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 efficacy of ixazomib combination treatment therapies for multiple myeloma (MM), and the influencing factors of prognosis.Methods:The clinical data of 80 MM patients admitted to Yancheng Third People's Hospital from January 2020 to January 2022 were retrospectively analyzed. All patients received 3 courses of ixazomib combination treatment therapies (28 d was 1 course). All combination treatment therapies included ID group (ixazomib + dexamethasone, 11 cases) and ID + immunomodulator group (ixazomib + dexamethasone + lenalidomide/thalidomide, 50 cases), ID + other chemotherapy drugs group (ixazzomib + dexamethasone + doxorubicin liposome/cyclophosphamide/bendamustine, 19 cases). The clinical efficacy of patients in different treatment regimens was compared, and the prognosis was followed up and recorded. The clinical characteristics between the survival and the dead patients were compared. Cox proportional risk model was used to make multivariate analysis of the overall survival of MM patients receiving ixazomib combination therapies.Results:The treatment was effective in 9 cases (81.82%) of the ID group, 32 cases (64.00%) of the ID + immunomodulator group, and 9 cases (47.37%) of the ID + other chemotherapy drugs group. There was no statistically significant difference in the effectiveness rate of 3 ixazomib combination regimens ( χ2 = 0.62, P = 0.432). All patients were followed up for 5 to 20 months, with an average follow-up time of (15±4) months. There were statistically significant differences in immunoglobulin type, Durie-Salmon stage, early treatment line and therapeutic efficacy between the survival group (49 cases) and the death group (31 cases) (all P < 0.05). Multivariate Cox regression analysis showed that the clinical effectiveness (effectiveness vs. ineffectiveness: OR = 0.242, 95% CI 0.103-0.567, P = 0.001) and the previous first-line treatment (the first-line vs. the other lines: OR = 0.577, 95% CI 0.452-0.736, P < 0.001) were independent protective factors for the overall survival of MM. Conclusions:The 3 ixazomib combination therapies have a certain efficacy in the treatment of MM; ID regimen has the best clinical efficacy and survival. The clinical effectiveness and the previous first-line treatment are independent protective factors for the survival of MM.

ObjectiveXenotransplantation is an effective way to address the shortage of human organ donors, but it faces serious immune rejection reactions, including hyperacute rejection caused by blood type differences. Establishing a stable, convenient, and reliable method for pig blood type identification can quickly screen suitable donor pigs for xenograft research.MethodsBanna miniature inbred pigs, Diannan small eared pigs, and Bama Xiang pigs were selected as the research objects. DNA was extracted from the blood, oral buccal mucosa, and fetal fibroblasts of the three strains of pigs using DNA extraction kits. The target fragment of the ABO homologous gene EAA intron 7 in pigs was amplified using PCR method. Blood agglutination reaction was used to detect hemolysis in pig anterior vena cava whole blood after adding anti A and B antibodies. Immunohistochemical method was used to detect the expression level of A antigen in pig heart, liver, spleen, lung, and kidney tissues. Immunofluorescence method was used to detect the expression level of A antigen in pig oral mucosa. By comparing the results of different methods for determining pig blood types, the stability and reliability of the PCR method were verified, and a convenient PCR based pigblood type identification method was established.Results Firstly, the blood PCR results of 69 inbred strains of Banna miniature pigs, 7 Diannan small eared pigs, and 34 Bama Xiang pigs showed 20 AO blood types, 66 AA blood types, and 24 O blood types. The PCR results of fetal fibroblasts from 47 Diannan small eared pigs showed that all 47 fetuses were O blood type. Among them, the oral mucosal PCR results of 8 gene edited cloned pigs were consistent with those of donor fetal fibroblasts, all of which were O blood type. The oral mucosal PCR results of 8 wild-type pigs (2 inbred lines of Banna miniature pigs, 4 Diannan small eared pigs, and 2 Bama Xiang pigs) were consistent with the blood PCR identification results. Then, 11 inbred lines of Banna miniature pigs, 4 Diannan small eared pigs, and 2 Bama Xiang pigs were randomly selected for blood agglutination reaction validation, and the results were consistent with the PCR identification results of both blood samples and oral mucosa samples. Moreover, immuno-histochemical analysis was performed on the heart, liver, lung, kidney, and spleen tissues of one Banna miniature pig inbred line and two Bama Xiang pigs, and the results were consistent with blood PCR identification and blood agglutination reaction results. Finally, oral mucosal samples were collected from 2 inbred strains of Banna miniature pigs and 1 Bama Xiang pig for immunofluorescence detection, and the results were consistent with the blood PCR identification results.ConclusionBy collecting fetal cells and oral mucosal samples from live pigs for PCR detection, the blood type of pigs can be accurately and efficiently identified, providing a convenient method for blood type screening of xenograft donor pigs.

Objective:To explore the status of depression symptoms and related factors of depressive symptoms in middle-aged and elderly disabled people in China and provide references on the preventive cares of depression.Methods:Research data were obtained from the 2018 follow-up data of China Health and Retirement Longitudinal Study(CHARLS).Disabled people were selected as the research object. The scores of center for epidemiological studies depression scale(CES-D) were used to evaluate depressive symptoms. Fourteen variables derived from the 3 dimensions of demographic characteristics, health status, and health related behaviors were utilized in this study.SAS 9.4 were used for descriptive analysis, univariate analysis and Logistic regression analysis.Results:A total of 1 708 middle-aged and elderly disabled people were included, among whom 932(54.5%)were diagnosed with depressive symptoms, and the score of CES-D was (11.45±7.19).Self-rated poor health, self-rated poor memory, life dissatisfaction, physical pain were the risk factors of depressive symptoms ( OR=1.87, 4.22, 1.83, 5.30, 1.75). Male, educational level (middle-high school, high school or above), sleeping duration (6-9 h) were the protective factors of depressive symptoms( OR=0.74, 0.72, 0.10, 0.56). Conclusion:Middle-aged and elderly disabled people in China have a high detection rate of depressive symptoms. Female, lower education background, inadequate amount of sleep, self-rated unsatisfactory healthy, self-rated poor memory, dissatisfaction towards life and suffering from physical pain may be more likely to have depressive symptoms in middle-aged and elderly people with disabilities.

Objective To explore the measurement accuracy of right coronary artery during coronary angiography (CAG)at 45Gdegree left oblique position.Methods The images of right coronary artery angiographic view of 45Gdegree left oblique position were divided into nine average areas.The images with marker segment of guiding wire located in the central area (n＝45)or subcentral area (n＝45)of CAG cases were collected retrospectively and randomly.The marker segment was measured by catheter calibration method,and the measure values were compared with the actual length (30 mm).Results A N OVA analysis suggested statistical differences among the three groups (F＝4.59,P＜0.05).By paired comparison,no significant differences were found in measured values between central areas (3 1.1 9± 4.12)mm or subcentral areas (29.55±2.75)mm and the actual length (P＞0.05).Significant differences were found in measured values between central areas and subcentral areas (P＜0.05).The measuring error of subcentral areas (－1.5%)was less than that of central areas (3.9%).Conclusion During CAG at 45Gdegree left oblique position,the values of subcentral area were more accurate than those of central area and the subcentral area was thus regarded as image area with less measurement error in interventional surgery.

Objective: To assess the association of single nucleotide polymorphisms of multidrug resistance gene 1 (MDR1) with refractory epilepsy in children. Methods: Peripheral blood samples were collected from 200 children with epilepsy and 100 healthy controls. Genomic DNA was extracted and subjected to PCR amplification, agarose gel electrophoresis and target site sequencing. Genotypes of rs1922242, rs2235048, rs10808072, rs868755 and rs1202184 loci of the MDR1 gene were analyzed. Results: No significant difference was found in genotypic distribution and allelic frequencies of the rs1922242, rs2235048, rs10808072 and rs868755 loci between the drug-resistant and drug-sensitive groups. For the rs1202184 locus, a significant difference in genotypic distribution was found (P = 0.008). No significant difference was found in the frequencies of various haplotypes between the two groups. Conclusion Genotypes of the rs1202184 locus of the MDR1 gene are associated with refractory epilepsy in children, for which the AA genotype plays a dominant role.

Objective To summarize the correlation between clinical manifestations and imaging characteristics of schizencephaly in order to provide a basis for the diagnosis and prognosis.Methods Thirty-five outpatients with schizencephaly diagnosed at Department of Pediatric Neurology of Henan Provincial People's Hospital from January 2009 to May 2015 were retrospectively selected,and they were divided into different groups (patients with unilateral lesions or bilateral lesions) according to their cranial magnetic resonance imaging (MRI) features.The clinical manifestations were compared between different groups.Results Of 35 patients,19 (54.3％) patients were male and 16 (45.7％) patients were female.Twenty-four patients were found with unilateral lesions (68.6％),including 16 cases (66.7％) with hemiparesis and 8 cases(33.3％) with no motor impairment.Mental retardation was observed in 8 patients (33.3％) and 6 patients (25.0％) showed speech impairment.Eleven patients were found the bilateral lesions (31.4％),including 5 patients (45.4％) with tetraparesis,4 patients (36.4％) with hemiparesis,and 2 paticnts (18.2％) with no motor impairment.Mental retardation was observed in 9 patients (81.8％) and 9 patients (81.8％) showed speech impairment.There were significant differences in motor impairment,mental retardation and speech impairment between the unilateral lesion group and bilateral lesion group (Z =-2.40,P =0.002;x2 =7.09,P =0.012;x2 =9.94;P =0.003).Epileptic seizure occurred in 18 patients (51.4％).Binary Logistic regression analysis indicated that cortical dysplasia beyond the cleft and open-lip lesions were the major risk factors for seizures (OR =4.44,2.73;P =0.005,0.029).Imaging characteristics:there were closed-lip lesions in 10 patients (28.6％),open-lip lesions in 21 patients (60.0％) and open/closed-lip lesions in 4 patients (11.4％).Anatomic localization of all clefts was found in the frontal lobes in 19 patients (54.3％),in frontoparietal lobes in 11 patients (31.4％),in parietal lobes in 2 patients (5.7％),and in occipital lobes in 3 patients (8.6％).After 1-6-year follow-up,12 patients treated with antiepileptic drug were seizure-free (all with unilateral lesions),and 6 patients had refractory epilepsy (3 patients with bilateral lesions).Conclusions Schizencephaly is a rare structural disorder of cerebral cortical development.Those with bilateral lesions are usually manifested with severe motor,speech impairment and mental retardation and their prognosis is poor.Schizencephaly patients complicated with cortical dysplasia beyond the cleft or open-lip lesions are more easily attacked by seizures.MRI plays an important role in the diagnosis and prognosis judgment of schizencephaly.

Objective To explore the differences of aortic diameter measured by catheter calibration method and centimeter sizing catheter calibration method in endovascular graft exclusion for Stanford type B aortic dissection.Methods A total of 30 patients with Stanford B type aortic dissections treated with endovascular graft exclusion were measured with the two calibration methods to measure aortic diameters at left subclavicular artery position.The measurement parameters were compared with CTA measurement results.Results Statistical differences of measurement parameters were found among catheter calibration,centimeter sizing catheter calibration and CTA (F=3.15,P＜0.05),and paired comparison showed statistical differences between catheter calibration method and CTA result (P＜0.05),and between the results of catheter calibration method and centimeter sizing catheter calibration method (P＜0.05),while no statistical difference was found between centimeter sizing catheter calibration method and CTA result (P＞0.05).Taking CTA as golden standard method,Bland-Altman analysis indicated that the centimeter sizing catheter calibration method exerted better consistency with CTA.Conclusion The measurement value of centimeter sizing catheter calibration method is identical to the CTA measurement result,and superior to catheter calibration method for precise stent selection in Stanford type B aortic dissection.

Degradation of composite resin during ageing always produces micro-cracks, the latter could gradually expand, leading to the fracture of tooth restoration. In order to extend the service life of composite resin by repairing cracks intelligently, self-healing microcapsule models have been introduced in resin-based dental materials. This paper will present the concept and classification of self-healing materials, and review the research progresses on self-healing dental resin.

Objective To investigate the type distribution of urogenital Chlamydia trachomatis(Ct)among STD clinic outpatients in Guangxi Zhuang Autonomous Region, and to estimate the prevalence of Ct infection among the patients during posttreatment follow?up. Methods Urethral and cervical swabs were collected from male and female outpatients with confirmed urogenital Ct infection, respectively, in Institute of Dermatology of Guangxi Zhuang Autonomous Region. The patients with positive results in preliminary screening tests were followed up after treatment, and specimens were collected at follow?up visits. General and clinical information was also obtained from these patients. DNA was extracted from these samples by using the QIAxtractor instrument. Nested PCR was performed to amplify the major outer membrane protein A(ompA)gene for ompA typing, and to amplify CT046(hctB), CT058, CT144, CT172 and CT682 (pbpB) genes for high?resolution multilocus sequence typing (hr?MLST). Then, PCR products were sequenced, and ompA and MLST types of Ct were determined by sequence alignment and MLST analysis, respectively. The obtained MLST sequence types (STs) were compared with those from an Italian population by using the BioNumerics7 software, and a minimum spanning tree(MST)was generated. Results Totally, 44 and 6 Ct?positive specimens were collected at first visits and follow?up visits respectively. Among the 50 specimens, 42 underwent successful ompA typing and hr?MLST, and 7 ompA genotypes and 15 hr?MLST STs were identified, including 3 first reported STs. The distribution of STs of Ct isolates from Guangxi Zhuang Autonomous Region was significantly different from that from the Italian population. Among the 6 followed patients with posttreatment Ct infection, 3 were confirmed to be reinfected with Ct, and the other 3 failed to be diagnosed because of unsuccessful genotyping. Conclusion The genotypes of Ct strains isolated from STD clinic outpatients in Guangxi Autonomous Region were characteristic, and Ct reinfection occurred in some patients during follow?up.