Activation of TAS2R4 signaling attenuates podocyte injury induced by high glucose.

Bitter taste receptors (TAS2Rs) Tas2r108 gene possesses a high abundance in mouse kidney; however, the biological functions of Tas2r108 encoded receptor TAS2Rs member 4 (TAS2R4) are still unknown. In the present study, we found that mouse TAS2R4 (mTAS2R4) signaling was inactivated in chronic high glucose-stimulated mouse podocyte cell line MPC, evidenced by the decreased protein expressions of mTAS2R4 and phospholipase C ß2 (PLCß2), a key downstream molecule of mTAS2R4 signaling. Nonetheless, agonism of mTAS2R4 by quinine recovered mTAS2R4 and PLCß2 levels, and increased podocyte cell viability as well as protein expressions of ZO-1 and nephrin, biomarkers of podocyte slit diaphragm, in high glucose-cultured MPC cells. However, blockage of mTAS2R4 signaling with mTAS2R4 blockers γ-aminobutyric acid and abscisic acid, a Gßγ inhibitor Gallein, or a PLCß2 inhibitor U73122 all abolished the effects of quinine on NLRP3 inflammasome and p-NF-κB p65 as well as the functional podocyte proteins in MPC cells in a high glucose condition. Furthermore, knockdown of mTAS2R4 with lentivirus-carrying Tas2r108 shRNA also ablated the effect of quinine on the key molecules of the above inflammatory signalings and podocyte functions in high glucose-cultured MPC cells. In summary, we demonstrated that activation of TAS2R4 signaling alleviated the podocyte injury caused by chronic high glucose, and inhibition of NF-κB p65 and NLRP3 inflammasome mediated the protective effects of TAS2R4 activation on podocytes. Moreover, activation of TAS2R4 signaling could be an important strategy for prevention and treatment of diabetic kidney disease.

Piperine, as a TAS2R14 agonist, stimulates the secretion of glucagon-like peptide-1 in the human enteroendocrine cell line Caco-2.

Piperine is reported to ameliorate common metabolic diseases, however, its molecular mechanism is still unclear. In the present study, we examined whether piperine could stimulate glucagon-like peptide-1 (GLP-1) secretion in a human enteroendocrine cell line, Caco-2, and explored the potential mechanisms from the activation of human bitter taste receptors (TAS2Rs). It was found that TAS2R14 was highly expressed in Caco-2 cells, far more than TAS2R4 and TAS2R10. Piperine and flufenamic acid (FA, a known TAS2R14 agonist) markedly increased intracellular calcium mobilization and significantly enhanced the GLP-1 secretion, accompanied by elevated levels of proglucagon mRNA in Caco-2 cells compared with the control. Moreover, piperine and FA activated TAS2R14 signaling as evidenced by the increased mRNA and protein levels of TAS2R14, and the protein expression of its downstream key molecules including phospholipase C ß2 (PLCß2) and a transient receptor potential channel melastatin 5 (TRPM5). On the other hand, a G protein ßγ subunit inhibitor Gallein or a PLC inhibitor U73122 alleviated piperine-stimulated GLP-1 secretion in Caco-2 cells. In the meantime, a flavanone hesperetin significantly attenuated piperine and FA induced the intracellular calcium mobilization and GLP-1 secretion. Furthermore, TAS2R14 knockdown reversed the piperine-triggered up-regulation of PLCß2 and TRPM5 as well as increased the GLP-1 secretion in Caco-2 cells by TAS2R14 shRNA transfection. In summary, our findings demonstrated that piperine promoted the GLP-1 secretion from enteroendocrine cells through the activation of TAS2R14 signaling. Moreover, TAS2R14 was likely a target of piperine in the alleviation of metabolic diseases.

Lack of miRNA-17 family mediates high glucose-induced PAR-1 upregulation in glomerular mesangial cells.

Upregulation of thrombin receptor protease-activated receptor 1 (PAR-1) is verified to contribute to chronic kidney diseases, including diabetic nephropathy; however, the mechanisms are still unclear. In this study, we investigated the effect of PAR-1 on high glucose-induced proliferation of human glomerular mesangial cells (HMCs), and explored the mechanism of PAR-1 upregulation from alteration of microRNAs. We found that high glucose stimulated proliferation of the mesangial cells whereas PAR-1 inhibition with vorapaxar attenuated the cell proliferation. Moreover, high glucose upregulated PAR-1 in mRNA level and protein expression while did not affect the enzymatic activity of thrombin in HMCs after 48 h culture. Then high glucose induced PAR-1 elevation was likely due to the alteration of the transcription or post-transcriptional processing. It was found that miR-17 family members including miR-17-5p, -20a-5p, and -93-5p were significantly decreased among the eight detected microRNAs only in high glucose-cultured HMCs, but miR-129-5p, miR-181a-5p, and miR-181b-5p were markedly downregulated in both high glucose-cultured HMCs and equivalent osmotic press control compared with normal glucose culture. So miR-20a was selected to confirm the role of miR-17 family on PAR-1 upregulation, finding that miR-20a-5p overexpression reversed the upregulation of PAR-1 in mRNA and protein levels induced by high glucose in HMCs. In summary, our finding indicated that PAR-1 upregulation mediated proliferation of glomerular mesangial cells induced by high glucose, and deficiency of miR-17 family resulted in PAR-1 upregulation.

High glucose upregulates PAR-1 in SH-SY5Y cells via deficiency of miR-20a and miR-190a.

Thrombin activity enhancement and its receptor protease-activated receptor 1 (PAR-1) activation play vital roles in neurologic deficits in the central nervous system. Our recent study showed that PAR-1 upregulation stimulated by chronic high glucose (HG) caused central neuron injury through neuroinflammation; however, the molecular mechanisms are far from clear. In the present study, we found that HG resulted in neuronal injury of SH-SY5Y cells as evidenced by decreased cell viability and increased lactate dehydrogenase release and elevated the mRNA level of PAR-1. Moreover, we predicted and determined several potential microRNAs (miRs) combining with the 3'-UTR of PAR-1 mRNA, finding that miR-20a-5p, miR-93-5p, and miR-190a-5p were significantly decreased in HG-cultured SH-SY5Y cells compared with control. Further, SH-SY5Y cells stably transfected with miR-20a-5p or miR-190a-5p mimic were established, and overexpression efficiency were confirmed. It was found that miR-20a-5p or miR-190a-5p overexpression markedly decreased PAR-1 mRNA level and protein expression in SH-SY5Y cells cultured with HG and normal glucose, indicating that miR-20a or miR-19a deficiency contributed to HG-induced PAR-1 upregulation. Together, our findings demonstrated that PAR-1 upregulation mediated HG-induced neuronal damage in central neurons, which was achieved through miR-20a or miR-190a deficiency.

Sarsasapogenin attenuates Alzheimer-like encephalopathy in diabetes.

BACKGROUND: A crosstalk exists between diabetes and Alzheimer's disease (AD), and diabetic encephalopathy displays AD-like disorders. Sarsasapogenin (Sar) has strong anti-inflammatory efficacy, showing neuroprotection and memory-enhancement effects. PURPOSE: This study aims to verify the ameliorative effects of Sar on diabetic encephalopathy in vivo and in vitro, and to clarify the mechanisms from attenuation of AD-like pathology. METHODS: Streptozotocin-induced type 1 diabetic rats and high glucose-cultured SH-SY5Y cells were used in this study. After Sar treatment (20 and 60 mg/kg) for consecutive 9 weeks, Morris water maze and novel object recognition tasks were performed. Hematoxylin-eosin staining was used for examining loss of neurons in CA1 area and ki67 expression for reflecting neurogenesis in DG area of hippocampus. Aß production pathway and tau phosphorylation kinase cascade were examined in these two models. RESULTS: Sar improved learning and memory ability, loss of neurons and reduction of neurogenesis in the hippocampus of diabetic rats. Moreover, Sar suppressed Aß overproduction due to up-regulation of BACE1 in protein and mRNA and tau hyperphosphorylation from inactivation of AKT/GSK-3ß cascade in the hippocampus and cerebral cortex of diabetic rats and high glucose-cultured SH-SY5Y cells, and PPARγ antagonism abolished the effects of Sar on key molecules in the two pathways. Additionally, it was found that high glucose-stimulated Aß overproduction was prior to tau hyperphosphorylation in neurons. CONCLUSION: Sar alleviated diabetic encephalopathy, which was obtained through inhibitions of Aß overproduction and tau hyperphosphorylation mediated by the activation of PPARγ signaling. Hence, Sar is a good candidate compound for AD-like disorders.