Effect of sulfasalazine on ferroptosis during intestinal injury in rats after liver transplantation.

Using a rat autologous orthotopic liver transplantation (AOLT) model and liver cold ischemia-reperfusion (I/R)-induced intestinal injury, we clarified whether ferroptosis occurred in rat AOLT cold I/R-induced intestinal injury. Additionally, the role and possible mechanism of the ferroptosis activator sulfasalazine (SAS) in intestinal injury-induced ferroptosis in rats with AOLT liver cold I/R were investigated. Sixty specific pathogen free (SPF)-grade adult male Sprague‒Dawley (SD) rats were randomly divided into 5 groups using the random number table method (n = 12). Six rats were randomly selected at 6 hour (h) and 24 h after I/R. Inferior vena cava blood specimens were collected from the portal vein (PV) opening at 6 h and 24 h. The concentrations of serum malondialdehyde (MDA), serum interleukin 6 (IL-6) were determined by enzyme-linked immunosorbent assay (ELISA). Ileal tissue was obtained from the PV opening in rats in each group at 6 h and 24 h, and ileal tissue sections were observed under light microscopy. The contents of intestinal MDA, superoxide dismutase (SOD), glutathione(GSH), glutathione peroxidase 4 (GPX4), and tissue iron were determined by ELISA, and the expression of GPX4 and the cysteine glutamate reverse transporter light chain protein (xCT) was determined by Western blot. The experimental results show that ferroptosis is involved in the pathophysiological process of intestinal injury induced by cold hepatic ischemia-reperfusion in AOLT rats. In addition, SAS (500 mg/kg) may inhibit the cystine/glutamate antiporters (System Xc¯)/GSH/GPX4 signal axis in intestinal injury induced by cold I/R in rat AOLT liver, or iron overload after reperfusion, causing a massive accumulation of L-ROS and activating cellular ferroptosis, further aggravate the intestinal injury.

Faecal Microbiota Transplantation Alleviates Ferroptosis after Ischaemic Stroke.

Ischaemic stroke can induce changes in the abundance of gut microbiota constituents, and the outcome of stroke may also be influenced by the gut microbiota. This study aimed to determine whether gut microbiota transplantation could rescue changes in the gut microbiota and reduce ferroptosis after stroke in rats. Male Sprague-Dawley rats (6 weeks of age) were subjected to ischaemic stroke by middle cerebral artery occlusion (MCAO). Fecal samples were collected for 16S ribosomal RNA (rRNA) sequencing to analyze the effects of FMT on the gut microbiota. Neurological deficits were evaluated using the Longa score. triphenyl tetrazolium chloride (TTC) staining was performed in the brain, and kits were used to measure malondialdehyde (MDA), iron, and glutathione (GSH) levels in the ipsilateral brain of rats. Western blotting was used to detect the protein expression levels of glutathione peroxidase 4 (GPX4), solute carrier family 7 member 11 (SLC7A11), and the transferrin receptor 2 (TFR2) in the ipsilateral brain of rats. Stroke induced significant changes in the gut microbiota, and FMT ameliorated these changes. TTC staining results showed that FMT reduced cerebral infarct volume. In addition, FMT diminished MDA and iron levels and elevated GSH levels in the ipsilateral brain. Western blot analysis showed that FMT increased GPX4 and SLC7A11 protein expression and decreased TFR2 protein expression in the ipsilateral brain after stroke. FMT can reverse gut microbiota dysbiosis, reduce cerebellar infarct volume, and decrease ferroptosis after stroke.

Neuroprotective potentials of ferulic acid against intracerebral hemorrhage COVID-19 through using network pharmacology approach and molecular docking analysis.

Intracerebral hemorrhage (ICH) refers to severe stroke subtype that may be life-threatening or even cause death. It is clinically observed that coronavirus disease 2019 (COVID-19) may be associated with the high mortality in ICH patients. Ferulic acid, one of the functional bioactive ingredients from medicinal herbs, has been preclinically proven with beneficial activities, including neuroprotection and anti-inflammation actions. Based on current findings, we assumed that ferulic acid may play the potentials against COVID-19 when ICH. In this study, preclinical approach including network pharmacology and molecular docking was applied to detect and identify the core targets and pharmacological mechanisms involved in ferulic acid on COVID-19 and ICH. The network pharmacology analysis identified total eleven core targets in ferulic acid and COVID-19/ICH. The molecular mechanisms of ferulic acid against COVID-19 and ICH were mostly involved in induction of antiviral activity, modulation of inflammatory reaction. Molecular docking model revealed that ferulic acid might effectively bind to epidermal growth factor receptor (EGFR) protein based on strong binding capability. Current findings reflected the preclinical pharmacological activities of ferulic acid that might use for management of COVID-19 and ICH. Although there are the limitations that are absence of experimental validation, these bioinformatic results underline that ferulic acid may exert simultaneous potentials against COVID-19 and ICH through modulating integrative mechanisms and key biotargets.

Excessive aggregation of fine particles may play a crucial role in adolescent spontaneous pneumothorax pathogenesis.

Background: The pathogenesis of primary spontaneous pneumothorax (PSP) is unclear. Fine particles aggregated in the lung can be phagocytosed by alveolar macrophages (AMs) to induce an inflammatory reaction and damage local pulmonary tissue, which could be a mechanism of PSP. This project aimed to explore the pathological association between fine particulate matter and PSP. Methods: Thirty pulmonary bullae tissues were obtained from surgery of PSP patients (B group). The adjacent normal tissues of the lungs were defined as the control S group. Another 30 normal lung tissues with nonpneumothorax disease (NPD) were applied as the control N group. Hematoxylin and eosin (H & E), Wright-Giemsa (W-G), Victoria blue, and immunohistochemical (IHC) staining experiments were performed to measure the levels of fine particulate matter, alveolar macrophages (AMs), pulmonary elastic fibers, monocyte chemoattractant protein-1 (MCP-1), and matrix metalloproteinase-9 (MMP-9) in the lung tissues. The serum levels of MCP-1 and MMP-9 were prospectively analyzed as well. Results: Histopathological examinations revealed obvious deposition of fine particulate matter and inflammatory reactions (proliferation of AMs) in the B group, compared with those in the S group and the N group. These alterations were significantly associated with PSP. The numbers of AMs and pulmonary elastic fibers, the positive area of the H-score, as well as the concentrations of MCP-1 and MMP-9 in the lungs of the experimental group were obviously raised compared with the controls (P < 0.05). Conclusions: Fine particulate matter aggregation, inflammation (macrophage hyperplasia), and overexpression of MCP-1 and MMP-9 may contribute to the pathogenesis of PSP. The overaccumulation of fine particulate matter may play a crucial part in the occurrence of adolescent and young adult PSP. Trial registration: This project was enrolled on the Chinese Clinical Trial Registry: ChiCTR2100051460.

HIF-1α-regulated lncRNA-TUG1 promotes mitochondrial dysfunction and pyroptosis by directly binding to FUS in myocardial infarction.

Myocardial infarction (MI) is a fatal heart disease that affects millions of lives worldwide each year. This study investigated the roles of HIF-1α/lncRNA-TUG1 in mitochondrial dysfunction and pyroptosis in MI. CCK-8, DHE, lactate dehydrogenase (LDH) assays, and JC-1 staining were performed to measure proliferation, reactive oxygen species (ROS), LDH leakage, and mitochondrial damage in hypoxia/reoxygenation (H/R)-treated cardiomyocytes. Enzyme-linked immunoassay (ELISA) and flow cytometry were used to detect LDH, creatine kinase (CK), and its isoenzyme (CK-MB) levels and caspase-1 activity. Chromatin immunoprecipitation (ChIP), luciferase assay, and RNA-immunoprecipitation (RIP) were used to assess the interaction between HIF-1α, TUG1, and FUS. Quantitative real-time polymerase chain reaction (qRT-PCR), Western blotting, and immunohistochemistry were used to measure HIF-1α, TUG1 and pyroptosis-related molecules. Hematoxylin and eosin (HE), 2,3,5-triphenyltetrazolium chloride (TTC), and terminal deoxynucleotidyl transferase dUTP risk end labelling (TUNEL) staining were employed to examine the morphology, infarction area, and myocardial injury in the MI mouse model. Mitochondrial dysfunction and pyroptosis were induced in H/R-treated cardiomyocytes, accompanied by an increase in the expression of HIF-α and TUG1. HIF-1α promoted TUG1 expression by directly binding to the TUG1 promoter. TUG1 silencing inhibited H/R-induced ROS production, mitochondrial injury and the expression of the pyroptosis-related proteins NLRP3, caspase-1 and GSDMD. Additionally, H/R elevated FUS levels in cardiomyocytes, which were directly inhibited by TUG1 silencing. Fused in sarcoma (FUS) overexpression reversed the effect of TUG1 silencing on mitochondrial damage and caspase-1 activation. However, the ROS inhibitor N-acetylcysteine (NAC) promoted the protective effect of TUG1 knockdown on H/R-induced cardiomyocyte damage. The in vivo MI model showed increased infarction, myocardial injury, ROS levels and pyroptosis, which were inhibited by TUG1 silencing. HIF-1α targeting upregulated TUG1 promotes mitochondrial damage and cardiomyocyte pyroptosis by combining with FUS, thereby promoting the occurrence of MI. HIF-1α/TUG1/FUS may serve as a potential treatment target for MI.

Hydrogen exerts neuroprotective effects on OGD/R damaged neurons in rat hippocampal by protecting mitochondrial function via regulating mitophagy mediated by PINK1/Parkin signaling pathway.

Cerebral ischemia/reperfusion injury (IRI) is a serious complication during the treatment of stroke patients with very few effective clinical treatment. Hydrogen (H2) can protect mitochondria function and have favorable therapeutic effects on cerebral IRI. Mitophagy plays an important role in eliminating damaged or dysfunctional mitochondria and maintaining mitochondria homeostasis. However, whether the protection of H2 on cerebral IRI is via regulating mitophagy is still unknown. In this study, OGD/R damaged hippocampal neurons were used to mimic cerebral IRI in vivo and we detected the effect of H2, Rap (autophagy activator) and 3-MA (autophagy inhibitor) on OGD/R neurons. The results of MTT indicated that H2 and RAP could increase cell viability after OGD/R treatment, while 3-MA further aggravated injury and inhibited the protection of H2 and RAP. Furthermore, the intracellular ROS and apoptosis ratio were determined, the results showed that ROS and apoptosis level significantly increased after OGD/R, H2 and RAP effectively restrained the increment of ROS level and apoptosis ratio but their protective effect can be weakened by 3-MA. Mitochondrial membrane potential (MMP) and mitophagy level were also determined, the data showed that H2 and RAP protected against the loss of MPP and increased the co-localization of mitochondria with GFP-LC3 while 3-MA exerted antagonistic effect. At last, the mitophagy-related factors LC3, PINK1 and Parkin expression were detected and analyzed. We found that the expression of LC3 was increased after OGD/R which can be further enhanced by H2 and RAP treatment, but treatment with 3-MA was opposite. The result revealed H2 and RAP could activate mitophagy while 3-MA inhibit mitophagy. In addition, the study found H2 and RAP could significantly induce the expression of PINK1 and Parkin in OGD/R neurons which was inhibited by 3-MA. Taken together, our findings demonstrated H2 had a neuroprotective effect on OGD/R damaged neurons by protecting mitochondrial function and the potential protection mechanism may closely related to enhancement of mitophagy mediated by PINK1/Parkin signaling pathway.

[Effects of nerve growth factor pretreatment on apoptosis of neurons and expression of Bcl-2, Bax protein in brain tissue following cerebral ischemia/reperfusion injury in gerbils].

OBJECTIVE: To study the effect of nerve growth factor (NGF) pretreatment on apoptosis of neurons and the expression of Bcl-2 and Bax protein in cerebral cortex and hippocampus CA1 zone following global cerebral ischemia/reperfusion (I/R) injury in gerbils and to explore the mechanism of protection and the best time window of NGF pretreatment. METHODS: Global cerebral I/R injury model was induced by occlusion of bilateral carotid arteries. NGF was injected into the lateral ventricle. Thirty gerbils were randomly divided into five groups, with six animals in each: sham operation group (A group), I/R injury group (B group), NGF pretreatment 12, 24 and 48 hours groups (C, D and E group). Gerbils in all groups were sacrificed after being subjected to 20 minutes of cerebral ischemia followed by 72 hours reperfusion, except A group. Neural apoptosis was identified by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL), and immunohistochemistry was used to detect the expression of Bcl-2 and Bax protein in cerebral cortex and hippocampus CA1 zone. RESULTS: Compared with B group, the number of apoptotic neurons and the expression of Bax positive cells in NGF pretreatment groups were decreased significantly (all P<0.05), while the expression of Bcl-2 positive cells was increased significantly (all P<0.05). The apoptotic rate in cerebral cortex and hippocampus CA1 zone and expression rate of Bax protein positive cells were the lowest, but the expression rate of Bcl-2 protein positive cells was the highest at 48 hours. CONCLUSION: NGF pretreatment can significantly decrease the neuronal apoptosis of the cerebral I/R injury in gerbils, and the best time window of NGF pretreatment is 48 hours. The mechanism of protection may be related to induction of Bcl-2 protein expression and inhibition of Bax protein expression by NGF pretreatment, thereby preventing neuronal apoptosis.

[Significance of detecting HBV-DNA by the fluorescence quantitative PCR].

BACKGROUND: To study the correlativity between HBV-DNA and the markers of hepatitis B virus infection and different clinical types of hepatitis B. METHODS: The fluorescence quantitation (FQ) of HBV-DNA of 105 patients with hepatitis B was performed by PCR, and the correlativity between the fluorescence quantitation of HBV-DNA and the markers of hepatitis B virus and different clinical types of hepatitis B was analyzed. RESULTS: Ninety-seven percent of the patients were found HBsAg(+), HBeAg(+), HBcAb(+); 75% were HBsAg(+), HBeAb(+), HBcAb(+); 60% were HBsAg(+), HBcAb(+); 40% were HBsAg(+); in HBsAb(+), HBeAb(+), HBcAb(+) (or both HBsAb and HBcAb were positive) group the HBV DNA was undetectable. The analysis indicated that there was a significant difference among different groups (P less than 0.05).HBV-DNA was detected in 72.2% in acute hepatitis B group, in 75% of chronic hepatitis B group, and in 70% of cases of liver cirrhosis with hepatitis B group. The analysis indicated that there was no significant difference among the different clinical types of hepatitis (P greater than 0.05). CONCLUSION: The levels of viral replication were not correlated with different clinical types of hepatitis B; the concentration of HBV-DNA in serum was related to hepatitis B antigen.