Objective To investigate the efficacy of histone deacetylase (HDAC) inhibitor chidamide combined with the PD-1 inhibitor on CD8⁺ T cells anti-cancer function in OVA-expressing MC38 (MC38-OVA) colorectal-bearing mice. Methods Animal experiments: C57BL/6 tumor models were constructed by subcutaneously injecting MC38-OVA colorectal cancer cells into the back of mice. We randomized mice into control group, chidamide group, anti-PD-1 group and chidamide+anti-PD-1 group (20 each group). We monitored the tumor growth and animal survival rate of each group; we employed a flow-based method to detect the number and ratio of tumor-infiltrating CD8⁺ T cells, CD8⁺IFN-γ⁺ T cells, OVA antigen-specific CD8⁺ T cells, and the expression changes of regulatory T cells (Treg), myeloid-derived suppressor cells (MDSC), and tumor-associated macrophages (TAM). Cell experiments: We used a flow-based method to detect the apoptosis of CD8⁺ T cells and MC38-OVA tumor cells after 0, 10, 25, 50, 100, or 200 nmol/L chidamide treatment. The proliferation of CD8⁺ T cells and MC38-OVA tumor cells treated with 0 and 100 nmol/L chidamide was detected by Ki-67 antibody labeling and cell counting. To evaluate CD8⁺ T cell killing ability, we treated CD8⁺ T cells with various conditions (control group, chidamide group, anti-PD-1 group and chidamide+anti-PD-1 group) followed by co-culture with MC38-OVA tumor cells, using the flow-based method. In the condition that CD8⁺ T cells treated with 0 and 100 nmol/L chidamide co-cultured with the same number of MC38-OVA tumor cells, the expression of CD107a was detected by flow cytometry. Results Compared with control group, the tumor growth was inhibited (P<0.05) while the survival rate was improved (P<0.01) in chidamide+anti-PD-1 group. The number of tumor-infiltrating CD8⁺ T cells was significantly higher in chidamide group, anti-PD-1 group and chidamide+anti-PD-1 group than that in control group (P<0.05). Nonetheless, the ratio and levels of CD8⁺IFN-γ⁺ and OVA antigen-specific CD8⁺ T cells were significantly higher in chidamide+anti-PD-1 group than those in other groups (P<0.05). The in vitro experiment results showed that chidamide could enhance the killing ability of CD8⁺ T cells and the expression of CD107a. Conclusion Chidamide combined with PD-1 inhibitor significantly enhanced the number and function of tumor-infiltrating CD8⁺ T cells and increased antigen-specific CD8⁺ T cells, which will provide a theoretical and experimental basis for the combination of chidamide in clinical solid tumor immunotherapy.

Helicobacter pylori (HP) infection is a Class Ⅰ carcinogen in gastric cancer, closely related to the occurrence of gastric cancer. Many studies have shown that HP eradication has a preventive effect on gastric cancer. However, 2.7%-6.1% of patients with early gastric cancer who have been eradicated after endoscopic submucosal dissection (ESD) can still develop metachronous gastric cancer (MGC), and the mechanism of its occurrence is still unclear. In this review, the atrophy of gastric mucosa and intestinal metaplasia cannot be completely reversed after HP eradication, the excessive proliferation of gastric mucosa epithelial cells, the accumulation of genetic abnormalities, the homeostasis imbalance of the epigenetic group, changes in immune microenvironment, the abnormality of stem cells in gastric mucosa, chromatin accessibility, and changes in chromosome remodeling were discussed in the mechanism of carcinogenesis caused by the above molecular changes after ESD and HP eradication in early gastric cancer.

Corona virus disease 2019 (COVID-19) epidemic has been effectively controlled, but its related complications still cannot be ignored, especially the cardiovascular circulatory system is the active site of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Angiotensin-converting enzyme 2 (ACE2) is a type Ⅰ transmembrane glycoprotein that is highly expressed in heart, kidney and testis. Spike protein of SARS-CoV-2 invades host cells by binding to the cell surface receptor ACE2. However, there are still many deficiencies in the clinical application of vaccines and drugs developed based on this target. As a molecular chaperone, cyclophilin A (CypA) promotes protein folding and T cell activation. CD147 is one of the most widely studied CypA receptors, and the interaction of CypA/CD147 plays an important role in the entry of SARS-CoV-2 into host cells. However, there are few reports on the invasion of SARS-CoV-2 into the cardiovascular system through the CypA/CD147 signaling pathway. Based on this, this article summarizes the previous research evidence and the research basis of the research group, and reviews the structure and function of CypA/CD147, the role of CypA/CD147 in cardiovascular disease, and the cardiovascular disease caused by SARS-CoV-2 targeting CypA/CD147 signal pathway, in order to provide reference for the diagnosis and treatment of the COVID-19 complicated with cardiovascular system diseases.

The prothrombin complex concentrate (PCC) is a blood product that mainly contains vitamin K-dependent coagulation factors. PCC has been used clinically for more than 70 years, and its safety has been significantly improved. It has been widely used in the treatment of coagulation disorders or severe bleeding in critically ill patients. If used improperly, it may weaken the hemostatic effect or increase the risk of thrombosis. In order to standardize the rational use of PCC, Chinese Society of Thrombosis, Hemostasis and Critical Care, Chinese Medicine Education Association, People's Liberation Army Professional Committee of Critical Care Medicine, Chinese Society of Thrombosis and Hemostasis, Chinese Research Hospital Association have organized the formulation of this expert consensus, including three parts: definition, indications, and monitoring and evaluation of efficacy, with a total of 10 suggestions, aiming to help clinicians reasonably use PCC and improve the treatment in critical illness.

Objective To investigate the preventive effect of schisandrin (SCH) on fetal neural tube defects (NTDs) of mice and its mechanism. Methods C57BL/6 mice were mated with female and male at a ratio of 2:1. Pregnant female mice with vaginal plug after mating were randomly divided into control group, model group, SCH group, and folic acid group, with 9 mice in each group. The NTDs fetal mice model was induced by intraperitoneal injection of all-trans retinoic acid (atRA) (7.5 mg/kg) on embryonic day 7.5 (E 7.5 d). During E 0.5 d-E 11.5 d, pregnant rats in folic acid group were given folic acid [61.0 μg/(kg·d)] by gavage once a day, and pregnant rats in SCH group were given SCH [8.0 mg/(kg·d)] by gavage once a day. Fetal mice were removed by cesarean section on E 11.5 d. PC12 cells were divided into control group, model group and SCH group. PC12 cells were treated with atRA (20 μmol/L) for 12 hours to establish cell damage model in model group, and treated with SCH (2.5 μmol/L) for 24 hours in SCH group. Fetuses were identified NTDs by stereoscopic microscopy. HE staining was used to observe the closure of the neural tube. The expression levels of p-PI3K, Akt and p-Akt molecules in PI3K/Akt signaling pathway were detected by Western Blotting. Results Compared with control group, the incidence of NTDs was significantly increased in mice of model group (P<0.01); compared with model group, the incidence of NTDs was decreased in folic acid group and SCH group (P<0.01); compared with folic acid group, SCH group had a lower incidence of NTDs (P<0.01). Western Blotting results showed that compared with control group, the expression of p-PI3K and p-Akt protein in fetal tissues of model group was significantly decreased (P<0.01, P<0.05); compared with model group, there was no significant difference in expression of p-PI3K and p-Akt in fetal tissues of folic acid group (P>0.05), while the expression of p-PI3K and p-Akt protein in SCH group was significantly higher (P<0.05). Compared with control group, PC12 cells in model group showed lower expression levels of p-PI3K and p-Akt (P<0.05); compared with model group, PC12 cells in SCH group showed higher expression levels of p-PI3K and p-Akt (P<0.05). Conclusions SCH can reduce the incidence of atRA-induced NTDs in fetal mice, and its preventive effect is better than folic acid, which may be related to the activation of the PI3K/Akt signaling pathway.

Objective Establishing animal model of Vibrio vulnificus (V. vulnificus) early infection via respiratory tract, to observe the survival, pathological changes of target organs and the expression changes of inflammatory cytokines in model mice, and to provide references for the relevant prevention and treatment strategies. Methods A total of 24 healthy female BALB/c mice (8 to 10 weeks old) were randomly divided into 4 groups (6 mice per group), including control group and three infection groups (low, medium and high concentration of infection, respectively), based on which the V. vulnificus infection models were established by intranasal administration. The survival statistics, status of defecation, hair and respiration as well as mental state of mice were monitored during 0-12 h early infection progress. Hematoxylin-eosin (HE) staining was performed on the important organs of mice to analyze the pathological manifestations, and immunohistochemical staining was used to detect the expression of interleukin (IL)-6, IL-10, interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α) in the target organs. Mouse inflammation panel combined with flow cytometry was used to further detect the changes of multiple inflammatory and anti-inflammatory cytokines in serum. Results V. vulnificus infection via respiratory tract caused mice deterioration in survival rate, defecation, hair status, respiratory tract and mental state within 12 h. HE pathological analysis showed inflammatory cell infiltration and cell necrosis in ileum, lung, liver and spleen of mice in different infection groups, and the injury was much severer in high concentration of infection group. Immunohistochemical results reflected the positive expression rates of IL-6, IL-10, IFN-γ and TNF-α in ileum, lung, liver and spleen of mice in medium and high concentration of infection groups were significantly higher than those in control group (P<0.05). The results of mouse inflammation panel and flow cytometry showed that compared with the control group, the expression levels of TNF-α and monocyte chemotactic protein (MCP)-1 in all three infection groups were significantly increased (P<0.05), and the expression level of IL-27 significantly decreased (P<0.05); the expression levels of IL-1β, IL-6, IL-17A and granulocyte-macrophage colony-stimulating factor (GM-CSF) were significantly increased in medium and high concentration of infection groups (P<0.05); the expression levels of IL-1α, IL-23, IL-12p70 and IFN-γ were significantly increased in high concentration of infection group (P<0.05). Conclusions V. vulnificus infection progressed rapidly through respiratory tract in mice, and lung, intestine, liver and spleen were major target organs. Accompanied by increased secretion of multiple serum inflammatory cytokines, V. vulnificus infection through respiratory tract might further causes severe inflammatory reaction in hosts.

Objective To investigate the possible mechanism of Xuebijing injection in regulating abnormal degradation of pulmonary vascular endothelial calyx to improve acute lung injury induced by severe heatstroke. Methods Forty-eight Wistar rats were randomly divided into control group, heatstroke group and Xuebijing group. Before heatstroke induction, rats in Xuebijing group were administrated with Xuebijing injection (2 ml/kg, 2 times/d) for 3 days. All rats were exposed to an environment with temperature of (40±2) ℃ and humidity of 65%±5% for 60 minutes to induce heatstroke. Two hours later, the lung wet/dry weight ratio was recorded; the concentration of proteins in BALF was measured; the pulmonary vascular permeability was measured by Evans blue (EB); HE staining was used to observe the pathological changes of lung tissue; the changes of hyaluronic acid (HA) on the surface of pulmonary vessels were observed by immunofluorescence; Western blotting was used to detect the expression of Syndecan-1, Glypican-1, VE-Cadherin, Occludin, VCAM-1 and E-selectin in lung tissues; Enzyme-linked immunosorbent assay (ELISA) was utilized to quantify the concentration of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) in serum and heparanase (HPA) in lung tissue. Results Xuebijing could decrease the lung wet/dry weight ratio, reduce protein exudation and improve pulmonary vascular permeability (P<0.01); reduce the histological injury (P<0.01); reduce the degradation of HA, Syndecan-1 and Glypican-1 on the surface of pulmonary vessels (P<0.01); increase the expression of VE-Cadherin and Occludin (P<0.01); regulate the overexpression of VCAM-1 and E-selectin (P<0.01); down-regulate the expression of TNF-α, IL-6 and HPA (P<0.01). Conclusion Xuebijing injection decrease the expression of HPA, improve the disintegration of pulmonary vascular endothelial calyx, repair the integrity of pulmonary vessels, reduce the damage of cell connections, down-regulate the expression of adhesion molecules, inhibit the inflammatory reaction, relieving acute lung injury caused by severe heat stroke.

Objective To explore the protective effects and mechanisms of cysteinyl leukotriene 2 (CysLT₂) receptor antagonist HAMI3379 on cerebral ischemia injury in rats. Methods 30 male SD rats were randomly divided into sham operation group, model group, and HAMI3379 group, with 10 rats in each group. The rats of model group were subjected to middle cerebral artery occlusion (MCAO) to construct the cerebral ischemia injury model, while HAMI3379 group received intraperitoneal injection of HAMI3379 (0.2 mg/kg) before and after MCAO 30 min. The rats after cerebral ischemia injury were scored for neurological symptoms. The infarction volume of rats was observed by TTC staining, the activation marker Iba1 of microglia was detected by immunofluorescence staining, the mRNA level of M1/M2 polarized phenotype molecules of microglia was detected by Real-time PCR, the number of neurons was observed by NeuN staining, and neuronal degeneration was observed by Fluoro-Jade B staining. Western blotting assay was used to detect the expression of CysLT₂ protein and nuclear factors κB-related protein Cα (PKCα), IκBα, p65 and p50 proteins in brain tissue. Results Compared with sham operation group, the neurological symptom score and cerebral infarction volume of model group were significantly increased (P<0.05). Compared with model group, the neurological symptom score and cerebral infarction volume of HAMI3379 group were significantly decreased (P<0.05). The results of immunofluorescence staining showed that the expression of microglia activation marker Iba1 was increased in brain tissue of rats after cerebral ischemia injury (P<0.05). Compared with sham operation group, mRNA expression of M1 polarized molecules (CD86, IL-1β, TNF-α) and M2 polarized molecules (CD206, TGF-β, IL-10) were significantly increased in the ischemic central brain tissue of model group (P<0.05). Compared with model group, the expression of M1 polarized molecules in HAMI3379 group was significantly downregulated (P<0.05), while the expression of M2 polarized molecules was significantly upregulated (P<0.05). Compared with sham operation group, the expressions of PKCα, IκBα, p65 and p50 in brain tissue of model group were significantly up-regulated (P<0.05); compared with model group, the expressions of PKCα, IκBα, p65, and p50 in HAMI3379 group were significantly down-regulated (P<0.05). The NeuN staining results showed that the number of neurons in the brain tissue of model group was decreased when compared with sham operation group (P<0.05), while the number of degenerated neurons was increased (P<0.05). Compared with model group, the number of neurons in HAMI3379 group was increased (P<0.05), while the number of degenerated neurons was decreased (P<0.05). Conclusions CysLT₂ receptor antagonist HAMI3379 may regulate PKCα/IκBα/NF-κB signaling pathway, inhibits M1 polarization activation of microglia and promotes its transition to M2 polarization, inhibits neuronal degeneration, and plays a neuroprotective role.

Objective To investigate the improvement effect and potential mechanism of m⁶A recognition protein YTHDF1 on Duchenne muscular dystrophy (DMD). Methods (1) We collected muscle tissue and peripheral blood from DMD patients (38 confirmed patients with surgery at Xi'an Children's Hospital from September 2016 to June 2017, DMD group) and non-DMD patients with orthopedic surgery (33 age-matched patients, 2-3 years old, control group). The expression levels of YTHDF1 and dystrophin in the muscle tissues were quantified by RT-qPCR and Western blotting. The serum creatine phosphokinase (CPK) content was determined using ELISA. (2) Take SkMC from skeletal muscle myoblasts and set up empty vector group (transfected with scrambled negative control), AAV-YTHDF1 group transfected with YTHDF1 overexpression vector and scrambled group (transfected with NC siRNA) and si-YTHDF1 group (transfected with YTHDF1 siRNA). Cell proliferation was analyzed using an EdU assay. The expressions of dystrophin, myogenin (MyoG), and myosin heavy chain (MHC) proteins were measured by Western blotting. A RIP assay was used to verify the interaction between YTHDF1 and YES-associated protein 1 (YAP1) mRNA and investigate the modification level of the m⁶A site on YAP1 mRNA. (3) Thirty Mdx mice were randomly divided into: the empty vector group (n=15, intraperitoneal injection of empty adenoviral empty vector) and the AAV-YTHDF1 group (n=15, intraperitoneal injection of YTHDF1 overexpression adenovirus vector). We profiled the body weights, muscle and organ wet weights, fiber diameter, and fiber type of the mice. The protein levels of YTHDF1 and dystrophin were detected using Western blotting. We used HE staining to observe the pathological changes in the gastrocnemius and quadriceps femoris muscles. Results (1) Compared with control group, the expression of YTHDF1 and dystrophin was significantly lower in DMD patients (P<0.01), and the serum CPK content was significantly increased (P<0.0001). (2) Compared with empty vector group, the EdU positive rate and the expression of dystrophin, MyoG, and MHC proteins significantly increased (P<0.01). In addition, overexpression of YTHDF1 prolonged the half-life and improved the stability of YAP1 mRNA. Compared with Scramble group, the EdU-positive rate and the expression of dystrophin, MyoG, and MHC proteins significantly decreased (P<0.05 or P<0.01). (3) Compared with empty vector group, the mice in AAV-YTHDF1 group developed more rapidly with higher muscle mass in the gastrocnemius muscle, quadriceps muscle, and triceps (P<0.05). We observed no difference in the weights of inguinal, gonadal, or retroperitoneal fat pads and a significant increase in the fiber areas of gastrocnemius and quadriceps femoris (P>0.05). Conclusion YTHDF1 regulates the stability of YAP1 mRNA by recognizing the m⁶A modification of YAP1 mRNA and thus promotes YAP1/dystrophin-mediated myocyte proliferation to inhibit DMD progression.

Objective To explore the pattern of early expression and secretion of tissue factor (TF) in vascular endothelial cells induced by heat stress. Methods Thirty SPF-rated C57BL/6 male mice were randomly divided into five groups: the control group and groups of indicated recovery time, including 0, 3, 6, and 9 h in room temperature after heat stress (n=6). Mice in the heat stress groups were exposed to an animal incubator to reach 42.5 ℃ for core body temperature for heat stroke. We analyzed the histopathological changes in the liver, lung, and kidney tissues with HE staining. We measured the TF mRNA in mice tissues by RT-qPCR and the plasma concentration of TF in mice with a commercial ELISA kit. Human umbilical vein endothelial cells (HUVECs) were placed in a culture incubator to build an in vitro heat stress model. HUVECs were divided into five groups, including a control group and groups of indicated recovery time, including 0, 3, 6, and 9 h after heat stress. We quantified the expression of TF mRNA and protein in HUVEC cells by RT-qPCR, Western blotting, and immunofluorescence and measured the secreted TF with a commercial ELISA kit. Results No significant pathological injury was observed in the tissues of the control group. Mice treated with heat stress had various degrees of structural injuries and hemorrhagic and inflammatory changes in multiple tissues. Compared to control group, the expression of TF mRNA significantly increased in the kidney of heat stress-treated mice with 0 and 3 h recovery time (1.719±0.018, 1.241±0.178 vs. 1.000±0.063), the lung with 3 h recovery time (2.444±0.511 vs. 1.000±0.106) and the liver with 6 h recovery time (7.312±0.618 vs. 1.000±0.147) (P<0.05). The concentration of TF in plasma also sustainedly elevated in mice with 0, 3, 6, and 9 h recovery time after heat stress as compared to control group [(132.426±17.920) pg/ml, (119.400±10.267) pg/ml, (107.374±13.495) pg/ml, (163.767±22.810) pg/ml vs. (75.479±13.831) pg/ml, respectively, P<0.01]. The expression levels of TF mRNA were higher in heat stress HUVECs with 6 h and 9 h recovery time than the control cells (1.905±0.354, 2.564±0.297 vs. 1.000±0.097, P<0.01). Secreted TF in the supernatant from HUVECs treated with heat stress and different recovery time also increased significantly [(36.309±4.101) pg/ml, (38.425±5.484) pg/ml, (41.655±4.380) pg/ml, (43.586±4.718) pg/ml vs. (14.996±0.254) pg/ml, P<0.01]. Conclusion Heat stress increased early expression and secretion of TF in vascular endothelial cells. Vascular endothelial cells may be a main source of circulating TF in heat stroke.