BACKGROUND:Intervertebral disc degeneration is clinically considered to be the main cause of low back pain,but due to the unclear pathogenesis of intervertebral disc degeneration,there is still a lack of effective means to delay the progression of the disease.Single-cell RNA sequencing technology can amplify and sequence mRNA at the single-cell level,reveal the gene expression intensity of a single cell,discover different cell subsets in tissues according to the heterogeneity of cells,study the pathogenesis of intervertebral disc degeneration at the molecular level,and provide a new theoretical basis for its early diagnosis and treatment. OBJECTIVE:To introduce the basic principles of single-cell RNA sequencing technology and review the research progress of single-cell RNA sequencing technology in intervertebral disc degeneration in recent years. METHODS:A computer was used to search PubMed,Web of Science,CNKI and WanFang databases for the literature published from 2012 to 2022.Key words were"single-cell RNA sequencing,intervertebral disc degeneration,sequencing Technology"in Chinese and English.Duplicate,poor-quality and irrelevant articles were excluded;a total of 70 articles were eventually included. RESULTS AND CONCLUSION:(1)We identified new cell subsets such as homeostatic chondrocytes,hypertrophy chondrocyte-like nucleus pulposus cells and fibrous nucleus pulposus cells,identified the marker genes and transcription factors of these cell subsets,and described the functions,differentiation paths and cell fate of these cell subsets during the development and progression of intervertebral disc degeneration,and proposed the concept of progenitor nucleus pulposus cells.A cell subpopulation with progenitor nucleus pulposus cells properties was identified and its effectiveness in treating intervertebral disc degeneration was verified in mice.(2)Fibro chondrocyte-like annulus fibrosus cells and annulus fibrosus stem cells with both cartilage and fiber properties were identified,and a new type of composite hydrogel was prepared by combining fibrous cartilage inducers silk fibroin and hyaluronic acid in vitro.Experiments in mice demonstrated that this hydrogel could repair both annulus fibrosus tissue and cartilage matrix,and was remarkably effective in the treatment of intervertebral disc degeneration.(3)Regulatory chondrocytes were found in endplate cartilage.Two distinct fates in the progression of intervertebral disc degeneration were analyzed and the differential genes in the two fates were identified.Intercellular communication analysis indicated that regulatory chondrocytes interact with endothelial cells to promote angiogenesis.(4)Immune cells such as macrophages,T cells,myeloid progenitor cells and neutrophils were identified in the degenerated intervertebral disc tissues,demonstrating the existence of immune response during intervertebral disc degeneration.It was found that apolipoprotein induced the polarization of macrophages M1 and M2 subtypes,and this polarization process affected the activity of progenitor nucleus pulposus cells by amplifying the inflammatory response through the MIF signaling pathway.

Objective This research aimed to investigate the expression and mechanism of miR-155 and suppressor of cytokine signaling 1(SOCS1)in rheumatoid arthritis(RA).Methods RT-PCR and flow cytometry were applied to detect the expression differences of miR-155,Th17,and Treg cells in peripheral blood of RA patients(RA group)and control group(HC group).Bioinformatics analysis and dual-luciferase reporter assay were conducted to investigate the regulatory relationship between miR-155 and SOCS1.CD4+T cells were isolated from peripheral blood of RA patients and transfected with miR-155 inhibitor,si-SOCS1,and their respective negative control sequences,and divided into four groups:miR-NC group,miR-155 inhibitor group,miR-155 inhibitor+si-NC group,and miR-155 inhibitor+si-SOCS1 group.The cells were treated with Th17-inducing differentiation medium,and flow cytometry was used to determine the ratio of Th17 cells in each group of CD4+T cells.Western blot was used to determine the ratio of p-STAT3/STAT3 in the cells.Results Compared to the HC group,RA patients showed increased expression of miR-155 and Th17 ratio(P<0.01),and decreased Treg cell ratio(P<0.01).MiR-155 could target and inhibit the expression of SOCS1.Compared to the miR-NC group,the miR-155 inhibitor group,miR-155 inhibitor+si-NC group,and miR-155 inhibitor+si-SOCS1 group showed decreased Th17 ratio and p-STAT3/STAT3 ratio(P<0.01).Compared to the miR-155 inhibitor group,the miR-155 inhibitor+si-SOCS1 group exhibited increased Th17 ratio and p-STAT3/STAT3 ratio in CD4+T cells(P<0.05).Conclusion Elevated expression of miR-155 in RA patients may mediate the differentiation of CD4+T cells into Th17 cells through the SOCS1/STAT3 pathway,contributing to the imbalance of Th17/Treg cells in peripheral blood of RA patients.

Objective To explore the value of energy spectrum CT based material imaging for quantitatively and qualitatively evaluating lumbar intervertebral disc degeneration.Methods Lumbar energy spectrum CT and MRI were collected from 82 patients with back and leg pain,and 74 keV CT,water(chondroitin sulfate[CS])and CS(water)material images were obtained.Intervertebral discs Pfirrmann grading(PG)was performed based on MRI.CT values,water(CS)and CS(water)concentration of nucleus pulposus(NP)and annulus fibrosus(AF)in lumbar intervertebral disc with different PG grades were compared,and the relations with PG grades were analyzed.Visual classification of lumbar intervertebral disc was performed based on water(CS)and CS(water)falsecolor map,while PG distribution of different visual classification were compared,and the relations with PG grades were observed.Results Totally 250 lumbar intervertebral discs were enrolled,including 22 of PG Ⅰ,49 of PG Ⅱ,83 of PG Ⅲ and 96 of PG Ⅳ.CT values,water(CS)and CS(water)concentration of anterior AF,NP and posterior AF of intervertebral disc with different PG grades were significantly different(all P＜0.001),and CT values were negatively correlated with PG grades(rs=-0.504,-0.399,-0.258,all P＜0.001),while water(CS)concentration was positively correlated(rs=0.476,0.753,0.324,all P＜0.001)but CS(water)concentration was negatively correlated with PG grades(rs=-0.486,-0.760,-0.329,all P＜0.001).Significant differences of PG grades were found among different water(CS)and CS(water)falsecolor images visual classification(all P＜0.001),while water(CS)and CS(water)falsecolor images visual classification were all positively correlated with PG grades(all rs≥0.700,all P＜0.001).Conclusion Energy spectrum CT based material imaging could be used for quantitatively and qualitatively evaluating lumbar intervertebral disc degeneration.

ObjectiveTo explore the effect of transcranial direct current stimulation (tDCS) combined with acupuncture on central and upper limb function in stroke patients at flaccid stage based on central-peripheral-central theory. MethodsFrom September, 2018 to December, 2021, 120 patients with upper limb dysfunction after stroke in Guangdong Work Injury Rehabilitation Hospital were selected and randomly divided into control group 1 (n = 40), control group 2 (n = 40) and experimental group (n = 40). All the groups received conventional rehabilitation treatment. In addition, the control group 1 received acupuncture treatment, the control group 2 received anodal tDCS, and the experimental group received combined treatment of both, for four weeks. They were assessed with Fugl-Meyer Assessment-Upper Extremities (FMA-UE) and modified Barthel Index (MBI) before and after treatment. Electroencephalograph (EEG) was used to detect brain symmetry index (BSI), and electromyography (EMG) was used to detect root mean square values (RMS) of triceps brachii, biceps brachii, extensor wrist and flexor wrist of the affected upper limbs. ResultsTwo cases in the control group 1, one in the control group 2 and one in the experimental group dropped off, respectively. After treatment, the scores of FMA-UE and MBI significantly increased in all the groups (t > 11.757, P < 0.001), and they were higer in the experimental group than in the control groups (P < 0.001); the BSI decreased in the control group 2 and the experimental group (t > 2.324, P < 0.05), and it was less in the experimental group than in the control group 2 (P < 0.05); the RMS of biceps increased in all the groups (t > 2.953, P < 0.01), and was higer in the experimental group than in the control groups (P < 0.05); the RMS of flexor wrist and triceps increased in the control group 1 and the experimental group (t > 2.230, P < 0.05), and were higher in the experimental group than in the control group 1 (P < 0.05); the RMS of wrist extensor muscle increased only in the experimental group (t = 3.350, P < 0.01). ConclusiontDCS combined with acupuncture based on central-peripheral-central theory could effectively improve the upper limb function of stroke patients at flaccid stage, with advantages in improving hemispheric asymmetry and enhancing the activation level of affected muscles.

OBJECTIVE: To investigate the protective effect and potential mechanism of tubastatin A (TubA), a specific inhibitor of histone deacetylase 6 (HDAC6), on renal and intestinal injuries after cardiopulmonary resuscitation (CPR) in swine. METHODS: Twenty-five healthy male white swine were divided into Sham group (n = 6), CPR model group (n = 10) and TubA intervention group (n = 9) using a random number table. The porcine model of CPR was reproduced by 9-minute cardiac arrest induced by electrical stimulation via right ventricle followed by 6-minute CPR. The animals in the Sham group only underwent the regular operation including endotracheal intubation, catheterization, and anesthetic monitoring. At 5 minutes after successful resuscitation, a dose of 4.5 mg/kg of TubA was infused via the femoral vein within 1 hour in the TubA intervention group. The same volume of normal saline was infused in the Sham and CPR model groups. Venous samples were collected before modeling and 1, 2, 4, 24 hours after resuscitation, and the levels of serum creatinine (SCr), blood urea nitrogen (BUN), intestinal fatty acid binding protein (I-FABP) and diamine oxidase (DAO) in serum were determined by enzyme-linked immunoadsordent assay (ELISA). At 24 hours after resuscitation, the upper pole of left kidney and terminal ileum were harvested to detect cell apoptosis by TdT-mediated dUTP-biotin nick end labeling (TUNEL), and the expression levels of receptor-interacting protein 3 (RIP3) and mixed lineage kinase domain-like protein (MLKL) were detected by Western blotting. RESULTS: After resuscitation, renal dysfunction and intestinal mucous injury were observed in the CPR model and TubA intervention groups when compared with the Sham group, which was indicated by significantly increased levels of SCr, BUN, I-FABP and DAO in serum. However, the serum levels of SCr and DAO starting 1 hour after resuscitation, the serum levels of BUN starting 2 hours after resuscitation, and the serum levels of I-FABP starting 4 hours after resuscitation were significantly decreased in the TubA intervention group when compared with the CPR model group [1-hour SCr (μmol/L): 87±6 vs. 122±7, 1-hour DAO (kU/L): 8.1±1.2 vs. 10.3±0.8, 2-hour BUN (mmol/L): 12.3±1.2 vs. 14.7±1.3, 4-hour I-FABP (ng/L): 661±39 vs. 751±38, all P < 0.05]. The detection of tissue samples indicated that cell apoptosis and necroptosis in the kidney and intestine at 24 hours after resuscitation were significantly greater in the CPR model and TubA intervention groups when compared with the Sham group, which were indicated by significantly increased apoptotic index and markedly elevated expression levels of RIP3 and MLKL. Nevertheless, compared with the CPR model group, renal and intestinal apoptotic indexes at 24 hours after resuscitation in the TubA intervention group were significantly decreased [renal apoptosis index: (21.4±4.6)% vs. (55.2±9.5)%, intestinal apoptosis index: (21.3±4.5)% vs. (50.9±7.0)%, both P < 0.05], and the expression levels of RIP3 and MLKL were significantly reduced [renal tissue: RIP3 protein (RIP3/GAPDH) was 1.11±0.07 vs. 1.39±0.17, MLKL protein (MLKL/GAPDH) was 1.20±0.14 vs. 1.51±0.26; intestinal tissue: RIP3 protein (RIP3/GAPDH) was 1.24±0.18 vs. 1.69±0.28, MLKL protein (MLKL/GAPDH) was 1.38±0.15 vs. 1.80±0.26, all P < 0.05]. CONCLUSIONS TubA has the protective effect on alleviating post-resuscitation renal dysfunction and intestinal mucous injury, and its mechanism may be related to inhibition of cell apoptosis and necroptosis.
Male ; Animals ; Swine ; Abdominal Injuries ; Apoptosis ; Cardiopulmonary Resuscitation ; Kidney Diseases

Gene editing tools with nucleases as the main component have now implemented programmable targeted mutagenesis or insertion or deletion of mammalian genomes.From zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), CRISPR/Cas system to safer and more accurate Cas9 fusion protein gene editing tools and other nuclease gene editing tools, this paper systematically describes the development and evolution of gene editing, with detailed introduction to the development and optimization of next-generation gene editing tools, and a prospect of the clinical application of and challenges for gene editing tools.

Background and Objectives: Human umbilical cord mesenchymal stem cells (HUC-MSCs) are promising candidates for cell-based therapy in regenerative medicine or other diseases due to their superior characteristics, including higher proliferation, faster self-renewal ability, lower immunogenicity, a noninvasive harvest procedure, easy expansion in vitro, and ethical access, compared with stem cells from other sources. Methods: and Results: In the present study, we knocked down the expression of SOX9 in HUC-MSCs by lentivirus interference and found that knockdown of SOX9 inhibited the proliferation and migration of HUC-MSCs and influenced the expression of cytokines (IL-6 and IL-8), growth factors (GM-CSF and VEGF) and stemness-related genes (OCT4 and SALL4). In addition, the repair effect of skin with burn injury in rats treated with HUC-MSCs transfected with sh-control was better than that rats treated with HUC-MSCs transfected with shSOX9 or PBS, and the accessory structures of the skin, including hair follicles and glands, were greater than those in the other groups. We found that knockdown of the expression of SOX9 obviously inhibited the expression of Ki67, CK14 and CK18. Conclusions In conclusion, this study will provide a guide for modifying HUC-MSCs by bioengineering technology in the future.

OBJECTIVE：To investiga te main risk factors for adverse drug reactions （ADR）of skin by intravenous injection of iodine contrast agent. METHODS ：From Jan. 2009 to Apr. 2020，the patients suffering from skin ADR after enhanced CT with iodine contrast agent were collected from our hospital. The basic information ，laboratory test results before using iodine contrast agent and ADR related information were collected through hospital information system （HIS）. The use of iodine contrast agent ，main manifestations of skin ADR and drug combination were analyzed statistically. Taking the sex ，age，body mass index （BMI），the dosage of iodine contrast agent ，length of stay ，laboratory examination ，tumor history ，basic disease ，allergy history ，drinking history as independent variables ，the incidence of skin ADR related to iodine contrast agent was analyzed by single factor analysis ，and the variables with statistically significant were selected for multivariate Logistic stepwise regression analysis. RESULTS ：There were 157 cases of skin ADR ，involving 79 males（50.3％）and 78 females（49.7％）. The age ranged from 19 to 86 years old ，being（52.68± 18.73）years old in average. BMI was 14.6-40.7 kg/m2，being（22.5±3.7） kg/m2. 67 cases（42.68％）were treated with iprodione ，34 cases（21.66％）with iodixanol ，31 cases（19.74％）with iohexol and 25 cases（15.92％）with iopamidol ；the dose of iodine contrast agent were 50-100 mL，being（73.06±13.29）mL in average. There was no significant difference among different dosage of 4 kinds of iodine contrast agents （P≤0.05）. Among 4 kinds of iodine contrast agents ，the incidence of skin ADR induced by iopromide was the highest（0.197％）. The skin ADR related to iodine contrast agent was mainly acute （89.2％），the severity was mild （75.2％），and urticaria（38.9％）was the most common. After symptomatic treatment ，135 cases were cured ，13 cases were improved and 9 cases were not improved. Among the patients with iodine contrast agent related skin ADR ，the incidence of ADR induced by combined use of anti infective drugs was the highest （33.1％）；however，the combined use of anti-tumor drugs was the main cause of severe skin ADR. The length of stay ｛11～20 d[OR＝1.21，95％CI（1.07，1.20），P＝0.042]、21～30 d[OR＝1.39，95％CI（1.12，1.52），P＝0.035]、31～40 d[OR＝1.15，95％CI（1.03，1.37），P＝0.008]、＞40 d[OR＝1.33，95％CI（1.28，1.53），P＝0.003]｝，respitatory and circulatory system tumor history[OR ＝1.51，95％CI（1.35，1.61），P＝0.037]，injection allergy history[OR ＝1.50，95％CI（1.37，1.59），P＝0.005] can significantly increase the incidence of iodine contrast agent related skin ADR. CONCLUSIONS ：The main manifestation of skin ADR related to iodine contrast agent was urticaria. The main risk factors of skin ADR related to iodine contrast agent were length of stay （＞ 10 d），respiratory and circulatory system tumor history and injection allergy history.

Objective:To discuss the clinical significance of structural chest zones for gynecomastia.Methods:A total of 687 gynecomastia patients, aged from 14 to 45 years old, with an average age of 27.0, were admitted to the Department of Plastic Surgery, Huashan Hospital Fudan University from January 2012 to December 2018. Simon classification was used to record. Conduct Simon classification according to the preoperative measurement of the chest. Surgical area design, photo analysis, breast ultrasound examination and Derriford appearance scale evaluation were conducted as well. Six zones can be divided as follows: zone 0 nipple-areola zone, zone 1 gynecomastia zone, zone 2 accessory-breast zone, zone 3 lateral-roll zone, zone 4 IMF(inframammary fold) zone, zone T relative-contraindication zone. Positive and lateral images of the chest were taken before and one week, one month, three months, six months, and 12 months after the operation, and the preoperative and postoperative changes of each structural zone were compared. At the follow-up of 12 months after the operation, patients’ satisfaction was counted.Results:687 patients, according to Simon classification, were divided toⅠ degreein 65 cases, Ⅱa degree in 257 cases, Ⅱb degreein 194 cases, Ⅲ degree in 171 cases. Before operation, except for Simon Ⅰ degree in 65 cases showed just raised to 0 zone, most of the rest of the patients contains 1 zone raised, 2-4 zones come in different patients. After liposuction and glandular stripping were performed in different zones, postoperative follow-up was conducted one week, one month, three months, six months, and 12 months after the operation. The dynamic observation was made of the change process of chest shape shaping during the follow-up, whether there were postoperative complications, poor local shape, or bad regional shape. Evaluation results of patient satisfaction 12 months after surgery: among the 687 patients, there were 634 patients with 10 points of satisfaction, and 53 patients with 8-9 points, including 48 patients of zone 0 and zone 1, three patients of zone 3, and 2 patients of zone 4.Conclusions:The structural chest zones of gynecomastia provides plastic surgeons and male patients with simple, intuitive, and highly recognized preoperative evaluation, which can provide appropriate, simplified, and accurate guidance for the selection of surgical methods.

Objective:To discuss the clinical significance of structural chest zones for gynecomastia.Methods:A total of 687 gynecomastia patients, aged from 14 to 45 years old, with an average age of 27.0, were admitted to the Department of Plastic Surgery, Huashan Hospital Fudan University from January 2012 to December 2018. Simon classification was used to record. Conduct Simon classification according to the preoperative measurement of the chest. Surgical area design, photo analysis, breast ultrasound examination and Derriford appearance scale evaluation were conducted as well. Six zones can be divided as follows: zone 0 nipple-areola zone, zone 1 gynecomastia zone, zone 2 accessory-breast zone, zone 3 lateral-roll zone, zone 4 IMF(inframammary fold) zone, zone T relative-contraindication zone. Positive and lateral images of the chest were taken before and one week, one month, three months, six months, and 12 months after the operation, and the preoperative and postoperative changes of each structural zone were compared. At the follow-up of 12 months after the operation, patients’ satisfaction was counted.Results:687 patients, according to Simon classification, were divided toⅠ degreein 65 cases, Ⅱa degree in 257 cases, Ⅱb degreein 194 cases, Ⅲ degree in 171 cases. Before operation, except for Simon Ⅰ degree in 65 cases showed just raised to 0 zone, most of the rest of the patients contains 1 zone raised, 2-4 zones come in different patients. After liposuction and glandular stripping were performed in different zones, postoperative follow-up was conducted one week, one month, three months, six months, and 12 months after the operation. The dynamic observation was made of the change process of chest shape shaping during the follow-up, whether there were postoperative complications, poor local shape, or bad regional shape. Evaluation results of patient satisfaction 12 months after surgery: among the 687 patients, there were 634 patients with 10 points of satisfaction, and 53 patients with 8-9 points, including 48 patients of zone 0 and zone 1, three patients of zone 3, and 2 patients of zone 4.Conclusions:The structural chest zones of gynecomastia provides plastic surgeons and male patients with simple, intuitive, and highly recognized preoperative evaluation, which can provide appropriate, simplified, and accurate guidance for the selection of surgical methods.