Objective: To optimize the technology and process for recovering immune cells from discarded leukocyte filter through reverse flushing, and to investigate the effects of different reverse flushing solutions and physical conditions (temperature, soaking, and ultrasound) on the cell recovery rate. Additionally, to evaluate the feasibility and quality of isolating and culturing NK cells from the recovered cells. Methods: Normal saline, PBS, and PBS-EDTA were used as reverse flushing solutions to backwash the filter, and the white blood cell recovery rates were compared. The reverse flushing efficiency was optimized by adjusting the temperature of the reverse flushing solution, filter soaking treatment and ultrasonic treatment. The Ficoll separation method was employed to isolate peripheral blood mononuclear cells (PBMCs) from the elution solution and conduct in vitro expansion and culture. Lymphocyte subsets were identified and quantified using flow cytometry. Results: The physiological saline served as the backflush fluid was the most stable. The reverse temperature had a significant impact on the recovery rate of white blood cell subgroups. The recovery rates of lymphocytes and monocytes at 37℃ were significantly higher than those at 4℃ [(35.56±5.55)% vs (22.88±0.31)%, (32.42±4.82)% vs (8.00±1.6)%, t=5.581 and 5.758, respectively, both P<0.001)], and at 25℃ [(35.56±5.55)% vs (22.42±6.13)%, t=5.904, P<0.001; (32.4±4.82)% vs (21.11±1.23)%, t=2.629, P=0.030)]. The recovery rates of white blood cell subgroups after 37℃ immersion were higher than those after direct reverse flow at 37℃, but the difference was not statistically significant (P>0.05). The recovery rate of cells after ultrasonic immersion at 25℃ was further increased compared to that after immersion at 25℃, but the difference was not statistically significant (P>0.05). The purity of the NK cells obtained through sorting reached (94.2±4.5)%, and the viability was (94.8±0.9)%. After 15 days of culture, the cell count was expanded approximately 450 times, yielding more than 10 /mL highly active NK cells. Conclusion: By optimizing the backflushing process, immune cells can be efficiently recovered from discarded filters. The 37℃ backflow liquid has the best backflow effect; soaking and ultrasonic treatment can be used as auxiliary methods to improve recovery rates. The NK cells from this source have high purity, good activity, and strong expansion ability, and can be used as a reliable cell source for immunotherapy research, and in line with the circular economy concept of resource reuse.

Vav guanine nucleotide exchange factor 3 (Vav3) protein is one of the guanine nucleotide exchange factors of the Rho family GTPases. It is encoded by the proto-oncogene Vav3 and is involved in the regulation of cell proliferation and apoptosis, differentiation, migration, etc. In recent years, Vav3 has been closely related to the development of a variety of malignant tumors. In glioma, breast cancer, non-small cell lung cancer, gastric cancer, pancreatic cancer, colorectal cancer, prostate cancer, ovarian cancer, osteosarcoma and acute leukemia, the expression of Vav3 is elevated to varying degrees, and it participates in regulating multiple signaling pathways, which promotes the progression of tumors and affects the prognosis of patients. Therefore, Vav3 is expected to be a potential therapeutic target for these malignant tumors.

Objective:To explore the clinical and genetic characteristics of a fetus diagnosed with Congenital myasthenic syndrome type 16 (CMS16).Methods:A couple who had visited Tianjin Medical University General Hospital in February 2018 due to "adverse outcome of two pregnancies" was selected as the study subject. Clinical data was gathered. Peripheral blood and amniotic fluid samples were collected and subjected to whole exome sequencing (WES). Candidate variant was verified by Sanger sequencing. Low-depth whole-genome sequencing was carried out to detect copy number variation (CNV) in the fetus.Results:The couple′s first pregnancy had resulted in a miscarriage at 27 + 5 weeks, when ultrasound had revealed pleural effusion and polyhydramnios in the fetus. Their second pregnancy was terminated at 30 + 5 weeks due to fetal hand malformations, polyhydramnios and pleural fluid. Both couple had denied family history of genetic conditions. For their third pregnancy, no CNV abnormality was detected, whilst a compound heterozygous variants, including a maternally derived c. 3172C>T (p.R1058W) and paternal c. 1431delG (p.K477fs*89) in the SCN4A gene were detected. Based on the guidelines from the American College of Medical Genetics and Genomics, the c. 3172C>T (p.R1058W) was predicted as a likely pathogenic variant (PM1+ PM2_supporting+ PP3+ PP4), whilst the c. 1431delG (p.K477fs*89) was predicted as a pathogenic variant (PVS1+ PM2_supporting+ PP4). Conclusion:The c. 3172C>T (p.R1058W) and c. 1431delG (p.K477fs*89) compound heterozygous variants of the SCN4A gene probably underlay the CMS16 in the third fetus.

Objective:To explore the clinical and molecular basis for a Chinese pedigree affected with Complete androgen insensitivity syndrome (CAIS).Methods:A CAIS pedigree presented at Tianjin Medical University General Hospital between 2019 and 2021 was selected as the study subject. Clinical data of the proband was collected, along with peripheral blood samples from the proband and her family members. Chromosomal karyotyping, sex-determining region of the Y chromosome ( SRY) testing, and next-generation sequencing (NGS) were carried out for the proband, and candidate variant was verified by Sanger sequencing of her family members. Prenatal diagnosis was provided for the sister of the proband. This study was approved by Medical Ethics Committee of the Tianjin Medical University General Hospital (Ethics No. IRB2023-WZ-070). Results:The 18-year-old proband, who has a social gender of female, underwent laparoscopic examination, which showed no presence of uterus and ovaries. The karyotype of peripheral blood sample was 46, XY, with SRY gene detected. NGS indicated that the proband has harbored a heterozygous c. 1988C>G (p.Ser663Ter) variant of the AR gene. Sanger sequencing confirmed that her mother and sister had both harbored the same variant, whilst her father and younger sister were of the wild-type. Prenatal diagnosis revealed that her sister′s first fetus had harbored carried the same variant, which had led to termination of pregnancy. Her second fetus did not carry the variant, and a healthy boy was born. Based on guidelines from the American College of Medical Genetics and Genomics (ACMG), the variant was classified as likely pathogenic (PM2_Supporting+ PM4+ PP3_Moderate+ PP4). Conclusion:The c. 1988C>G (p.Ser663Ter) variant of the AR gene probably underlay the CAIS in the proband. The accurate diagnosis of sex development disorders will rely on the physicians′ thorough understanding of the clinical symptoms and pathogenic genes. Genetic testing and counseling can enable precise diagnosis, prenatal diagnosis, and guidance for reproduction

Benign gallbladder diseases are common in general surgery practice, and laparoscopic cholecystectomy (LC) has become the standard treatment for these conditions. With the widespread adoption of LC and the improved understanding of gallbladder disease, it is necessary to re-evaluate the diagnosis and treatment of benign gallbladder disease. Based on the latest expert consensus on the diagnosis and treatment of benign gallbladder disease, this review aims to provide an overview of the diagnostic and treatment strategies for benign gallbladder diseases. Specifically, it focuses on enhancing our understanding of the role of benign gallbladder diseases in the development of gallbladder cancer, the impact of congenital biliary anomalies on the diagnostic and treatment processes of benign gallbladder diseases, and the significance of the preoperative examinations.

Objective:The epidemiological and clinical characteristics of 18 cases of Japanese spotted fever (JSF) in Zigui County were analyzed in order to improve the prevention and treatment of JSF.Methods:This is a case series analysis. The epidemiological and clinical data, laboratory tests and imaging characteristics of 18 JSF cases with median age of 60 years (54, 68) identified by The People′s Hospital of Zigui from April 2021 to August 2022 were collected and analyzed retrospectively.Results:Most (17/18) of the patients were farmers and all had a field exposure history. The patient′s onset was from April to October. Spring and autumn were the seasons with the highest incidence of JSF. The first symptoms of patients were high fever, headache, and fatigue. Of the 18 cases, 15 had a rash and 12 presented an eschar and 3 had neither rash nor eschar. In addition, 10 of 18 cases experienced edema of both lower extremities, and 3 got disturbance of consciousness. Laboratory tests found that 15 patients had abnormal white blood cells and 11 patients had decreased platelets. C-reactive protein, procalcitonin, D-dimer, lactate dehydrogenase, and alpha-hydroxybutyrate dehydrogenase were elevated in all patients; 13 patients with elevated alanine aminotransferase, 14 patients with elevated aspartate transamination. Kidney damage caused by Rickettsia japonica infection showed by abnormal proteinuria in 11 of the patients. Conclusions:The most common clinical manifestations of JSF are non-specific indications such as high fever, chills, fatigue, headache. The eschar and rash, which are the main features of Rickettsia infection, are not present in all patients, resulting delay of diagnosis or misdiagnosis. Medical workers should be more alert to rickettsial infections in patients with fever of unknown origin, especially in seasons of high incidence of spotted fever. Early diagnosis and correct antibiotic treatment shall be given according to the patient′s clinical manifestations, laboratory results and imaging test to control disease progression.

Objective:To study the risk factors of lymph node metastases in patients with intrahepatic cholangiocarcinoma (ICC) and to establish a risk prediction model of lymph node metastases in ICC.Methods:The clinicopathological data of 587 ICC patients who underwent radical hepatectomy and lymph node dissection at Third Affiliated Hospital of Naval Medical University (Shanghai Eastern Hepatobiliary Surgery Hospital) from January 2007 to December 2011 were retrospectively analyzed. There were 395 males and 192 females with ages which ranged from 20 to 82 (54.7±10.8) years. Independent risk factors of lymph node metastases were studied using univariate and multivariate logistic regression analysis, and a risk prediction model was established. Receiver operating characteristic (ROC) curve was used to evaluate the accuracy of this model.Results:Of 587 patients, 158 (26.9%) had lymph node metastases. Multivariate logistic regression analysis showed that platelet count >300×10 9/L ( OR=1.985, 95% CI: 1.030-3.824, P=0.041), carbohydrate antigen 19-9 >37 U/ml ( OR=2.978, 95% CI: 1.994-4.448, P<0.001), tumor situated in left hemiliver ( OR=1.579, 95% CI: 1.065-2.341, P=0.023), multiple tumors ( OR=1.846, 95% CI: 1.225-2.783, P=0.003), and absence of cirrhosis ( OR=2.125, 95% CI: 1.192-3.783, P=0.011) were independent risk factors for lymph node metastases in ICC. The area under the ROC curve was 0.714, with a cutoff value of 0.215, and the sensitivity and specificity being 75.9% and 58.3%, respectively. Conclusions:The risk prediction model of ICC lymph node metastases was established using readily available clinical data obtained before operation. This model has good predictive values and can provide a reference for treatment decision on patients with ICC.

Cholangiocarcinoma (CCA) is a group of solid tumors with high malignant degree and poor prognosis. Surgical resection has still been the main therapy options. Targeting therapy and immunotherapy are the main anti-tumor methods that have been paid more and more attention in recent years, especially immunotherapy. The tumor microenvironment of CCA is complex, which encompasses not only interstitial and endothelial cells, but also a large number of immune cells. In addition, the innate and adaptive immune systems also play a role. This article summarizes the immune-related studies of cholangiocarcinoma and the latest clinical trials of immunotherapy.

Gallbladder cancer is a high malignancy which is predisposed to invade adjacent organs and have lymph node metastasis. Gallbladder cancer is not sensitive to radiotherapy or chemotherapy with the worst prognosis among biliary tract cancers. At present, radical resection is the only possible method to cure gallbladder cancer. However, there are still many controversies about the surgical strategies, the extent of liver resection and lymph node dissection, and the treatment of incidental gallbladder cancer. In addition, under the background of the great success of immunotherapy and targeted therapy in a variety of solid tumors, it is also a question worthy of further considerations that whether the status of surgery in the treatment of advanced gallbladder cancer will be changed in the near future.

Major obstetric hemorrhage (MOH) may endanger the life of maternal during the perioperative period. Its highly efficient management involves multidisciplinary co-operation and is a complex clinical emergency situation which needs teamwork to complete. Anesthesiologist, as the key member of the obstetric MOH management team, should actively cooperate with the obstetricians once they decide to perform an emergency procedure for the parturient. They should possess perfect skill in resuscitation and rich experiences for assessment of MOH. They also should be professional in monitoring and caring for the critically ill patient. Constantly simulation training also should be actively carried out for MOH. In this review, the definition and etiology of MOH, drug and surgical treatment, anesthesia management (including preparation before anesthesia, hemorrhagic shock assessment, choice of anesthesia methods, blood transfusion, blood cell salvage, coagulation function and hemodynamic monitoring) were reviewed, and the conclusions were just for references.