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 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 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 explore the changes in the expression of Tau protein and phosphorylated Tau (p-Tau) protein in neurons after spinal cord ischemia-reperfusion injury (SCII).Methods:Ninety-six healthy adult SD rats were randomly divided into a sham operation group ( n=48) and a SCII group ( n=48). Based on the reperfusion time of 3 h, 6 h, 12 h, 24 h, 48 h and 72 h, the SCII group was divided into 6 subgroups ( n=8 per subgroup). Immunohistochemical staining was used to observe the apoptosis of spinal cord neurons in the L 4-L 5 segments and the expression of Tau protein and p-Tau protein. Results:In the sham operation group, the neuron cells were intact, mainly concentrated in the gray matter. Tau protein was seen in a small number of neuron cells, and a small amount of filamentous p-Tau protein in the pernucleus and cytoplasm. There was no significant difference between Tau protein and p-Tau protein expression in neurons at each time point ( P>0.05). In the SCII group, scattered Tau protein was seen in the apoptotic cells while there was a strong positive expression of Tau protein in the non-apoptotic cells. The expression of Tau protein in the SCII group gradually increased after injury, reaching a peak at 48h and plateauing at 72 h, and was significantly different between any 2 time points (except for 72 h) ( P<0.05). In the SCII group, the positive expression of p-Tau protein was observed in the cytoplasm of the apoptotic cells in strips and sheets. It increased rapidly within 6 h but did not change significantly after 6 h, showing no significant difference between any 2 time points afterwards ( P>0.05). There was a statistically significant difference in the expression of Tau protein and p-Tau protein between the SCII group and the sham operation group at each time point ( P<0.05). Conclusion:It is hopeful to reduce the severity of spinal cord injury by regulating the expression of Tau protein and p-Tau protein within 6 to 48 hours after SCII.

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.

OBJECTIVE: To investigate the effect of exosomes derived from Epstein-Barr virus (EBV)-positive nasopharyngeal carcinoma (NPC) cells on lymphangiogenesis and lymph node metastasis of NPC. METHODS: Exosomes from NP69 cells and EBV-positive HK1 (HK1-EBV) cells were obtained by ultracentrifugation and identified by Western blotting and nanoparticle tracking analysis. Dio dye phagocytosis test was performed to observe exosome uptake by lymphatic endothelial cells. Lymphatic endothelial cells were treated with exosomes from nasopharyngeal epithelium (NP69), HK1-EBV, and C666-1 cells or exosome-free supernatant of HK1-EBV and C666-1 cells, and tube formation and migration of the cells were observed. In a nude mouse model of popliteal lymph node metastasis of NPC, the effects of normal saline, NP69 cell-derived exosomes, HK1-EBV cell-derived exosomes, exosome-free supernatant of HK1-EBV cells, and HK1-EBV exosome-free supernatant protein on lymphangiogenesis and lymph node metastasis of the tumor were observed. RESULTS: The exosomes obtained by ultracentrifugation contained abundant exosome-specific proteins and showed a normal size range. The exosomes from NPC cells and NP69 cells could be taken up by lymphatic endothelial cells. Compared with the blank control and exosomes form NP69 cells, exosomes derived from HK1-EBV and C666-1 cells significantly promoted tube formation and migration of lymphatic endothelial cells ( CONCLUSIONS Exosomes from EBV-positive NPC cells can significantly promote lymphangiogenesis and lymph node metastasis of NPC.
Animals ; Cell Line, Tumor ; Endothelial Cells ; Epstein-Barr Virus Infections ; Exosomes ; Herpesvirus 4, Human ; Humans ; Lymphangiogenesis ; Lymphatic Metastasis ; Mice ; Mice, Nude ; Nasopharyngeal Carcinoma ; Nasopharyngeal Neoplasms

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 explore and clarify the correlation between short-term aggressive intrasegmental recurrence (AIR) and functional magnetic resonance imaging after radiofrequency ablation of hepatocellular carcinoma (HCC). Methods: A retrospective analysis of 1 262 patients with HCC who underwent radiofrequency ablation (RFA) in our hospital from January 2012 to June 2018, all patients were confirmed by pathology as HCC, of which 30 patients were found to have AIR during radiographic follow-up within 3 months after surgery, another 35 patients with disease progression who were controlled in a short period of time were randomly selected as the control group. All the enrolled patients underwent dynamic enhanced magnetic resonance imaging (DCE-MRI) and diffusion weighted imaging (DWI) scanning before surgery, and the differences in clinical data, lesion location, and functional magnetic resonance parameters between the two groups were compared, and their correlation with AIR after RFA was analyzed. Chi-square test, t test and Pearson test were used. Results: The lesions in the AIR group were significantly more in the Ⅰand Ⅳsegments than that in the control group (P<0.05), and there was no significant difference in other liver segments (P>0.05). There was no significant difference in the lesion morphology between the two groups (P>0.05), and there was a significant difference in the early enhancement pattern of the lesions (P<0.05), and the early arterial enhancement rate and apparent diffusion coefficient (ADC) value of the AIR group were significantly lower than the control group (P<0.05). Further correlation analysis found that early enhancement of the arterial artery, early arterial enhancement rate, ADC value and lesion location were associated with AIR and were positively correlated, r values were 0.455, 0.633, 0.518, 0.375 and 0.287 (P<0.05). Conclusion The short-term AIR and functional imaging parameters (arterial early enhancement, early arterial enhancement rate, ADC value) and the liver segment (Ⅰ and Ⅳ) were highly correlated with radiofrequency ablation.

Objective To investigate the clinical effect of infective foot ulcer and soft tissue defects of small area with free peroneal artery perforator flap.Methods From February,2016 to Apirl,2017,9 cases of infective foot ulcer and soft tissue defect of small area were repaired with peroneal artery perforator flap.Cause of infection:injury in 6 cases,diabetes in 2 cases,spinal cord injury subsequent pressure sore in 1 case.Infective ulcer part:4 cases on plantar metatarsophalangeal joint,2 cases on dorsal metatarsophalangeal joint,2 cases on interior of the first metatarsophalangeal joint and 1 case on the heel.Defect area:3.0 cm×2.0 cm-5.0 cm×3.0 cm;flap area:4.0 cm×3.5 cm-7.0 cm×4.5 cm.The donor sites were sutured directly.All the flaps contained lateral sural cutaneous nerve.In order to rebuild the sensation of flaps,nerves between surgical area and receiving area were received end-to-side anastomosis.Outpatient follow-up was used in all patients.Results All the flaps survived,and all the wounds were primary healed,without recurrence of infection.All patients were followed-up from 6 to 24 months with an average of 16 months.The appearance of flaps was good with slight bloated.The texture and color of the flaps were close to the recipient site.Flap feel was good except the cases of diabetes and spinal cord injury pressure sore.The AOFAS score at last follow-up time was 80.05±7.80,which was excellent in 5 cases,good in 2 cases,and fair in 2 cases.Conclusion The free peronerl artery perforator flap has many advantages,such as vascular anatomy constant,blood supply is reliable,thickness moderate,easy to get,strong resistance to infection,small district damage,etc.It is a useful clinical method to repair foot infection ulcer and soft issue defects of small area.