ObjectiveTo explore the effect of serum containing Zhenwutang on myocardial mast cell apoptosis induced by angiotensin Ⅱ （AngⅡ） and the mechanism of the correlation between apoptosis and mitochondrial autophagy. MethodsIn this experiment， AngⅡ and serum containing Zhenwutang with different concentrations were used to interfere with H9C2 cardiomyocytes for 24 h， and the survival rate of H9C2 cardiomyocytes was detected by cell counting kit-8 （CCK-8） to screen the optimal concentration for the experiment. Enzyme-linked immunosorbent assay （ELISA） was used to detect the content of B-type natriuretic peptide （BNP） in cell culture supernatant， and immunofluorescence was used to detect the cell surface area to verify the construction of the myocardial mast cell model. Subsequently， the experiment was divided into a blank group （20% blank serum）， a model group （20% blank serum + 5×10^-5 mol·L^-1 AngⅡ）， low-， medium-， and high-dose （5%， 10% and 20%） serum containing Zhenwutang groups， an autophagy inhibitor group （1×10^-4 mol·L^-1 3-MA）， and autophagy inducer group （1×10^-7 mol·L^-1 rapamycin）. The apoptosis level of H9C2 cells and the changes of mitochondrial membrane potential were detected by flow cytometry. The lysosomal probe （Lyso Tracker） and mitochondrial probe （Mito Tracker） co-localization was employed to detect autophagy. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） was used to detect Caspase-3， Caspase-9， B-cell lymphoma 2 （Bcl-2）， Bcl-2-related X protein （Bax）， and cytochrome C （Cyt C） in apoptosis-related pathways and the relative mRNA expression of ubiquitin ligase （Parkin）， phosphatase and tensin homolog （PTEN）-induced kinase 1 （PINK1）， and p62 protein in mitochondrial autophagy-related pathways. Western blot was used to detect cleaved Caspase-3， cleaved Caspase-9， Bax， Bcl-2， and Cyt C in apoptosis-related pathways， phosphorylated ubiquitin ligase （p-Parkin）， phosphorylated PTEN-induced kinase 1 （p-PINK1）， p62， and Bcl-2 homology domain protein Beclin1 in mitochondrial autophagy-related pathways， and the change of microtubule-associated protein 1 light chain 3 （LC3） Ⅱ/Ⅰ ratio. ResultsCCK-8 showed that when the concentration of AngⅡ was 5×10^-5 mol·L^-1， the cell activity was the lowest， and there was no cytotoxicity. At this concentration， the surface area of cardiomyocytes was significantly increased （P<0.01）， and the content of BNP in the supernatant of culture medium was significantly increased （P<0.05）. Therefore， AngⅡ with a concentration of 5×10^-5 mol·L^-1 was selected for the subsequent modeling of myocardial mast cells. Compared with the blank group， the model group and the autophagy inhibitor 3-MA group had a significantly increased apoptosis rate （P<0.01） and significantly decreased mitochondrial membrane potential （P<0.01）. The results of immunofluorescence co-localization showed that compared with the blank group， the model group had a significantly decreased number of red and green fluorescence spots. The results of Real-time PCR showed that compared with that in the blank group， the relative mRNA expression of Bax， Caspase-3， Caspase-9， Cyt C， and p62 in the model group was significantly up-regulated （P<0.01）， while the relative mRNA expression of Bcl-2， Parkin， and PINK1 was significantly down-regulated （P<0.01）. In addition， the relative protein expression of Bax， cleaved Caspase-3， cleaved Caspase-9， Cyt C， and p62 was significantly up-regulated （P<0.01）. The LC3Ⅱ/Ⅰ was significantly decreased， and the relative protein expression of Bcl-2， p-Parkin， p-PINK1， and Beclin1 was significantly down-regulated （P<0.01）. Compared with the model group， the serum containing Zhenwutang groups and the autophagy inducer group had significantly decreased apoptosis rate （P<0.01）， and the decrease ratio of mitochondrial membrane potential is significantly lowered （P<0.01） in a dose-dependent manner. Additionally， both red and green fluorescence spots became more in these groups. In the 3-MA group， the number of red and green fluorescence spots decreased significantly. The relative mRNA expression of Bax， Caspase-3， Caspase-9， Cyt C， and p62 was significantly down-regulated （P<0.05， P<0.01）， while that of Bcl-2， Parkin， and PINK1 was significantly up-regulated （P<0.01）. In the serum containing Zhenwutang groups， the relative protein expression levels of Bax， cleaved Caspase-3， cleaved Caspase-9， Cyt C， and p62 were significantly down-regulated （P<0.05，P<0.01）. The LC3Ⅱ/Ⅰ was significantly increased， and the relative protein expression levels of Bcl-2， p-Parkin， p-PINK1， and Beclin1 were significantly up-regulated （P<0.01）. ConclusionThe serum containing Zhenwutang can reduce the apoptosis of myocardial mast cells and increase mitochondrial autophagy. This is related to the inhibition of intracellular Bax/Bcl-2/Caspase-3 apoptosis pathway and regulation of Parkin/PINK1 mitochondrial autophagy pathway.

Objective To investigate the risk factors for postoperative numbness syndrome secondary to lumbar disc herniation (LDH) based on propensity score matching (PSM). Methods A retrospective analysis was conducted on the clinical data of 429 LDH patients who underwent surgical treatment. The patients were divided into numbness syndrome group (85 patients) and non-numbness syndrome group (344 patients) according to whether they developed postoperative numbness syndrome. A 1∶1 match was performed between the numbness syndrome group and the non-numbness syndrome group based on PSM, with 67 patients in each group after matching. The clinical data of the two groups before and after matching were compared, and Cox regression analysis was performed on the matched data to screen risk factors for postoperative numbness syndrome secondary to LDH. Results The incidence of postoperative numbness syndrome in LDH patients was 19.81%(85/429). Before matching, the numbness syndrome group had a higher or longer proportion of males, age, body mass index (BMI), duration of disease, proportion of patients with L₃ to L₄ or L₄ to L₅ lesion segments, proportion of patients with grade Ⅳ to Ⅴ disc degeneration, proportion of patients with prolapsed or free-fragment herniation, operation time, proportion of patients with incomplete nucleus pulposus removal during surgery, proportion of patients with annulus fibrosus rupture, proportion of patients with a history of hyperlipidemia, proportion of patients who did not undergo postoperative rehabilitation exercises, and postoperative activity intensity scores compared to the non-numbness syndrome group (P < 0.05). After matching, the numbness syndrome group had higher or longer age, BMI, duration of disease, proportion of patients with grade Ⅳ to Ⅴ disc degeneration, proportion of patients with prolapsed or free-fragment herniation, proportion of patients with incomplete nucleus pulposus removal during surgery, proportion of patients with annulus fibrosus rupture, proportion of patients with a history of diabetes, proportion of patients who did not undergo postoperative rehabilitation exercises, and postoperative activity intensity scores compared to the non-numbness syndrome group (P < 0.05). The results of multivariate Cox regression analysis showed that high BMI, grade Ⅳ to Ⅴ disc degeneration, incomplete nucleus pulposus removal during surgery, annulus fibrosus rupture, history of diabetes, lack of postoperative rehabilitation exercises, and high postoperative activity intensity were independent risk factors for postoperative numbness syndrome secondary to LDH (HR=4.395, 7.183, 4.404, 5.633, 3.521, 4.929, 3.710, P < 0.05). Conclusion After controlling for confounding factors based on PSM, the risk factors for postoperative numbness syndrome secondary to LDH include high BMI, grade Ⅳ to Ⅴ disc degeneration, incomplete nucleus pulposus removal during surgery, annulus fibrosus rupture, a history of diabetes, lack of postoperative rehabilitation exercises, and high postoperative activity intensity, which is helpful for clinical healthcare professionals to early identify high-risk patients with postoperative numbness syndrome secondary to LDH and provide a reference for subsequent patient treatment.

ObjectiveTo investigate whether Jiedu Huoxue prescription can induce macrophage autophagy and inhibit inflammatory response to stabilize vulnerable plaques of atherosclerosis （AS） by regulating phosphatidylinositol 3-kinase （PI3K）/protein kinase B （Akt）/mammalian target of rapamycin （mTOR） signaling pathway. MethodThirty ApoE^-/- mice fed with high-fat diet were randomly assigned into model， low-， medium-， and high-dose （5.35， 10.7， and 21.4 g·kg^-1·d^-1， respectively） Jiedu Huoxue prescription （Chinese medicine）， and rapamycin （2 mg·kg^-1·d^-1） groups. Six ApoE^-/- mice fed with common diet were used as the control group， and 6 C57BL/6J mice fed with common diet as the blank group. The drugs or equal volume of normal saline were administrated by gavage after 7 weeks of modeling， and the treatment lasted for 4 weeks. The serum levels of lipids and inflammatory cytokines ［monocyte chemoattractant protein-1 （MCP-1） and interleukin-6 （IL-6）］ were measured. Hematoxylin-eosin （HE） staining was employed to observe the pathological changes of the vascular wall of the aortic root. Immunohistochemistry was employed to detect the expression of macrophages/monocytes monoclonal antibody （MOMA-2） and α-smooth muscle actin （α-SMA）. Transmission electron microscopy was employed to count the autophagosomes in the aorta， and Western blot to determine the protein levels of Beclin-1， LC3， PI3K， Akt， and mTOR. ResultCompared with the control group， the model group showed elevated serum levels of lipids， MCP-1， and IL-6 （P<0.05）， inhibited expression of MOMA-2 and α-SMA （P<0.05， P<0.01）， up-regulated protein level of Beclin-1 （P<0.05）， and down-regulated protein levels of PI3K， Akt， and mTOR （P<0.05， P<0.01）. The model group presented obvious atherosclerotic plaques on the inner wall of the aorta， infiltration of inflammatory cells in the plaque， thickened and disarranged vascular intima where the plaque was attached， decreased autophagosomes and mitochondria， and destroyed mitochondrial structure. Chinese medicine and rapamycin groups showed lower levels of total cholesterol， triglycerides， low-density lipoprotein cholesterol， MCP-1， and IL-6 （P<0.05）， higher level of high-density lipoprotein cholesterol （P<0.05）， inhibited expression of MOMA-2 and α-SMA （P<0.05， P<0.01）， higher protein levels of Beclin-1 and LC3Ⅱ （P<0.05， P<0.01）， and lower protein levels of PI3K， Akt， and mTOR （P<0.05， P<0.01） than the model group. Moreover， Chinese medicine and rapamycin groups showed only a small number of atherosclerotic plaques on the inner wall of the aorta， reduced infiltration of inflammatory cells and thickness of the blood vessel wall， and increased autophagosomes and autophagic lysosomes. ConclusionJiedu Huoxue prescription can improve lipid metabolism， enhance macrophage autophagy， and reduce AS-induced inflammation to improve the stability of vulnerable plaques in AS mice by inhibiting the PI3K/Akt/mTOR signaling pathway.

OBJECTIVE: To detect the expression of plasma exosomal microRNA (miRNA) in systemic sclerosis (SSc), and to investigate its clinical significance. METHODS: A total of 20 patients who were initially diagnosed with SSc and did not receive medication in Department of Rheumatology and Immunology of Meizhou People' s Hospital from January 2020 to January 2022 were recruited, as well as 15 healthy individuals whose gender and age matched with those of the SSc patients. Plasma exosomes were isolated using ultracentrifugation method. The expression levels of exosomal miR-34-5p, miR-92-3p and miR-142-3p were detected by quantative real-time polymerase chain reaction (qRT-PCR). Correlations between the expression levels of exosomal miRNAs and clinical characteristic were analyzed by Spearman's rank correlation coefficient test. RESULTS: The mean age of 20 patients with SSc was (52.6±12.6) years, including 7 males and 13 females. Among the 20 SSc patients, 13 cases were diagnosed as limited cutaneous systemic sclerosis (lcSSc) and 7 cases were diagnosed as diffuse cutaneous systemic sclerosis (dcSSc) according to the extent of skin involvement. According to the findings of high resolution chest CT, 7 of 20 SSc patients were diagnosed with interstitial lung disease (ILD) and 13 SSc patients were diagnosed with non-ILD. The expression levels of exosomal miR-34-5p, miR-92-3p and miR-142-3p were significantly elevated in the SSc patients compared with those in the healthy controls group (P=0.003, P=0.000 1, and P=0.016, respectively). Compared with the SSc patients without ILD, the expression levels of miR-34-5p and miR-142-3p were significantly lower in the SSc patients with ILD (P=0.037 and P=0.015, respectively). The expression levels of exosomal miR-34-5p and miR-142-3p showed negative correlation with ILD (r=-0.48, P=0.031 and r=-0.55, P=0.011, respectively), and arthritis (r=-0.46, P=0.040 and r=-0.48, P=0.032, respectively). The expression levels of exosomal miR-142-3p showed a negative correlation with erythrocyte sedimentation rate (ESR) (r=-0.55, P=0.012). CONCLUSION Plasma exosomal miR-34-5p, miR-92-3p and miR-142-3p were dysregulated in SSc. The dyregulation of exosomal miR-34-5p and miR-142-3p showed correlation with SSc associated ILD (SSc-ILD).
Male ; Female ; Humans ; Young Adult ; Adult ; Clinical Relevance ; MicroRNAs/genetics* ; Scleroderma, Systemic/genetics* ; Lung Diseases, Interstitial

Objective:To explore the expression of miR-6883-5p in the serum of bladder cancer patients and its effect on the migration and proliferation ability of bladder cancer cells.Methods:The real-time fluorescent quantitative polymerase chain reaction (qRT-PCR) method was used to detect the expression level of miR-6883-5p in the serum of 39 patients with bladder cancer and 39 healthy subjects, as well as the expression level of miR-6883-5p in normal bladder mucosal epithelial cells and bladder cancer cells. The bladder cancer cells with the lowest expression of miR-6883-5p were transfected with miR-6883-5p mimics or negative control (NC), namely miR-6883-5p group and NC group. Transwell migration experiment and cell counting kit-8 (CCK-8) colorimetric method were used to detect the migration and proliferation ability of cells transfected with miR-6883-5p. Bioinformatics software and dual luciferase reporter gene were used to predict and test the target genes of miR-6883-5p. qRT-PCR and Western blot were used to detect the expression levels of target genes in cells transfected with miR-6883-5p.Results:The expression level of miR-6883-5p in the serum of bladder cancer patients was significantly lower than that of healthy persons ( P<0.01). The expression level of miR-6883-5p in bladder cancer cells was significantly lower than that in normal bladder mucosal epithelial cells (all P<0.05), and the lowest expression of miR-6883-5p was in 5637 cells ( P<0.01). After transfection with miR-6883-5p, the migration and proliferation of cells in miR-6883-5p group were significantly lower than those in NC group (all P<0.05). Phosphatidyl alcohol proteoglycan-6 (GPC6) was the target gene of miR-6883-5p. After transfection with miR-6883-5p, the expression levels of GPC6 mRNA and protein in miR-6883-5p group significantly decreased ( P<0.01). Conclusions:miR-6883-5p is significantly low expressed in the serum of bladder cancer patients, and miR-6883-5p is related to the occurrence and development of bladder cancer; overexpression of miR-6883-5p can inhibit the migration and proliferation of bladder cancer cells; miR-6883-5p can inhibit the occurrence and development of bladder cancer by down-regulating the expression of GPC6.

Objective: Surgical operation is the main treatment for advanced adenocarcinoma of esophagogastric junction (AEG). Due to its special anatomic location and unique lymph node reflux mode, the surgical treatment of Siewert II AEG is controversial. Lower mediastinal lymph node dissection is one of the most controversial points and a standard technique has not yet been established. This study is aim to explore the safety and feasibility of five-step maneuver of transthoracic single-port assisted laparoscopic lower mediastinal lymph node dissection for Siewert type II AEG. Methods: A descriptive case series study was conducted. The intraoperative and postoperative data of 25 patients with Siewert type II AEG who underwent five-step maneuver of transthoracic single-port assisted laparoscopic lower mediastinal lymph node dissection in Guangdong Provincial Hospital of Traditional Chinese Medicine from January 2019 to April 2021 were retrospectively analyzed. Five-step maneuver was as follows: In the first step, the subcardiac sac was exposed; the right pulmonary ligament lymph nodes and the anterior thoracic paraaortic lymph nodes were dissected cranial to inferior pericardium, left to left edge of thoracic aorta. In the second step, the left diaphragm was opened, and a 12 mm trocar was placed through the 6-7 rib in the left anterior axillary line. The supra-diaphragmatic nodes were dissected through the thoracic operation hole. In the third step, the left inferior pulmonary ligament was severed. The anterior fascia of thoracic aorta was incised to join the anterior space of thoracic aorta formed in the first step and then the lymphatic tissue was dissected upward until the exposure of left inferior pulmonary vein. In the fourth step, the posterior pericardium was denuded retrogradely from ventral side to oral side to the level of left inferior pulmonary vein, right to right pleura, and then the right pulmonary ligament lymph nodes were completely removed. In the fifth step, the esophagus was denuded, and the esophagus was transected 5 cm above the tumor using a linear stapler to complete the dissection of lower thoracic paraesophageal lymph nodes. Results: Operations were successfully completed in 25 patients without conversion, intra-operative complication and perioperative death. Total gastrectomy was performed in 19 cases and proximal gastrectomy in 6 cases. The mean operative time was (268.7±85.6) minutes, the mean estimated blood loss was (90.4±44.2) ml, the mean time of lower mediastinal lymph node dissection was (38.6±10.3) minutes, and the mean harvested number of lower mediastinal lymph node was 5.9±2.9. The length of esophageal invasion was >2 cm in 7 cases and ≤ 2 cm in 18 cases. Eight patients (33.0%) had lower mediastinal lymph node metastasis, including 3 cases with esophageal invasion >2 cm and 5 cases with esophageal invasion ≤ 2 cm. The mean time to postoperative first flatus was (5.5±3.1) days. The average time of postoperative thoracic drainage was (5.9±2.9) days. The mean hospital stay was (9.7±3.1) days. Two patients (8.0%) developed postoperative grade IIIa complications according to the Clavien-Dindo classification, including 1 case of pancreatic fistula and 1 case of pleural effusion, both of whom were cured by puncture drainage. Conclusions: Five-step maneuver of transthoracic single-port assisted laparoscopic lower mediastinal lymph nodes dissection for Siewert type II AEG is safe and feasible. Which can ensure sufficient lower mediastinal lymph node dissection to the level of left inferior pulmonary vein.
Adenocarcinoma/surgery* ; Esophagogastric Junction ; Humans ; Laparoscopy ; Lymph Node Excision ; Retrospective Studies

AIM: To explore the value of frequency threshold optical coherence tomography(OCT)in the diagnosis of primary open angle glaucoma optic nerve injury.

METHODS: Eighty patients with early primary glaucoma who were admitted to the hospital between January 2018 and March 2020 and 100 healthy subjects were selected as the study subjects. Patients with primary open angle glaucoma were divided into early group, middle group and late group. OCT was used to measure the thicknesses of upper, lower, nasal, bitemporal peri-papillary retinal nerve fiber layer(pRNFL)and upper and lower macular ganglion cell complex(mGCC)in each group. The mean deviation(MD)of visual field was determined through perimetry. The OCT parameters and perimetry parameters were compared among groups. Spearman correlation analysis was performed to analyze the correlation between OCT parameters and visual field defects, and the receiver operating characteristic(ROC)curve was used to calculate the value of OCT parameters in diagnosing primary open angle glaucoma.

RESULTS: There were 24 cases, 39 cases and 17 cases in the early group, middle group and late group, respectively. There were statistically significant differences in pRNFL and mGCC among 3 groups(P<0.05).The upper, lower, average pRNEL and the upper, lower, and average mGCC of patients in the early group of a third-class hospital were significantly lower than those of the control group, and the parameters of pRNFL and mGCC in each position of the mid-stage and late group were significantly lower than those of the control group. The upper, lower, nasal pRNFL, average pRNFL, upper, lower and average mGCC of the late group of glaucoma patients were significantly lower than those of the early group and the middle group. The indicators of the middle group were significantly lower than those of the early group(P<0.05). Spearman correlation analysis showed that pRNFL and mGCC parameters were negatively correlated with glaucoma severity(P<0.05)ROC curve analysis showed that the areas under the curves of the upper, lower, nasal, bitamporal and average pRNFL for diagnosis of primary open angle glaucoma optic nerve injury were 0.693, 0.846, 0.676, 0.579 and 0.844, respectively. The areas under the curves of upper, lower and average mGCC for diagnosis of primary open angle glaucoma optic nerve injury were 0.542, 0.677 and 0.676 respectively. The area under the curve of average pRNFL combined with average mGCC for the diagnosis of primary open angle glaucoma optic nerve injury was 0.883.

CONCLUSION:The pRNFL and mGCC measured by OCT are closely related to the degree of primary open angle glaucoma optic nerve injury. Both have high diagnostic value for glaucoma and can be used clinically for diagnosis and condition assessment.

Mitochondrial dysfunction is closely related to the occurrence and development of benign and malignant diseases of the pancreas. Mitochondrial membrane of the respiratory chain electron transfer and energy transfer plays an important role in maintaining normal cellular function. When the respiratory chain was disrupted, the oxidative stress was increased in the cell, and produced a large number of oxide intermediate products which target mitochondrial protein, DNA, etc, lead to mitochondrial dysfunction finally induced acute pancreatitis, pancreatic cancer and other diseases. In addition, mitochondrial homeostasis plays an indispensable role in maintaining the normal function of islet cells. This paper reviewed the research status of mitochondrial dysfunction in pancreatic diseases.

Non-alcoholic steatohepatitis (NASH) is an important link for the progression of metabolic-related fatty liver disease to end-stage liver disease such as cirrhosis and hepatocellular carcinoma, which seriously endangers human health. NASH pathogenesis is complex, and involves the interaction between hepatic parenchymal and non-parenchymal cells (NPCs), sinusoidal endothelial cells, Kupffer cells, hepatic stellate cells, and so on. Herein, the relevant research progress of NPCs in the pathogenesis of NASH is reviewed in order to further understand the role of NPCs in NASH.

OBJECTIVE: To study the expression of BIRC6 in renal cancer tissues and investigate the effect of BIRC6 silencing on apoptosis and autophagy of 786-O cells. METHODS: Twenty surgical specimens of renal cancer tissues and adjacent renal tissues were collected from Meizhou People's Hospital between February, 2016 and December, 2018 for detection of BIRC6 protein expression using immunohistochemistry. Renal cancer 786-O cells were transfected with a control small interfering RNA (siRNA) or BIRC6 siRNA RESULTS: The expression of BIRC6 protein was significantly higher in renal cancer tissues than in the adjacent renal tissues. Western blotting showed that siRNA-mediated silencing of BIRC6 significantly lowered the expression of BIRC6 in 786-O cells. In the cells with BIRC6 silencing, treatment with 12.5, 25, 50, 100 and 200 μg/mL 5-FU resulted in significantly higher proliferation inhibition rates than in the cells transfected with the control siRNA ( CONCLUSIONS Interference of BIRC6 mediated by siRNA can inhibit autophagy and promote 5-FU-induced apoptosis to enhance the sensitivity of 786-O cells to 5-FU.
Apoptosis ; Autophagy ; Cell Line, Tumor ; Cell Proliferation ; Humans ; Inhibitor of Apoptosis Proteins/genetics* ; Kidney Neoplasms/genetics* ; RNA, Small Interfering/genetics*