OBJECTIVE:To develop HPLC method for quantitative determination of ti anepine sodium in blood plasma METHODS:Tianepine was extracted with ethyl acet ate The residues were analyzed with a reverse phase HPLC system(DiamonsilTM C18 column,4 6mm?250mm,5?m);Mobile phase,MeOH-C2H3OONH4(80∶20);UV detecti on 220nm RESUL_TS:The average recoveries for tianepine sodium in high,medium and low concentr ations were 97 63%,97 60% and 93 67%,respectively The within-day and b etween-day relative standard deviations were less than 5%(n=5) The calibratio n curves had good linearity within a concentration range of 1 0～83 3?g/ml T he regression equation was Y=6 05?10-2X+8 18?10-1,r=0 9 994(n=12) The limit of quantitation for tianepine sodium was 0 2?g/ml CONCLUSION:The metho d provides a se_nsitive,accurate,precise and reliable analytical procedure for clinical monito ring of tianepine sodium blood in plasma and its pharmacokinetic studies

OBJECTIVETo investigate the change and correlation of carbon monoxide/heme oxygenase and nitrogen monoxide/nitric oxide synthase system in atherosclerosis and the influence of the two systems on atherosclerotic progress.

METHODSThe rabbits received 1% cholesterol diet (chol group, n = 8), or 1% cholesterol diet plus L-arginine (L-arg group, n = 8) or L-NAME (L-NAME group, n = 8) by drinking water, or 1% cholesterol diet plus heme-L-lysinate (Heme group, n = 8) or ZnPP-IX (ZnPP group, n = 8) by injection in abdominal cavity for 10 weeks.

RESULTSCompared with those in control group, aortic NO production and expression of NOS decreased markedly; while CO production (P < 0.01) and HO-1 activity increased obviously in chol group. The aortic plaques area was (40.2 +/- 8.9)% in chol group. Compared with those in chol group, aortic areas [(26.6 +/- 9.2)%] reduced distinctly in heme group, aortic CO production and NOS activity increased obviously (P < 0.01) in L-arg group. However, compared with those in control group, HO-1 expression and CO production decreased markedly (P < 0.01) in heme group, while they were not different from those in chol group. Compared with those in chol group, aortic cNOS activity and NO production increased obviously and aortic plaques area [(28.1 +/- 7.7)%] greatly reduced (P < 0.01) in L-arg group. However, HO-1 expression and CO production of L-arg group decreased distinctly compared with those of control group, but they were similar to those of chol group. The aortic c-myc and c-fos expressions in both heme group and L-arg group reduced markedly compared with those in chol group, while they were similar to those in ZnPP and L-NAME group.

CONCLUSIONThe reciprocal relationship between heme oxygenase/carbon monoxide and nitric oxide synthase/nitrogen monoxide system in atherosclerosis may play the inhibitory role against atherosclerotic lesion.

Animals ; Atherosclerosis ; metabolism ; pathology ; physiopathology ; Carbon Monoxide ; metabolism ; Heme Oxygenase-1 ; metabolism ; Male ; Nitric Oxide ; metabolism ; Nitric Oxide Synthase ; metabolism ; Rabbits

Objective To explore the role of viral infection in the development of drug eruption in patients with HIV infection, and to evaluate the efficacy of antiviral treatment. Methods This study enrolled 87 HIV-positive patients, including 11 with and 76 without drug eruption, all of whom received highly active antiretroviral therapy(HAART). Clinical data on, baseline CD4+ and CD8+ T cell counts and CD4/CD8 ratio in these subjects were retrospectively analyzed. Results The severity of drug eruption was mild in the 11 HIV-positive patients, with a mean latency period of (14.00 ± 8.10)(range, 8 - 34)days. Of the 11 patients with drug eruption, 7 had liver function impairment, which was not in accordance with the severity of skin lesions. Drug eruption was controlled in all the 11 patients after anti-anaphylactic treatment without withdrawal of antiviral drugs. Compared with 75 HIV-positive patients without drug eruption, the 11 HIV-positive patients with drug eruption showed significantly increased baseline CD4 + T cell counts (493.00 ± 245.68 (range, 42 - 810)/μl vs. 347.81 ± 167.00 (range, 11 - 814)/μl, t = 647.50, P < 0.05), but decreased proportion of patients with baseline CD4+ T cell counts below the lower limit of normal(3/11 vs. 48/75(64.00%), X2 = 3.95, P < 0.05). There were no significant differences between 10 patients with drug eruption and 69 patients without drug eruption in the baseline CD8+ T cell count(1472.30 ± 858.55/μl vs. 1356.59 ± 684.06/μl, P > 0.05), CD4/CD8 ratio(0.40 ± 0.27 vs. 0.29 ± 0.16, P > 0.05), or percentage of patients with a CD4/CD8 ratio below the lower limit of normal (9/10 vs. 68/69 (98.55%), P >0.05). Conclusions The latency period of drug eruption seems to be long in HIV-positive patients receiving HAART, and mild drug eruption can be complicated by liver function impairment in the patients. Relatively high CD4 + counts may be a risk factor for the development and aggravation of drug eruption in HIV-positive patients.

BACKGROUNDIn recent years, many progresses have been made in molecular target therapy for lung cancer, in which anti-angiogenic target therapy is a hot spot drawing researchers' attention. The aim of this study is to explore the expression of canstatin gene transfected into human lymphocytes and its inhibitory effect on growth and metastasis of Lewis lung carcinoma.

METHODSThe eukaryotic expression vector of pCMV-Script and the recombinant pCMV-Script/Canstatin vector were separately transfected into lymphocytes by electroporation. The expression of canstatin protein in supernatant of lymphocyues was examined by SDS-PAGE assay. Furthermore, Lewis lung carcinoma cells were subcutaneously inoculated to C57BL mice to make animal model of tumor. When the transplanted tumors on the mice developed to 1cm³, the 30 mice were randomized into 3 groups, which were injected with 0.2mL supernatant of lymphocytes transfected with recombinant vector or naked vector, or 0.2mL NS respectively. After the treatment for 14 days, the size and pathological section of subcutaneous tumors were observed, and the number of pulmonary metastatic node was calculated.

RESULTSCanstatin protein was found in supernatant of the lymphocytes in the recombinant vector group by SDS-PAGE assay. After the treatment, the tumor size in the recombinant vector group (1.49cm³±0.18cm³) was significantly smaller than that in the naked vector group (2.44cm³± 0.19cm³) and NS group (2.53cm³±0.18cm³) (P=0.000). The numbers of pulmonary metastatic node were 3.40±1.14, 7.60±2.61 and 7.60±2.41 in the recombinant vector group, naked vector group and NS group respectively (recombinant vector group vs the other two groups, P=0.013).

CONCLUSIONSThe pCMV-Script/Canstatin vector can express canstatin protein in human lymphocytes. Canstatin has strongly inhibitory effect on growth and metastasis of mouse Lewis lung carcinoma.

AIMTo explore the effects of sodium hydrogen exchanger (NHE-1) specific hammerhead ribozyme on the expression and activity of NHE-1 and pHi in pulmonary artery smooth muscle cells (PASMCs) in rats, and its role in PASMCs proliferation.

METHODSAccording to the secondary structure of NHE-1 mRNA in rats, NHE-1 specific hammerhead ribozyme was designed with the assistance of computer. The recombinant vector of retroviral plasmid pLXSN and NHE-1 hammerhead ribozyme, PRZ, was transfected into the cultured PASMCs. G418 resistant cell clones were screened with 60 microg/ml G418. Then, the expression of NHE-1 mRNA was detected by RT-PCR, intracellular pH was measured with fluorescent probe-BCECF, 22Na and 3H-TdR incorporation were determined respectively.

RESULTSCompared with the cells transfected with pLXSN and non-transfected cells, NHE-1 mRNA, pHi value, 3H-TdR and 22Na incorporation decreased significantly in cells transfected with recombinant vector PRZ. No significance was found between the pLXSN transfected group and non-transfected group.

CONCLUSIONNHE-1 hammerhead ribozyme can cleave the target RNA specifically, reduce the expression of NHE-1 mRNA, induce intracellular acidosis and consequently prohibit the proliferation of PASMCs.

Animals ; Cell Proliferation ; Muscle, Smooth, Vascular ; Myocytes, Smooth Muscle ; cytology ; Pulmonary Artery ; cytology ; pathology ; RNA, Catalytic ; genetics ; RNA, Messenger ; genetics ; Rats ; Sodium-Hydrogen Exchanger 1 ; Sodium-Hydrogen Exchangers ; metabolism ; Transfection

OBJECTIVE: To investigate the classification and incidence of acid-base disturbance (ABD) in the patients with post-traumatic multiple organ dysfunction syndrome (MODS). METHODS: A total of 119 patients with MODS were examined with arterial blood gas analysis and serum electrolytes detection for 675 times in this study. RESULTS: Different types of ABD existed in 647 times out of 675 times (95.9%) of blood-gas analyses. There were 270 times (41.7%) of simple ABD, 271 times (41.9%) of double ABD and 106 times (16.4%) of triple ABD. Among which, 404 times (62.4%) were in respiratory alkalosis (RAL), 332 times (51.3%) in metabolic acidosis (MA), 227 times (35.1%) in metabolic alkalosis (MAL) and 167 times (25.8%) in respiratory acidosis (RA). In this study, 79 cases (66.4%) out of 119 cases with MODS died from these kinds of ABD. CONCLUSIONS: It suggests that in the early stage of MODS, RAL with or without hypoxemia may exist, and later on, MA or even triple ABD may occur. In order to detect and correct the primary disorders as early as possible, it is important to keep the balance of hydrolyte. The treatment of primary diseases is also important. Disorders of acid-base balance were corrected according to pH standard values, anion gap (AG) and the potential [HCO(3)(-)] were also calculated simultaneously. When pH was more than 7.50 or lower than 7.20, it is necessary to give drugs of acidity or alkalinity to the patients with ABD to maintain pH value within a normal range.

OBJECTIVETo investigate the effect of modified bone morphogenetic protein-2 polylactic acid nanospheres (BMP-2-PLA-Ns) sustained-release system on rabbit mandibular defect repair.

METHODSThe polylactic acid s nanospheres (PLA-Ns) and BMP-2-PLA-Ns were prepared by ultrasonic emulsification after graft polymerization. Forty-five rabbits were randomly divided into 3 groups: Blank group, PLA-Ns gel group(control group), and BMP-2-PLA-Ns gel group (experimental group). The rabbit mandibular defect models were established. The defect area of control group was implanted with PLA-Ns gel, meanwhile, the experimental group was implanted with BMP-2-PLA-Ns gel, the blank group experienced no special handling. Rabbits were killed in 1, 2, 4 weeks after operation and the iconography, hematine eosin(HE) staining and PCNA immunohistochemistry were used to detect the reparative effect on rabbit mandible defects.

RESULTSImage observation showed that bone defect repair in the experimental group was well and the shadow was not obvious. Better repair effect was seen compared with the control group and blank group. HE staining showed that the experimental group and the control group had a large number of neovascularization and secondary callus formation, callus in experimental group was obviously higher than that of control group and blank group. Immunohistochemical observation showed that the experimental group's PCNA positive chondrocytes were more than those in the control group and the blank group in the first 2 weeks; all groups of PCNA positive cells were rare in the fourth week, PCNA positive expression rate of the fourth week was lower than that of the first 2 weeks.

CONCLUSIONThe modified BMP-2-PLA-Ns sustained-release system promotes mandibular defect repair obviously.

Animals ; Bone Morphogenetic Protein 2 ; Delayed-Action Preparations ; Lactic Acid ; Mandible ; Nanospheres ; Polyesters ; Polymers ; Rabbits ; Reconstructive Surgical Procedures

OBJECTIVETo determine the role and related mechanisms of heme oxygenase-1/carbon monoxide (HO-1/CO) on VSMCs proliferation induced by insulin-like growth factor-I (IGF-I).

METHODSVSMCs isolated from rabbit aorta were cultured in vitro and proliferation was induced by IGF-I. Hemin (a substrate and inducer of HO-1) or zinc protoporphyrin-IX (Znpp-IX, an inhibitor of HO-1) was added to stimulate or inhibit the expression of HO-1. The mRNA and protein expressions of HO-1 were detected by RT-PCR and Western blot analysis. CO released into the culture media was quantitated by measuring carbon monoxide hemoglobin (COHb), VSMCs proliferation and cell cycle were determined by (3)H-TdR incorporation assay and flow cytometry, respectively.

RESULTSThe HO-1 mRNA and protein expressions in VSMCs and the amount of COHb in the culture media were significantly increased and the IGF-I-induced (3)H-TdR incorporations of VSMCs significantly reduced by hemin in a dose-dependent manner (P < 0.01). Furthermore, VSMCs in the G(0)/G(1) phase were increased and in the S and G(2)/M phase decreased by hemin (P < 0.01). Opposite results were observed in VSMCs treated with Znpp-IX.

CONCLUSIONSEndogenous HO-1 and CO are important mediators for inhibiting IGF-I induced VSMCs proliferation by reducing VSMCs DNA synthesis and decelerating cell cycle progression.

Animals ; Carbon Monoxide ; metabolism ; Cell Proliferation ; Cells, Cultured ; Heme Oxygenase-1 ; metabolism ; Insulin-Like Growth Factor I ; pharmacology ; Muscle, Smooth, Vascular ; cytology ; Myocytes, Smooth Muscle ; cytology ; metabolism ; RNA, Messenger ; genetics ; Rabbits

BACKGROUNDAngiogenesis is essential for tumor's growth and metastasis. Canstatin, a newly found potent endogenous angiogenesis inhibitor, has drawn researcher's attention to its powerful biological activities on endothelial cells. The aim of this experiment is to explore the expression and effects of canstatin gene in lung cancer A549 cells and HUV-ECC cells.

METHODSExpression vector of pCMV- Script/Canstatin was transfected into A549 and ECC cells by electroporation, and the positive clone was screened by G418. Growth characteristics of the two cell lines were compared before and after transfection. Expression of canstatin protein in supernatant was examined by SDS-PAGE assay, and cell cycles of the two cell lines were analysed by flow cytometry.

RESULTSThe expression of canstatin gene was found in supernatant of the transfected A549 cells and ECC cells. The apoptotic rate in the transfected ECC cells (16.04%) was significantly increased compared with that of the naked plasmid control group (0.43%) and parental cell group (2.92%) (P < 0.01). The growth of the transfected ECC cells was significantly inhibited (P < 0.01). The apoptotic rate in the transfected A549 cells (0.19%) showed no marked difference from the naked plasmid control group (0.13%) and parental cell group (0.07%) (P > 0.05). No significant difference in cell growth was found among the transfected A549 cell, naked plasmid control and parental cell groups.

CONCLUSIONSThe results indicate that canstatin gene can express in lung cancer A549 cell line and HUV-ECC cell line, and it can specifically inhibit proliferation of endothelial cell and induce its apoptosis.

BACKGROUND/AIMS: Upregulated CD64 expression on neutrophils is the most useful marker for acute bacterial infections and systemic inflammation. However, it is unknown whether CD64 is involved in the pathogenesis of acute pancreatitis (AP). This study was designed to determine whether CD64 is implicated in severe acute pancreatitis (SAP), and thus, is a suitable marker for SAP. METHODS: SAP was induced in rats with an intraperitoneal injection of L-arginine. CD64 expression in the rat pancreas was determined by quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemistry. Additionally, the CD64 mRNA expression in peripheral blood leukocytes from 21 patients with mild acute pancreatitis (MAP) and 10 patients with SAP was investigated at the time of admission and during remission by qRT-PCR. RESULTS: CD64 mRNA and protein expression in the pancreas was significantly higher in rats with SAP, compared to the controls. The CD64 expression was higher in the patients with SAP than in the patients with MAP. During remission, CD64 mRNA decreased in both the MAP and SAP patients. The area under the curve of CD64 expression for the detection of SAP was superior to both the Ranson and the Acute Physiology and Chronic Health Evaluation II scores. CONCLUSIONS: The CD64 level was significantly increased in correlation with the disease severity in SAP and may act as a useful marker for predicting the development of SAP.
Acute Disease ; Adolescent ; Adult ; Aged ; Animals ; Arginine/toxicity ; Female ; History, Ancient ; Humans ; Immunohistochemistry ; Male ; Middle Aged ; Pancreatitis/*metabolism ; RNA, Messenger/metabolism ; Rats, Sprague-Dawley ; Receptors, IgG/*metabolism ; Up-Regulation ; Young Adult