[Hmga2 knockdown enhances osteogenic differentiation of adipose-derived mesenchymal stem cells and accelerates bone defect healing in mice].

OBJECTIVE: To investigate the role of high-mobility group AT-hook 2 (HMGA2) in osteogenic differentiation of adipose-derived mesenchymal stem cells (ADSCs) and the effect of Hmga2 knockdown for promoting bone defect repair. METHODS: Bioinformatics studies using the GEO database and Rstudio software identified HMGA2 as a key factor in adipogenic-osteogenic differentiation balance of ADSCs. The protein-protein interaction network of HMGA2 in osteogenic differentiation was mapped using String and visualized with Cytoscape to predict the downstream targets of HMGA2. Primary mouse ADSCs (mADSCs) were transfected with Hmga2 siRNA, and the changes in osteogenic differentiation of the cells were evaluated using alkaline phosphatase staining and Alizarin red S staining. The expressions of osteogenic markers Runt-related transcription factor 2 (RUNX2), osteopontin (OPN), and osteocalcein (OCN) in the transfected cells were detected using RT-qPCR and Western blotting. In a mouse model of critical-sized calvarial defects, mADSCs with Hmga2-knockdown were transplanted into the defect, and bone repair was evaluated 6 weeks later using micro-CT scanning and histological staining. RESULTS: GEO database analysis showed that HMGA2 expression was upregulated during adipogenic differentiation of ADSCs. Protein-protein interaction network analysis suggested that the potential HMGA2 targets in osteogenic differentiation of ADSCs included SMAD7, CDH1, CDH2, SNAI1, SMAD9, IGF2BP3, and ALDH1A1. In mADSCs, Hmga2 knockdown significantly upregulated the expressions of RUNX2, OPN, and OCN and increased cellular alkaline phosphatase activity and calcium deposition. In a critical-sized calvarial defect model, transplantation of mADSCs with Hmga2 knockdown significantly promoted new bone formation. CONCLUSION: HMGA2 is a crucial regulator of osteogenic differentiation in ADSCs, and Hmga2 knockdown significantly promotes osteogenic differentiation of ADSCs and accelerates ADSCs-mediated bone defect repair in mice.

[Clinical features and follow-up study on 55 patients with adolescence-onset methylmalonic acidemia].

Objective: To investigate the clinical features and outcomes of adolescence-onset methylmalonic acidemia (MMA) and explore preventive strategies. Methods: This was a retrospective case analysis of the phenotypes, genotypes and prognoses of adolescence-onset MMA patients. There were 55 patients diagnosed in Peking University First Hospital from January 2002 to June 2023, the data of symptoms, signs, laboratory results, gene variations, and outcomes was collected. The follow-ups were done through WeChat, telephone, or clinic visits every 3 to 6 months. Results: Among the 55 patients, 31 were males and 24 were females. The age of onset was 12 years old (range 10-18 years old). They visited clinics at Tanner stages 2 to 5 with typical secondary sexual characteristics. Nine cases (16%) were trigged by infection and 5 cases (9%) were triggered by insidious exercises. The period from onset to diagnosis was between 2 months and 6 years. Forty-five cases (82%) had neuropsychiatric symptoms as the main symptoms, followed by cardiovascular symptoms in 12 cases (22%), kidney damage in 7 cases (13%), and eye disease in 12 cases (22%). Fifty-four cases (98%) had the biochemical characteristics of methylmalonic acidemia combined with homocysteinemia, and 1 case (2%) had the isolated methylmalonic acidemia. Genetic diagnosis was obtained in 54 cases, with 20 variants identified in MMACHC gene and 2 in MMUT gene. In 53 children with MMACHC gene mutation,1 case had dual gene variants of PRDX1 and MMACHC, with 105 alleles. The top 5 frequent variants in MMACHC were c.482G>A in 39 alleles (37%), c.609G>A in 17 alleles (16%), c.658_660delAAG in 11 alleles (10%), c.80A>G in 10 alleles (10%), c.567dupT and c.394C>T both are 4 alleles (4%). All patients recovered using cobalamin, L-carnitine, betaine, and symptomatic therapy, and 54 patients (98%) returned to school or work. Conclusions: Patients with adolescence-onset MMA may triggered by fatigue or infection. The diagnosis is often delayed due to non-specific symptoms. Metabolic and genetic tests are crucial for a definite diagnosis. Treatment with cobalamin, L-carnitine, and betaine can effectively reverse the prognosis of MMA in adolescence-onset patients.

Risk of major congenital heart disease in pregestational maternal diabetes is modified by hemoglobin A1c.

OBJECTIVES: The association between pregestational diabetes mellitus (PDM) and risk of congenital heart disease (CHD) is well recognized; however, the importance of glycemic control and other coexisting risk factors during pregnancy is less clear. We sought to determine the relative risk (RR) of major CHD (mCHD) among offspring from pregnancies complicated by PDM and the effect of first-trimester glycemic control on mCHD risk. METHODS: We determined the incidence of mCHD (requiring surgery within 1 year of birth or resulting in pregnancy termination or fetal demise) among registered births in Alberta, Canada. Linkage of diabetes status, maximum hemoglobin A1c (HbA1c) at < 16 weeks' gestation and other covariates was performed using data from the Alberta Perinatal Health Program registry. Risk of mCHD according to HbA1c was estimated as an adjusted RR (aRR), calculated using log-binomial modeling. RESULTS: Of 1412 cases of mCHD in 594 773 (2.37/1000) births in the study period, mCHD was present in 48/7497 with PDM (6.4/1000; RR, 2.8 (95% CI, 2.1-3.7); P < 0.0001). In the entire cohort, increased maternal age (aRR, 1.03 (95% CI, 1.02-1.04); P < 0.0001) and multiple gestation (aRR, 1.37 (95% CI, 1.1-1.8); P = 0.02) were also associated with mCHD risk, whereas maternal prepregnancy weight > 91 kg was not. The stratified risk for mCHD associated with HbA1c ≤ 6.1%, > 6.1-8.0% and > 8.0% was 4.2/1000, 6.8/1000 and 17.1/1000 PDM/gestational diabetes mellitus births, respectively; the aRR of mCHD associated with PDM and HbA1c > 8.0% was 8.5 (95% CI, 5.0-14.4) compared to those without diabetes and 5.5 (95% CI, 1.6-19.4) compared to PDM with normal HbA1c (≤ 6.1%). CONCLUSIONS: PDM is associated with a RR of 2.8 for mCHD, increasing to 8.5 in those with HbA1c > 8%. These data should facilitate refinement of referral indications for high-risk pregnancy screening. © 2023 The Authors. Ultrasound in Obstetrics & Gynecology published by John Wiley & Sons Ltd on behalf of International Society of Ultrasound in Obstetrics and Gynecology.

Single-cell and bulk RNA sequencing reveal heterogeneity and diagnostic markers in papillary thyroid carcinoma lymph-node metastasis.

PURPOSE: Papillary thyroid carcinoma (PTC) is characterized by lymph-node metastasis (LNM), which affects recurrence and prognosis. This study analyzed PTC LNM by single-cell RNA sequencing (scRNA-seq) data and bulk RNA sequencing (RNA-seq) to find diagnostic markers and therapeutic targets. METHODS: ScRNA-seq data were clustered and malignant cells were identified. Differentially expressed genes (DEGs) were identified in malignant cells of scRNA-seq and bulk RNA-seq, respectively. PTC LNM diagnostic model was constructed based on intersecting DEGs using glmnet package. Next, PTC samples from 66 patients were used to validate the two most significant genes in the diagnostic model, S100A2 and type 2 deiodinase (DIO2) by quantitative reverse transcription-polymerase chain reaction (RT-qPCR) and immunohistochemical (IHC). Further, the inhibitory effect of DIO2 on PTC cells was verified by cell biology behavior, western blot, cell cycle analysis, 5-ethynyl-2'-deoxyuridine (EdU) assay, and xenograft tumors. RESULTS: Heterogeneity of PTC LNM was demonstrated by Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) analysis. A total of 19 differential genes were used to construct the diagnostic model. S100A2 and DIO2 differ significantly at the RNA (p < 0.01) and protein level in LNM patient tissues (p < 0.001). And differed in PTC tissues with different pathologic typing (p < 0.001). Further, EdU (p < 0.001) and cell biology behavior revealed that PTC cells overexpressed DIO2 had reduced proliferative capacity. Cell cycle proteins were reduced and cells are more likely to be stuck in G2/M phase (p < 0.001). CONCLUSIONS: This study explored the heterogeneity of PTC LNM using scRNA-seq. By combining with bulk RNA-seq data, diagnostic markers were explored and the model was established. Clinical diagnostic efficacy of S100A2 and DIO2 was validated and the treatment potential of DIO2 was discovered.

Development and validation of a nomogram for urinary tract infection in geriatric patients with hip fracture: a retrospective study.

OBJECTIVE: This study aims to develop and validate a risk nomogram for urinary tract infections (UTIs) in geriatric patients with hip fractures. PATIENTS AND METHODS: A total of 900 geriatric patients who underwent hip fracture surgery at Dandong Central Hospital between June 2017 and June 2023 were systematically collected. The cohort was randomly divided into a training set (70%, n=632) and a validation set (30%, n=268) for model development and validation, respectively. Univariate and multivariate logistic regression analyses were conducted to identify the independent risk factors associated with UTIs. Based on the results of the multivariate analysis, a UTI nomogram prediction model was developed and evaluated in the training and validation sets using the C-index, ROC curve, calibration curve, and decision curve analysis to assess discrimination, calibration, and clinical utility, respectively. RESULTS: Out of the 900 participants, 24.6% were diagnosed with UTIs. The nomogram was developed based on 9 predictors that were found to be independently associated with UTI. The area under the curve (AUC) for predicting UTI in geriatric patients with hip fractures was 0.829 in the training set and 0.803 in the validation set. Following internal verification, the modified C-index remained at 0.829. Furthermore, the nomogram's calibration plot and decision curve analysis demonstrated good performance in both the training and validation sets. CONCLUSIONS: The established and validated nomogram provides a reliable and convenient tool for predicting UTI risk in geriatric patients with hip fractures. This model facilitates the early identification of high-risk patients and offers guidance for implementing targeted preventive interventions.

Development and validation of a nomogram for 30-day readmission after hip fracture surgery in geriatric patients.

OBJECTIVE: 30-day readmission after hip fracture surgery in the elderly is common and costly. A predictive tool to identify high-risk patients could significantly improve outcomes. This study aims to develop and validate a risk nomogram for 30-day readmission after hip fracture surgery in geriatric patients. PATIENTS AND METHODS: We retrospectively analyzed 1,249 geriatric hip fracture patients (≥60 years) undergoing surgery at Dandong Central Hospital from October 2011 to October 2023. Using a 7:3 ratio, patients were randomly divided into training (n=877) and validation (n=372) sets. Independent risk factors for 30-day readmission were identified using LASSO regression and logistic regression in the training set. A nomogram was constructed using the identified predictors. Finally, the C-index, ROC curve, calibration curve, and decision curve analysis were used to validate the model in the training and validation sets respectively. RESULTS: The nomogram was developed based on the 8 predictors of age, prior stroke, chronic liver disease, treatment, uric acid (UA), total protein (TP), albumin (ALB), and pneumonia that were found to be independently associated with 30-day readmission. The nomogram showed good discrimination with a C-index of 0.88 in the training set and 0.84 in the validation set. Calibration curves exhibited good agreement between predicted and observed outcomes. Decision curve analysis demonstrated clinical utility. CONCLUSIONS: We developed and validated a nomogram incorporating eight clinical variables to accurately predict the individualized risk of 30-day readmission after hip fracture surgery in elderly patients. The model demonstrated favorable discrimination, calibration, and clinical utility. It can help to identify high-risk patients needing additional interventions to prevent avoidable hospital readmissions.

[Perspectives on molecular diagnosis and targeted therapy for complex vascular malformations in pediatrics].

Vascular malformations are due to abnormal development of blood and/or lymphatic vessels during embryonic life without endothelial cell proliferation. Most of the previous treatments were symptomatic methods as surgery and sclerotherapy because the pathogenic mechanism was not clearly understood. With advances in molecular biology, the pathogenesis of vascular malformations is thought to be related to inherited and/or somatic mutations that eventually activate the PI3K/ATK/mTOR, Ras/Raf/MEK/ERK pathways. Also, related studies have promoted the use of targeted inhibitors. This article provides a review of current causative genes and targeted drugs for pediatric vascular malformations, aiming to provide a basis for promoting accurate molecular diagnosis and precision targeted therapy for these diseases.

Global burden of neoplasms attributable to specific occupational carcinogens over 30 years: a population-based study.

OBJECTIVES: The study aimed to analyze the global burden of occupational neoplasms from various epidemiological perspectives. STUDY DESIGN: In this cross-sectional study, secondary analyses were conducted to assess the burden of neoplasms attributable to occupational carcinogens and their distribution characteristics using data from GBD 2019 and the World Bank database. METHODS: Based on the GBD 2019 and the World Bank database, we analyzed the global burden of occupational neoplasms including the age-period-cohort model, decomposition analysis, health inequality analysis, and panel model. All analyses were conducted in R (version 4.0.3) and Joinpoint (version 4.9.1). RESULTS: The absolute number of neoplasms burden attributable to occupational carcinogens has continued to rise over 30 years. In 2019, occupational neoplasms caused 333,867 [95% uncertainty interval (UI): 263,491 to 404,641] mortalities and 6,964,775 (95% UI: 5,467,884 to 8,580,431) disability-adjusted life years (DALYs) globally. Greenland, Monaco, the Netherlands, and Andorra suffered the highest burden. The burden was higher in countries with a higher sociodemographic index. The age effect was prominent in the elderly, and the 1925 birth cohort had the highest cohort effect. Population growth was the most significant driver of the mortalities (89%) and DALYs (111%) change. Moreover, the proportion of urban population was significantly positively associated with the disease burden, while GDP per capita was negatively correlated with the disease burden. CONCLUSIONS: The burden of occupational neoplasms was unevenly distributed across locations and populations. The need for rational allocation of healthcare resources was urgent.

Development of the first PCR for detection of Psoroptes ovis var. cuniculi infection and its comparison to microscopic examination and serological assay in rabbits.

Psoroptes mites are the common ecto-parasites of wild and domestic animals worldwide, which causes considerable economic losses in livestock industry. Microscopy is deemed to be the 'gold standard' for the diagnosis of Psoroptes mite infection but it has poor sensitivity for low mite infections and/or sub-clinical infections. To overcome these shortcomings, we screened four genes to develop a sensitive and specific PCR for the detection of Psoroptes mite infection in rabbits, and confirmed its practicability in detecting early infection and monitoring treatment outcome with traditional microscopy and serological tests. Results showed that PCR assay targeting ITS2 (ITS2-PCR) had a high specificity and sensitivity (detection limit: 40.3 pg/µL DNA) for detecting P. ovis DNA. In rabbits artificially infected with P. ovis, all three diagnostic tests showed the same detection rate from 14 days post infection (dpi) to 42 days dpi. However, these diagnostic tests behave differently at 7 dpi and after treatment: at 7 dpi, the detection rate of ITS2-PCR was higher than rPsoSP3-based iELISA and traditional microscopy (ITS2-PCR: 88.9%, rPsoSP3-iELISA: 77.7%, microscopy: 33.3%); at 7 days post treatment (dpt), positivity rates of ITS2-PCR and microscopy rapidly decreased to 0.00% and 11.1%, whereas rPsoSP3-iELISA remained 100% positive rate. Furthermore, the comprehensive comparisons of diagnostic performance and features of three diagnostic tests at 7 dpi were performed. Compared to ITS2-PCR or rPsoSP3-iELISA, microscopy had the lowest sensitivity, and the agreement between these assays was low (κ < 0.3). Field study showed that ITS2-PCR showed a higher detection rate than microscopy (19.4% and 11.1%, respectively). Our results suggested that the ITS2-PCR developed in this study provided a new laboratory tool for diagnosis of P. ovis var. cuniculi infection, and it had advantages over microscopic examination in detection low-level mite infections and serological assay in monitoring treatment outcome.

Hepatitis C virus genotype diversity in Ningxia Hui autonomous region, Northwestern China.

Hepatitis C virus (HCV) genotypes vary greatly in different regions. The aim of this study is to investigate the distribution of HCV genotypes in HCV infected patients, in Ningxia Hui Autonomous Region. Nucleic acid extraction and amplification were performed with test kits on 153 HCV infected patients serum samples. The HCV viral load was measured using reverse transcriptase PCR (RT-PCR) and HCV genotypes were determined. Among the 153 HCV-infected patients, 56 had genotype (GT)1b (36.60%), 45 had GT2a (29.40%), 23 had GT3a (15.00%), 14 had GT3b (9.20%),13 had GT6a (8.50%), 1 had GT1g (0.70%), 1 had GT6xa (0.70%). In GT1b, 21.40% were female and 78.60% were male; in GT2a, 42.20% were female and 57.80% were male;Males were most prevalent in genotypes 1b(39.30%), while female were most prevalent in genotype 2a(46.30%). Rare GT1g and GT6xa were also detected in males. The 41-50 year age group had the highest HCV prevalence of 32.00%. HCV GT1b is the predominant HCV genotype in Ningxia Hui Autonomous Region.

[Intelligent identification of livestock, a source of Schistosoma japonicum infection, based on deep learning of unmanned aerial vehicle images].

OBJECTIVE: To develop an intelligent recognition model based on deep learning algorithms of unmanned aerial vehicle (UAV) images, and to preliminarily explore the value of this model for remote identification, monitoring and management of cattle, a source of Schistosoma japonicum infection. METHODS: Oncomelania hupensis snail-infested marshlands around the Poyang Lake area were selected as the study area. Image datasets of the study area were captured by aerial photography with UAV and subjected to augmentation. Cattle in the sample database were annotated with the annotation software VGG Image Annotator to create the morphological recognition labels for cattle. A model was created for intelligent recognition of livestock based on deep learning-based Mask R-convolutional neural network (CNN) algorithms. The performance of the model for cattle recognition was evaluated with accuracy, precision, recall, F1 score and mean precision. RESULTS: A total of 200 original UAV images were obtained, and 410 images were yielded following data augmentation. A total of 2 860 training samples of cattle recognition were labeled. The created deep learning-based Mask R-CNN model converged following 200 iterations, with an accuracy of 88.01%, precision of 92.33%, recall of 94.06%, F1 score of 93.19%, and mean precision of 92.27%, and the model was effective to detect and segment the morphological features of cattle. CONCLUSIONS: The deep learning-based Mask R-CNN model is highly accurate for recognition of cattle based on UAV images, which is feasible for remote intelligent recognition, monitoring, and management of the source of S. japonicum infection.

[Improvement of China's legal system for public health emergency management from the perspective of lifecycle management].

A crucial lesson gained through the pandemic preparedness and response to COVID-19 is that all measures for epidemic control must be law-based. The legal system is related not only to public health emergency management per se but also to all aspects of the institutional supporting system throughout the lifecycle. Based on the lifecycle emergency management model, this article analyses the problems of the current legal system and the potential solutions. It is suggested that the lifecycle emergency management model shall be followed to establish a more comprehensive public health legal system and to gather the intelligence and consensus of experts with different expertise, including epidemiologists, sociologists, economists, jurist and others, which will collaboratively promote the science-based legislation in the field of epidemic preparedness and response for the establishment of a comprehensive legal system for public health emergency management and with Chinese characteristics.

[Oncologic outcomes of surgical treatments of advanced sinonasal malignancies].

Objective: To investigate the prognoses of advanced (T3-T4) sinonasal malignancies (SNM). Methods: The clinical data of 229 patients with advanced (T3-4) SNM who underwent surgical treatments in the First Affiliated Hospital of Sun Yat-sen University from 2000 to 2018 were retrospectively analyzed, including 162 males and 67 females, aged (46.8±18.5) years old. Among them, 167 cases received endoscopic surgery alone, 30 cases received assisted incision endoscopic surgery, and 32 cases received open surgery. The Kaplan-Meier method was used to estimate the 3-year and 5-year overall survival (OS) and event-free survival (EFS). Univariate and multivariate Cox regression analyses were performed to explore significant prognostic factors. Results: The 3-year and 5-year OS were respectively 69.7% and 64.0%. The median OS time was 43 months. The 3-year and 5-year EFS were respectively 57.8% and 47.4%. The median EFS time was 34 months. The 5-year OS of the patients with epithelial-derived tumors was better than that of the patients with mesenchymal-derived tumors and malignant melanoma (5-year OS was respectively 72.3%, 47.8% and 30.0%, χ2=36.01, P<0.001). Patients with microscopically margin-negative resection (R0 resection) had the best prognosis, followed by macroscopically margin-negative resection (R1 resection), and debulking surgery was the worst (5-year OS was respectively 78.4%, 55.1% and 37.4%, χ2=24.63, P<0.001). There was no significant difference in 5-year OS between the endoscopic surgery group and the open surgery group (65.8% vs. 53.4%, χ2=2.66, P=0.102). Older patients had worse OS (HR=1.02, P=0.011) and EFS (HR=1.01, P=0.027). Patients receiving adjuvant therapy had a lower risk of death (HR=0.62, P=0.038). Patients with a history of nasal radiotherapy had a higher risk of recurrence (HR=2.48, P=0.002) and a higher risk of death (HR=2.03, P=0.020). Conclusion: For patients with advanced SNM, the efficacy of endoscopic surgery can be comparable to that of open surgery when presence of safe surgical margins, and a treatment plan based on transnasal endoscopic surgery as the main comprehensive treatment is recommended.

[Revision rate of periprosthetic joint infection post total hip or knee arthroplasty of 34 hospitals in China between 2015 and 2017: a multi-center survey].

Objective: To investigate the rate of periprosthetic joint infection (PJI) revision surgeries and clinical information of hip-/knee- PJI cases nationwide from 2015 to 2017 in China. Methods: An epidemiological investigation. A self-designed questionnaire and convenience sampling were used to survey 41 regional joint replacement centers nationwide from November 2018 to December 2019 in China. The PJI was diagnosed according to the Musculoskeletal Infection Association criteria. Data of PJI patients were obtained by searching the inpatient database of each hospital. Questionnaire entries were extracted from the clinical records by specialist. Then the differences in rate of PJI revision surgery between hip- and knee- PJI revision cases were calculated and compared. Results: Total of 36 hospitals (87.8%) nationwide reported data on 99 791 hip and knee arthroplasties performed from 2015 to 2017, with 946 revisions due to PJI (0.96%). The overall hip-PJI revision rate was 0.99% (481/48 574), and it was 0.97% (135/13 963), 0.97% (153/15 730) and 1.07% (193/17 881) in of 2015, 2016, 2017, respectively. The overall knee-PJI revision rate was 0.91% (465/51 271), and it was 0.90% (131/14 650), 0.88% (155/17 693) and 0.94% (179/18 982) in 2015, 2016, 2017, respectively. Heilongjiang (2.2%, 40/1 805), Fujian (2.2%, 45/2 017), Jiangsu (2.1%, 85/3 899), Gansu (2.1%, 29/1 377), Chongqing (1.8%, 64/3 523) reported relatively high revision rates. Conclusions: The overall PJI revision rate in 34 hospitals nationwide from 2015 to 2017 is 0.96%. The hip-PJI revision rate is slightly higher than that in the knee-PJI. There are differences in revision rates among hospitals in different regions.

[Construction of TRAF6 ubiquitin site 331 mutant colorectal cancer cell stable line and its effect on biological behavior of colorectal cancer cells].

Objective: To investigate the effect of ubiquitin mutation at position 331 of tumor necrosis factor receptor related factor 6 (TRAF6) on the biological characteristics of colorectal cancer cells and its mechanism. Methods: lentivirus wild type (pCDH-3×FLAG-TRAF6) and mutation (pCDH-3×FLAG-TRAF6-331mut) of TRAF6 gene expression plasmid with green fluorescent protein tag were used to infect colorectal cancer cells SW480 and HCT116, respectively. The infection was observed by fluorescence microscope, and the expressions of TRAF6 and TRAF6-331mut in cells was detected by western blot. Cell counting kit-8 (CCK-8) and plate cloning test were used to detect the proliferation ability of colorectal cancer cells in TRAF6 group and TRAF6-331mut group, cell scratch test to detect cell migration, Transwell chamber test to detect cell migration and invasion, immunoprecipitation to detect the ubiquitination of TRAF6 and TRAF6-331mut with ubiquitinof lysine binding sites K48 and K63. Western blot was used to detect the effects of TRAF6 and TRAF6-331mut over expression on the nuclear factor kappa-B (NF-κB) and mitogen activated protein kinase mitogen-activated protein kinase (MAPK)/activating protein-1(AP-1) signal pathway. Results: The successful infection of colorectal cancer cells was observed under fluorescence microscope. Western blot detection showed that TRAF6 and TRAF6-331mut were successfully expressed in colorectal cancer cells. The results of CCK-8 assay showed that on the fourth day, the absorbance values of HCT116 and SW480 cells in TRAF6-331mut group were 1.89±0.39 and 1.88±0.24 respectively, which were lower than those in TRAF6 group (2.09±0.12 and 2.17±0.45, P=0.036 and P=0.011, respectively). The results of plate colony formation assay showed that the number of clones of HCT116 and SW480 cells in TRAF6-331mut group was 120±14 and 85±14 respectively, which was lower than those in TRAF6 group (190±21 and 125±13, P=0.001 and P=0.002, respectively). The results of cell scratch test showed that after 48 hours, the percentage of wound healing distance of HCT116 and SW480 cells in TRAF6-331mut group was (31±12)% and (33±14)%, respectively, which was lower than those in TRAF6 group [(43±13)% and (43±7)%, P=0.005 and 0.009, respectively]. The results of Transwell migration assay showed that the migration numbers of HCT116 and SW480 cells in TRAF6-331mut group were significantly lower than those in TRAF6 group (P<0.001 and P<0.002, respectively). The results of Transwell invasion assay showed that the number of membrane penetration of HCT116 and SW480 cells in TRAF6-331mut group was significantly lower than those in TRAF6 group (P=0.008 and P=0.009, respectively). The results of immunoprecipitation detection showed that the ubiquitin protein of K48 chain pulled by TRAF6-331mut was lower than that of wild type TRAF6 in 293T cells co-transfected with K48 (0.57±0.19), and the ubiquitin protein of K63 chain pulled down by TRAF6-331mut in 293T cells co-transfected with K63 was lower than that of wild type TRAF6 (0.89±0.08, P<0.001). Western blot assay showed that the protein expression levels of NF-κB, p-NF-κB and p-AP-1 in TRAF6-331mut-HCT116 cells were 0.63±0.08, 0.42±0.08 and 0.60±0.07 respectively, which were lower than those in TRAF6-HCT116 cells (P=0.002, P<0.001 and P<0.001, respectively). The expression level of AP-1 protein in TRAF6-HCT116 cells was 0.89±0.06, compared with that in TRAF6-HCT116 cells. The difference was not statistically significant (P>0.05). The protein expression levels of NF-κB, p-NF-κB and p-AP-1 in TRAF6-331mut-SW480 cells were 0.50±0.06, 0.51±0.04, 0.48±0.02, respectively, which were lower than those in TRAF6-SW480 cells (all P<0.001). There was no significant difference in AP-1 protein expression between TRAF6-331mut-SW480 cells and TRAF6-SW480 cells. Conclusion: The ubiquitin site mutation of TRAF6 gene at 331 may prevent the binding of TRAF6 and ubiquitin lysine sites K48 and K63, and then affect the expressions of proteins related to downstream NF-κB and MAPK/AP-1 signal pathways, and inhibit the proliferation, migration and invasion of colorectal cancer cells.

Phenotype and genetic characteristics in 20 Chinese patients with 46,XY disorders of sex development.

PURPOSE: 46,XY disorders of sex development (DSD) is the most complicated and common type of DSD. To date, more than 30 genes have been identified associated with 46,XY DSD. However, the mutation spectrum of 46,XY DSD is incomplete owing to the high genetic and clinical heterogeneity. This study aims to provide clinical and mutational characteristics of 18 Chinese patients with 46,XY DSD. METHODS: A total of 20 unrelated individuals with 46,XY DSD were recruited. Whole-exome sequencing (WES) or custom-panel sequencing combined Sanger sequencing were performed to detect the pathogenic mutations. The pathogenicity of the variant was assessed according to the American College of Medical Genetics and Genomics (ACMG) guidance and technical standards recommended by the ACMG and the Clinical Genome Resource (ClinGen). RESULTS: Six patients harbored NR5A1 mutations; two patients harbored NR0B1 mutations; six patients harbored SRD5A2 mutations; six patients harbored AR mutations. Six novel genetic variants were identified involved in three genes (NR5A1, NR0B1, and AR). CONCLUSION: We determined the genetic etiology for all enrolled patients. Our study expanded the mutation spectrum of 46,XY DSD and provided diagnostic evidence for patients with the same mutation in the future.

[Phenotypes and genotypes of 78 patients with propionic acidemia].

Objective: Propionic acidemia is a rare inherited metabolic disorder caused by propionyl CoA carboxylase (PCC) deficiency. This study aims to analyze the clinical characteristics and gene variations of Chinese patients with propionic acidemia, and to explore the correlation between clinical phenotypes and genotypes. Methods: Single-center, retrospective and observational study. Seventy-eight patients of propionic acidemia (46 males and 32 females) from 20 provinces and autonomous regions were admitted from January 2007 to April 2022. Their age of initial diagnosis ranged from 7 days to 15 years. The clinical manifestations, biochemical and metabolic abnormalities, genetic variations, diagnosis, treatment and outcome were studied. Chi-Square test or Mann-Whitney U test were used for statistical analysis. Results: Among 78 cases, 6 (7.7%) were identified by newborn screening; 72 (92.3%) were clinically diagnosed after onset, and the age of onset was 2 hours after birth to 15 years old; 32 cases had early-onset disease and 40 cases had late-onset disease. The initial manifestations included lethargy, hypotonia, vomiting, feeding difficulties, developmental delay, epilepsy, and coma. Among the 74 cases who accepted gene analysis, 35 (47.3%) had PCCA variants and 39 (52.7%) had PCCB variants. A total of 39 PCCA variants and 32 PCCB variants were detected, among which c.2002G>A and c.229C>T in PCCA and c.838dupC and c.1087T>C in PCCB were the most common variants in this cohort. The variants c.1228C>T and c.1283C>T in PCCB may be related to early-onset type. The variants c.838dupC, c.1127G>T and c.1316A>G in PCCB, and c.2002G>A in PCCA may be related to late-onset disease. Six patients detected by newborn screening and treated at asymptomatic stage developed normal. The clinically diagnosed 72 cases had varied complications. 10 (12.8%) cases of them died. 62 patients improved after metabolic therapy by L-carnitine and diet. Six patients received liver transplantation because of recurrent metabolic crisis. Their clinical symptoms were markedly improved. Conclusion: The clinical manifestations of propionic acidemia are complex and lack of specificity. Newborn screening and high-risk screening are keys for early treatment and better outcome. The correlation between the genotype and phenotype of propionic acidemia is unclear, but certain variants may be associated with early-onset or late-onset propionic acidemia.

Avian predators taste reject mimetic prey in relation to their signal reliability.

Aposematic organisms defend themselves through various means to increase their unprofitability to predators which they advertise with conspicuous warning signals. Predators learn to avoid aposematic prey through associative learning that leads to lower predation. However, when these visual signals become unreliable (e.g., through automimicry or Batesian mimicry), predators may switch from using visual signals to taste sampling prey to choose among them. In this experiment, we tested this possibility in a field experiment where we released a total of 4800 mealworm prey in two clusters consisting of either: (i) undefended prey (injected with water) and (ii) model-mimics (injected with either quinine sulphate [models] or water [mimics]). Prey were deployed at 12 sites, with the mimic frequency of the model-mimics ranging between 0 and 1 (at 0.2 intervals). We found that taste rejection peaked at moderate mimic frequencies (0.4 and 0.6), supporting the idea that taste sampling and rejection of prey is related to signal reliability and predator uncertainty. This is the first time that taste-rejection has been shown to be related to the reliability of prey signals in a mimetic prey system.

[Characteristic analysis of patients with type 2 diabetes mellitus combined with advanced stage chronic liver disease: a community population-based cross-sectional study in Heping District, Shenyang City (CHESS-LN 2101)].

Objective: To investigate the prevalence and independent risk factors of non-alcoholic fatty liver disease (NAFLD) and advanced chronic liver disease among the type 2 diabetes mellitus (T2DM) population in the Shenyang community, so as to provide evidence for the prevention and control of T2DM combined with NAFLD. Methods: This cross-sectional study was conducted in July 2021. 644 T2DM cases from 13 communities in Heping District, Shenyang City were selected. All the surveyed subjects underwent physical examination (measurements of height, body mass index, neck circumference, waist circumference, abdominal circumference, hip circumference, and blood pressure), infection screening (excluding hepatitis B and C, AIDS, and syphilis), random fingertip blood glucose, controlled attenuation parameter (CAP), and liver stiffness measurement (LSM). The study subjects were divided into the non-advanced chronic liver disease group and the advanced chronic liver disease group according to whether the LSM value was greater than 10 kPa. Cirrhotic portal hypertension development was indicated in patients with LSM ≥ 15 kPa. The comparison of multiple mean values among the sample groups was performed by analysis of variance when the normal distribution was met. Results: In the T2DM community population, there were 401 cases (62.27%) combined with NAFLD, 63 cases (9.78%) combined with advanced chronic liver disease, and 14 cases (2.17%) combined with portal hypertension. There were 581 cases in the non-advanced chronic liver disease group and 63 cases (9.78%) in the advanced chronic liver disease group (LSM ≥10 kPa), including 49 cases (7.61%) with 10 kPa≤LSM<15 kPa, 11 cases (1.71%) with 15 kPa ≤LSM<25 kPa, and 3 cases (0.47%) with LSM ≥ 25 kPa. Age, body mass, body mass index, neck circumference, waist circumference, hip circumference, waist-to-height ratio, systolic blood pressure, and CAP were all statistically different between the non-advanced chronic liver disease group and the advanced chronic liver disease group (F=-1.983,-2.598,-4.091,-2.062,-3.909, -4.581,-4.295,-2.474, and -5.191, respectively; P<0.05). There was a statistically significant difference in terms of whether or not there was combined cerebrovascular disease (2=4.632, P=0.031); however, there were no statistically significant differences in terms of lifestyle, diabetes complications, and other complications (P>0.05). Conclusion: Patients with T2DM have a higher prevalence of NAFLD (62.27%) than those with advanced chronic liver disease (9.78%). 2.17% of T2DM cases in the community may not have had early diagnosis and early intervention, and they might have been combined with cirrhotic portal hypertension. So, the management of these patients should be strengthened.