PHLDA1 contributes to hypoxic ischemic brain injury in neonatal rats via inhibiting FUNDC1-mediated mitophagy.

Hypoxia-ischemia (HI) is one of the main causes of neonatal brain injury. Mitophagy has been implicated in the degradation of damaged mitochondria and cell survival following neonatal brain HI injury. Pleckstrin homology-like domain family A member 1 (PHLDA1) plays vital roles in the progression of various disorders including the regulation of oxidative stress, the immune responses and apoptosis. In the present study we investigated the role of PHLDA1 in HI-induced neuronal injury and further explored the mechanisms underlying PHLDA1-regulated mitophagy in vivo and in vitro. HI model was established in newborn rats by ligation of the left common carotid artery plus exposure to an oxygen-deficient chamber with 8% O2 and 92% N2. In vitro studies were conducted in primary hippocampal neurons subjected to oxygen and glucose deprivation/-reoxygenation (OGD/R). We showed that the expression of PHLDA1 was significantly upregulated in the hippocampus of HI newborn rats and in OGD/R-treated primary neurons. Knockdown of PHLDA1 in neonatal rats via lentiviral vector not only significantly ameliorated HI-induced hippocampal neuronal injury but also markedly improved long-term cognitive function outcomes, whereas overexpression of PHLDA1 in neonatal rats via lentiviral vector aggravated these outcomes. PHLDA1 knockdown in primary neurons significantly reversed the reduction of cell viability and increase in intracellular reactive oxygen species (ROS) levels, and attenuated OGD-induced mitochondrial dysfunction, whereas overexpression of PHLDA1 decreased these parameters. In OGD/R-treated primary hippocampal neurons, we revealed that PHLDA1 knockdown enhanced mitophagy by activating FUNDC1, which was abolished by FUNDC1 knockdown or pretreatment with mitophagy inhibitor Mdivi-1 (25 µM). Notably, pretreatment with Mdivi-1 or the knockdown of FUNDC1 not only increased brain infarct volume, but also abolished the neuroprotective effect of PHLDA1 knockdown in HI newborn rats. Together, these results demonstrate that PHLDA1 contributes to neonatal HI-induced brain injury via inhibition of FUNDC1-mediated neuronal mitophagy.

Protective effect of electroacupuncture at ST36 against damage of intestinal mucosa, oxidative stress and apoptosis induced by 5-FU chemotherapy in mice with colon cancer. / 电针"足三里"å‡è½»å¤§è‚ ç™Œå°é¼ åŒ–ç–—åŽè‚ é»è†œæŸä¼¤åŠå ¶å¯¹æ°§åŒ–åº”æ¿€å’Œç»†èƒžå‡‹äº¡çš„å½±å“.

OBJECTIVES: To observe the effect of electroacupuncture (EA) at "Zusanli"(ST36) on intestinal mucosal damage, intestinal mucosal oxidative stress injury and apoptosis induced by 5-fluorouraeil (5-FU) chemotherapy in colorectal cancer-bearing mice. METHODS: Thirty male BALB/c mice were randomly divided into normal control, colorectal cancer (CT26), 5-FU, non-acupoint and ST36 groups, with 6 mice in each group. Except for those of the normal control group, mice of the remaining 4 groups received subcutaneous implantation of colorectal CT26 cell suspension (0.1 mL) in the right armpit for establishing colorectal cancer model. Rats of the 5-FU group, non-acupoint group and ST36 group were given with 5 mg/mL 5-FU solution once every 3 days for a total of 21 days. For mice of the non-acupoint group and ST36 group, EA (2 Hz, 1-2 mA) was applied to bilateral ST36 or non-acupoints (the bilateral sunken spots about 3 mm to the midpoint between the tail root and the anus) for 5 min after each intraperitoneal infusion of 5-FU, once every 3 days, for a total of 21 days. After the intervention, the diarrhea index was assessed. The length of colon (from the endpoint of cecum to the anal orifice) was measured. Histopathological changes of colonic mucosa were observed by H.E. staining, and the length of colonic villi was measured. The content of malondialdehyde (MDA), and activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) of colonic tissue were detected by thibabituric acid, xanthine oxidase and colorimetric method, respectively. The rate of cell apoptosis in the colonic tissue was measured by TUNEL assay. The positive expressions of Bax and Bcl-2 in colonic tissue were determined by immunohistochemistry. RESULTS: The CT26 model group didn't show any significant changes in the diarrhea index, colon length, colon villus length, MDA content, SOD and GSH-Px activities, colonic cell apoptosis rate, and Bax and Bcl-2 expression levels when compared with the normal group. Compared with the CT26 group, the 5-FU group had a remarkable increase in the diarrhea index, MDA content, colonic cell apoptosis rate and Bax expression level (P<0.01, P<0.05), and a marked decrease in the colon length, colon villus length, SOD and GSH-Px activities and Bcl-2 expression level (P<0.01), suggesting the side effects of administration of 5-FU. Compared with the 5-FU group, the diarrhea index, MDA content, colonic cell apoptosis rate and Bax expression level were markedly decreased (P<0.05, P<0.01) and those of the colon length, colon villus length, SOD and GSH-Px activities and Bcl-2 expression level were obviously increased (P<0.01) in the ST36 group. Compared with the 5-FU group, the non-acupoint group also had an increase in the colon villus length, SOD and GSH-Px activities (P<0.01, P<0.05) and a decrease in the cell apoptosis rate (P<0.01). CONCLUSIONS: EA at ST36 has a positive effect in reducing intestinal mucosal damage induced by 5-FU chemotherapy in cancer-bearing mice, which may be related to its function in relieving oxidative stress injury and inhibiting apoptosis of colonic tissue.

RP58 knockdown contributes to hypoxia-ischemia-induced pineal dysfunction and circadian rhythm disruption in neonatal rats.

Hypoxia-ischemia (HI) of the brain not only impairs neurodevelopment but also causes pineal gland dysfunction, which leads to circadian rhythm disruption. However, the underlying mechanism of circadian rhythm disruption associated with HI-induced pineal dysfunction remains unknown. The zinc finger protein repressor protein with a predicted molecular mass of 58 kDa (RP58) is involved in the development and differentiation of nerve cells. In this study, we established an HI model in neonatal rats to investigate the expression of RP58 and its role in pineal dysfunction and circadian rhythm disruption induced by HI. We demonstrated that RP58 was highly expressed in the pineal gland under normal conditions and significantly downregulated in the pineal gland and primary pinealocytes following HI. Knockdown of RP58 decreased the expression of enzymes in the melatonin (Mel) synthesis pathway (tryptophan hydroxylase 1 [TPH1], acetylserotonin O-methyltransferase [ASMT], and arylalkylamine N-acetyltransferase [AANAT]) and clock genes (circadian locomotor output cycles kaput [CLOCK] and brain and muscle ARNT-like 1 [BMAL1]), and it also reduced the production of Mel, caused pineal cell injury, and disrupted circadian rhythms in vivo and in vitro. Similarly, HI reduced the expression of Mel synthesis enzymes (TPH1, ASMT, and AANAT) and clock genes (CLOCK and BMAL1), and caused pineal injury and circadian rhythm disruption, which were exacerbated by RP58 knockdown. The detrimental effect of RP58 knockdown on pineal dysfunction and circadian rhythm disruption was reversed by the addition of exogenous Mel. Furthermore, exogenous Mel reversed HI-induced pineal dysfunction and circadian rhythm disruption, as reflected by improvements in Mel production, voluntary activity periods, and activity frequency, as well as a diminished decrease in the expression of Mel synthesis enzymes and clock genes. The present study suggests that RP58 is an endogenous source of protection against pineal dysfunction and circadian rhythm disruption after neonatal HI.