Researchers commonly use cyclization recombination enzyme/locus of X-over P1 (Cre/loxP) technology-based conditional gene knockouts of model mice to investigate the functional roles of genes of interest in Sertoli and Leydig cells within the testis. However, the shortcomings of these genetic tools include high costs, lengthy experimental periods, and limited accessibility for researchers. Therefore, exploring alternative gene silencing techniques is of great practical value. In this study, we employed adeno-associated virus (AAV) as a vector for gene silencing in Sertoli and Leydig cells. Our findings demonstrated that AAV serotypes 1, 8, and 9 exhibited high infection efficiency in both types of testis cells. Importantly, we discovered that all three AAV serotypes exhibited exquisite specificity in targeting Sertoli cells via tubular injection while demonstrating remarkable selectivity in targeting Leydig cells via interstitial injection. We achieved cell-specific knockouts of the steroidogenic acute regulatory ( Star ) and luteinizing hormone/human chorionic gonadotropin receptor (Lhcgr) genes in Leydig cells, but not in Sertoli cells, using AAV9-single guide RNA (sgRNA)-mediated gene editing in Rosa26-LSL-Cas9 mice. Knockdown of androgen receptor ( Ar ) gene expression in Sertoli cells of wild-type mice was achieved via tubular injection of AAV9-short hairpin RNA (shRNA)-mediated targeting. Our findings offer technical approaches for investigating gene function in Sertoli and Leydig cells through AAV9-mediated gene silencing.
Animals ; Male ; Leydig Cells/metabolism* ; Mice ; Dependovirus/genetics* ; Sertoli Cells/metabolism* ; Gene Silencing ; Genetic Vectors ; Testis/cytology*

OBJECTIVES: To explore the measures to improve the follow-up rate of preterm infants after discharge, and to evaluate the effectiveness of these measures using quality improvement methodology. METHODS: The follow-up status of preterm infants discharged from March to May 2017 was used as the baseline before quality improvement, and a specific quality improvement goal for the follow-up rate was proposed. The Pareto chart was used to analyze the causes of follow-up failure, and a key driver diagram was constructed based on the links involved in improving follow-up rate. The causes of failure were analyzed to determine the key links and intervention measures for quality improvement, and the follow-up rate was monitored weekly using a control chart until the quality improvement goal was achieved. RESULTS: The follow-up rate of preterm infants after discharge was 57.92% (117/202) at baseline before quality improvement, and the quality improvement goal was set to increase the follow-up rate of preterm infants from baseline to more than 80% within 12 months. The Pareto chart analysis showed that the main causes of follow-up failure were deficiencies in follow-up file management and irregular follow-up times (33.70%, 31/92), insufficient follow-up education and poor communication (25.00%, 23/92), and the inability to meet the diverse needs of parents (18.48%, 17/92). Based on the key links for quality improvement and the main causes of follow-up failure, the following intervention measures were adopted: (1) strengthen follow-up publicity and education; (2) build a follow-up team; and (3) establish a follow-up platform and system. The control chart indicated that with the implementation of the above intervention measures, the weekly follow-up rate increased to 74.09% (306/413) in July 2017 and 83.09% (511/615) in December 2017, finally achieving the quality improvement goal. During the COVID-19 pandemic, the follow-up rate of preterm infants fluctuated between 23.54% (460/1 954) and 70.97% (1 931/2 721), and subsequently, it returned to pre-pandemic levels starting in February 2023. CONCLUSIONS The application of quality improvement methodology can help to formulate intervention measures based on the main causes of follow-up failure, thereby improving the follow-up rate of preterm infants after discharge. This quality improvement method is feasible and practical and thus holds promise for clinical application.
Humans ; Quality Improvement ; Infant, Premature ; Infant, Newborn ; Patient Discharge ; Follow-Up Studies ; Female ; Male

OBJECTIVES: To investigate the clinical features and prognosis of children with non-Down-syndrome-related acute megakaryoblastic leukemia (non-DS-AMKL). METHODS: A retrospective analysis was conducted on the medical data of 17 children with non-DS-AMKL who were admitted to Children's Hospital of The First Affiliated Hospital of Zhengzhou University from January 2013 to December 2023, and their clinical features, treatment, and prognosis were summarized. RESULTS: Among the 17 children with non-DS-AMKL, there were 8 boys and 9 girls. Fourteen patients had an onset age of less than 36 months, with a median age of 21 months (range:13-145 months). Immunophenotyping results showed that 16 children were positive for CD61 and 13 were positive for CD41. The karyotype analysis was performed on 16 children, with normal karyotype in 6 children and abnormal karyotype in 9 children, among whom 5 had complex karyotype and 1 had no mitotic figure. Detected fusion genes included EVI1, NUP98-KDM5A, KDM5A-MIS18BP1, C22orf34-BRD1, WT1, and MLL-AF9. Genetic alterations included TET2, D7S486 deletion (suggesting 7q-), CSF1R deletion, and PIM1. All 17 children received chemotherapy, among whom 16 (94%) achieved complete remission after one course of induction therapy, and 1 child underwent hematopoietic stem cell transplantation (HSCT) and remained alive and disease-free. Of all children, 7 experienced recurrence, among whom 1 child received HSCT and died of graft-versus-host disease. At the last follow-up, six patients remained alive and disease-free. CONCLUSIONS Non-DS-AMKL primarily occurs in children between 1 and 3 years of age. The patients with this disorder have a high incidence rate of chromosomal abnormalities, with complex karyotypes in most patients. Some patients harbor fusion genes or gene mutations. Although the initial remission rate is high, the long-term survival rate remains low.
Humans ; Male ; Female ; Leukemia, Megakaryoblastic, Acute/etiology* ; Child, Preschool ; Infant ; Child ; Retrospective Studies ; Prognosis ; Down Syndrome/complications*

Objective Little is known about the association between whole-blood nicotinamide adenine dinucleotide(NAD+)levels and nabothian cysts.This study aimed to assess the association between NAD+levels and nabothian cysts in healthy Chinese women. Methods Multivariate logistic regression analysis was performed to analyze the association between NAD+levels and nabothian cysts. Results The mean age was 43.0±11.5 years,and the mean level of NAD+was 31.3±5.3 μmol/L.Nabothian cysts occurred in 184(27.7%)participants,with single and multiple cysts in 100(15.0%)and 84(12.6%)participants,respectively.The total nabothian cyst prevalence gradually decreased from 37.4%to 21.6%from Q1 to Q4 of NAD+and the prevalence of single and multiple nabothian cysts also decreased across the NAD+quartiles.As compared with the highest NAD+quartile(≥34.4 μmol/L),the adjusted odds ratios with 95%confidence interval of the NAD+Q1 was 1.89(1.14-3.14)for total nabothian cysts.The risk of total and single nabothian cysts linearly decreased with increasing NAD+levels,while the risk of multiple nabothian cysts decreased more rapidly at NAD+levels of 28.0 to 35.0 μmol/L. Conclusion:Low NAD+levels were associated with an increased risk of total and multiple nabothian cysts.

Objective To explore the current application of high-throughput drug sensitivity(HDS)testing in children with relapsed and refractory acute leukemia(RR-AL)and analyze the feasibility of salvage treatment plans.Methods A retrospective collection of clinical data from children with RR-AL who underwent HDS testing at the Department of Children's Hematology and Oncology of the First Affiliated Hospital of Zhengzhou University from November 2021 to October 2023 was conducted,followed by an analysis of drug sensitivity results and treatment outcomes.Results A total of 17 children with RR-AL underwent HDS testing,including 7 cases of relapsed refractory acute myeloid leukemia and 10 cases of relapsed refractory acute lymphoblastic leukemia.The detection rate of highly sensitive chemotherapy drugs/regimens was 53%(9/17),while the detection rate of moderately sensitive chemotherapy drugs/regimens was 100%(17/17).Among the 17 RR-AL patients with highly and moderately sensitive chemotherapy drugs and regimens,the MOACD regimen(mitoxantrone+vincristine+cytarabine+cyclophosphamide+dexamethasone)accounted for 100%,with the highest inhibition rate for single-agent mitoxantrone(94%,16/17),and the highest inhibition rate for targeted therapy being bortezomib(94%,16/17).Nine patients adjusted their chemotherapy based on HDS testing results,with 4 undergoing hematopoietic stem cell transplantation.Four patients achieved disease-free survival,while 5 died.Eight patients received empirical chemotherapy,with 2 undergoing hematopoietic stem cell transplantation;4 achieved disease-free survival,while 4 died.Conclusions HDS testing can identify highly sensitive drugs/regimens for children with RR-AL,improving the rate of re-remission and creating conditions for subsequent hematopoietic stem cell transplantation.

Studies have shown that a variety of diseases such as cardiovascular disease, renal disease and cancer are closely related to trimethylamine oxide (TMAO). Clinically, abnormal elevation of TMAO has been used as an evaluation index of atherosclerosis (AS) prior to imaging. In this study, we investigated the effects of lipid metabolism disorders as well as pharmacological interventions on urinary TMAO using a hyperlipidemic golden gopher model. The study used 48 Syrian golden hamster modeled with a high-fat diet for 2 weeks, and then ezetimibe, simvastatin, ezetimibe and simvastatin groups were administered for 4 consecutive weeks, as well as the clinical trial drug, IMM-H007, for pharmacological intervention. The animal experiment was conducted in accordance with the regulations of the Ethics Committee for Experimental Animal Management and Animal Welfare of Institute of Materia Medica, Chinese Academy of Medical Sciences (approval number: SCXK (Beijing) 2021-0011). Urine from rats was analyzed for 2D band selective heteronuclear single quantum coherence (2D bs-HSQC) at week 2 and 4 after drug administration. The results indicated that, in comparison to the control group, the high-fat diet significantly elevated urinary TMAO levels in the model group of hamsters after both 2 and 4 weeks of treatment (P < 0.05). Urinary TMAO levels were significantly reduced (P < 0.05) in the model group after 2 or 4 weeks of intervention with ezetimibe, simvastatin, combination therapy, and IMM-H007, showing a marked decrease even after 2 weeks of treatment. The detection of TMAO could precede the measurement of serum biochemical indicators, facilitating earlier efficacy assessment. This study evaluated the modulatory effects of clinical drugs and clinical trial drugs on TMAO, which provides useful information for clinical drug use and drug research. It also provides a means of TMAO detection based on 2D NMR technology, which is helpful for the clinical application of TMAO detection index.

Objective: To analyze the incidence and the related risk factors of retropharyngeal lymph node metastasis in patients with hypopharyngeal squamous cell carcinoma, evaluate the accuracy of preoperative enhanced CT in judging retropharyngeal lymph node metastasis, and investigate the impact of retropharyngeal lymph node metastasis on the prognosis. Methods: Retrospective analyses were made on 398 patients with hypopharyngeal squamous cell carcinoma who underwent surgery as the primary therapy and accepted retropharyngeal lymph node exploration and clearance during surgery in Shandong Provincial ENT Hospital from January 2014 to December 2019. Multivariate logistic regression analysis was used to clarify the related risk factors of retropharyngeal lymph node metastasis. Multivariate Cox regression analysis was used to investigate the impact of retropharyngeal lymph node metastasis on prognosis. The retropharyngeal lymph nodes of 218 cases with available preoperative enhanced CT images were evaluated by two experienced radiologists and compared with postoperative pathological results. Results: Retropharyngeal lymph node metastasis were confirmed in 54 of 398 (13.6%) cases according to postoperative pathology. The sensitivity and specificity of preoperative enhanced CT in the diagnosis of retropharyngeal lymph node metastasis were 34.6% and 91.1%, respectively, and the overall accuracy was 84.4%. Multivariate logistic regression analysis showed that the site of the primary lesion and pathological N stage were independent risk factors for retropharyngeal lymph node metastasis in hypopharyngeal squamous cell carcinoma. Patients with primary lesion located in the posterior wall of hypopharynx (OR=4.83, 95% CI: 1.27-18.40), N2 stage (OR=6.30, 95% CI: 2.25-17.67), and N3 stage (OR=26.89, 95% CI: 5.76-125.58) were prone to retropharyngeal lymph node metastasis. The 5-year overall survival rate of the 398 patients was 50.4%, and the 5-year disease-free survival rate was 48.3%. Multivariate Cox regression analysis showed that T stage, N stage, retropharyngeal lymph node metastasis, and radiotherapy were independent influencing factors for overall survival (T stage: HR=1.28, 95% CI: 1.06-1.54; N stage: HR=1.26, 95% CI: 1.14-1.40; retropharyngeal lymph node metastasis: HR=2.13, 95% CI: 1.47-3.08; radiotherapy: HR=0.54, 95% CI: 0.38-0.76) and disease-free survival of patients with hypopharyngeal squamous cell carcinoma (T stage: HR=1.26, 95% CI: 1.06-1.51; N stage: HR=1.25, 95% CI: 1.13-1.37; retropharyngeal lymph node metastasis: HR=2.24, 95% CI: 1.56-3.21; radiotherapy: HR=0.55, 95% CI: 0.40-0.77). Conclusions: Metastasis of retropharyngeal lymph nodes in hypopharyngeal squamous cell carcinoma is not rare. Enhanced CT is of low accuracy and limited value in diagnosing retropharyngeal lymph node metastasis. Primary lesions located in the posterior wall of the hypopharyngx, N2 stage, and N3 stage are independent high-risk factors for retropharyngeal lymph node metastasis. The prognosis of hypopharyngeal cancer patients with retropharyngeal lymph node metastasis is worse, and active surgical exploration and clearance can effectively reduce the mortality caused by retropharyngeal lymph node metastasis.
Humans ; Squamous Cell Carcinoma of Head and Neck/pathology* ; Lymphatic Metastasis/pathology* ; Retrospective Studies ; Carcinoma, Squamous Cell/surgery* ; Lymph Nodes/pathology* ; Hypopharyngeal Neoplasms/surgery* ; Prognosis ; Head and Neck Neoplasms/pathology* ; Neoplasm Staging

OBJECTIVE: To investigate the involvement of endothelial cells (ECs)-derived exosomes in the anti-apoptotic effect of Danhong Injection (DHI) and the mechanism of DHI-induced exosomal protection against postinfarction myocardial apoptosis. METHODS: A mouse permanent myocardial infarction (MI) model was established, followed by a 14-day daily treatment with DHI, DHI plus GW4869 (an exosomal inhibitor), or saline. Phosphate-buffered saline (PBS)-induced ECs-derived exosomes were isolated, analyzed by miRNA microarray and validated by droplet digital polymerase chain reaction (ddPCR). The exosomes induced by DHI (DHI-exo), PBS (PBS-exo), or DHI+GW4869 (GW-exo) were isolated and injected into the peri-infarct zone following MI. The protective effects of DHI and DHI-exo on MI hearts were measured by echocardiography, Masson's trichrome staining, and TUNEL apoptosis assay. The Western blotting and quantitative reverse transcription PCR (qRT-PCR) were used to evaluate the expression levels of miR-125b/p53-mediated pathway components, including miR-125b, p53, Bak, Bax, and caspase-3 activities. RESULTS: DHI significantly improved cardiac function and reduced infarct size in MI mice (P<0.01), which was abolished by the GW4869 intervention. DHI promoted the exosomal secretion in ECs (P<0.01). According to the results of exosomal miRNA microarray assay, 30 differentially expressed miRNAs in the DHI-exo were identified (28 up-regulated miRNAs and 2 down-regulated miRNAs). Among them, DHI significantly elevated miR-125b level in DHI-exo and DHI-treated ECs, a recognized apoptotic inhibitor impeding p53 signaling (P<0.05). Remarkably, treatment with DHI and DHI-exo attenuated apoptosis, elevated miR-125b expression level, inhibited capsase-3 activity, and down-regulated the expression levels of proapoptotic effectors (p53, Bak, and Bax) in post-MI hearts, whereas these effects were blocked by GW4869 (P<0.05 or P<0.01). CONCLUSION DHI and DHI-induced exosomes inhibited apoptosis, promoted the miR-125b expression level, and regulated the p53 apoptotic pathway in post-infarction myocardium.
Mice ; Animals ; Tumor Suppressor Protein p53/metabolism* ; Endothelial Cells/metabolism* ; Exosomes/metabolism* ; bcl-2-Associated X Protein/metabolism* ; Myocardium/metabolism* ; Myocardial Infarction/drug therapy* ; Apoptosis ; MicroRNAs/metabolism*

Objective: To explore the dynamic changes in serum lipid levels and nutritional status during BCMA-CAR-T-cell therapy in patients with refractory or relapsed multiple myeloma (R/R MM) based on LEGEND-2. Methods: The data of patients with R/R MM who underwent BCMA-CAR-T therapy at our hospital between March 30, 2016, and February 6, 2018, were retrospectively collected. Serum lipid levels, controlled nutritional status (CONUT) score, and other clinical indicators at different time points before and after CAR-T-cell infusion were compared and analyzed. The best cut-off value was determined by using the receiver operator characteristic (ROC) curve. The patients were divided into high-CONUT score (>6.5 points, malnutrition group) and low-CONUT score groups (≤6.5 points, good nutrition group), comparing the progression-free survival (PFS) and total survival (OS) of the two groups using Kaplan-Meier survival analysis. Results: Before the infusion of CAR-T-cells, excluding triglycerides (TG), patients' serum lipid levels were lower than normal on average. At 8-14 d after CAR-T-cell infusion, serum albumin (ALB), total cholesterol (TC), high-density lipoprotein (HDL), low-density lipoprotein (LDL), and apolipoprotein A1 (Apo A1) levels dropped to the minimum, whereas CONUT scores reached the maximum. In addition to TG, apolipoprotein B (Apo B) levels increased compared with baseline. After CAR-T-cell therapy, the patients' serum lipid levels significantly increased with well-improved nutritional status. Spearman's related analysis showed that TC, HDL, and ApoA1 levels after CAR-T-cell injection were significantly negatively correlated with the grade of cytokine-release syndrome (CRS) (r=-0.548, P=0.003; r=-0.444, P=0.020; r=-0.589, P=0.001). Furthermore, survival analysis indicated that the CONUT score was unrelated to PFS, and the median OS of patients with R/R MM in the high-CONUT score group was shorter than that in the low-CONUT score group (P=0.046) . Conclusions: During CAR-T-cell therapy, hypolipidemia and poor nutritional status were aggravated, which is possibly related to CRS. The patients' serum lipid levels and nutritional status were significantly improved after CAR-T-cell treatment. The CONUT score affected the median OS in patients treated with CAR-T-cells. Therefore, specific screening and intervention for nutritional status in patients receiving CAR-T-cell therapy are required.
Humans ; Multiple Myeloma/drug therapy* ; Nutritional Status ; Retrospective Studies ; Receptors, Chimeric Antigen/therapeutic use* ; B-Cell Maturation Antigen/therapeutic use* ; Cell- and Tissue-Based Therapy ; Lipids/therapeutic use*

Since the end of 2019,severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)infection has swept the world,bringing great harm to human society and significantly increasing the health burden.Due to stron-ger infectivity,faster transmission,and higher reinfection rate of the Omicron variant,it has now replaced the Delta variant as the main epidemic strain for both imported and local outbreaks in China.Chinese Diagnosis and treatment protocol for SARS-CoV-2 infection(10th trial version)emphasizes"strengthening the protection of key popula-tions,"which includes the increasing number of immunocompromised population.These people have a high inci-dence of severe diseases and a high fatality rate after infected with SARS-CoV-2,and belong to the high-risk popula-tions of severe or critical diseases.Moreover,due to underlying diseases,these people take immunosuppressants and other related drugs chronically.The interactions between anti-SARS-CoV-2 infection treatment drugs and origi-nal drugs are complicated,thus bring significant challenges to the treatment after the SARS-CoV-2 infection.Cur-rently,there is a lack of guidelines or consensus on the diagnosis and treatment of SARS-CoV-2 infection among im-munocompromised population.Therefore,the Guangzhou Institute of Respiratory Health and National Center for Respiratory Medicine organized experts from multiple disciplines(respiratory and critical care medicine,organ transplantation,rheumatology and immunology,hematology,infection,critical care medicine,etc.)in China.Af-ter multiple rounds of discussions,13 items of recommendations are made as the reference for peers based on evi-dence-based medical evidence,so as to provide a theoretical and practical reference for the diagnosis and treatment strategies of this population.