Objective　To investigate the changes of free calcium distribution in hepatocytes after administration of phenylephedrine. Methods The changes of fluorescence intensity were observed by confocal laser scanning microscopy after administration of phenylephedrine alone or pretreated with phentolamine before phenylephedrine administration. Results　The fluorescence intensity increased rapidly after administration of phenylephedrine to hepatocytes. When liver cells were pretreated by phentolamine before phenylephedrine administration, the changes of fluorescence intensity not obvious. Meanwhile, the inconformity of the fluorescence intensity in hepatocytes suggested the existence of the second subarea of free calcium distribution. Conclusion　Ca2+ signal can be arisen by phenylephedrine via the α-receptor in hepatocytes in vitro. The distribution and dynamic changes of free calcium in hepatocytes display some characteristics.

BACKGROUND:Conventional surgical repair can cause large traumas in patients with knee injuries, and patients often recover slowly after implant fixation, most of whom can appear to have poor recovery of knee function. OBJECTIVE: To explore the folow-up effect of arthroscopic double-steel wire clip fixation on tibial eminence avulsion fracture of anterior cruciate ligament. METHODS: A retrospective analysis was performed on the clinical data of 23 patients with tibial eminence avulsion fractures, who were given arthroscopic double-steel wire clip fixation. The patients were folowed up for 1-6 months. Short- and middle-term therapeutic effect as wel as IKDC and Lysholm scores before and after treatment were observed and analyzed. RESULTS AND CONCLUSION:The operation time was 35-65 minutes, and no complications, such as blood, nerve and anterior cruciate ligament injuries occurred. Moreover, no infection and other poor biocompatible reactions occurred after internation fixation. Al patients were folowed up for 1-6 months. The excelent and good rate was 87% at 1 month after treatment and 96% at 6 months after treatment. Al the patients had improved IKDC score and Lysholm score after treatment (P < 0.05), indicating that the knee function of patients was improved significantly.

OBJECTIVETo investigate surgical outcomes of strip nail internal fixation with bone graft in treating tibial plateau fracture.

METHODSFrom May 2012 to May 2014,36 patients with tibial plateau fracture were retrospectively analyzed, including 25 males and 11 females with an average age of 43.5 (ranged from 17 to 65) years old. The time from injury to operation ranged from 3 to 10 days with an average of 5.8 days. All patients were treated with L-shaped and T-shaped strip nail internal fixation with bone graft. It was evaluated by the Knee Functional therapy assessment method of the Special Surgical Hospital of American at final following-up. Varus angle, caster angle and femorotibial angle were recorded and compared at 3 days and 1 year.

RESULTSOperative time was (2.2 ± 0.6) h on average, blood loss was (310.5 ± 36.2) ml on average, hospital stay was (14.8 ± 2.7) days on average. Thirty-six patients were followed up from 12 to 30 months with an average of 18.2 months. Fracture healing time ranged from 4 to 8 months with an average of 6.2 months. The difference is not significant among varus angle, caster angle and femorotibial angle at 3 days and 1 year. According to the knee functional therapy assessment method of the Specialized Surgical Hospital of American, 18 cases got excellent results, 13 good, 4 moderate and 1 poor.

CONCLUSIONStrip nail internal fixation with bone graft for the treatment of tibial plateau fracture could effectively prevent the joint surface from secondary collapse, and achieve anatomic reduction, stable fixation and earlier functional exercise in further to get satisfied clinical effects.

Adolescent ; Adult ; Aged ; Bone Nails ; Bone Transplantation ; Female ; Fracture Fixation, Internal ; methods ; Fracture Healing ; Humans ; Male ; Middle Aged ; Retrospective Studies ; Tibial Fractures ; physiopathology ; surgery

Objective To prepare an attenuated Listeria vaccine Lmdd-LMP2A expressing the Ep-stein-Barr virus latent membrane protein 2A ( EBV-LMP2A) and evaluate its specific anti-tumor effects on nasopharyngeal carcinoma.Methods The gene fragment encoding EBV-LMP2A was amplified by PCR analysis and then subcloned into the shuttle vector p1565.PCR restriction enzyme digestion and DNA se-quencing were performed to identify the recombinant shuttle vector p1565-LMP2A.The p1565-LMP2A vector was then transformed into competent strains of the attenuated Listeria monocytogenes ( Lmdd) .The recombi-nant attenuated Listeria vaccine strain Lmdd-LMP2A was verified by Western blot assay.The histological sections of spleen and liver tissues were stained by Haematoxylin and eosin ( H&E) for analysis of inflamma-tion.A tumor-bearing HLA-A2 transgenic mouse model was established by subcutaneous injection of CNE-1/HLA-A2/LMP2A nasopharyngeal carcinoma cell line.The prepared Lmdd-LMP2A vaccine was inoculated into the mice via tail intravenous injection for the evaluation of specific CTL induction and the in vivo anti-tumor effects.Results The shuttle vector p1565-LMP2A and the recombinant attenuated Listeria vaccine strain Lmdd-LMP2A with stable expression of LMP2A protein were successfully constructed.The immunized mice showed mild inflammations with no structural damage and necrosis as indicated by H&E staining.The growth of tumors in tumor-bearing HLA-A2 transgenic mice was significantly inhibited by the immunization of Lmdd-LMP2A vaccine as compared with mice without inoculation.The survival time of mice was prolonged with the immunization of Lmdd-LMP2A vaccine.Conclusion The prepared attenuated Listeria vaccine Lm-dd-LMP2A showed specific anti-tumor effects with the safety advantage, suggesting the possibility of using it for anti-tumor therapy in clinic.

BACKGROUND: The occurrence and severity of acute high-altitude disease(AHAD) are determined by the speed entering the highland, the altitude of highland and seasons. The association between obesity and AHAD has not been fully investigated.OBJECTIVE: To investigate the association between obesity and acute high-altitude in people exposed rapidly to the highland.DESIGN: Comparative observation.SETTING: Department of Cardiology, General Hospital of Chinese PLA and Department of Cardiology, Tibetan General Hospital of Chinese PLA.PARTICIPANTS: The experiment was conducted at the Department of Cardiology, General Hospital of Tibetan Military Area Command of Chinese PLA in August 2006. Totally 82 Henan subjects aged 28-45 years were selected from the male workers constructing the Qinghai-Tibet Railway and their managers with acute high-altitude exposure. They had never been to the highland before, and all agreed to the detection.METHODS: ①Every subject completed the AHAD self-report questionnaire at sea level and 12 hours and 24 hours after ascending high-altitude. The items in the questionnaire included symptoms of headache, gastrointestinal symptoms,fatigue or weakness, dizziness, and insomnia. Each symptom was graded from 0-3 with 0 as no symptoms, 1 as mild symptoms, 2 as moderate symptoms, and 3 as severe symptoms and a total score of 15. A score of 4 or more could by identified as AHAD. ②The height and body mass were measured to calculate the body mass (BMI). Those with BMI≥28 kg/m2 served as the obesity group [n =39, mean age (35±8) years], and those with BMI of 18.5-23.9 kg/m2 as the normal body mass group [n =43, mean age (35±8) years]. ③Arterial blood was taken to evaluate arterial oxygen saturation (SO2), arterial oxygen pressure (PaO2) and arterial carbon dioxide pressure (PaCO2) at baseline and 24 hours after ascending high-altitude. ④The measurement data was compared by t test.MAIN OUTCOME MEASURES: BMI, vital capacity of lungs, SO2, PaO2 and PaCO2 levels of obese and normal people.RESULTS: Totally 39 obese people and 43 normal people were involved in the result analysis. ①AHAD score: No symptom was reported at sea level in all participants (scored 0), but the AHAD scores in the obesity group were significantly higher than those in normal group 12-hour and 24-hour after ascending high-altitude. ②Blood gas analysis:At sea level, there were no statistical differences in the levels of SO2, PaO2, and PaCO2 between two groups (P ＞ 0.05).But 24 hours after ascending high-altitude, SO2 and PaO2 of the obesity group were much lower than in the normal group (P＜ 0.01), and PaCO2 was significantly higher than in the normal group (P＜ 0.01).CONCLUSION: Obese men are more vulnerable to high-altitude hypoxia than people with normal body mass. Obesity is an important risk factor for the development of acute high-altitude disease.

Objective:To investigate the expression levels of PPARα/γin RAW264. 7 cells in the early stages of co-cultivation with Echinococcus granulosus in vitro. Methods:RAW264. 7 cells were co-cultured with E. granulosus and collected at 12,24,36,48, 72 h. The mRNA levels of PPAR-γ,PPAR-α,M1 macrophages-associated cytokines including TNF-α,MCP-1 and IL-1β,and M2 mac-rophages-associated cytokines including Arg-1,TGF-β and Fizz-1 were detected by qRT-PCR. The protein levels of Arg-1 and MR were analyzed by ELISA. Results:The expression levels of PPAR-γ, PPAR-αand the M2 macrophages-associated cytokines including Arg-1,TGF-β,Fizz-1 and MR were significantly increased,especially at 72 h (P<0. 05). M1 macrophages-associated cytokines including TNF-α,MCP-1 and IL-1β were decreased at 72h although increased at first. Conclusion:During the early stages of co-cultivation with Echinococcus granulosus in vitro, the levels of PPAR-γ/α are up-regulated in RAW264. 7 cells, which may drive macrophage polarization and play a role in the immune escape.

Objective To explore the biological basis of TCM syndromes from a biomolecules network perspective with qi deficiency syndrome as the breakthrough point. Methods A data dictionary of neuro-endocrine-immune (NEI) related genes and qi deficiency syndrome characterization terminology thesaurus were established. Literature about qi deficiency syndrome characterization was retrieved by using Genclip, to excavate the characteristic NEI gene, thereby to explore different bioactive substances of syndromes. Results The analysis of the genetic data, showed qi deficiency related cluster with the relevance of endocrine, signal transduction, hematopoietic cell and immune deficiencies etc. It is confirmed that the intrinsic biological features of TCM syndrome can effectively identify in the NEI level. Conclusion Literature mining method as a new way to discover syndromes biological indicators has certain feasibility, and it is recommended to be further expanded into other studies on syndromes to validate the universality and reliability of this method.

This research explored the synergistic effects and the potential mechanisms of RCE-4 and various nonsteroidal anti-inflammatory drugs (NSAIDs) on the proliferation of cervical cancer Ca Ski cells. The MTT assay and CalcuSyn V2.0 software were used to detect cell proliferation and calculate the combination index (CI); the expression levels of various proteins were analyzed using Western blot assay; mitochondrial membrane potential (MMP) was assessed using JC-1 staining; acridine orange/ethidium bromide (AO/EB) double-fluorescence staining was used to detect the apoptosis of Ca Ski cells; a co-immunoprecipitation (Co-IP) assay was used to analyze the relative content of Bcl-2-Beclin 1 complex in Ca Ski cells. The results demonstrate that the combination of RCE-4 and NSAIDs increases the inhibition of Ca Ski cells compared to the single-RCE-4 group, and celecoxib provided the best synergistic effect among the four NSAIDs tested, with a CI of 0.32. The combination of RCE-4 and celecoxib significantly down-regulated the expression of cyclooxygenase-2 (COX-2) and nuclear transcription factor-κB (NF-κB), and promoted the expression of non-steroidal anti-inflammatory drugs activity gene-1 (NAG-1). In addition, autophagy induced by RCE-4 was markedly inhibited in combination with celecoxib, which was associated with down-regulation of the expression of microtubule-associated protein 3 (LC3)-II, Beclin 1, p62 and autophagy-related gene (ATG) 3/4B/5/7/14. RCE-4-induced apoptosis was significantly enhanced by altering the depolarization of mitochondrial membrane potential and the expression of B cell lymphoma-2 (Bcl-2), B cell lymphoma-xl (Bcl-xl), Bcl-2 associated X protein (Bax), Bcl-2/Bcl-xl-associated death promoter (Bad) and cleaved cysteinyl aspartate specific proteinase (cleaved-caspase) 3/7/9. Furthermore, the formation of the Bcl-2-Beclin 1 complex was significantly inhibited in Ca Ski cells treated with RCE-4 in combination with celecoxib. Taken together, this research shows that the combination of RCE-4 and celecoxib has a significant synergistic effect on the proliferation of Ca Ski cells by promoting apoptosis, inhibiting autophagy and disturbing the formation of the Bcl-2-Beclin 1 complex, which may be a novel strategy to increase the sensitivity of anti-cervical cancer drugs.

OBJECTIVETo observe the effect and safety of Shuxuetong Injection (SXT) in treating patients with type 2 diabetes mellitus accompanying lower extremity atherosclerotic obliterans.

METHODSThe selected patients were assigned by randomized digital table to the treated group (180 cases treated with SXT) and the control group (80 cases treated with alprostadil). The clinical efficacy was assessed after 4 weeks of treatment, and the inner diameters and blood flow of the superficial femoral artery, the posterior tibial artery and the foot dorsal artery were measured and compared before and after treatment using Doppler's ultrasonography.

RESULTSThe total effective rate in the treated group was 94.4% (170/180), better than that in the control group, 85.0% (68/80, P < 0.01). Ultrasonic examination showed that the inner diameter and the blood flow of all the arteries measured in the treated groups increased after treatment (P < 0.01), with the increase superior to those in the control group (P < 0.05). No adverse reaction was found during the treatment.

CONCLUSIONSXT has the effect of activating blood circulation, dissolving stasis and dredging collaterals, it could reduce or eliminate the formation of thrombi, being effective and safe when applied in clinical practice for the treatment of lower extremity diabetic atherosclerotic obliterans.

Adult ; Aged ; Arteriosclerosis Obliterans ; drug therapy ; etiology ; Diabetes Mellitus, Type 2 ; complications ; drug therapy ; Drugs, Chinese Herbal ; administration & dosage ; Female ; Humans ; Infusions, Intravenous ; Lower Extremity ; blood supply ; diagnostic imaging ; Male ; Middle Aged ; Phytotherapy ; Ultrasonography

OBJECTIVETo establish and evaluate a BA-ELISA method for the quantitative detection of cystic fibrosis transmembrane conductance regulator (CFTR) protein.

METHODSWe deliberately selected three tables of CFTR and made the synthetic peptide be expressed in E. coli, then used the antigen to immunize rabbits to obtain the anti-CFTR polyclonal serum. After that, 96 well plates were coated with the purified antibody against CFTR. The antigen CFTR which was extracted from human sperm was detected by anti-CFTR antibody labeled with biotin, horseradish peroxidase conjugated avidin, and the substrate. The concentrations of two kinds of antibodies and the experiment parameters were optimized. Thereby, the double antibody sandwich BA-ELISA method for the quantitative detection of CFTR protein was established. Furthermore, the reproducibility, specificity and so on were evaluated by clinical specimens of sperm.

RESULTSThe optimal concentration of coated anti-CFTR IgG was 4 µg/ml, while the biotin labeled anti-CFTR IgG was 10 µg/ml; the optimal blocking buffer was 1% BSA-PBST, the optimal time of the reaction between antigen and antibody was 60 min, the optimal chromogenic time was 15 min, the intra-assay and inter-assay coefficient were 2.16%-9.23% and 2.29%-11.71% respectively; The lowest detectable limit was 0.15 ng/ml; the standard curve had a good linear correlation of R2 = 0.962.

CONCLUSIONThe BA-ELISA method for the quantitative detection of CTFR protein is successfully established, and it is demonstrated that the method has strong specificity, high sensitivity and good reproducibility. It provides the basis and evidence of the further application of the method.

Animals ; Antibodies ; Cystic Fibrosis Transmembrane Conductance Regulator ; analysis ; Enzyme-Linked Immunosorbent Assay ; methods ; Escherichia coli ; Humans ; Peptides ; Rabbits ; Reproducibility of Results ; Sensitivity and Specificity