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 explore the clinical characteristics, causes, diagnosis and treatment of transplant renal artery stenosis after pediatric-to-adult kidney transplantation.Methods:Between July 2014 and March 2019, clinical data were retrospectively reviewed for 25 en-bloc and 27 single kidney transplant cases.Results:One en-bloc(4.0%)and two single kidney recipients(7.4%)were diagnosed as renal artery stenosis at Month 13-23 months post-transplantation.It was higher than the rate of stenosis in adult-to-adult transplant cases(1.1%)during the same period.As compared with recipients without stenosis, stenotic ones had younger pediatric donors( P<0.05)and yet similar body weight of donors as well as recipients( P>0.05). The inner diameters of stenonotic sites were(1.40-1.63)mm and predominant stenotic site was proximal renal artery rather than anastomotic site.The remaining parts of major renal arteries varied from 2.31 to 4.93 mm in diameter.It was normal in children with a corresponding age.All three cases responded well to percutaneous transluminal angioplasty and stenting. Conclusions:The cause of stenosis may be an undeveloped local artery diameter due to extensive tissue dissection around artery.Therefore cautious selections of infantile single renal graft for adult recipients and preserving surrounding tissue of renal artery assist in the prevention of graft arterial stenosis.

Objective:To survey the public attitude towards xenotransplantation and examine its influencing factors.Methods:A survey form with 46 multiple-choice questions is offered. It is composed of general profiles of respondent and scale. The questionnaire is distributed online through the platform of Wenjuanxing(https: //www.wjx.cn). All adult respondents filled in anonymously online. Statistical processing included descriptive analysis, reliability and validity testing and variance and correlation analysis.Results:A total of 4 414 valid questionnaires are obtained between December 1, 2021 and January 31, 2022. Cronbach's alpha coefficient is 0.912 and the scale has decent reliability. Based upon the results of exploratory factor analysis, the items are grouped into five main factors, namely organ source, decision, psychosocial change, infection risk and other risks. If pig organs are proven feasible, the risks and prognosis are basically the same as human organs, 65.4% of the respondents definitely supported xenotransplantation. Among the respondents, individuals aged 31~50 years, male, born or resident in Chinese western region, higher education, non-medical institution practitioners, self/family members/friends have done or awaiting organ transplantation, self/partners supporting organ donation, future needs for organ transplantation, previous discussion of organ donation or organ transplantation with family/friends, blood donation, volunteer social worker, atheist or Buddhist/Christianc and hearing about xenotransplantation are more inclined to support xenotransplantation.Correlation analysis showed significant correlations among five main factors.Conclusions:Despite differences in attitudes towards xenotransplantation among different populations, overall attitude is favorable. Respondents are more concerned about their associated risks, especially psychosocial changes. The related researches should be stressed. And for different groups of people, corresponding stratified tutoring should be carried out. Strengthening clinical trials, heightening public attention and training medical staff are expected to further popularize this new technology.

Objective:To summarize the clinical experiences of pediatric en-bloc kidney transplantation (EBKT) at a single center and explore the measures of improving its efficacy.Methods:Clinical data and outcomes retrospectively analyzed for 38 EBKT children between September 2014 and November 2019 from Department of Urology Affiliated Union Hospital Tongji Medical College Huazhong University of Science & Technology. The pediatric donors were aged (63.6±5.7) days with a weight of (4.1±0.2) kg. And the transplant recipients were aged (28.1±1.4) years with a weight of (48.7±4.9) kg. Serum levels of creatinine and basic profiles of both donors and recipients were recorded at Day 0/7/30/90/80/360 post-EBKT. The postoperative occurrences of such complications such as thrombosis, urine leakage, delayed graft function (DGF), proteinuria and hematoma were measured.Results:The one-year graft survival rate was 76.3%(29/38) and the recipient survival rate 100.0%(38/38). Among 29 recipients with long-term graft survival, no dialysis was required at Week 2 post-EBKT and the serum level of creatinine dropped to normal at Year 1. Thrombosis was a major post-EBKT complication with an incidence of 18.4%(7/38). The other complications included urine leakage (20.7%, 6/29), hematoma (6.9%, 2/29) and primary non-functioning kidney (2.6%, 1/38).Conclusions:As an effective way of expanding the pool of donors, pediatric EBKT is clinically feasible.

Allografts ; COVID-19 ; China/epidemiology* ; Disease Outbreaks ; Humans ; Kidney Transplantation/adverse effects* ; SARS-CoV-2

Objective:To explore the short-term outcomes of dual kidney transplantation and summarize its safety and feasibility.Methods:From September 2018 to September 2019, a total of 7 dual kidney transplantations were performed. And retrospective analysis was performed for baseline profiles, clinical data and postoperative complications.Results:The mean age was (62.7±8.5) years for donors and (43.9±9.3) years for recipients. The Remuzzi score of 6 paired kidneys ranged from 4 to 6 points. During follow-ups, the survival rate of 7 dual kidney transplantation grafts and recipients was 100%. The median follow-up period was 16 months. Renal function of 6 recipients normalized within 1 week and delayed graft function (DGF) occurred in one case. All of them underwent unilateral kidney transplantation with an average operative duration of (5.6±1.4) hours. There was no onset of operative complications. One case of rejection was not confirmed by biopsy. Among three patients of lung infections, there was one case of severe pneumonia. In 3 cases, lateral plasma flow of transplanted kidney exceeded that of medial plasma flow.Conclusions:Dual kidney transplantation in adults is both safe and feasible so as to expand the availability of donated kidney.

Objective:To explore the clinicopathologic features and risk factors of IgA nephropathy (IgAN) after kidney transplantation.Methods:A retrospective analysis was conducted for biopsy-confirmed IgAN patients after transplantation from January 2016 to September 2019. The clinicopathological characteristics and risk factors of IgAN after expanded criteria donor (ECD) donor for kidney transplantation were examined by statistical analysis of general clinical data and Oxford classification during and after transplantation.Results:The diagnostic rate of IgAN after transplantation was around 12.77%. The clinical symptoms occurred at an average of 22.5 months after transplantation. 66.67%(12/18) patients were in stage 3-4 chronic kidney disease (CKD). There were microscopic hematuria (76.92%) and varying degrees of proteinuria (88.89%). The scores of pathological parameters were as follows: 78% patients with mesangial cell proliferation (M1), 17% with endothelial cell proliferation (E1), 65% with segmental sclerosis (S1), 39% with moderate-to-severe tubules atrophy/interstitial fibrosis (T1/T2) and 17% with crescent formation (C1) respectively. Tubuloatrophy/interstitial fibrosis was associated with estimated glomerular filtration rate (eGFR) at biopsy ( P<0.05). Group of urine protein quantification >1g/d and group of <1g/d group had statistically significant differences in segmental sclerosis and adhesion lesion ( P<0.05). Conclusions:Oxford classification is valuable in the clinical evaluations of allograft IgAN: Tubular atrophy/interstitial fibrosis is associated with a decline of glomerular filtration. And segmental sclerosis and adhesion lesion are correlated with the severity of proteinuria.

Organ transplantation is the most effective method to treat end-stage organ failure. As the increase of transmission risk of donor-derived diseases, the quality, safety and selection criteria of transplanted organs become more and more important. Chapter 7 of the European Union's Guide to the Quality and Safety of Organs for Transplantation (6th Edition) proposed basic requirements in terms of donor and organ quality assessment, selection criteria and procedures, which were worthy of study and practice in clinical practice.

Objective To explore the correlation between pre-transplantation donor kidney biopsy and short-term renal function after transplantation.Methods This study include 240 kidney transplantation of donation after cardiac death (DCD) from July 2016 to April 2018.Banff's score of donor kidney biopsy was employed for estimating kidney status.Results No significant correlation existed between rate of glomerulosclerosis and delayed graft function (DGF) (P =0.815).The rate of glomerulosclerosis was significantly correlated with 1-week estimated glomerular filtration rate (eGFR) and discharge eGFR (P＜0.05).Based upon the glomerulosclerosis rate,the patients were divided into two groups ＜ 20％ (n =220) and ≥20％ (n =20),there was no significant inter-group difference in DGF,1-week eGFR or discharge eGFR (P＞0.05).Arterial fibrosis was significantly positively correlated with DGF and negatively with 1-week eGFR and discharge eGFR (P＜0.05).Statistically significant inter-group differences existed in 1-week eGFR and discharge eGFR that arterial fibrosis scores ＜ 2 (n =19) and ≥2 (n =41) (P＜0.05).Arteriolar hyalinosis score was negatively correlated with 1-week eGFR and discharge eGFR (P＜0.05).Based upon arteriolar hyalinosis scores,they were divided into two groups ＜ 2 (n =193) and ≥2 (n =47).There were significant inter-group differences in DGF,1-week eGFR and discharge eGFR (P＜0.05).Remuzzi scores were negatively correlated with 1-week eGFR and discharge eGFR (P＜0.05).Interstitial fibrosis was significantly positively correlated with DGF (P＜0.05) and negatively with 1-week eGFR and discharge eGFR (P＜0.05).Conclusions Donor kidney glomerulosclerosis rate affects short-term renal function of recipients after transplantation.However,using 20％ as a threshold value is limited in clinical practice.Arterial intimal fibrosis and arteriolar hyalinosis are important factors affecting short-term eGFR.Recipient kidneys with Remuzzi score ＞ 4 had poor renal function after transplantation.Interstitial fibrosis score may be used as a predictor of postoperative DGF and shortterm renal function recovery.It is expected to be discussed more extensively in literature.

OBJECTIVETo explore the effect difference between the skin needle embedding therapy and western medication for obese impaired glucose tolerance (IGT).

METHODSA total of 300 cases of obese IGT were assigned into an embedding group and a western medication group by random number table, 150 cases in each one. Standardized diagnosis and treatment programs were applied to reduce blood pressure, lipid, weight, and exercise and scientific diet management were used. 0.25 g oral deltamine was prescribed three times a day in the western medication group. Thumb-tack needle for subcutaneous embedding was at bilateral Weiwanxiashu (EX-B 3), Ganshu (BL 18), Pishu (BL 20), Tianshu (ST 25) and Zusanli (ST 36) for 36 to 48 hours on Monday and Thursday, 3 months as a session, with other acupoints differentiated. All the treatment was given for 2 years. The indexes included the blood sugar indexes [fasting plasma glucose (FPG), 2 h postprandial blood glucose (2 h PG), glycosylated hemoglobin (HbA1c)], incidence of hypoglycemia, obesity indexes [waist circumference, body mass index (BMI)], blood lipid indexes [serum total cholesterol (TC), serum triacylglycerol (TG), low density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C)], liver and kidney function indexes [serum creatinine (Scr), blood urea nitrogen (BUN) and blood uric acid (UA), glutamic-pyruvic transaminase (ALT)] and TCM symptom score. The effects and the incidence of type 2 diabetes were evaluated.

RESULTSAfter treatment, 2 h PG and HbA1c reduced in the two groups (<0.01,<0.05) and the results in the embedding group were better (both<0.05). After treatment, the incidence of hypoglycemia in the embedding group was 0.7% (1/150), and that in the western medication group was 1.3% (2/150), without statistical difference (>0.05). After treatment, waist circumference and BMI reduced in the two groups (both<0.01) and the improvements in the embedding group were better (both<0.05). TC, TG and LDL-C after treatment were lower than those before treatment, and HDL-C were higher in the two groups (all<0.05), without statistical different values before and after treatment between the two groups (all>0.05). Scr, BUN, UA and ALT before and after treatment in the two groups had no statistical difference (all>0.05), without statistical difference after treatment between the two groups (all>0.05). The TCM score after treatment was lower than that before treatment in the embedding group (<0.05), and the difference was not statistical in the western medication group (>0.05). The different value of TCM score in the embedding group was better than that in the western medication group (<0.01). The total effective rate in the embedding group was 98.0% (147/150), which was superior to 92.7% (139/150) in the western medication group (<0.05). The incidence of type 2 diabetes was 2.0% (3/150) in the embedding group, and that was better than 7.3% (11/150) in the western medication group (<0.05).

CONCLUSIONThumb-tack needle for subcutaneous embedding for 2 years could apparent improve the indexes of IGT, which is better than western medication, without liver and kidney damage.