotoxic changes of endothelial cells. The ROP model demonstrated a significantly lower neovascular cell count of 58% in the study group compared to the control group (6.35 (SD 2.1) cells per histological section versus 14.9 (SD 5.3) cells; p<0.005). Conclusions: Triamcinolone acetonide inhibits bFGF induced proliferation of retinal endothelial cells in vivo and in vitro. These findings contribute to understanding the mode of action and effects of triamcinolone acetonide on retinal neovascularisation.

Accumulating evidence points to a causal role for advanced glycation end products (AGEs) in the development of diabetic vascular complications, including retinopathy. Possible pathogenic mechanisms linking AGEs to diabetic retinopathy include protein kinase C (PKC) activation, oxidative stress, and vascular endothelial growth factor (VEGF) expression. In the present study, we investigated the effect of AGEs on VEGF expression in bovine retinal endothelial cells (BRECs) and determined the role of PKC and oxidative stress in this effect. Incubation of BRECs with AGEs led to enhanced VEGF mRNA and protein expression. This treatment also induced PKC translocation in these cells. The AGE-induced increases in VEGF expression and PKC activation were inhibited by the pan-specific PKC inhibitor, calphostin C, and by the antioxidant drug and compounds, gliclazide, N-acetylcysteine, and vitamin E. In contrast, glyburide which does not exhibit antioxidant properties, did not affect the AGE-induced VEGF expression. Exposure of BRECs to AGEs resulted in a significant increase of nuclear protein binding to the NF-κB consensus sequence of the VEGF promoter region. Induction of DNA binding activity for NF-κB by AGEs was prevented by gliclazide. Treatment of BRECs with AGEs also increased the proliferation of these cells. This effect was abrogated by incubating the cells with an anti-VEGF antibody and was inhibited in the presence of gliclazide. Overall, these data demonstrate that AGEs increase VEGF expression in retinal endothelial cells through generation of oxidative stress and downstream activation of the PKC pathway. Targeting VEGF expression with specific pharmacological agents, such as antioxidants and PKC inhibitors, may prove efficacious for the treatment of diabetic retinopathy.

According to the N.C. Cooperative Extension, which has an office at the New Hanover County Arboretum, several hollies and a few species of cactuses thrive in the area.

Dorothie M. Pierce ROLLING MEADOWS - Dorothie M. Pierce (nee French), 93, of Arlington Heights, formerly of Rolling Meadows, died Sept. 24, 2016. Born Oct. 3, 1922, in Huron, South Dakota.

\"Our set is supposedly haunted,\" she explains. \"So one day after [Coy Stewart] went home, Haley Tju [who plays Pepper] and I made it look like a ghost had been in his dressing room. We even got Coy's parents in on it.\" \"Bella didn't really have a love interest in season l for more than an episode,\" she says. \"It would be cool if she found someone to keep around for a while.\"

Of the 500 Methodist churches in the New York Annual Conference, which includes western Connecticut, the Hudson River Valley and Long Island, about one church per year is closed, [Pastor Dennis Winkleblack] said. \"Hopefully, in the next year we will come to a decision as to what we are going to do,\" said Winkleblack, who took the part-time job after lay leaders decided not to pay a full-time clergyman to supervise the church's affairs. Winkleblack, who is also part-time assistant to Jeremiah Park, resident bishop of the New York Annual Conference, said this was the first time in more than 100 years that the church has not had a full-time pastor. [Elaine Deysenroth] and Winkleblack said a merger agreement with another church is still possible, but [Brec Morgan] remains doubtful.