BACKGROUND: The primary limitation to kidney transplantation is organ shortage. Recent progress in gene editing and immunosuppressive regimens has made xenotransplantation with porcine organs a possibility. However, evidence in pig-to-human xenotransplantation remains scarce, and antibody-mediated rejection (AMR) is a major obstacle to clinical applications of xenotransplantation. METHODS: We conducted a kidney xenotransplantation in a brain-dead human recipient using a porcine kidney with five gene edits (5GE) on March 25, 2024 at Xijing Hospital, China. Clinical-grade immunosuppressive regimens were employed, and the observation period lasted 22 days. We collected and analyzed the xenograft function, ultrasound findings, sequential protocol biopsies, and immune surveillance of the recipient during the observation. RESULTS: The combination of 5GE in the porcine kidney and clinical-grade immunosuppressive regimens prevented hyperacute rejection. The xenograft kidney underwent delayed graft function in the first week, but urine output increased later and the single xenograft kidney maintained electrolyte and pH homeostasis from postoperative day (POD) 12 to 19. We observed AMR at 24 h post-transplantation, due to the presence of pre-existing anti-porcine antibodies and cytotoxicity before transplantation; this AMR persisted throughout the observation period. Plasma exchange and intravenous immunoglobulin treatment mitigated the AMR. We observed activation of latent porcine cytomegalovirus toward the end of the study, which might have contributed to coagulation disorder in the recipient. CONCLUSIONS 5GE and clinical-grade immunosuppressive regimens were sufficient to prevent hyperacute rejection during pig-to-human kidney xenotransplantation. Pre-existing anti-porcine antibodies predisposed the xenograft to AMR. Plasma exchange and intravenous immunoglobulin were safe and effective in the treatment of AMR after kidney xenotransplantation.
Transplantation, Heterologous/methods* ; Kidney Transplantation/methods* ; Heterografts/pathology* ; Immunoglobulins, Intravenous/administration & dosage* ; Graft Survival/immunology* ; Humans ; Animals ; Sus scrofa ; Graft Rejection/prevention & control* ; Kidney/pathology* ; Gene Editing ; Species Specificity ; Immunosuppression Therapy/methods* ; Plasma Exchange ; Brain Death ; Biopsy ; Male ; Aged

Objective:To study the effects of cathepsin K(CTSK)on the healing process of tooth extraction socket and type H blood vessel angiogenesis in mice.Methods:Ctsk knockout(Ctsk-/-)mice were generated by CRISPR/Cas9 technology,and genotype sequen-cing,general observation,Micro-CT and immunohistochemistry were performed to confirm successful knockout of Ctsk.Then 8 week-old WT and Ctsk-/-mice were used to establish the tooth extraction modle by extracting the left maxillary first molars,and the mice were sac-rificed at the day 7,10,14,21,28 and 35 respectively(n=3)after tooth extraction.Then samples were subjected to stereo microscope and Micro-CT examination.Immunofluorescence staining was used to study the effect of Ctsk knockout on type H blood vessel angiogene-sis.Results:Ctsk knockout did not affect the soft tissue healing of tooth extraction socket,but significantly promoted the bone healing process,and Ctsk deficency significantly enhanced type H blood vessel angiogenesis in the tooth extraction socket.Conclusion:Ctsk knockout can enhance type H vessel angiogenesis,and promote bone healing process of tooth extraction socket in mice.

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.

Objective:To investigate the changing trends in cardiac function following xenogeneic heterotopic heart transplantation of multi-gene edited pig hearts and assess the impact of recipient immune responses on donor heart, laying experimental groundwork for the clinical application of gene editing technology.Methods:On December 16, 2023, xenogeneic heterotopic heart transplantation was performed between pigs and rhesus monkeys. Functional status of the graft under post-transplantation load conditions and recipient immune indicators were observed.Results:The recipient monkeys survived for 40 days with satisfactory functionality of both donor and recipient hearts, and no hyperacute or acute immune rejection reactions were observed.Conclusion:Multi-gene editing technology provides potential for xenotransplantation, yet further exploration is needed for its clinical application.

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.

Purpose: Mixed-lineage leukemia protein 4 (MLL4/KMT2D) is a histone methyltransferase, and its mutation has been reported to be associated with a poor prognosis in many cancers, including lung cancer. We investigated the function of MLL4 in lung carcinogenesis. Materials and Methods: RNA sequencing (RNA-seq) in A549 cells transfected with control siRNA or MLL4 siRNA was performed. Also, we used EdU incorporation assay, colony formation assays, growth curve analysis, transwell invasion assays, immunohistochemical staining, and in vivo bioluminescence assay to investigate the function of MLL4 in lung carcinogenesis. Results: We found that MLL4 expression was downregulated in non–small cell lung cancer (NSCLC) tissues compared to adjacent normal tissues and tended to decrease with disease stage progression. We analyzed the transcriptomes in control and MLL4- deficient cells using high-throughput RNA deep sequencing (RNA-seq) and identified a cohort of target genes, such as SOX2, ATF1, FOXP4, PIK3IP1, SIRT4, TENT5B, and LFNG, some of which are related to proliferation and metastasis. Our results showed that low expression of MLL4 promotes NSCLC cell proliferation and metastasis and is required for the maintenance of NSCLC stem cell properties. Conclusion Our findings identify an important role of MLL4 in lung carcinogenesis through transcriptional regulation of PIK3IP1, affecting the PI3K/AKT/SOX2 axis, and suggest that MLL4 could be a potential prognostic indicator and target for NSCLC therapy.

Objective:To investigate the effects of types of ulnar styloid process fracture on the treatment of distal radius fracture.Methods:The 80 patients were analyzed retrospectively who had been treated at The First Department of Hand Surgery, Honghui Hospital from January 2019 to January 2020 for fracture of distal radius complicated with fracture of ulnar styloid process. They were 25 males and 55 females, aged from 30 to 85 years (average, 58.6 years). According to the types of ulnar styloid process fracture, 40 patients were assigned into a Hauck type Ⅰ group and the other 40 into a Hauck type Ⅱ group. The 2 groups were compared in terms of operation, postoperative complications, hospital stay, bone union, visual analogue scale (VAS) on postoperative 1 to 3 days, and modified Mayo wrist function score, wrist range of motion and quality of life by WHOQOL-BREF at the last follow-up.Results:The 2 groups were comparable because there was no significant difference in age, gender, American Society of Anesthesiologists (ASA) rating, or time from injury to operation between them ( P>0.05). All the patients were followed up for 12 to 24 months (average, 17 months). There was no significant difference between Hauck type Ⅰ group and Hauck type Ⅱ group in operation time, intraoperative blood loss, hospital stay, rate of postoperative complications, fracture union, modified Mayo wrist function score or VAS on postoperative 1 to 3 days ( P>0.05). At the last follow-up, the palm tilt and ulnar inclination angles were 13.8°±1.9° and 21.6°±2.8° in Hauck type Ⅰ group, significantly larger than those in Hauck type Ⅱ group (11.9°±1.6° and 18.8°±2.3°) ( P<0.05). At the last follow-up, Hauck Ⅰ group scored 85.3±6.4,85.6±6.5, 84.7±6.3 and 85.0±6.7 respectively in the domains of physical health, psychology, environment and social relationships, significantly higher than those Hauck type Ⅱ group did (78.5±6.5, 78.9±6.5, 77.8±6.1 and 77.9±6.3) ( P<0.05). Conclusions:In open reduction and internal fixation for distal radius fracture, Hauck Type Ⅰ fracture of ulnar styloid process has no significant effect on the functional recovery of the wrist but Hauck Type Ⅱ fracture of ulnar styloid process may. Therefore, surgical fixation needs to be strengthened if Hauck Type Ⅱ fracture of ulnar styloid process is complicated.

Objective:To investigate the efficacy of different doses of vitamin C (VC) in prevention and treatment of non-alcoholic fatty liver disease (NAFLD) in mice.Methods:C57BL/6 mice were fed with high-fat diet to establish NAFLD models. The experimental animals were divided into early prevention and later treatment groups. Both of these two experimental processes had five subgroups, including control, high-fat diet (HFD), low-dose vitamin C (LD-VC, 15 mg/kg per day), medium-dose vitamin C (MD-VC, 30 mg/kg per day) and high-dose vitamin C (HD-VC, 90 mg/kg per day) subgroup, with six mice in each subgroup. In the early prevention group, the mice were prophylactically received VC for 12 weeks. In the later treatment group, the mice were treated with different dose of VC for 12 weeks after fed with HFD for six weeks and confirmed NAFLD by liver pathology. The differences in body weight, perirenal adipose tissue mass and serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), total cholesterol (TC), and triacylglycerol (TG) were observed among different groups. The scores of hepatocyte steatosis, lobular inflammation and ballooning in liver histopathology of mice in each group were evaluated by non-alcoholic fatty liver disease activity score (NAS) scoring system. Tukey′s multiple comparison test and Kruskal-Wallis H test were performed for statistical analysis. Results:In the early prevention group, the body weight, perirenal adipose tissue mass, TG level and the score of liver steatosis of LD-VC subgroup were all lower than those of HFD subgroup ((30.27±0.94) g vs. (32.18±1.35) g, (0.25±0.05) g vs. (0.32±0.02) g, (0.25±0.02) mmol/L vs. (0.30±0.03) mmol/L, 0 vs. 1.0(1.0)). The body weight, perirenal adipose tissue mass, blood glucose level, TG level and score of liver steatosis of MD-VC subgroup were all lower than those of HFD subgroup ( (29.72±0.58) g vs. (32.18±1.35) g, (0.24±0.05) g vs. (0.32±0.02) g, (6.93±0.59) mmol/L vs. (8.33±1.02) mmol/L, (0.24±0.04) mmol/L vs. (0.30±0.03) mmol/L, 0 vs. 1.0(1.0)); meanwhile, the blood glucose level and TG level of HD-VC subgroup were both lower than those of HFD subgroup ((6.72±0.59) mmol/L vs. (8.33±1.02) mmol/L, (0.23±0.04) mmol/L vs. (0.30±0.03) mmol/L), and the differences were statistically significant (all P<0.05). In the later treatment group, TG level of LD-VC subgroup was lower than that of HFD subgroup ((0.25±0.07) mmol/L vs. (0.37±0.06) mmol/L); the body weight, perirenal adipose tissue mass, blood glucose level, TG level and liver steatosis score of MD-VC subgroup were lower than those of HFD subgroup ((29.93±1.28) g vs. (33.24±2.45) g, (0.29±0.08) g vs. (0.53±0.14) g, (7.63±0.57) mmol/L vs. (9.13±1.52) mmol/L, (0.23±0.03) mmol/L vs. (0.37±0.06) mmol/L, 0.5(1.0) vs. 2.0(1.0)); the blood glucose level and TG level of HD-VC subgroup were both lower than those of HFD subgroup ((7.20±0.72) mmol/L vs. (9.13±1.52) mmol/L, (0.19±0.03) mmol/L vs. (0.37±0.06) mmol/L); however the body weight, liver weight, perirenal adipose tissue mass and lobular inflammation score of HD-VC subgroup were all high than those of HFD subgroup( (36.34±2.44) g vs. (33.24±2.45) g, (1.18±0.07) g vs. (1.06±0.09) g, (0.78±0.17) g vs. (0.53±0.14) g, 1.0(1.0) vs.0(1.0)), and the differences were statistically significant (all P<0.05). The body weight, perirenal adipose tissue mass and the score of liver steatosis, lobular inflammation and ballooning of LD-VC subgroup of the early prevention group were all lower than those of LD-VC subgroup of the later treatment group ((30.27±0.94) g vs. (34.75±1.64) g, (0.25±0.05) g vs. (0.61±0.14) g, 0 vs.1.5(1.0), 0 vs. 0.5(1.0), 0 vs. 1.0(0)); and the body weight, liver weight, perirenal adipose tissue mass, ALT level, AST level and scores of liver steatosis and lobulor inflammation of HD-VC subgroup of the early prevention group were all lower than those of HD-VC subgroup of the late treatment group ((31.78±0.71) g vs. (36.34±2.44) g, (1.01±0.02) g vs. (1.18±0.07) g, (0.30±0.05) g vs. (0.78±0.17) g, (8.83±0.98) U/L vs. (12.75±2.05) U/L, (29.00±4.19) U/L vs. (41.88±14.36) U/L, 1.0(0) vs. 2.5(1.0), 0 vs. 1.0(1.0)), and the differences were statistically significant (all P<0.05). Conclusions:MD-VC can prevent the occurrence of NAFLD in mice at an early stage, and it is also benefit to the later treatment of NAFLD in mice. However, HD-VC has potential risks in early prevention and later treatment of NAFLD in mice.

Objective:To study the timing of radiofrequency ablation (RFA) in treatment of complicated pyogenic liver abscesses (PLA).Methods:A retrospective analysis was performed on 49 patients with complicated PLA who were treated with the RFA-based treatment modality from August 2010 to January 2020 at Beijing Chaoyang Hospital, West Campus, Capital Medical University, Institute of Hepatobiliary, Pancreas and Spleen Surgery and the Second Hospital, Binzhou. The patients were divided into the early RFA group (≤ 72 h, n=27) and the delayed RFA group (>72 h, n=22) according to the timing of RFA. RFA was guided by laparoscopy combined with Ultrasound or CT. The safety and effective rates of RFA, and the total expenses were evaluated. Results:All patients in both the early and the delayed RFA groups were successfully cured (100%). No serious complications, including biliary leakage and massive hemorrhage, happened in the 2 groups. Significantly longer operating time and hospital stays [(8.1±1.6)d vs. (9.5±1.5)d], and higher hospital costs [(3.4±0.2) ten thousand yuan vs. (3.8±0.4) ten thousand yuan] were found in the delayed RFA group when compared with the early RFA group ( P<0.05). Conclusion:RFA treatment of complicated PLA should be completed within 72 hours of onset of PLA.