[摘 要] 目的：探讨异位表达血红蛋白亚基（HBA/HBB）对缺氧条件下嵌合抗原受体T细胞（CAR-T细胞）功能障碍的改善作用及其对肿瘤细胞的杀伤效应。方法：全基因合成技术合成靶向HER2的CAR序列，构建共表达HBA或HBB的CAR慢病毒载体，包装慢病毒后感染人原代T淋巴细胞，制备异位表达HBA/HBB的CAR-T细胞，命名为HBA CAR-T和HBB CAR-T。采用缺氧探针检测小鼠实体瘤缺氧状态。通过流式细胞术检测瘤内CAR-T细胞占比、异位表达血红蛋白亚基的CAR-T细胞阳性率及CAR-T细胞的活性氧、凋亡水平。WB法检测HBA CAR-T和HBB CAR-T内相关血红蛋白亚基表达情况，采用细胞计数板计数检测细胞增殖水平，通过萤光素酶报告基因法检测CAR-T细胞对肿瘤细胞的杀伤能力，qPCR检测CAR-T细胞中缺氧诱导因子-1α（HIF-1α）表达水平，利用MitoXpress Intra试剂盒检测CAR-T细胞内氧气含量。结果：不同细胞构建的实体瘤模型均存在明显缺氧情况，且CAR-T细胞浸润水平与缺氧程度呈显著负相关（P < 0.000 1）。HBA CAR-T与HBB CAR-T构建成功（阳性率 > 60%），相应血红蛋白亚基可稳定表达。缺氧环境下HBA CAR-T和HBB CAR-T的ROS水平、凋亡水平显著下降，增殖、对肿瘤细胞的体外杀伤能力显著强于传统CAR-T细胞（均P < 0.05）。HBA CAR-T与HBB CAR-T内HIF-1α表达降低（均P < 0.001），且缺氧程度显著降低（均P < 0.001）。结论：异位表达血红蛋白亚基可改善缺氧条件下CAR-T细胞功能障碍并增强其对肿瘤细胞的体外杀伤作用。

Amyotrophic lateral sclerosis (ALS) is a chronic, progressive degenerative disease affecting both upper and lower motor neurons, primarily characterized by skeletal muscle weakness and atrophy. Notably, the same muscle group may exhibit asynchronous involvement. This study aims to investigate the involvement patterns of the orbicularis oculi (OOc) and orbicularis oris (OOr) in ALS patients, compare the findings with healthy controls (HCs) and myasthenia gravis (MG) patients, and explore the characteristics and clinical significance of facial muscle involvement in ALS.

Objective To gain insights into the biological characteristics of different populations of Culex tritaeniorhynchus within and around the Yellow Sea wetland from the perspective of the occurrence of resistance, we investigated the levels of resistance to deltamethrin and kdr gene mutation in the wetland and its peripheral areas. Methods Specimens were collected from Cx. tritaeniorhynchus populations at two monitoring sites in the Rare Bird National Nature Reserve and Tiaozi Ni Wetland Scenic Area, and also from two populations in Yancheng City and the Liuhe District of Nanjing, and the resistance of these mosquitoes to deltamethrin was determined using the CDC biotest bottle method. For each concentration of deltamethrin assessed, a random subset of exposed specimens was selected for amplification of the kdr gene fragment, followed by Sanger sequencing to identify and analyze resistance-associated mutations. Results The LC50 levels of deltamethrin among mosquitoes from the four populations in Luhe, Yancheng, the Rare Bird National Nature Reserve and the Tiaozi Ni Wetland Scenic Area were 2.048 5, 7.798 2, 3.473 3, and 17.695 5 mg/mL, respectively, with corresponding concentrations of deltamethrin ranging from 0.005 to 5.000,0.050 to 50.000,0.050 to 25.000 and 0.050 to 50.000 mg/mL, respectively. Furthermore, the ranges of the KT50 values were 11.76-107.43, 67.05-216.30,29.77-107.43 and 28.40-329.51 min; the 1-h knockdown rates were 34.58%-99.15%, 9.52%-43.80%, 55.09%-73.01%, and 10.09%-68.07%; and the 24-h mortality rates were 12.15%-67.52%,9.52%-79.56%,13.17%-82.21%, and 11.01%-78.99%, respectively. With respect to kdr gene mutation, we assayed a total of 63,70,59, and 57 mosquitoes for the four populations, for which we detected L1014F mutation frequencies of 14.29%, 35.00%, 20.34%, and 31.58%, respectively, with a majority of these mutations being heterozygous for resistance. In addition, five adult mosquitoes were identified has having synonymous mutations at site 1011[i. e. , AAT(asparagine)mutation to AAC(asparagine)]. Conclusions Our findings revealed the clear resistance of Cx. tritaeniorhynchus to deltamethrin in the Yancheng region of the Yellow Sea wetland, and the resistance phenotype and kdr frequency of Cx. tritaeniorhynchus in the wetland environment were comparable to those of Cx. tritaeniorhynchus in the wetland environment, thereby indicating that the resistance of different populations of Cx. tritaeniorhynchus was homogeneous under the pressure of different insecticide selection within and around the wetland. However, the underlying mechanisms need to be further studied.

Abstract Hyperuricemia (HUA) is a metabolic disorder syndrome caused by purine metabolism dysregulation, and its prevalence increases year by year. The development and progression of HUA are accompanied by significant alterations in the composition of intestinal microbiota, making probiotics a potential and safe method to reduce serum uric acid. Probiotics ameliorate HUA through three pathways: competing with intestinal epithelial cells for purine absorption to decrease uric acid synthesis, inhibiting xanthine oxidase activity through modulation of inflammatory cytokines to reduce the conversion of purine to uric acid, as well as restoring and maintaining an orderly state of the gut microbiota to facilitate normal uric acid excretion. This article reviews the role of probiotics in ameliorating HUA, so as to provide the reference for the application of probiotics in the prevention and intervention of HUA.

This study aims to integrate data mining techniques of single cell transcriptomics and spatial transcriptomics, along with animal experiment validation, so as to systematically characterize the protective effects of Jianpi Tongluo Formula(JTF) on the cartilage in knee osteoarthritis(KOA) and elucidate the underlying molecular mechanisms. Single cell transcriptomics and spatial transcriptomics datasets(GSE254844 and GSE255460) of the cartilage tissue obtained from KOA patients were analyzed to map the single cell-spatial heterogeneity and identify key pathogenic factors. After that, a KOA rat model was established via knee joint injection of papain. The intervention effects of JTF on the expression features of these key factors were assessed through real-time quantitative polymerase chain reaction(PCR), Western blot, and immunohistochemical staining. As a result, the integrated single cell and spatial transcriptomics data identified distinct cell subsets with different pathological changes in different regions of the inflamed cartilage tissue in KOA, and their differentiation trajectories were closely related to the inflammatory fibrosis-like pathological changes of chondrocytes. Accordingly, the expression levels of the two key effect targets, namely nuclear receptor coactivator 4(NCOA4) and high mobility group box 1(HMGB1) were significantly reduced in the articular surface and superficial zone of the inflamed joints when JTF effectively alleviated various pathological changes in KOA rats, thus reversing the abnormal chondrocyte autophagy level, relieving the inflammatory responses and fibrosis-like pathological changes, and promoting the repair of chondrocyte function. Collectively, this study revealed the heterogeneous characteristics and dynamic changes of inflamed cartilage tissue in different regions and different cell subsets in KOA patients. It is worth noting that NCOA4 and HMGB1 were crucial in regulating chondrocyte autophagy and inflammatory reaction, while JTF could reverse the regulation of NCOA4 and HMGB1 and correct the abnormal molecular signal axis in the target cells of the inflamed joints. The research can provide a new research idea and scientific basis for developing a personalized therapeutic schedule targeting the spatiotemporal heterogeneity characteristics of KOA.
Animals ; Drugs, Chinese Herbal/administration & dosage* ; Rats ; Osteoarthritis, Knee/pathology* ; Humans ; Male ; Cartilage, Articular/metabolism* ; Chondrocytes/metabolism* ; Rats, Sprague-Dawley ; Female ; Protective Agents/administration & dosage* ; Single-Cell Analysis ; Middle Aged ; HMGB1 Protein/metabolism*

OBJECTIVE: To explore the application value and clinical effect of 3D printing combined with customized bone plate in the treatment of acetabular fracture. METHODS: From June 2020 to June 2022, 11 patients with acetabular fractures underwent preoperative planning using 3D printing technology and were treated with customized bone plates including 8 males and 3 females, aged 25 to 66 years old. The fractures were classified according to Letournel-Judet:4 posterior wall fractures, 2 T-type fractures, 2 transverse posterior wall fractures, 2 double column fractures, and 1 anterior column with posterior semi-transverse fractures. The operative time, intraoperative blood loss, intraoperative fluoroscopy times, postoperative drainage volume, postoperative fracture healing time, and hip function score were recorded and analyzed. RESULTS: The operation time of 11 patients was 80 to 150 min, intraoperative blood volume was 150 to 700 ml, fluoroscopy frequency was 2 to 6, postoperative drainage flow was 60 to 195 ml, and the fracture healing time was 2.5 to 6.0 months. Fracture reduction was evaluated according to Matta score:anatomical reduction in 3 cases and satisfactory reduction in 8 cases. Eleven patients were followed up for 7 to 18 months. The hip Merle d'Aubigne function scores were excellent in 6 cases, good in 3 cases, fair in 1 case and poor in 1 case. Incision fat liquefaction occurred in 1 case and obturator nerve traction in 1 case. CONCLUSION The application of 3D printing technology combined with customized bone plates in the treatment of acetabular fracture is effective. In addition, the printed model can provide the operator with the results of the three-dimensional shape of the fracture, which is convenient for surgical reduction and effectively improves the efficiency of surgery.
Humans ; Female ; Male ; Middle Aged ; Acetabulum/surgery* ; Printing, Three-Dimensional ; Adult ; Aged ; Bone Plates ; Fractures, Bone/surgery* ; Fracture Fixation, Internal/methods*

OBJECTIVE: To explore prevalence, incidence and possible factors of immediate herniated discs after posterior cervical expansive open-door laminoplasty (EODL). METHODS: Totally 29 patients with cervical spinal stenosis and intervertebral disc herniation who underwent EODL from October 2020 to December 2021 were collected, including 24 males and 5 females, aged from 43 to 81 years old with an average of (61.3±9.0) years old;the courses of disease ranged from 1 to 120 months with an average of (36.4±37.0) months. Three or more intervertebral discs on C₃-C₇ were observed. The clinical efficacy was evaluated according to Japanese Orthopaedic Association (JOA) score before operation, 3 days and 1, 3, 6 and 12 months after operation, respectively. The changes of herniated disc before and after operation were measured by multipoint area method and two-dimensional distance method, and incidence and percentage of herniated disc regression were further calculated. Cervical imaging parameters such as Cobb angle (C₃-C₇), intervertebral angle, T₁ slope (T₁S), spinal canal sagittal diameter, K-line angle, dural sac sagittal diameter were measured and compared before and after operation. Pearson correlation was used to analyze correlation between cervical sagittal imaging parameters and disc herniation changes before and after operation. RESULTS: All patients obtained grade A wound healing, and 14 of them were followed up for 3(1.00, 5.25) months. There were no immediate or long-term postoperative complications. Totally 101 herniated intervertebral discs were measured, of which 79 regression numbers were obtained by area measurement. The number of intervertebral disc regressions by distance measurement was 77. There was no statistically significant difference in Cobb angle, intervertebral angle, T₁S and K-line angle of C₃-C₇ (P>0.05), however, there were statistically significant differences in sagittal diameter of spinal canal, sagittal diameter of dural sac, and JOA score before and after operation(P<0.05). The regression ratio of disc herniation ranged from 5% to 50%, and regression ratio of disc herniation was greater than 25% in 45.57%(36/79). Disc herniation in C_4,5 was positively correlated with sagittal plane diameter in C₅(r=0.423, P=0.028). There was a negative correlation between changes of C_3,4 and C_3,4 intervertebral angle (r=-0.450, P=0.041). The improvement rate of cervical JOA score immediately after operation was (59.54±15.07) %, and postoperative follow-up improved to (76.57±14.66) %. CONCLUSION Herniated disc regression immediately after EODL is a common occurrence, and EODL should be selected as far as possible under the premise of satisfying surgical indications. The regression of disc herniation is positively correlated with spinal canal sagittal diameter, and spinal canal should be enlarged as far as possible in the appropriate scope during EODL, so as to create more opportunities and conditions for disc regression and achieve better clinical results.
Humans ; Female ; Male ; Intervertebral Disc Displacement/diagnostic imaging* ; Spinal Stenosis/diagnostic imaging* ; Laminoplasty/methods* ; Middle Aged ; Aged ; Cervical Vertebrae/diagnostic imaging* ; Adult ; Aged, 80 and over ; Intervertebral Disc/surgery*

OBJECTIVE: To explore the differences in clinical efficacy between enhanced workflows and express workflows in robotic-assisted total hip arthroplasty(THA). METHODS: A retrospective analysis was conducted on 46 patients who underwent robotic-assisted THA between November 2020 and May 2021. They were divided into the enhanced workflows group and the express workflows group based on the surgical methods. There were 20 patients in the enhanced workflows group, including 11 males and 9 females;aged from 51 to 78 years old with an average of (67.30±7.52) years old. The BMI ranged from 18.24 to 24.03 kg·m^-2 with an average of(23.80±3.01) kg·m^-2. There were 26 patients in the express workflows group, including 12 males and 14 females;aged from 57 to 84 years old with a mean age of (67.58±7.29) years old, and their BMI ranged from 19.72 to 30.08 kg·m^-2 with an average of (24.41 ±2.92) kg·m^-2. The operation time, hospital stay, and perioperative complications of the patients were recorded. The postoperative acetabular prosthesis anteversion angle, abduction angle, limb length, and offset distance data were measured. The Harris hip score at the latest follow-up was recorded. RESULTS: All patients completed the surgery as planned and were followed up, with the follow-up period ranging from 47 to 54 months with a mean of (49.78±1.85) months and the length of hospital stay ranging from 2 to 11 days with an average of (6.57±1.82 ) days. The operation time of enhanced workflows group was (93.41±16.41) minutes, which was longer than that of the express workflow groups (75.19±18.36) minutes, and the difference was statistically significant (P<0.05). In enhanced workflows group, the postoperative acetabular anteversion angle was (19.20±4.46)°, the limb length discrepancy was (-1.55±9.13) mm, and changes of the offset was (-5.15±6.77) mm. The corresponding values in express workflows group were (20.46±3.29)°, (2.19±4.39) mm, and (-2.39±4.34) mm, respectively. There was no statistically significant difference in these indicators between the two groups(P>0.05). One patient in the enhanced workflows group developed deep venous thrombosis after surgery. No cases of dislocation or periprosthetic infection. At the latest follow-up, all patients had well-positioned prostheses without loosening. Harris hip score was (90.50±1.67) points in enhanced workflows group and (90.73±2.36) points in the express workflows group, with no statistically significant difference between the two groups (P>0.05). CONCLUSION The clinical efficacy of robot assisted total hip arthroplasty technology is satisfactory. The enhanced workflows will increase the surgical time. For patients with normal anatomical hip joint disease, this study did not find significant advantages in joint stability and functional scoring for the enhanced workflows.
Humans ; Arthroplasty, Replacement, Hip/methods* ; Male ; Female ; Aged ; Middle Aged ; Robotic Surgical Procedures/methods* ; Retrospective Studies ; Aged, 80 and over ; Workflow ; Treatment Outcome

PURPOSE: To investigate the protective effect of sub-hypothermic mechanical perfusion combined with membrane lung oxygenation on ischemic hypoxic injury of yorkshire brain tissue caused by traumatic blood loss. METHODS: This article performed a random controlled trial. Brain tissue of 7 yorkshire was selected and divided into the sub-low temperature anterograde machine perfusion group (n = 4) and the blank control group (n = 3) using the random number table method. A yorkshire model of brain tissue injury induced by traumatic blood loss was established. Firstly, the perfusion temperature and blood oxygen saturation were monitored in real-time during the perfusion process. The number of red blood cells, hemoglobin content, NA⁺, K⁺, and Ca²⁺ ions concentrations and pH of the perfusate were detected. Following perfusion, we specifically examined the parietal lobe to assess its water content. The prefrontal cortex and hippocampus were then dissected for histological evaluation, allowing us to investigate potential regional differences in tissue injury. The blank control group was sampled directly before perfusion. All statistical analyses and graphs were performed using GraphPad Prism 8.0 Student t-test. All tests were two-sided, and p value of less than 0.05 was considered to indicate statistical significance. RESULTS: The contents of red blood cells and hemoglobin during perfusion were maintained at normal levels but more red blood cells were destroyed 3 h after the perfusion. The blood oxygen saturation of the perfusion group was maintained at 95% - 98%. NA⁺ and K⁺ concentrations were normal most of the time during perfusion but increased significantly at about 4 h. The Ca²⁺ concentration remained within the normal range at each period. Glucose levels were slightly higher than the baseline level. The pH of the perfusion solution was slightly lower at the beginning of perfusion, and then gradually increased to the normal level. The water content of brain tissue in the sub-low and docile perfusion group was 78.95% ± 0.39%, which was significantly higher than that in the control group (75.27% ± 0.55%, t = 10.49, p < 0.001), and the difference was statistically significant. Compared with the blank control group, the structure and morphology of pyramidal neurons in the prefrontal cortex and CA1 region of the hippocampal gyrus were similar, and their integrity was better. The structural integrity of granulosa neurons was destroyed and cell edema increased in the perfusion group compared with the blank control group. Immunofluorescence staining for glail fibrillary acidic protein and Iba1, markers of glial cells, revealed well-preserved cell structures in the perfusion group. While there were indications of abnormal cellular activity, the analysis showed no significant difference in axon thickness or integrity compared to the 1-h blank control group. CONCLUSIONS Mild hypothermic machine perfusion can improve ischemia and hypoxia injury of yorkshire brain tissue caused by traumatic blood loss and delay the necrosis and apoptosis of yorkshire brain tissue by continuous oxygen supply, maintaining ion homeostasis and reducing tissue metabolism level.
Animals ; Perfusion/methods* ; Disease Models, Animal ; Brain Injuries/etiology* ; Swine ; Male ; Hypothermia, Induced/methods*