Objective To explore the construction of α-1,3-galactosyltransferase (GGTA1) gene-knockout (GTKO) Diannan miniature pigs and the kidney xenotransplantation from pigs to rhesus macaques, and to assess the effectiveness of GTKO pigs. Methods The GTKO Diannan miniature pigs were constructed using the CRISPR/Cas9 gene-editing system and somatic cell cloning technology. The phenotype of GTKO pigs was verified through polymerase chain reaction, Sanger sequencing and immunofluorescence staining. Flow cytometry was used to detect antigen-antibody (IgM) binding and complement-dependent cytotoxicity. Kidney xenotransplantation was performed from GTKO pigs to rhesus macaques. The humoral immunity, cellular immunity, coagulation and physiological indicators of the recipient monkeys were monitored. The function and pathological changes of the transplanted kidneys were analyzed using ultrasonography, hematoxylin-eosin staining, immunohistochemical staining and immunofluorescence staining. Results Single-guide RNA (sgRNA) targeting exon 4 of the GGTA1 gene in Diannan miniature pigs was designed. The pGL3-GGTA1-sgRNA1-GFP vector was transfected into fetal fibroblasts of Diannan miniature pigs. After puromycin selection, two cell clones, C59# and C89#, were identified as GGTA1 gene-knockout clones. These clones were expanded to form cell lines, which were used as donor cells for somatic cell nuclear transfer. The reconstructed embryos were transferred into the oviducts of trihybrid surrogate sows, resulting in 13 fetal pigs. Among them, fetuses F04 and F11 exhibited biallelic mutations in the GGTA1 gene, and F04 had a normal karyotype. Using this GTKO fetal pig for recloning and transferring the reconstructed embryos into the oviducts of trihybrid surrogate sows, seven surviving piglets were obtained, all of which did not express α-Gal epitope. The binding of IgM from the serum of rhesus monkey 20# to GTKO pig PBMC was reduced, and the survival rate of GTKO pig PBMC in the complement-dependent cytotoxicity assay was higher than that of wild-type pig. GTKO pig kidneys were harvested and perfused until completely white. After the left kidney of the recipient monkey was removed, the pig kidney was heterotopically transplanted. Following vascular anastomosis and blood flow restoration, the pig kidney rapidly turned pink without hyperacute rejection (HAR). Urine appeared in the ureter 6 minutes later, indicating successful kidney transplantation. The right kidney of the recipient was then removed. Seven days after transplantation, the transplanted kidney had good blood flow, the recipient monkey's serum creatinine level was stable, and serum potassium and cystatin C levels were effectively controlled, although they increased 10 days after transplantation. Seven days after transplantation, the levels of white blood cells, lymphocytes, monocytes and eosinophils in the recipient monkey increased, while platelet count and fibrinogen levels decreased. The activated partial thromboplastin time, thrombin time and prothrombin time remained relatively stable but later showed an upward trend. The recipient monkey survived for 10 days. At autopsy, the transplanted kidney was found to be congested, swollen and necrotic, with a small amount of IgG deposition in the renal tissue, and a large amount of IgM, complement C3c and C4d deposition, as well as CD68⁺ macrophage infiltration. Conclusions The kidneys of GTKO Diannan miniature pigs may maintain normal renal function for a certain period in rhesus macaques and effectively overcome HAR, confirming the effectiveness of GTKO pigs for xenotransplantation.

Objective To observe the effect of ultrasound-guided iliopsoas plane block(IPB)on the quality of postoperative recovery in patients undergoing hip arthroplasty.Methods Sixty patients who underwent hip arthroplasty were selected,37 males and 23 females,aged 40-79 years,BMI 18-30 kg/m2,ASA physical status Ⅰ-Ⅲ.The patients were divided into two groups by random number table method:the iliopsoas plane block group(group IPB)and the femoral nerve block(FNB)group(group FNB),30 pa-tients in each group.Before anesthesia induction,IPB was performed with 0.5%ropivacaine 10 ml and lat-eral femoral cutaneous nerve block was performed with 0.5%ropivacaine 5 ml in group IPB.And FNB was performed with 0.5%ropivacaine 10 ml and lateral femoral cutaneous nerve block was performed with 0.5%ropivacaine 5 ml in group FNB.The dosages of propofol,remifentanil,and cis-atracurium during operation were recorded.The quality of recovery-15(QoR-15)scale was evaluated preoperatively and postoperatively 1 day,2 and 3 days.The max VAS(VASmax)pain score and manual muscle test(MMT)score of quadri-ceps muscle were recorded 12,24,and 48 hours after surgery.The time of getting out of bed for the first time,opioid dosage,and patient satisfaction were recorded.The incidence of nerve injury,vascular injury,puncture site infection,and local anesthetic poisoning were recorded.The postoperative complications of diz-ziness,nausea and vomiting,deep vein thromboses,and elirium were also recorded.Results There was no significant difference in the dosage of propofol,remifentanil,and cis-atracurium between the two groups.Compared with group FNB,the QoR-15 scale score in group IPB was significantly higher 1 day,2 and 3 days after operation(P＜0.05).Compared with group FNB,the MMT scores of quadriceps muscle was sig-nificantly higher in group IPB 12 and 24 hours after surgery(P＜0.05),and the first time of getting out of bed was shortened in group IPB(P＜0.05).However,there were no significant differences in the VASmax pain score,MMT score of quadriceps muscle 48 hours after surgery,opioid dosage,and patient satisfaction between the two groups.No nerve block related complications were found in both groups.There were no sig-nificant differences in postoperative complications between the two groups.Conclusion The iliopsoas plane block can improve the quality of postoperative recovery and accelerate the recovery of patients with hip re-placement,and the effect is better than that of femoral nerve block.

Objective:To evaluate the pin-bone interface surface culture in the diagnosis of pin tract infection in external fixation.Methods:A prospective observational study was conducted to enroll the patients who underwent either partial or complete removal of external fixators after external fixation at Department of Orthopaedic Trauma, Nanfang Hospital, Southern Medical University from June 2023 to September 2023. The secretions from the pin track (pin-soft tissue interface) were plated for bacterial culture. Additionally, the surface of the pins placed within the bone (pin-bone interface) was cultured directly with tryptic soy agar (TSA). Positive cases were subjected to additional analysis using qualitative microbial culture and antibiotic susceptibility testing. Comparisons were made between the cultural results derived from both interfaces.Results:The present study enrolled 23 patients [18 males and 5 females with an age of (37.3±17.6) years] and a duration of bearing external fixation of 8.1 (4.0, 11.3) months. A total of 212 samples were cultured. The positive rate of bacterial culture at the pin-soft tissue interface was 53.8% (57/106), significantly higher than that at the pin-bone interface [17.9% (19/106)] ( P<0.05). No correlation was found in the results of bacterial culture between the pin-bone interface and the pin-soft tissue interface ( r=-0.011, P=0.913). In terms of bacterial strains, single pathogenic bacteria were found in all the 19 positive samples cultured at the pin-bone interface, with Staphylococcus aureus as the most common pathogenic bacteria (7); of the 57 positive samples cultured at the pin-soft tissue interface, single pathogen infection was found in 51 and mixed bacterial infection in 6. Positive culture was found at both interfaces in 10 samples, of which identical bacterial strains were found in 4 and partially identical bacterial strains in 1. A total of 82 bacterial samples were subjected to drug sensitivity testing, of which 74.4% (61/82) were infected with Staphylococcus. The drug sensitivity test of Staphylococcus showed that the top 3 resistant drugs were ampicillin, oxacillin, and penicillin. The top 3 sensitive drugs were vancomycin, teicoplanin, and linezolid, all of which showed little resistance. Conclusions:Pin-bone interface culture of external fixators is a necessary evaluation of the infection of deep bone tissue. Simultaneous culture of pin-bone interface and pin-soft tissue interface can provide more comprehensive basis for the treatment of pin tract infection.

Objective:To evaluate the effect of heavy smoking on the dose-effect relationship of ciprofol for painless gastroscopy when combined with alfentanil.Methods:This was a prospective study. American Society of Anesthesiologists Physical Status classification Ⅰ or Ⅱ male patients, aged 18-64 yr, with body mass index of 18-28 kg/m 2, undergoing elective painless gastroscopy from October 2023 to February 2024 in Baoding First Central Hospital, were divided into non-smoking group and heavy smoking group (smoking index>400) according to the status of smoking. Alfentanil 5 μg/kg and ciprofol were intravenously injected, and gastroscopy was performed after the patient′s eyelash reflex disappeared and the jaw was relaxed. The study was performed by the Dixon′s up-and-down method, and the initial dose of ciprofol was 0.4 mg/kg. The dose of ciprofol increased or decreased by 0.04 mg/kg each time based on the positive or negative response of the previous patient. A positive response was defined as the Modified Observer′s Assessment of Alertness/Sedation Scale score being greater than 1 point at 3 min after ciprofol injection or the occurrence of coughing, swallowing, body movement, or other responses that affected the operation during the insertion of the endoscope. The median effective dose (ED 50) and 95% confidence interval of propofol for painless gastroscopy were determined by the probit analysis. Results:Twenty-five patients were finally included in non-smoking group and 23 patients in heavy smoking group. The ED 50 (95% confidence interval) of ciprofol when combined with alfentanil was 0.205 (0.159, 0.244) mg/kg in non-smoking group and 0.252 (0.184, 0.295) mg/kg in heavy smoking group. The ED 50 was significantly higher in heavy smoking group than in non-smoking group ( u=390, P=0.009). Conclusions:Heavy smoking can weaken the sedative potency of propofol for sedation when used for painless gastroscopy when combined with alfentanil.

Objective:To evaluate the effect of age on the sedative potency of remimazolam.Methods:This was a prospective study. Patients of either sex, aged 40-80 yr, with body mass index of 18-28 kg/m 2, of American Society of Anesthesiologists Physical Status classification Ⅰ or Ⅱ, scheduled for elective surgery with general anesthesia, were divided into middle age group (40-64 yr) and elderly group (65-80 yr) based on the age of the patients. The test was performed by the Dixon′s up-and-down method. The initial dose of remimazolam was 0.3 mg/kg. The Modified Observer′s Assessment of Alertness/Sedation Scale score ≤1 and bispectral index value≤60 within 3 min after administration were considered as an effective sedation. The dose of remimazolam was increased/decreased by 0.03 mg/kg based on the sedative efficacy in the previous patient. The 50% effective dose (ED 50) and 95% confidence interval ( CI) of remimazolam were calculated by the probit method. The time to the loss of consciousness was recorded. Results:The ED 50 (95% CI) of remimazolam was 0.345 (0.306-0.384) mg/kg in middle age group, while the ED 50 (95% CI) of remimazolam was 0.322 (0.303-0.339) mg/kg in elderly group. The ED 50 was significantly higher in middle age group than in elderly group ( u=417.00, P=0.022). The time to loss of consciousness was significantly longer in elderly group than in middle age group ( t=-2.96, P=0.008). Conclusions:For middle-aged and elderly patients aged 40-80 yr, the sedative potency of remimazolam increases with age.

Objective To investigate the effect of early glucose metabolism abnormality on skeletal muscle content of young men.Methods 88 males who underwent physical examinations in our hospital from September 2020 to September 2021 were divided into normal blood glucose group(NGT,n=47)and IGR group(n=41),according to their FPG and 2 hPG levels.FPG,FIns and visceral fat area(VFA)were measured.HOMA-IR,HOMA-β,skeletal muscle index(SMI)and body fat ratio(BFR)were calculated.Results Compared with NGT group,the males in IGR group showed elevated BMI,WC,FPG,2 hPG,HOMA-IR and VFA(P＜0.05),but decreased HDL-C and SMI(P＜0.05).Pearson correlation analysis showed that SMI was negatively correlated with BMI,WC,FPG,2 hPG,HOMA-IR,VFA and BFR in young men(P＜0.05 or P＜0.01).Logistic regression analysis showed that BMI was the influencing factor of FPG,while SMI was the influencing factor of 2 hPG.Conclusions In young men with abnormal glucose metabolism,the postprandial blood glucose significantly increase with the decrease of skeletal muscle content,and fasting blood glucose is mainly affected by BMI.

Objective:To determine the median effective dose (ED 50) of ciprofol inhibiting responses to insertion of laryngeal mask airway in the patients when combined with alfentanil. Methods:American Society of Anesthesiologists Physical Status classification Ⅰ or Ⅱ patients of either sex, aged 40-64 yr, with body mass index of 20-30 kg/m 2, undergoing elective general anesthesia, were enrolled. Midazolam 0.025 mg/kg was intravenously injected for anesthesia induction, the baseline mean arterial pressure and heart rate were recorded 5 min later, and the average value of three times was considered as the baseline value. Ciprofol and alfentanil 10 μg/kg were intravenously injected in sequence, rocuronium 0.6 mg/kg was intravenously injected when BIS value < 60, 2 min later a laryngeal mask airway was placed, and mechanical ventilation was performed. Positive response was defined as increase in the maximum mean arterial blood pressure or heart rate more than or equal to 20% of the baseline value within 3 min after placement of the laryngeal mask airway or as the occurrence of body movement, bucking, frowning, mouth and face twitching, tearing, laryngospasm or the BIS value failing to drop below 60. The study was performed by the Dixon′s up-and-down method. The initial dose of ciprofol was 0.4 mg/kg, and the ratio between the two successive doses was 1.1. If a positive response occurred, the dose was increased in the next patient, otherwise the dose was reduced. The ED 50 and 95% confidence interval of ciprofol inhibiting responses to insertion of laryngeal mask airway were calculated by the probit method. Results:The ED 50(95% confidence interval) of ciprofol inhibiting responses to insertion of laryngeal mask airway was 0.291(0.231-0.318) mg/kg when combined with alfentanil 10 μg/kg. Conclusions:The ED 50 of ciprofol inhibiting responses to insertion of laryngeal mask airway is 0.291 mg/kg in the patients when combined with alfentanil 10 μg/kg.

Objective:To evaluate the value of endoclip-assisted and submucosal injection-assisted cannulation techniques for difficult cannulation during endoscopic retrograde cholangiopancreatography (ERCP).Methods:Data of 12 458 patients treated with ERCP for the first time in the First Affilated Hospital of Naval Medical University from June 2015 to September 2020 were retrospectively analyzed. Twenty eight (0.22%) were identified as difficult cannulation where metal clip- or submucosal injection-assisted cannulation was used. The selective cannulation success rate, intubation time and complication incidence of the two techniques in difficult cannulation patients were analyzed.Results:Difficult cannulation was performed in 18 males (64.3%) and 10 females (35.7%) with an age of 69.6±14.1 years assisted by metal clips or submucosal injection. Five cases (17.9%) were type Ⅱ, 5 cases (17.9%) type Ⅲ, and 18 cases (64.3%) type Ⅴ according to papilla classification. Sixteen patients (57.1%) received metal clip-assisted cannulation, and 12 cases (42.9%) submucosal injection-assisted cannulation. Twenty-five (89.3%) patients successfully underwent selective cannulation with the cannulation time of 9.9±4.3 min. One case (3.6%) of mild post-ERCP pancreatitis and 3 cases (10.7%) of post-ERCP hyperamylasemia occurred. No postoperative bleeding or perforation occurred. All patients were cured and discharged after conservative treatment.Conclusion:When selective cannulation is difficult due to poor papilla exposure or deflection, endoclip- or submucosal injection-assisted cannulation can effectively improve the successful selective cannulation rate during ERCP with low complication incidence, which is worth of clinical promotion.

Objective:To evaluate the dose-response relationship of alfentanil in combination with midazolam-etomidate inhibiting cardiovascular responses to laryngeal mask airway implantation in elderly patients.Methods:American Society of Anesthesiologists Physical Status Ⅰ or Ⅱ patients of either sex, aged 65-85 yr, with body mass index of 20-30 kg/m 2, undergoing elective operation under general anesthesia, were enrolled in this study.Midazolam 0.025 mg/kg was intravenously injected for adequate sedation, 5 min later mean arterial pressure and heart rate were recorded for 3 consecutive times at 3-min interval, the mean value was collected and considered as the baseline value.Etomidate 0.2 mg/kg was intravenously injected, and alfentanil and rocuronium 0.6 mg/kg were intravenously injected when bispectral index value < 60.A laryngeal mask airway was inserted at 1.4 min after intravenous injection of alfentanil, and mechanical ventilation was performed.The dose of alfentanil was determined by the Dixon′s up-and-down method.The initial dose of alfentanil was set at 6.83 μg/kg.The dose of alfentanil in the next patient was determined according to the development of cardiovascular response to laryngeal mask airway placement.If the cardiovascular response to laryngeal mask airway placement occurred, the dose was increased for the next patient, and if cardiovascular response to laryngeal mask airway placement did not occur, the dose was decreased, and the ratio between the two successive doses was 1.0∶1.1.The cardiovascular response to laryngeal mask airway placement was defined as increase in maximum mean arterial pressure or maximum heart rate by≥20% of baseline values within 2 min after laryngeal mask airway placement.The median effective dose (ED 50), 95% effective dose (ED 95) and 95% confidence interval (95% CI) of alfentanil inhibiting cardiovascular responses to laryngeal mask airway placement in elderly patients were calculated by the Probit method. Results:When combined with midazolam and etomidate, the ED 50 (95% CI) of alfentanil inhibiting the cardiovascular responses to laryngeal mask airway placement in elderly patients were 5.605 (5.036-6.082) μg/kg, and the ED 95 (95% CI) were 6.625 (6.125-9.763) μg/kg. Conclusions:When combined with midazolam and etomidate, the ED 50 and ED 95 of alfentanil inhibiting the cardiovascular responses to laryngeal mask airway placement are 5.605 and 6.625 μg/kg, respectively, in elderly patients.

Objective:To investigate the dose-effect relationship of alfentanil inhibiting cardiovascular responses to tracheal intubation when combined with midazolam and etomidate.Methods:American Society of Anesthesiologists Physical Status classification Ⅰ or Ⅱ patients of either sex, aged 18-64 yr, with body mass index<32 kg/m 2, undergoing elective operation under general anesthesia with endotracheal intubation, were enrolled in this study.Midazolam 0.025 mg/kg was intravenously injected for adequate sedation, and 5 min later mean arterial pressure and heart rate were recorded for 3 consecutive times at an interval of 3 min, and the mean value was calculated and served as the baseline value.Etomidate 0.3 mg/kg was intravenously injected, and alfentanil and rocuronium 0.6 mg/kg were intravenously injected when bispectral index value < 60, and then 1.4 min later tracheal intubation was performed.The dose of alfentanil was determined by the Dixon′s up-and-down method.The initial dose of alfentanil was set at 20 μg/kg.The dose of alfentanil in the next patient was determined according to the development of cardiovascular responses to tracheal intubation, and the ratio between the two successive doses was 1.0∶1.1.The cardiovascular response was defined as as positive when the maximum value of mean arterial pressure or heart rate increased by ≥20% of the baseline value within 2 min after endotracheal intubation.Probit method was used to determine the ED 50, ED 95 and 95% confidence interval of alfentanil inhibiting cardiovascular responses to tracheal intubation. Results:When combined with midazolam and etomidate, the ED 50 (95% confidence interval) of alfentanil inhibiting cardiovascular responses to tracheal intubation was 21.343 (19.105-24.516) μg/kg, and the ED 95 (95% confidence interval) was 25.043 (22.983-48.983) μg/kg. Conclusions:When combined with midazolam and etomidate, the ED 50 and ED 95 of alfentanil inhibiting cardiovascular responses to tracheal intubation are 21.343 and 25.043 μg/kg, respectively.