OBJECTIVE: To explore the genetic basis for a child featuring global developmental disorder with epilepsy. METHODS: A child who had presented at Guangzhou Women and Children's Medical Center in July 2022 was selected as the study subject. Clinical data was collected. Potential variant was detected by whole exome sequencing (WES). Candidate variant was validated by Sanger sequencing and bioinformatic analysis. RESULTS: The child, a three-year-old ethnic Zhuang Chinese girl, had presented with global developmental disorder and epilepsy, for which rehabilitation therapy was ineffective. Genetic testing revealed that she has harbored a homozygous c.821T>C (p.Leu274Pro) missense variant of the PIGW gene, for which both of her parents and sister were heterozygous carriers. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), the variant was classified as variant of uncertain significance. CONCLUSION The homozygous c.821T>C (p.Leu274Pro) variant of the PIGW gene probably underlay the onset of disease in this child. Above finding has enriched the mutational spectrum of the PIGW gene.
Child, Preschool ; Female ; Humans ; Computational Biology ; Developmental Disabilities ; Epilepsy/genetics* ; Genetic Testing ; Homozygote

【Objective】 To investigate the effect of total flavone of oldenlandia diffusa(FOD) on the stemness, proliferation and apoptosis of breast cancer(BC) stem cells sorted from MDA-MB-231. 【Methods】 Human BC cell lines MDA-MB-231 was cultured in vitro; MDA-MB-231 was stimulated by different concentrations(0 μg/mL, 100 μg/mL, 200 μg/mL and 400 μg/mL) of FOD for different time (24 h, 48 h and 72 h). CCK8 and plate cell cloning assay were used to detect the effect of FOD on MDA-MB-231 proliferation; CD44⁺/CD24^-MDA-MB-231 cell line were tested by flow cytometry and stem cell markers such as Nanog, Oct4 and Sox2 were tested by Western blotting; Annexin V-PE/7-AAD was used to detect the effect of FOD on MDA-MB-231 apoptosis and Bcl2, cleaved-caspase3 and Bax were tested by Western blotting. 【Results】 Cell proliferation of MDA-MB-231 was significantly inhibited by FOD, with the significant suppression at concentrations of 400 μg/mL for 72 h compared with negative control group(P<0.05). The apoptosis rate was significantly upregulated than the negative control group (P<0.05). The protein expression of Bcl2 decreased while Bax and cleaved-caspae3 increased, and stemness markers such as Nanog, Sox2 and Oct4 decreased in FOD-treated cells. Moverover, Akt-GSK3β-β-catenin axis was inhibited in FOD-treated cells. 【Conclusion】 FOD could significantly inhibit the stemness and proliferation and promote the apoptosis of MDA-MB-231.

【Objective】 To investigate the effects of total flavone of oldenlandia diffusa (FOD) on the proliferation and apoptosis of hepatocellular carcinoma (HCC) stem cells sorted from Huh7. 【Methods】 Human HCC cell lines Huh7 was cultured in vitro; CD133 positive (CD133⁺) stem cells in Huh7 cell line were sorted by flow cytometry, and stem cell markers such as Nanog, Oct4 and Sox2 were tested by Western blotting. CD133⁺-Huh7 was stimulated by different concentrations (0 μg/mL, 50 μg/mL, 100 μg/mL and 400 μg/mL) of FOD for different time (24 h, 48 h, 72 h and 96 h). CCK8 and plate cell cloning assay were used to detect the effect of FOD on CD133⁺-Huh7 proliferation while Annexin V-PE/7-AAD was used to detect the effect of FOD on CD133⁺-Huh7 apoptosis. Western blotting was used to detect the protein expressions of protein 53 (P53), factor associated suicide-Fas-associating protein with a novel death domain (Fas-FADD), B-cell lymphoma-2 (Bcl-2), Cleaved-Caspase3, and Bcl-2 associated X protein (Bax). 【Results】 More than 95% of stem cells were purified for further experiments. Cell proliferation of CD133⁺-Huh7 was significantly inhibited by FOD, with the significant suppression at the concentration of 100 μg/mL for 72 h compared with negative control group (P<0.05). The apoptosis rate was significantly upregulated than that in the negative control group (P<0.05). The protein expression of Bcl2 decreased while Bax and Cleaved-Caspae3 increased via FAS/FADDD and P53 axis. 【Conclusion】 FOD can significantly inhibit the proliferation and promote the apoptosis of CD133⁺-Huh7.

Pharmaceutical water is the most widely used raw material and solvent in the process of pharmaceutical products, which is widely used in the cleaning and disinfecting process of appliances, equipment, facilities and systems, and is involved in the whole process of pharmaceutical products. Pharmaceutical water is one of the main sources of microbial contamination of pharmaceutical products, so the microbial safety of pharmaceutical water is crucial for pharmaceutical quality. This paper mainly introduces the characterization of the pharmaceutical water system, and conducts biofilm analysis. Due to the characteristics of oligotrophic and slow-growing of microorganisms in the pharmaceutical water system, this paper also further discusses the microbiological testing and culture methods of pharmaceutical water, so as to provide reference for the establishment of a scientific and reasonable microbiological testing system, as well as provide an important theoretical basis and new ideas for further research on hard-to-culture microorganisms.

Objective:To explore the feasibility of a stable pig-to-monkey orthotopic liver transplantation (LT) model and provide a favorable experimental tool for preclinical studies of xenotransplantation.Methods:In this retrospective analysis, the authors reviewed the perioperative conditions and outcomes of 7 pig-to-monkey orthotopic liver transplants performed by a xenotransplantation research team of Second Xiangya Hospital.Gene-edited Banna miniature pigs were selected as donors and rhesus monkeys with similar anatomical characteristics, physiology, biochemistry and immune mechanism to humans were selected as recipients for pig-monkey xenogeneic orthotopic LT.Based upon classic transplantation procedures, whole liver xenogeneic orthotopic transplantation was performed.Surgical processes were modified for minimizing intraoperative hemorrhage and shortening anhepatic period.A bile duct drainage tube was implanted for observing bile secretion.ATG + anti-CD20 + snake venom factor and FK-506 were utilized for immunoinduction pre-operation while tacrolimus, mycophenolate mofetil (MMF) and methyl prednisolone for postoperative immunomaintenance therapy.Antibiotics and antiviral agents were also applied and thrombin complex for improving coagulation functions.Results:All procedures were successfully completed.After the stability and maturity of our model, in case No.7, donor's acquisition operative duration was 42 min without heat ischemic time, donor's trimming time 87 min, donor cold retention time 128 min, recipient's operative duration 123 min and anhepatic phase 27 min.Subhepatic inferior vena cava was occluded for 38 min.Blood loss was around 10 ml.And 4/7 models survived post-operation and the longest survival time was 27 h. Among 3 non-survival cases, the causes were anesthesia accident (n=1) and immaturity of early operation (n=2). Model No.7 had a biliary secretion volume of 86 ml post-operation.Conclusions:Qualified donor acquisition, high-quality vascular anastomosis, intraoperative reduction of blood loss, shortening of anhepatic period, strict fluid replenishing and careful monitoring are essential for boosting the success rate of pig-monkey liver xenotransplantation model.Optimization of donor gene combination and advanced immunosuppression protocol help to further achieve the long-term survival of pig-monkey liver xenotransplantation model.

Objective To investigate the efficacy and safety of programmed cell death protein-1 (PD-1) monoclonal antibody on the treatment of malignant tumor after solid organ transplantation (SOT). Methods The relevant literatures in 7 databases were searched. The data on 54 cases of recipients with malignant tumors treated with PD-1 monoclonal antibody after SOT were collected, and the clinical effects and rejection of SOT recipients treated with PD-1 monoclonal antibody were analyzed. Results Total 32 acceptable articles including 54 SOT recipients were incorporated, including 43 males and 11 females aged 14-79 years old. There are 29 renal transplant recipients, 19 liver transplant recipients and 6 heart transplant recipients. The types of PD-1 monoclonal antibody agent used by SOT recipients included pembrolizumab for 28 patients and nivolumab for 26 patients. The overall remission rate, disease progression rate and fatality rate of PD-1 monoclonal antibody for postoperative malignant tumors of SOT recipients were 32% (17/54), 44% (24/54) and 36% (19/54), respectively. After treatment with PD-1 monoclonal antibody for postoperative malignant tumors of SOT recipients, the incidence of rejection was 39% (21/54), indicating no significant correlation between rejection and type of PD-1 monoclonal antibody (P > 0.05). Conclusions PD-1 monoclonal antibody can effectively treat postoperative malignant tumors of SOT recipients, and may induce rejection during the treatment. But rejection is not the most common cause for death of recipients.

Objective To evaluate the effect of donor risk index (DRI) on the early prognosis of liver transplantation for acute-on-chronic liver failure (ACLF). Methods Clinical data of 159 ACLF recipients undergoing liver transplantation were retrospectively analyzed. According to the calculation formula of DRI, all recipients were divided into DRI < 1.65 group (n=96) and DRI≥1.65 group (n=63). Based on the Chronic Liver Failure Consortium acute-on-chronic liver failure score (CLIF-C ACLFs), all recipients were divided into CLIF-C ACLFs < 48 group (n=78) and CLIF-C ACLFs≥48 group (n=81). The early prognosis indexes including the length of intensive care unit (ICU) stay and the length of postoperative hospital stay of the recipients in each group were observed after liver transplantation. The 90 dsurvival rate of the recipients after liver transplantation was analyzed by Kaplan-Meier survival curve. The risk factors affecting the early prognosis of ACLF recipients after liver transplantation were analyzed by Cox's hazards regression model. Results The length of ICU stay and the length of postoperative hospital stay did not significantly differ between the DRI < 1.65 group and DRI≥1.65 group (both P > 0.05). The length of postoperative hospital stay did not significantly differ between the CLIF-C ACLFs < 48 group and CLIF-C ACLFs≥48 group (P > 0.05). The length of ICU stay in the CLIF-C ACLFs < 48 group was 4 (3-14) d, significantly shorter than 7 (1-33) d in the CLIF-C ACLFs≥48 group (P < 0.05). The CLIF-C ACLFs was a risk factor of the early prognosis of ACLF recipients after liver transplantation (P < 0.05). The postoperative 90 d survival rate did not significantly differ between the DRI < 1.65 group and DRI≥1.65 group (P > 0.05). The postoperative 90 d survival rate in the CLIF-C ACLFs < 48 group was 94%, significantly higher than 79% in the CLIF-C ACLFs≥48 group (P < 0.05). Conclusions The early prognosis of ACLF recipients after liver transplantation is correlated with the severity of the disease rather than the DRI. Liver transplantation should be performed early and promptly.

Objection: To analyze the factors affecting the prognosis of patients with gastric neuroendocrine neoplasms (G-NENs) by using the surveillance of National Cancer Institute (NCI) of America, Epidemiology and End Results (SEER) database, and to construct a prognostic Nomogram model for individualized prediction of prognosis in patients with G-NENs. Methods: The clinical data of 2720 G-NENs patients with complete follow-up data from 2010 to 2015 in the SEER database were collected. The prognostic Nomogram model was constructed based on independent risk factors determined by survival analysis. The consistency index (C-index) and calibration curve were used to evaluate its accuracy.Area under the curve (AUC) was used to compare the evaluation value between the Nomogram and the 7th edition of AJCC TNM staging. Results: The 1-, 3-, and 5-year survival rates of 2,720 patients with G-NENs were 88.14%, 79.09%, and 71.86%, respectively. Multivariate COX regression analysis showed that gender, age, marital status, other associated tumors, histological type, tumor grade, T stage, M stage, and surgery were independent risk factors affecting survival time of GNENs patients. The C-index of newly constructed Nomogram prediction model was 0.816, which was significantly higher than 0.702 of the 7thAJCC TNM staging (P<0.001), and the 1-, 3- and 5-year calibration curves showed a good agreement between predicted survival and actual survival. The AUC for 1-, 3- and 5-year survival by Nomogram prognostic model was 0.800, 0.811, and 0.820, which was higher than 0.650, 0.688 and 0.698 of the 7th AJCC TNM staging, and the differences were statistically significant (Z= 6.600, 8.085, 9.632, all P<0.0001). Conclusion: The Nomogram prediction model drawn in this study has a high prognostic value and can individually predict the survival rate of G-NENs patients, which is helpful for clinical treatment decision-making and clinical research options.

Objective To evaluate the outcome of 1iver transplantation for acute-on-chronic liver failure (ACLF) patients .Methods We included 453 consecutive patients with previously cirrhosis who underwent liver transplantation between January 2013 and December 2017 .Patients were categorized as no ACLF (n=294) and ACLF(n=159) according to EASL-CLIF consortium criteria .Furthermore ,we used ACLF grades to categorize the ACLF patients .Their clinical data were reviewed and their 90-days survival outcomes were compared .Results Compared with the no ACLF group ,the length of stay in the ICU was significantly prolonged for all patients with ACLF ,and the 90-days survival rate after transplantation was significantly reduced in ACLF group .The length of stay in the ICU was shorter in Grade 1 and Grade 2 group when compared to Grade 3 group .The 90-days survival rate of no ACLF ,Grade 1 ,Grade 2 and Grade 3 group were 93 .20％ ,92 .59％ ,93 .33％ and 73 .68％ ,respectively .There were no statistically significant differences in 90-days survival rate among the no ACLF ,Grade 1 and Grade 2 group .However , the 90-days survive rate of Grade 3 group was lower than that of other groups .Conclusions Liver transplantation has been shown to be safe and effective with good outcome in patients with ACLF and should be offered in early course of ACLF before onset of multi-organ failure .

Objective To construct the orthotopic mouse liver transplantation model and cover troubleshooting,in order to provide experimental techniques support for organ transplantation pathology and immunology studies.Methods Male C57BL/6 mice,10-12 weeks,were selected as the allograft donors.Male C3H mice with same age were selected as the allograft recipients.The orthotopic mouse liver transplantation model consisted of 3 stages,including harvesting the donor liver,back-table preparation of the liver graft and transplantation of the donor liver into the recipient.The average time for harvesting the donor livers was (40 ± 8.8) min,(23 ± 4.7) min for preparing the donor livers and (75 ± 9.6) min for transplanting the donor livers into the recipient.Results Seventy pairs of mice were used for the preliminary experiments.For the formal experiments,the allograft transplantation was established on 220 pairs with 90.4％ successful rate.Conclusion It is the skillful and high quality microsurgical technique that is the guarantee of establishing the orthotopic mouse liver transplantation model successfully.