Objective: To investigate the role of platelet derived growth factor receptor alpha (PDGFRα) on bidirectional differentiation of glioma-associated oncogene homolog 1-positive mesenchymal stem cells (Gli1+-MSC). Methods: Breeding double reporter transgenic mice ROSAmT/mG/Gli1-Cre^ERt2/PDGFRα^fl (Experimental group) and ROSA^mT/mG/Gli1-Cre^ERt2 (Control group), 20 mice in each of the two groups at four weeks of age were selected, MSC were isolated from the mouse aortic epithelium. After tamoxifen inducement, the two groups of Gli1⁺-MSC were screened by green fluorescent protein (GFP) labeling and flow cytometry sorting. PDGFRα was conditionally knocked out in the experimental group, and the control group Gli1⁺-MSC expressed PDGFRα normally. The two groups of Gli1⁺-MSC were subjected to adipogenic induction and fibrogenic induction, the Western blotting was performed to detect PDGFRα, adipocyte markers [perilipin and CCAAT/enhancer binding protein alpha (C/EBPα)] and fibrogenic markers [alpha smooth muscle actin (α-SMA) and fibroblast-specific protein 1 (FSP-1)] and semi-quantitative analysis was performed. The degree of cellular adipose differentiation after bidirectional induction of Gli1+-MSC in both groups was observed by oil red O staining and analyzed semi-quantitatively. Results: After tamoxifen induction, Gli1⁺-MSC could be accurately isolated from flow cytometry by GFP labeling. Via adipogenic differentiation, the expression of PDGFRα in the experimental group (0.017±0.002) was significantly lower than that in the control group (0.184±0.012) (t=25.48，P=0.002). The protein expressions of perilipin (3.138±0.414) and C/EBPα (3.565±0.289) were significantly higher than those in the control group (2.312±0.218 and 2.179±0.103, respectively) (t=6.21，P=0.025；t=6.69，P=0.022). Thus, the knock-out of PDGFRα enhanced the adipogenic differentiation ability of Gli1⁺-MSC. After fibrogenesis induction, the protein expressions of PDGFRα, α-SMA and FSP-1 in the experimental group (0.030±0.001, 0.932±0.177 and 0.276±0.020, respectively) were significantly lower than those in the control group (0.439±0.006, 1.352±0.170 and 0.835±0.097, respectively) (t=149.40, P<0.001; t=66.38,P<0.001; t=11.41,P<0.08). This suggested that the knock-out of PDGFRα significantly inhibited Gli1⁺-MSC differentiation toward fibroblasts. After bidirectional induction, significantly less adipocyte formation was seen in the control group and more in the experimental group. Quantitative analysis showed that the amount of oil red O staining in the experimental group (0.461±0.042) was significantly higher than that in the control group (0.017±0.007) after bidirectional induction (t=23.20, P<0.01). Conclusions: PDGFRα plays an important role in the regulation of bidirectional differentiation of vascular adventitial Gli1+-MSC.

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 evaluate the efficacy and safety of oral anisodine hydrobromide tablets in the treatment of nonarteritic anterior ischemic optic neuropathy (NAION).Methods:A multicenter nonrandomized controlled trial was conducted.A total of 282 acute NAION patients (282 eyes) were recruited from 16 hospitals in China from July 2020 to May 2021.Patients were divided into two groups according to treatment methods, which were control group (124 cases, 124 eyes) receiving regular treatment including citicoline sodium plus Ginkgo biloba leaf liquid extract or Ginkgo biloba leaf extract tablets plus mecobalamin, and experimental group (158 cases, 158 eyes) receiving treatment in control group plus oral anisodine hydrobromide tablets 1 mg, twice daily for 2 to 3 months.Best corrected visual acuity (BCVA), visual field index (VFI), peripapillary retinal nerve fiber layer (pRNFL) and radial peripapillary capillary vessel density (RPC) were assessed at 1, 2, 3, and 6 months after enrollment using the standard decimal visual acuity chart, 750i Humphery visual field analyzer, Cirrus HD-OCT 4000/Cirrus HD-OCT 5000, RTVue-XR optical coherence tomography respectively.The primary outcomes were BCVA and VFI, and the secondary outcomes were pRNFL, RPC, and the side effects during the follow-up.The study adhered to the Declaration of Helsinki.All patients were fully informed about the treatment and purpose of this study and voluntarily signed the informed consent form.The study protocol was approved by Chinese PLA General Hospital (No.S2020-021-01). Results:In all, 242 patients (242 eyes) completed the follow-up of BCVA, and 98 patients (98 eyes) completed the VFI follow-up.In terms of visual function, BCVA and VFI improved significantly over time in the two groups, and BCVA and VFI were better in experimental group than in control group at various follow-up time points (all at P<0.05). In terms of structure, pRNFL gradually decreased in both groups with the extension of treatment, and pRNFL was significanthy thinner in experimental group than in control group at various follow-up time points (all at P<0.05). There was no significant difference in RPC between the two groups at the last follow-up ( P>0.05). There were two cases with side effects and one case was discontinued due to side effects 25 days after enrollment. Conclusions:Oral anisodine hydrobromide can improve visual acuity and visual field in NAION and accelerate the regression of optic disc edema, with good safety.

OBJECTIVE: To observe the inhibitory effect of Shenbai Jiedu Fang (SBJDF, a compound recipe of traditional Chinese herbal drugs) on chemically induced carcinogenesis of colorectal adenoma in mice and explore the role of PTEN/PI3K/AKT signaling pathway in mediating this effect. METHODS: Four-week-old male C57BL/6 mice were randomly divided into control group (n=10), AOM/DSS model group (n=20), low-dose (14 g/kg) SBJDF group (n=10) and high-dose (42 g/kg) SBJDF group (n= 10). In the latter 3 groups, the mice were treated with azoxymethane (AOM) and dextran sodium sulphate (DSS) to induce carcinogenesis of colorectal adenoma. In the two SBJDF treatment groups, SBJDF was administered daily by gavage during the modeling. The survival rate, body weight, general condition of the mice, and intestinal adenoma formation and carcinogenesis were observed. The expressions of proteins associated with the PTEN/PI3K/AKT signaling pathway in the intestinal tissue were detected using immunohistochemistry. RESULTS: Compared with those in the model group, the mice treated with SBJDF, especially at the high dose, showed a significantly lower incidence of intestinal carcinogenesis and had fewer intestinal tumors with smaller tumor volume. Pathological examination showed the occurrence of adenocarcinoma in the model group, while only low-grade and high-grade neoplasia were found in low-dose SBJDF group; the mice treated with high-dose SBJDF showed mainly normal mucosal tissues in the intestines with only a few lesions of low-grade neoplasia of adenoma. Compared with those in the control group, the mice in the model group had significantly elevated plasma miRNA-222 level (P < 0.05), which was obviously lowered in the two SBJDF groups (P < 0.01). The results of immunohistochemistry revealed that compared with the model group, the two SBJDF groups, especially the high-dose group, had significantly up-regulated expressions of PTEN, P-PTEN and GSK-3β and down-regulated expressions of p-GSK-3 β, PI3K, AKT, P-AKT, β-catenin, c-myc, cyclinD1 and survivin in the intestinal tissues. CONCLUSION SBJDF can significantly inhibit colorectal adenoma formation and carcino-genesis in mice possibly through regulating miRNA-222 and affecting PTEN/PI3K/AKT signaling pathway.
Animals ; Male ; Mice ; Adenoma/prevention & control* ; Azoxymethane/adverse effects* ; Carcinogenesis/drug effects* ; Colorectal Neoplasms/prevention & control* ; Dextran Sulfate/adverse effects* ; Disease Models, Animal ; Glycogen Synthase Kinase 3 beta/metabolism* ; Mice, Inbred C57BL ; MicroRNAs/metabolism* ; Phosphatidylinositol 3-Kinases/metabolism* ; Proto-Oncogene Proteins c-akt/metabolism* ; Signal Transduction ; Drugs, Chinese Herbal/therapeutic use*

ObjectiveTo explore the differences in the protective effects of five formulas for promoting blood circulation and removing blood stasis on the aortic endothelial cells of New Zealand rabbits with heart blood stasis syndrome. MethodEighty New Zealand rabbits were randomly divided into a normal group （n=10） and an experimental group （n=70）. The heart blood stasis syndrome model was induced by starvation combined with a high-fat diet and adrenaline in the rabbits of the experimental group. Subsequently， the model rabbits were randomly divided into a model group， a Xuefu Zhuyutang group （3.55 g·kg^-1·d^-1）， a Taohong Siwutang group （2.66 g·kg^-1·d^-1）， a Danshenyin group （1.962 g·kg^-1·d^-1）， a Huoluo Xiaolingdan group （2.80 g·kg^-1·d^-1）， a Shixiaosan group （0.56 g·kg^-1·d^-1）， and a c-Jun N-terminal kinase （JNK） inhibitor （SP600125， 5 μg·kg^-1）group. The normal group and the model group received the same amount of distilled water. The rabbits in five Chinese medicine groups were treated correspondingly by gavage， and those in the SP600125 group were injected with 0.5 mL of SP600125-dimethyl sulfoxide diluent. After the treatment， the aorta was collected， and terminal deoxynucleotidyl transferase dUTP nick end labeling （TUNEL） assay was used to detect the apoptosis of aortic endothelial cells. The enzyme-linked immunosorbent assay （ELISA） was used to detect serum levels of tumor necrosis factor-α （TNF-α） and interleukin-6 （IL-6）. Western blot was used to detect the protein expression of JNK， phosphorylated JNK （p-JNK）， B-cell lymphoma-2 （Bcl-2）， Bcl-2-associated X protein （Bax）， cysteinyl aspartate-specific protease-9 （Caspase-9）， and cysteinyl aspartate-specific protease-3 （Caspase-3） in aortic tissues. Real-time fluorescence-based quantitative polymerase chain reaction （Real-time PCR） was used to detect the mRNA levels of JNK， Bcl-2， Bax， Caspase-9， and Caspase-3 in aortic tissues. ResultFive formulas could improve the apoptosis of aortic endothelial cells to varying degrees. To be specific， Xuefu Zhuyutang and Taohong Siwutang were optimal in efficacy， followed by Huoluo Xiaolingdan， Shixiaosan， and Danshenyin， and SP600125 was the worst （P<0.05， P<0.01）. Five formulas could reduce the content of TNF-α and IL-6 （P<0.05， P<0.01）， down-regulate the protein expression levels of JNK， p-JNK， Bax， Caspase-9， and Caspase-3 （P<0.05， P<0.01）， decrease the mRNA expression levels of JNK， Bax， Caspase-9， and Caspase-3 （P<0.05， P<0.01）， and up-regulate the protein and mRNA expression levels of Bcl-2 （P<0.05， P<0.01）. ConclusionFive formulas can all reduce the apoptosis of aortic endothelial cells in New Zealand rabbits with heart blood stasis syndrome with different efficacies. It may be related to the different effects of five formulas on the JNK signaling pathway.

OBJECTIVE: To explore the characteristics and rules of acupoint sensitization phenomena based on knee osteoarthritis (KOA), one of the clinical dominant diseases of acupuncture-moxibustion. METHODS: In combination with literature and expert experiences, the acupoints with the highest use frequency in treatment of KOA were screened, e.g. Heding (EX-LE 2), Liangqiu (ST 34), Mingmen (GV 4), Neixiyan (EX-LE 4), Ququan (LR 8) and Dubi (ST 35). In 814 patients with KOA and 217 healthy subjects, the acupoint temperature, mechanic pain threshold and pressure pain threshold were detected separately. Using machine learning method, the sensitization was judged at each acupoint. RESULTS: Compared with healthy subjects, the acupoint temperature was increased and the mechanic pain threshold and pressure pain threshold were reduced in KOA patients (P<0.05). Besides, the cut-off value was presented to distinguish whether the acupoint was sensitized or not. The results of machine learning showed that the highest prediction accuracy of acupoint sensitization was 86.7% (Shenshu [BL 23]) and the lowest one was 73.9% (Heding [EX LE 2]). The prediction accuracy at the third clinical stage trial was higher, the highest was 93.3% (Ququan [LR 8]) in KOA patients. CONCLUSION It is confirmed that the acupoint sensitization reflects the characteristics of disease and is correlative with the conditions of illness, which may provide the reference for the auxiliary diagnosis and condition assessment of KOA.
Acupuncture Points ; Acupuncture Therapy ; Humans ; Moxibustion ; Osteoarthritis, Knee/therapy* ; Treatment Outcome

OBJECTIVE: To determine the effects of Chinese medicine (CM) involving triple rehabilitation therapy on the progression of knee osteoarthritis (KOA). METHODS: A total of 722 patients recruited from 38 community health service centers located in China from March 2013 to March 2017 were randomly divided into treatment and control groups equally, using a cluster randomization design. Health education combined with CM involving triple rehabilitation therapy for KOA (electro-acupuncture, Chinese medicinal herb fumigating-washing, and traditional exercises) was administered in the treatment group while conventional rehabilitation therapy (physical factor therapy, joint movement training, and muscle strength training) was administered in the control group. Patients with a visual analog scale (VAS) scores ≽4 were treated with dispersible meloxicam tablets (7.5 mg, once daily). The Lequesne index scores, VAS scores, range of motion (ROM), lower limb muscle strength, knee joint circumference, quantitative scores of KOA symptoms, and the short-form 36 item health survey questionnaire (SF-36) scores were measured for each patient at 5 checkpoints (before treatment, at the 2nd week and the 4th week during the 4-week treatment period, at 1 month and 3 months after end of treatment), and adverse reactions were observed also. RESULTS: A total of 696 patients completed the entire process, with 351 in the treatment group and 345 in the control group. At all treatment checkpoints, the treatment group demonstrated better outcomes than the control group with regard to the total Lequesne index scores, effective rate and improvement rate of the total Lequesne index scores, VAS scores, lower limb muscle strength, knee circumference, quantitative scores of KOA symptoms, and SF-36 scores as well (P<0.05 or P<0.01). No adverse reactions were encountered in this study. CONCLUSIONS CM involving triple rehabilitation therapy can alleviate KOA-related pain and swelling, improve lower limb muscle strength, promote flexion and activity of the knee and improve the quality of life in patients undergoing KOA. It is suitable for patients with early or mid-stage KOA. (Registration No. ChiCTR-TRC-12002538).
Humans ; Medicine, Chinese Traditional ; Osteoarthritis, Knee/therapy* ; Outpatients ; Quality of Life ; Treatment Outcome

Aim To clarify the mechanism of Gonglaoye and Xianhecao herbal pair in the treatment of ischemic stroke so as to obtain the substantive evidence using network pharmacology data mining and molecular docking. Methods The main compounds of traditional Chinese medicine were obtained by TCMSP platform and consulting literature, the drug action targets were obtained by TCMSP, and the known genes about ischemic stroke were collected by searching Drugbank, Disgenet, TTD, Genecards, OMIM database, thus the drug-compound-target network map was constructed, and the common target proteins and main compounds were screened. The visual protein-protein interaction network map (PPI) was constructed by string. With the help of Cytoscape software, the original target network of active components was constructed and analyzed, and the gene ontology GO and Jingdu gene and genome encyclopedia KEGG analysis were carried out to analyze the GO function and KEGG pathway enrichment of the common targets of drugs and diseases. Finally, the molecular docking of the core protein and the core compound was carried out according to the relevant node parameters of the compound and protein. Results Seventeen active components and 296 potential targets of Gonglao leaf and crane herbs in the treatment of ischemic stroke were screened. GO enrichment was mainly concentrated in the response to oxides, cell response to chemical stimulation, positive regulation of cell metabolism, constant effect, active regulation of stimulus response, cell communication and so on. KEGG was mainly involved in signaling pathways such as PI3K-Akt, Ras, neuron ligand receptor interaction and so on. Molecular docking showed that quercetin and other active components had high affinity and tight connection with core targets such as AKT1. Conclusions The treatment of ischemic strokec is mainly through the mechanism of ursolic acid, hyperin and other active components, AKT1, cMAPK3 and other multi-targets, PI3K-AKT and other multi-pathway interaction mechanisms. Through this study the theoretical support can be provided for the further clinical application of Gonglaye and crane herbs, providing basic ideas for future experimental research and new drug research and development.

OBJECTIVE: To assess the trends in characteristics, treatments, and outcomes of acute myocardial infarction (AMI) patients in tertiary Chinese medicine (CM) hospitals in China between 2006 and 2013. METHODS: This retrospective study was based on two nationwide epidemiological surveys of AMI in tertiary CM hospitals during 2 years (2006 and 2013). Patients admitted to the hospital for AMI were enrolled. Hospital records were used as the data source. Case data were derived regarding baseline characteristics, treatments, and outcomes of patients to assess changes from 2006 to 2013. Logistic regression was used to analyze the relationship between prognosis, general influencing factors of disease, and various treatment measures. RESULTS: Totally 26 tertiary CM hospitals in 2006 and 29 tertiary CM hospitals in 2013 (18 were repetitive) were surveyed. A total of 2,311 patients with AMI were enrolled (1,094 cases in 2006 and 1,217 cases in 2013). From 2006 to 2013, the mean age did not significantly change, but the proportion of patients younger than 65 years increased. The prevalence of risk factors such as hypertension, diabetes, and hyperlipidemia also increased. Significant increases were observed in primary percutaneous coronary intervention [20.48% (2006) vs. 24.90% (2013)] and revascularization [36.11% (2006) vs. 52.42% (2013)]. In-hospital mortality decreased from 11.15% in 2006 to 10.60% in 2013. A mortality logistic regression analysis identified reperfusion therapy [odds ratio (OR), 0.222; 95% confidence interval (CI), 0.106-0.464], Chinese patent medicines (OR, 0.394; 95% CI, 0.213-0.727), and CM decoctions (OR, 0.196; 95% CI, 0.109-0.353) as protective factors. CONCLUSION Reperfusion and revascularization capabilities of tertiary CM hospitals have improved significantly, but in-hospital mortality has not significantly decreased. Efforts are needed to improve medical awareness of AMI and expand the use of CM to reduce in-hospital mortality in China.

OBJECTIVE: To observe the distribution characteristics and rules of pain sensitivity points on body surface in patients with knee osteoarthritis (KOA). METHODS: A total of 916 patients with KOA were selected in this study, the pain sensitivity points of local site of knee joint were probed by thumb palpation. Tape was used to measure the distance between the pain sensitivity points and the most nearby acupoints. The Wagner tenderness measuring instrument was used to measure the tenderness threshold of pain sensitivity points. RESULTS: A total of 3618 pain sensitivity points were probed, among them, 3338 pain sensitivity points were sensitized. The minimum sensitization degree was 1.00, the maximum sensitization degree was 3.39, while the average sensitization degree was (2.16±0.60). Pain sensitivity points were distributed 0.37-1.73 CONCLUSION The pain sensitivity points of patients with KOA may be the expansion effect of acupoint areas in the disease states, pain sensitivity points are more likely to appear on the medial side of knee joint.
Acupuncture Points ; Humans ; Knee Joint ; Osteoarthritis, Knee/therapy* ; Pain Threshold