Objective To validate whether the expression of human cluster of differentiation 55 (hCD55) protein in porcine islet cells could inhibit the activation of complement components in human serum. Methods Four adult pigs with WT (wild type), GTKO [α-1, 3-galactosyltransferase (GGTA1) knockout], GTKO/hCD55 and hCD55 genotypes were selected. Islet cells were isolated from WT, GTKO and GTKO/hCD55 pigs, and the purity and insulin secretion function were detected. The expression of hCD55 at the DNA, RNA and protein levels was analyzed by agarose gel electrophoresis, reverse transcription polymerase chain reaction (RT-PCR) and flow cytometry, respectively. Complement-dependent cytotoxicity assay and complement deposition assay were performed under the incubation conditions with fresh human serum. Results The purity of isolated porcine islet cells from three genotype pigs was > 75%, and the glycemic index was > 1. The expression of hCD55 messenger RNA(mRNA) and protein in GTKO/hCD55 porcine islet cells decreased the deposition of human complement component C3c and membrane-attacking complex C5b-9, and reduced the cytotoxicity. Conclusions The expression of hCD55 protein in porcine islet cells could inhibit the activation of human complement and reduce complement-mediated killing effect, indicating that hCD55 protein could exert complement protection effect on porcine islet cells. These findings provide theoretical basis for the application of hCD55 in islet xenotransplantation.

Objective:To evaluate the clinical effect of Guanxin Tongmai plaster combined with conventional western medicine in the treatment of phlegm and blood stasis syndrome of coronary heart disease and angina pectoris.Methods:A total of 60 patients in the Department of Cardiology of the Affiliated Hospital of Liaoning University of Traditional Chinese Medicine (TCM) from February to August 2020 who met the inclusion criteria were randomly divided into two groups with 30 in each group. Both groups were treated with conventional western medicine. On this basis, Guanxin Tongmai plaster was applied at the acupoints in the treatment group and placebo plaster was applied in the control group. TCM syndrome score was performed before and after treatment, angina score was evaluated from three aspects of angina attack frequency, duration and pain degree, and blood lipid TG, TC, LDL-C and HDL-C were detected by enzyme quantitative method. The blood homocysteine (Hcy) was detected by enzyme circulation method, the ECG and the nitroglycerin reduction rate were recorded, and the safety index was detected according to the ECG changes.Results:In the treatment period, 2 patients in the treatment group fell off, 3 in the control group. A total of 28 patients in the treatment group and 27 in the control group were analyzed. The total effective rate of ECG efficacy in the treatment group was 67.9% (19/28) and the control group was 48.1% (13/27). There was significant difference between the two groups ( χ2=4.46, P=0.040). After treatment, the TCM syndrome score and angina score in the treatment group were significantly lower than those in the control group ( t values were 9.12 and 4.45, P values were 0.004 and 0.042, respectively). The reduction rate of nitroglycerin in the treatment group was 82.1% (23/28) and 55.6% (15/27) in the control group. There was significant difference between the two groups ( χ 2=4.72, P=0.030). After treatment, the plasma TG, TC, LDL-C in the treatment group were significantly lower than those in the control group ( t values were 4.17, 6.57 and 6.52, P<0.05 or P<0.01), the level of HDL-C was significantly higher than that of the control group ( t=7.07, P=0.010), and the level of plasma Hcy was significantly lower than that in the control group ( t=6.70, P=0.012). There was no significant difference in liver, kidney and coagulation function between the two groups. Conclusion:Guanxin Tongmai plaster combined with conventional western medicine can improve the clinical symptoms of patients with coronary heart disease and angina pectoris, reduce the level of blood lipid and Hcy, and improve the clinical curative effect.

Objective To investigate the establishment of a six-gene-edited pig-to-non-human primate kidney xenotransplantation model. Methods The kidney of humanized genetically-edited pig (GTKO/β4GalNT2KO/CMAHKO/hCD55/hCD46/hTBM) was transplanted into a cynomolgus monkey. The survival of the recipient and kidney condition after blood perfusion were observed. The parenchymal echo, blood flow changes, and size of the kidney were monitored on a regular basis. Routine blood test, kidney function test and electrolyte assessment were carried out. Dynamic changes of urine, feces and body mass were monitored. At the end of life, the transplant kidney, heart, liver, spleen, lung, and cecum were collected for pathological examination. Results The recipient died at postoperative 7 d. After blood flow was restored, the kidney was properly perfused, the organ was soft and the color was normal. At the end of the recipient's life, a slight amount of purulent secretion was attached to the ventral side of the kidney, with evident congestion and swelling, showing the appearance of "red kidney". Postoperatively, the echo of renal parenchyma was increased, blood flow was decreased, the cortex was gradually thickened, and a slight amount of effusion surrounded the kidney and abdominal cavity over time. In the recipient, the amount of peripheral red blood cells, hemoglobin, albumin, and platelets was progressively decreased, and serum creatinine level was increased to 308 μmol/L at postoperative 7 d, whereas the K⁺ concentration did not significantly change. Light yellow urine was discharged immediately after surgery, diet and drinking water were resumed within postoperative 3 h, and light yellow and normal-shape stool was discharged. The reddish urine was gradually restored to normal color within postoperative 1 d, which were consistent with the results of the routine urine test. A large amount of brown bloody stool was discharged twice in the morning of 2 d after surgery. Omeprazole was given for acid suppression, and the stool returned to normal at postoperative 4 d. The β₂-microglobulin level was increased to 0.75 mg/L at postoperative 7 d. The body mass was increased by 1.7 kg. Autopsy pathological examination showed interstitial edema and bleeding of the transplant kidney, a large amount of infiltration of lymphocytes and macrophages, infiltration of lymphocytes in the arteriole wall and arterial cavity, accompanied by arteritis changes, lymphocyte infiltration in the cecal stroma and congestion in the spleen tissues. No significant abnormal changes were observed in other organs. Conclusions The humanized genetically-edited pig-to-non-human primate kidney xenotransplantation model is successfully established, and postoperative survival of the recipient is 1 week.

As one of the classic prescriptions of traditional Chinese medicine， Didangtang is an effective prescription for breaking and expelling blood stasis. It was considered that Didangtang damage the healthy qi and contained toxic Chinese medicinal materials such as leeches and gadflies， and thus it was rarely used. However， as the attention to classic prescriptions increases， Didangtang has been widely used in clinical practice and demonstrated definite efficacy in treating diabetes mellitus and its complications， malignant tumors， Alzheimer's disease， stroke， renal diseases， cardiovascular diseases， peripheral vascular diseases， gynecological disease， and male diseases according to the disease location， pathogenesis， symptoms， and pharmacological effects. Didangtang has the effects of mitigating insulin resistance， improving microvascular and peripheral vascular circulation， delaying diabetic macrovascular lesions， preventing vascular fibrosis， improving immunity， inhibiting tumor growth， protecting the brain tissue， nerve cells， vascular endothelial function， and kidney， reducing inflammation， and delaying aging. This paper summarizes the clinical application of Didangtang and initially explores the underlying mechanism.

Liver transplants are in huge demand in China, but facing the problem of extreme shortage of donor organs. Xenogeneic (pig) liver transplantation may be a potential way to alleviate the shortage of donors. The liver is a detoxifying organ with synthetic functions, and when genetically modified pigs are used as donors, it faces the dual problem of overcoming immune rejection and resolving physiological function mismatches. Therefore, suppressing the innate immune response can better alleviate the immune rejection of xenotransplantation. In addition, the use of chimeras of humanized cell livers may resolve the problem of physiological function mismatch. Moreover, with the development of gene editing technology, it has become possible to obtain multiple gene edited pigs and chimeras. Therefore, exploring and researching from these two aspects will hopefully solve the current problems of liver xenotransplantation and promote the further development of the field of liver xenotransplantation.

Objective To investigate the differences and the immunocompatibility of wild-type (WT), four-gene modified (TKO/hCD55) and six-gene modified (TKO/hCD55/hCD46/hTBM) pig erythrocytes with human serum. Methods The blood samples were collected from 20 volunteers with different blood groups. WT, TKO/hCD55, TKO/hCD55/hCD46/hTBM pig erythrocytes, ABO-compatible (ABO-C) and ABO-incompatible (ABO-I) human erythrocytes were exposed to human serum of different blood groups, respectively. The blood agglutination and antigen-antibody binding levels (IgG, IgM) and complement-dependent cytotoxicity were detected. The immunocompatibility of two types of genetically modified pig erythrocytes with human serum was evaluated. Results No significant blood agglutination was observed in the ABO-C group. The blood agglutination levels in the WT and ABO-I groups were higher than those in the TKO/hCD55 and TKO/hCD55/hCD46/hTBM groups (all P<0.001). The level of erythrocyte lysis in the WT group was higher than those in the ABO-C, TKO/hCD55 and TKO/hCD55/hCD46/hTBM groups. The level of erythrocyte lysis in the ABO-I group was higher than those in the TKO/hCD55 and TKO/hCD55/hCD46/hTBM groups (both P<0.01). The pig erythrocyte binding level with IgM and IgG in the TKO/hCD55 group was lower than those in the WT and ABO-I groups. The pig erythrocyte binding level with IgG and IgM in the TKO/hCD55/hCD46/hTBM group was lower than that in the WT group and pig erythrocyte binding level with IgG was lower than that in the ABO-I group (all P<0.05). Conclusions The immunocompatibility of genetically modified pig erythrocytes is better than that of wild-type pigs and close to that of ABO-C pigs. Humanized pig erythrocytes may be considered as a blood source when blood sources are extremely scarce.

OBJECTIVES: To investigate the mechanism of comorbidity between non-alcoholic fatty liver disease (NAFLD) and atherosclerosis (AS) based on metabolomics and network pharmacology. METHODS: Six ApoE^－/－ mice were fed with a high-fat diet for 16 weeks as a comorbid model of NAFLD and AS (model group). Normal diet was given to 6 wildtype C57BL/6J mice (control group). Serum samples were taken from both groups for a non-targeted metabolomics assay to identify differential metabolites. Network pharmacology was applied to explore the possible mechanistic effects of differential metabolites on AS and NAFLD. An in vitro comorbid cell model was constructed using NCTC1469 cells and RAW264.7 macrophage. Cellular lipid accumulation, cell viability, morphology and function of mitochondria were detected with oil red O staining, CCK-8 assay, transmission electron microscopy and JC-1 staining, respectively. RESULTS: A total of 85 differential metabolites associated with comorbidity of NAFLD and AS were identified. The top 20 differential metabolites were subjected to network pharmacology analysis, which showed that the core targets of differential metabolites related to AS and NAFLD were STAT3, EGFR, MAPK14, PPARG, NFKB1, PTGS2, ESR1, PPARA, PTPN1 and SCD. The Kyoto Encyclopedia of Genes and Genomes showed the top 10 signaling pathways were PPAR signaling pathway, AGE-RAGE signaling pathway in diabetic complications, alcoholic liver disease, prolactin signaling pathway, insulin resistance, TNF signaling pathway, hepatitis B, the relax in signaling pathway, IL-17 signaling pathway and NAFLD. Experimental validation showed that lipid metabolism-related genes PPARG, PPARA, PTPN1, and SCD were significantly changed in hepatocyte models, and steatotic hepatocytes affected the expression of macrophage inflammation-related genes STAT3, NFKB1 and PTGS2; steatotic hepatocytes promoted the formation of foam cells and exacerbated the accumulation of lipids in foam cells; the disrupted morphology, impaired function, and increased reactive oxygen species production were observed in steatotic hepatocyte mitochondria, while the formation of foam cells aggravated mitochondrial damage. CONCLUSIONS Abnormal lipid metabolism and inflammatory response are distinctive features of comorbid AS and NAFLD. Hepatocyte steatosis causes mitochondrial damage, which leads to mitochondrial dysfunction, increased reactive oxygen species and activation of macrophage inflammatory response, resulting in the acceleration of AS development.
Animals ; Mice ; Non-alcoholic Fatty Liver Disease/metabolism* ; Cyclooxygenase 2/metabolism* ; PPAR gamma/metabolism* ; Reactive Oxygen Species/metabolism* ; Mice, Inbred C57BL ; Hepatocytes ; Macrophages/metabolism* ; Liver

Cardiovascular diseases (CVDs) and metabolic disorders are major components of noncommunicable diseases, causing an enormous health and economic burden worldwide. There are common risk factors and developmental mechanisms among them, indicating the far-reaching significance in exploring the corresponding therapeutic targets. MST1/2 kinases are well-established proapoptotic effectors that also bidirectionally regulate autophagic activity. Recent studies have demonstrated that MST1/2 influence the outcome of cardiovascular and metabolic diseases by regulating immune inflammation. In addition, drug development against them is in full swing. In this review, we mainly describe the roles and mechanisms of MST1/2 in apoptosis and autophagy in cardiovascular and metabolic events as well as emphasis on the existing evidence for their involvement in immune inflammation. Moreover, we summarize the latest progress of pharmacotherapy targeting MST1/2 and propose a new mode of drug combination therapy, which may be beneficial to seek more effective strategies to prevent and treat CVDs and metabolic disorders.