Chlorogenic acid promotes angiogenesis and attenuates apoptosis following cerebral ischaemia-reperfusion injury by regulating the PI3K-Akt signalling.

CONTEXT: Chlorogenic acid (CGA) has good antioxidant effects, but its explicit mechanism in cerebral ischaemia-reperfusion injury is still uncertain. OBJECTIVE: We studied the effect of CGA in human brain microvascular endothelial cells (HBMECs) under OGD/R damage. MATERIALS AND METHODS: HBMECs in 4 groups were treated with oxygen-glucose deprivation/re-oxygenation (OGD/R) (4 + 24 h), normal no CGA treatment and different concentrations (20, 40 or 80 µM) of CGA. Male C57BL/6J mice were classified as sham, middle cerebral artery occlusion (MCAO), and MCAO + CGA (30 mg/kg/day) groups. Mice in the sham group were not subjected to MCAO. Cell viability, apoptosis, angiogenesis and related protein levels were investigated by CCK-8, flow cytometry, TUNEL staining, tube formation and western blot assays. Infarct volume of brain tissues was analyzed by TTC staining. RESULTS: CGA curbed apoptosis (from 32.87% to 13.12% in flow cytometry; from 34.46% to 17.8% in TUNEL assay) but accelerated cell angiogenesis of HBMECs with OGD/R treatment. Moreover, CGA augmented activation of the PI3K-Akt signalling (p-PI3K/PI3K level, from 0.39 to 0.49; p-Akt/Akt level, from 0.52 to 0.81), and the effect of CGA on apoptosis and angiogenesis was abolished by an inhibitor of PI3K-Akt signalling. Furthermore, CGA attenuated infarct (from 41.26% to 22.21%) and apoptosis and promoted angiogenesis and activation of the PI3K/Akt signalling in MCAO-induced mice. CONCLUSIONS: CGA effectively repressed apoptosis and promoted angiogenesis in OGD/R-treated HBMECs and MCAO-treated mice by modulating PI3K-Akt signalling. Our research provides a theoretical basis for the use of CGA in the treatment of ischaemic stroke.

Chlorogenic Acid Prevents Microglia-Induced Neuronal Apoptosis and Oxidative Stress under Hypoxia-Ischemia Environment by Regulating the MIR497HG/miR-29b-3p/SIRT1 Axis.

Background: Chlorogenic acid (CGA) is a polyphenolic compound with antioxidant and anti-inflammatory properties. CGA has been shown to improve neuroinflammation. This study is aimed at elucidating the exact mechanism by which CGA reduces neuroinflammation. Methods: Oxygen and glucose deprivation (OGD) was utilized to treat BV2 microglia and HT-22 hippocampal neurons to engineer an in vitro model of hypoxic ischemia reperfusion. The levels of inflammatory factors (IL-1ß, IL-6, TNF-α, IL-4, and IL-10) and oxidative stress factors (MDA, SOD, and GSH-PX) in microglia were determined by ELISA kits. The neuron proliferation was assessed by CCK-8 assay, and LDH kit was used to determine LDH release in neurons. The fluorescent dye DCF-DA was employed to measure ROS levels in neurons. Correlation of MIR497HG, miR-29b-3p, and SIRT1/NF-κB in neurons and microglia was determined by qRT-PCR. Expressions of inflammatory proteins (COX2, iNOS), oxidative stress pathways (Nrf2, HO-1), and apoptosis-related proteins (Bcl-2, Bax, caspase3, caspase8, and caspase9) in microglia or neurons were determined by western blot. The interactions between MIR497HG and miR-29b-3p, as well as between miR-29b-3p and SIRT1, were determined by dual luciferase assay and RIP assay. Results: CGA attenuated OGD-mediated inflammation and oxidative stress in microglia and inhibited microglia-mediated neuronal apoptosis. CGA increased the levels of MIR497HG and SIRT1 and suppressed the levels of miR-29b-3p in BV2 and HT-22 cells. MIR497HG knockdown, miR-29b-3p upregulation, and SIRT1 inhibition inhibited CGA-mediated anti-inflammatory and neuronal protective functions. There is a targeting correlation between MIR497HG, miR-29b-3p, and Sirt1. MIR497HG sponges miR-29b-3p to regulate SIRT1 expression in an indirect manner. Conclusion: CGA upregulates MIR497HG to curb miR-29b-3p expression, hence initiating the SIRT1/NF-κB signaling pathway and repressing OGD-elicited inflammation, oxidative stress, and neuron apoptosis.

The effect of group IV chitinase, HaCHT4, on the chitin content of the peritrophic matrix (PM) during larval growth and development of Helicoverpa armigera.

BACKGROUND: Extensive research has been conducted on insect chitinases. However, little is known about the function of chitinase in the regulation of the surface structure of the peritrophic matrix (PM) in larval midguts. The aim of this study was to analyze the effect of HaCHT4 on the chitin content and surface structure of the PM during larval growth and development of Helicoverpa armigera. RESULTS: The expression level of HaCHT4 was lower and the chitin content was higher in the early stages of fourth to sixth instar larvae, but they were reversed in the corresponding late stages. The correlation coefficient between the expression level of HaCHT4 and the chitin content was -0.585 (P < 0.05), with a higher negative correlation of -0.934 for the fourth instar (P < 0.01). Scanning electron microscopy (SEM) showed that the surface structure of PM was multi-laminated with small pores in the early stages of fourth to sixth instar larvae, and more and bigger pores in the late stages. Low expression of HaCHT4 caused by RNA interference (RNAi) resulted in the increase of chitin content in the PM, and the surface structure of PM became multilayered with smaller pore size in the late stage of fourth instar larvae. Also, induction of HaCHT4 by application of 2-tridecanone (2-TD), decreased the chitin content of PM, caused larger pores to form and lots of food bolus to attach to the PM surface, and also increased the larval susceptibility to chlorantraniliprole. CONCLUSION: These results provided strong evidence that HaCHT4 plays an important role by regulating the chitin content of the PM and its surface structure, thereby affecting the sensitivity of H. armigera to chlorantraniliprole.