OBJECTIVE: To observe the effects of Xingnao Kaiqiao acupuncture technique (for regaining consciousness and opening orifice) on methylation of N6-methyladenosine (m6A), and key methyltransferases and demethylases, so as to clarify the mechanism of acupuncture on cerebral ischemia-reperfusion injury (CIRI). METHODS: Of 68 male Sprague-Dawley rats of SPF grade, 15 rats were randomly selected as a sham-operation group, and the remaining rats were subjected to the model of middle cerebral artery occlusion using the suture ligation. CIRI was induced by ischemia for 2 h followed by reperfusion. Rats that failed to modeling or died were excluded. The rest 45 rats were randomly divided into three groups, i.e. model group, acupuncture group, and non-point acupuncture group, with 15 rats in each group. The rats in the acupuncture group were treated with acupuncture at bilateral "Neiguan" (PC6) and "Shuigou" (GV26). In the non-point acupuncture group, acupuncture was delivered at three non-points, located 3 mm below the bilateral midaxillary line and 3 mm lateral to the tip of the coccyx. One intervention was operated in these two acupuncture groups and the needles were retained for 30 min. Before modeling start and 2 h after ischemia, a laser speckle flowmeter was used to monitor the cerebral blood perfusion. In 2 h of ischemia and 24 h of reperfusion, the neurological behavioral score was evaluated. The volume of rat cerebral infarction was determined by triphenyltetrazolium chloride (TTC) staining, and the level of m6A methylation in ischemic cortical area was detected by Dot blot, and the protein and mRNA expression of the demethylase i.e. fat mass and obesity-associated protein (FTO), AlkB homolog 5 (ALKBH5) and key methyltransferases, i.e. methyltransferase-like 3 (METTL3), methyltransferase-like 14 (METTL14), and Wilms' tumor 1-associated protein (WTAP) in ischemic cortical area were detected by Western blot and real-time PCR. RESULTS: Cerebral blood perfusion decreased by>70% after 2 h ischemia. Compared with the sham-operation group, the neurobehavioral score and the percentage of cerebral infarction volume increased in the model group (P<0.01); the level of m6A methylation in the ischemic cortical area increased (P<0.01), the protein and mRNA expression of FTO decreased (P<0.01), and that of ALKBH5, METTL3, and METTL14 increased (P<0.01, P<0.05) in the model group. When compared with the model group and the non-point acupuncture group, the acupuncture group showed a decrease in the neurobehavioral score and the percentage of cerebral infarction volume (P<0.01), the level of m6A methylation in the ischemic cortical area was reduced (P<0.01, P<0.05), and the protein and mRNA expression of FTO was elevated (P<0.01). CONCLUSION Xingnao Kaiqiao acupuncture technique presents its protective effect on the brain in the rats with CIRI, which is related to up-regulating the expression of FTO and modulating m6A methylation.
Animals ; Rats, Sprague-Dawley ; Male ; Acupuncture Therapy ; Reperfusion Injury/genetics* ; Rats ; Brain Ischemia/genetics* ; Humans ; Adenosine/metabolism* ; Methylation ; Acupuncture Points ; Cerebral Cortex/metabolism*

OBJECTIVE: To observe the clinical efficacy of acupoint thread-embedding therapy of regulating governor vessel, dispersing lung, and suppressing reflux for gastroesophageal reflux cough (GERC). METHODS: A total of 120 GERC patients were randomly assigned to an observation group (60 cases, 1 case dropped out) and a control group (60 cases, 1 case was eliminated). The observation group received acupoint thread-embedding treatment at positive response points of governor vessel. If no such points were detected, the following acupoints were used: Dazhui (GV14), Fenghu (Extra), Shendao (GV11), Lingtai (GV10), and Zhiyang (GV9). Treatment was administered once every two weeks. The control group received oral rabeprazole enteric capsules at 20 mg twice daily. All the treatment was given for 6 weeks. Clinical outcomes were assessed using cough symptom score, reflux disease questionnaire (RDQ) score, and Leicester cough questionnaire (LCQ) score before and after treatment in the two groups. Clinical efficacy was also compared between the two groups. RESULTS: After treatment, both groups showed decreased cough symptom scores and the each item scores and total scores of RDQ (P<0.001), and increased LCQ scores (P<0.001) compare with those before treatment. The observation group exhibited lower cough symptom score and chest pain, reflux and total score of RDQ, and higher LCQ score compared to those in the control group (P<0.05). The total effective rate in the observation group was 94.9% (56/59), which was higher than 84.7% (50/59) in the control group (P<0.05). CONCLUSION Acupoint thread-embedding therapy of regulating governor vessel, dispersing lung, and suppressing reflux could effectively alleviate cough and reflux symptoms in patients with GERC and improve their quality of life.
Humans ; Acupuncture Points ; Gastroesophageal Reflux/physiopathology* ; Male ; Female ; Cough/physiopathology* ; Middle Aged ; Aged ; Acupuncture Therapy ; Adult ; Treatment Outcome ; Lung/physiopathology* ; Meridians

ObjectiveBased on the experience of traditional quality evaluation， the quality of Atractylodis Macrocephalae Rhizoma（AMR） with different production methods such as direct seeding， transplanting after seedling raising， topping and non-topping， and difference in growth years was compared. MethodVernier caliper was used to measure the trait data of AMR in different production methods. Paraffin sections of AMR with different production methods were made by saffron solid green staining， and the microstructure was observed. The contents of water-soluble and alcohol-soluble extracts in AMR with different production methods were determined according to the 2020 edition of Chinese Pharmacopoeia. The content of water-soluble total polysaccharides in AMR with different production methods was detected by sulfuric acid-anthrone method. Fiber analyzer was used to detect the content of fiber components in AMR with different production methods. The contents of monosaccharides， oligosaccharides and some secondary metabolites in AMR with different production methods were detected by ultra performance liquid chromatography（UPLC）， and the differences of chemical components were compared by multivariate statistical analysis methods such as principal component analysis（PCA） and partial least squares-discriminant analysis（PLS-DA）. ResultIn terms of traits， the 3-year-old AMR with direct seeding and without topping was close to the high-quality AMR with "phoenix-head and crane-neck， strong sweetness and clear aroma" recorded in ancient materia medica， followed by the 3-year-old AMR with topping after transplanting， while the 2-year-old AMR with topping after transplanting with high market circulation rate was generally fat and strong with mild odor. In the microscopic aspect， the arrangement of xylem vessels and fiber bundles in the 3-year-old samples formed two obvious rings. Compared with the 2-year-old samples cultivated in Bozhou and Zhejiang， the 3-year-old samples without topping after transplanting had more wood fibers. In terms of chemical composition， the contents of 70% ethanol extract， fructose， glucose， sucrose， 1-kestose， atractylenolide Ⅰ， chlorogenic acid， neochlorogenic acid， cryptochlorogenic acid and other components in 3-year-old AMR with direct seeding and without topping were significantly higher than those in the other three samples（P<0.05）. The contents of cellulose， 70% ethanol extract， sucrose， atractylenolide Ⅰ， atractylone and other components in 3-year-old AMR with topping after transplanting were significantly higher than those in the 2-year-old AMR with high market circulation rate（P<0.05）， while the contents of water-soluble extract and water-soluble total polysaccharides in 2-year-old samples with topping after transplanting were significantly higher than those in the 3-year-old AMR with topping after transplanting， direct seeding and without topping（P<0.05）. ConclusionUnder the current mainstream production mode， too much manual intervention makes AMR heavily enriched in polysaccharides and increased the yield， but the accumulation of sweet substances， fragrant substances and fiber substances is insufficient， which affects its quality. The current quality standard of AMR has some shortcomings in guiding the high quality production of it， it is suggested to revise the quality standard of AMR， supplement the quantitative analysis of secondary metabolites， and strengthen the production of imitation wild AMR.

Objective:To explore the methods, complications and follow-up results of limb salvage for primary malignant tumors of the femoral shaft.Methods:From October 2006 to October 2019, 41 cases of primary malignant tumors of femoral shaft were analyzed retrospectively, 37 cases were followed up, including 22 males and 15 females, aged 8-46 years with an average age of 20.56±4.72 years old. All the lesions were located in the femoral shaft, and The Enneking stage was IIB. Tumor lesion ranged in the femur from 10 cm to 24 cm, and there was no pathological fracture. Pre-operative puncture biopsy was performed in all cases.22 cases were confirmed as osteosarcoma, 8 as chondrosarcoma, 5 as Ewing sarcoma, 2 as malignant fibrous histiocytoma. Neoadjuvant chemotherapy has been used to treat both osteosarcoma and Ewing sarcoma. Among the 37 patients, 8 cases of patients received total femoral prosthesis replacement, 14 cases of intramedullary nail fixation with allogenic bone graft, 8 cases of allograft-prosthetic composite (APC), and 7 cases of 3D printed prosthesis. The limb function was graded according to MSTS 93 system.Results:37 out of 41 patients were followed up, and the average follow-up time was 23.15±16.74 months (6-62 months). There were 9 patients with pulmonary metastases (26.47%), among which 6 patients passed away due to multiple metastases. 28 patients survived without tumor at last follow-up. The MSTS score was18-28 (average of 22.55±2.57). The average score of 3D printing prosthesis group was 27±1.74, allogeneic bone transplantation group was 22.85±2.59, total femoral prosthesis replacement group was 20.25±2.25, and the APC prosthesis group was 20.5±2.07. There was statistical difference between 3D printing group and other groups. The overall excellent and good rate of lower limb function was 79.41%, in which the 3D group was 100%, allogeneic bone group was 78.5%, APC prosthesis group was 87.5%, total femoral prosthesis replacement group was 62.5%. There was no statistical difference in the excellent and good rate of limb function among the groups. There weren't any cases of hip joint dislocation after total femoral prosthesis replacement. Delayed union of bone healing was seen in 3 cases of allogeneic bone transplantation and 2 cases APC patients. One patient suffered from the postoperative hemorrhage-related diseases-sciatic nerve compression, and the nerve recovered after emergency debridement. One patient suffered from poor wound healing. The overall complication rate was 24.32% (9/37).Conclusion:The patients with primary malignant tumors of the femoral shaft can effectively recover the lower extremity function by choosing the appropriate surgical scheme, in order to achieve the purpose of limb preservation and improve the quality of life of the patients. 3D printing prosthesis provides a new choice in the treatment of femoral shaft tumors.

Objective To adjust the layout of secondary medicine shelves in hospital pharmacy for outpatient services and optimize the varieties of automated equipment to improve working process and promote human-machine cooperation.Methods This article discussed the adjustment of the layout of secondary medicine shelves in hospital pharmacy for outpatient services and the optimization of varieties of automated equipment for storing by analyzing actual problems in automated system and clinical drug uses and the requirements of the automated equipment for drug storage respectively to improve working process and to adapt to the operation of outpatient pharmacy automated system.Results Dispensing efficiency of outpatient pharmacy automated system was improved, patients′ waiting time was significantly shortened, and the workload of pharmacists was decreased.Conclusion Optimization of hospital pharmacy drug delivery system and its operation can obviously improve work efficiency, shorten patients′ waiting time and reduce dispensing error, thus ensuring the safety of medication.

Objective To explore the clinical efficacy and complications of the surgical treatment of sternal tumor resection and titanium mesh reconstruction.Methods From 2008 January to 2012 June,there were 8 cases of sternal tumor patients in our hospital,including 5 male and 3 female,with an average age of 50.4 (37-66) years old.The histological morphology included 2 cases of chondrosarcoma,1 case of osteosarcoma,2 cases of malignant fibrous histiocytoma,eosinophilic granuloma in 1 case,and 2 cases of sternal metastasis of breast cancer.Tumor invasion sites included the sternal manubrium in 3 cases,the body in 2 cases,and both in 3 cases.All patients had undergone preoperative puncture or incision biopsy.8 cases of sternal tumor patients were treated with sternal tumor resection and reconstruction of the thorax using titanium mesh.The clinical effect and complications were retrospectively analyzed.Results All patients were followed up for 9 months to 4 years.The operations went well in all cases,with no intraoperative crisis or operative death.Deep wound hematoma occurred in 1 patient 1 week postoperatively,who healed 2 weeks after drainage and debridement.There was no abnormal breathing,subcutaneous emphysema,pneumothorax,infection or other complications in other cases.1 case of malignant fibrous histiocytoma died of lung metastasis at 9 months follow-up,and 1 died of liver metastasis at 14 months,while other patients got no tumor recurrence,with good thoracic shape,free breathing,no titanium mesh loosening,dyspnea,chest tightness,pain,or abnormal respiratory discomfort during follow-up period.The chest radiograph showed no chest deformity,no loosening or fracture of the fixation device.Conclusion Sternal tumor resection and reconstruction with titanium mesh has the advantages of good shaping effect,fewer complications,and simple operation,showing that titanium mesh is an ideal material for the reconstruction of sternum.

Objective To investigate the growth inhibition of human osteosareoma cell line(MG-63) intervened by Hydralazine and 5'-aza-2'-deoxycytidine (5-Aza-CdR),and the effect on the mRNA expression of gene WW domain-containing oxidoreductase (WWOX).Methods Certain volume of 5 × 104/ml of human osteosarcoma cell line MG-63 in logarithmic growth phase were added into 96-well plate.There were Hydralazine group (drug concentration,0.1,1.0,10 μmol/L),5-Aza-CdR group (drug concentration,5,10,20 μmol/L),Hydralazine combined with 5-Aza-CdR group (drug concentration,0.1 μmo/L + 5 μmol/L,1.0 μmol/L + 10 μmol/L,10 μmol/L + 20 μmol/L) and control group (culture medium).Methyl thiazol tetrazolium(MTT) colorimetric methods were used to test the growth inhibition of MG-63 cells intervened by different concentrations of Hydralazine and 5'-aza -2'-deoxycytidine (5-Aza-CdR).Flow cytometry AnnexinV-FITC/PI methods were used to assay the effects of Hydralazine and 5-Aza-CdR inducing apoptosis in osteosarcoma cells in vitro.Real-time polymerase chain reaction (Real-Time PCR)methods were used to detect amplification of WWOX mRNA induced by Hydralazine combined with 5-Aza-CdR or alone.Western-blotting methods were used to examine the expression of WWOX in MG-63 cells.Results Hydralazine and 5-Aza-CdR effectively inhibited the growth of MG-63 cells in a concentration and time-dependent manner.Combined effect was more obvious.Further more the expression levels of WWOX mRNA and protein were increased significantly in combined groups as compared with other groups.Conclusion Hydralazine and 5-Aza-CdR could effectively inhibit the proliferation of MG-63 cells and induce apoptosis which is concurrent with the promotion of the expression of WWOX.The mechanism may be that Hydralazine/5-Aza-CdR effectively cause the demethylated of WWOX gene CpG-rich promoter regions,leading to the high expression of WWOX and inhibit the growth of MG-63 cells.The use of hydralazine in the treatment of osteosarcoma is worthy of further investigation.

ObjectiveTo investigate the effect of arsenic trioxide (As2O3) on the metastasis capability of Ewing's sarcoma ceils. MethodsMTT assays were performed to choose appropriate concentrations of As2O3 (＜ 2 μmol/L) for the experiments.Migration and invasion assays were performed to assess the effect of As2O3 on the metastasis of Ewing's sarcoma cells. Changes in matrix metalloproteinase(MMP)-9 expressions were detected by gel zymography assay and the phosphoinositide 3-kinase/AKT(PI3K-AKT)pathway was investigated using Western blot. ResultsThe amount of Ewing's sarcoma cells across basal membrane of Transwell in migration and invasion assay decreased gradually with the increase in As2O3 concentration. The average quantities of A-673 across the membrane after treatment by gradual concentrations accounted for 54.3 ％,49.0 ％ and 17.0 ％ of that of untreated group respectively in migration assay (F=112.78,P ＜ 0.01), while 52.7 ％, 32.3 ％ and 10.3 ％ in invasion assay(F =183.76, P ＜ 0.01). Similarly, the percentage of RD-ES was 46.0 ％,39.0 ％ and 8.0 ％ in migration assay (F =408.25,P ＜ 0.01) and 58.7 ％,22.3 ％ and 9.0 ％ in invasion assay (F =373.25, P ＜ 0.01)respectively. The difference had statistics significance.The expression of MMP-9 was suppressed by As2O3 treatment according to gel zymography assay.Western blot assay showed that PI3K-AKT pathway was inhibited and nuclear factor kappa B(NF-κB)was inactivated.ConclusionLow-concentration As2O3 may inhibit metastasis capability of Ewing's sarcoma cells.

Objective To detect the different expression of Runx2 in dedifferentiated chondrosarcoma and conventional chondrosarcoma, and to investigate the role of Runx2 in the occurrence and development of dedifferentiated chondrosarcoma. Methods Dedifferentiated chondrosarcoma cell line NDCS-1 and normal chondrosarcoma cell line SW1353 were cultured, then mRNA and total cellular protein were extracted.RT-PCR Western blotting, and immunocytochemistry were used to detect the expression of Runx2.Immunohistochemistry was used to test Runx2's expression in the dedifferentiated chondrosarcoma specim ens that confirmed by pathology. Results Runx2 was high expression in dedifferentiated chondrosarcoma cell line and high-grade component of dedifferentiated chondrosarcoma tissues. Conclusion The high expression of Runx2 in dedifferentiated chondrosarcoma is involved in the occurrence and development of dedifferentiated chondrosarcoma.

Objective To study the effect and mechanism of arsenic trioxide on the motility, metastasis and invasion of osteosarcoma in vitro. Methods Wound healing assay, migration assay, invasion assay and Western-blot were performed to study the effect of arsenic trioxide on metastasis of osteosarcoma. Results Through screening MNNG cell was selected to perform the following research. After treatment of As_2O_3, the ability of MNNC cell flattening and spreading along the edges of the wound was inhibited, and the number of MNNG cells with migration and invasion in As_2O_3 - treated group was significantly less than in control group. Arsenic trioxide treatment also resulted in down-regulation of MMP-9. Conclusion This study is the first to report the effectiveness of arsenic trioxide as an inhibitor of osteosarcoma migration and invasion and the mechanism may be down-regulation of MMP-9.