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Objective To verify the performance characteristics of the human SLCO1B1 & APOE gene detection kit (PCR-fluorescent probe technique).Methods The verification protocol for the human SLCO1B1 & APOE gene detection kit manufactured by Wuhan YZY Medical Science and Technology Co.,Ltd.was designed by referring to the requirements of professional standard and pertinent literatures,and its accuracy,specificity,lowest detection limit and anti-interference ability were compared with those of sequencing technique (gold standard).Results The results coincidence of 20 clinical samples from fluorescence PCR and sequencing technique was 100％,which met the requirement of accuracy.Forty clinical samples with wild type loci determined by fluorescence PCR were verified by sequencing and no any mutation was detected,which met the requirement of specificity.The lowest detection limit of the kit was 10 ng/μL.Three interfering substances,including hemoglobin,bilirubin and triglyceride,were individually added into the blood samples with known genotype,and the results determined by the kit were coincident with that without interfering substances,which met the requirement of anti-interference ability.Conclusion The verification parameters of the human SLCO1B1 & APOE gene detection kit are basically consistent with the manufacturer's statement,indicating that the kit meets the requirements of the relevant quality management and may be applied to the detection of clinical samples.
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<p><b>BACKGROUND</b>Skeletal muscle has recently been recognized as an endocrine organ that can express, synthesize and secrete a variety of bioactive molecules which exert significant regulatory effects. Hydrogen sulfide (H2S) is endogenously produced in mammalian tissues and participates in a number of physiological and pathophysiological processes. We aimed to verify whether H2S could be endogenously generated and released by rat skeletal muscle, and determine the biological effects of H2S in rat skeletal muscle.</p><p><b>METHODS</b>The study was divided into two parts: detection of endogenous H2S generation and release in rat skeletal muscle and determination of antioxidative activity of skeletal muscle-derived H2S. H2S content and production in tissues were detected by sensitive sulfur electrode method. The expressions of H2S producing enzymes cystathionine β-synthase, cystathionine γ-lyase and mercaptopyruvate sulfurtransferase were detected by real-time PCR and western blotting and their tissue distributions were observed by immunohistochemical and immunofluorescent analysis. Rat skeletal muscular ischemia-reperfusion (I-R) injury model was created and evaluated by histological analysis under microscope. The malondialdehyde (MDA) contents, hydrogen peroxide levels, superoxide anion and superoxide dismutase (SOD) activities were detected using spectrophotometer.</p><p><b>RESULTS</b>H2S could be endogenously generated and released by skeletal muscle of Sprague-Dawley rats (H2S content: (2.06 ± 0.43) nmol/mg; H2S production: (0.17 ± 0.06) nmol×min(-1)×mg(-1)). Gene and protein expressions of the three H2S producing enzymes were detected in skeletal muscle, as well as the liver and kidney. Endogenous H2S content and production were decreased in skeletal muscles of rats with I-R skeletal muscle injury (P < 0.05). Furthermore, H2S significantly protected rat skeletal muscle against I-R injury and resulted in decreased MDA content, reduced hydrogen peroxide and superoxide anion levels, but increased SOD activity and protein expression in skeletal muscles (all P < 0.01).</p><p><b>CONCLUSION</b>H2S generation pathway exists in rat skeletal muscle and it acts as an antioxidant in skeletal muscle.</p>
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Animals , Male , Rats , Blotting, Western , Hydrogen Peroxide , Metabolism , Hydrogen Sulfide , Metabolism , Immunohistochemistry , Malondialdehyde , Metabolism , Muscle, Skeletal , Metabolism , Oxidative Stress , Physiology , Rats, Sprague-Dawley , Superoxide Dismutase , Metabolism , Superoxides , MetabolismABSTRACT
Objective To investigate interventional procedures and polygene feasibility for the treatment of liver carcinoma.Methods pCMV-p53 plasmid-liposome complex and concentrated TKCD retrovirus of supernatant liquid were prepared along with rabbit VX2 liver tumor models of 50 adult New Zealand rabbits.VX2 liver tumors about 2 cm in diameter from 45 adult rabbits were randomly divided into 5 groups of 9.Group 1 was the control group that used 0.9％ sodium chloride as a placebo.Group 2 had transcatheter arterial embolization with lipiodol as treatment.Group 3 was the lipiodol and p53 group.Group 4 was the lipiodol and TK/CD group.Group 5 was the lipiodol,p53,and TK/CD group.The microtubule(1.2f)was inserted from the femoral to hepatic artery,tumor supply arteries were demonstrated by angiograms,and the drug was slowly injected under x-ray.The VX2 liver tumors were examined with B-ultrasound and computed tomography for maximum diameter (a) and minimum diameter (b) before and 10 days after interventional therapy.Gross tumor volume (V=ab2/2) and tumor growth rate were calculated.All the adult rabbits were euthanized 8 weeks after interventional therapy (including natural deaths).Histopathological examination was taken and survival time was observed.Results The tumor volume among the 5 groups had no significant difference before interventional therapy (P＞0.05).Ten days after interventional therapy,analysis of the tumor volume for variance and the T-test were carried out.The results showed that each group compared to the control showed a significant difference in inhibiting cancer growth (P＜0.05).The lipiodol,p53,and TK/CD group showed the best effect.According to factorial statistic analysis (2x2),p53 or TK/CD combined with lipiodol therapy can control the tumor obviously,but no mutual synergism effect was found (P=0.793).Each treatment group showed a significant difference of prolonged survival time compared to the control group (P＜0.01).The multi-treatment or multi-gene group showed the best curative effects.Conclusions Interventional therapy can be the ideal path for administering medications for gene therapy.Transcatheter arterial embolization with lipiodol,wild-type p53 gene,TK/GCV,and CD/5-Fc applied in combination can control tumor growth and prolong survival time.
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Objective To evaluate the effcacy and side effects of [~(90)Y-1,4,7,10-tetraazacyclododecane-N,N',N",N'"-tetraacetic acid (DOTA)~0,Tyr3]octreotate (~(90)Y-DOTATATE) combined with lysine as a renal protective agent.Methods Twenty-five patients with advanced neuroendocrine tumors were confirmed as somatostatin receptor subtype-2 (SSTR2)-positive by somatostatin receptor scintigraphy (SRS).Each patient received 1-5 cycles of treatment and the interval between two cycles of treatment was 6-9 weeks.~(90)Y-DOTATATE was administered intravenously within 30 min.Lysine was injected before and after the administration of ~(90)Y-DOTATATE.After each treatment cycle.the side effects were assessed according to National Cancer Institute Grading Criteria(NCIGC).The etticacy was evaluated by the WHO criteria 8 weeks after the last treatment.Results Pattial remission was found in 1 patient (4%).minor response in 3 patients(12%),stable disease in 16 patients (64%)and tumor progression in 5 patients (20%).Two patients suffered from renal functional injuries and 3 patients developed leukocytopenia.Three patients showed nausea while another 3 patients presented vomiting.Conclusions ~(90)Y-DOTATATE in association with lysine may be a promising treatment method for the patients with metastatic and inoperable neuroendocrine tumors.More research work may be directed to reduce renal injury.
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Objective:To investigate the inhibitory effect of breast cancer-associated antigen 1(BRCAA1)gene silencing on gastric cancer MGC-803 cells and the related mechanism.Methods:Plasmid shRNA-BRCAA1 and shRNA-N were constructed and transfected with FuGene HD into gastric cancer cell line MGC-803.The transfection efficiency was examined using fluorescent microscope 24 h later.The total RNAs was extracted 48 h 'after transfection and the expression of BRCAA1 and GAPDH gene were analyzed by real-time PCR.The cell proliferation was assessed by MTT assay 24 h,48 h,and 72 h after transfection.The cell apoptosis was determined by Annexin V-PE/TAAD.The expression of Rb,Bax, Bcl-2 and BRCAA1 proteins was analyzed by Western blotting 48 h after transfection.Results:We found that the transfection efficiency of shRNA-BRCAA1 was(81.2?2.6)%24 h after transfection.Forty-eight hours after transfection with shRNA-BRCAA1 the expression of BRCAAI mRNA decreased by 61.4%;the inhibition rate of MGC-803 cells growth was 45.0%.The cell apoptosis rate of shRNA-BRCAA1 transfection group was significantly higher than those of untransfected group and mock plasmid transfected group([14.4?1.6]%vs[5.4?2.0]%,[4.4?2.5]%,P
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Objective:To investigate the feasibility of targeted imaging and therapy of prostate cancer using nanocomposite probes composed of fluorescent magnetic nanoparticles(FMCNPs) and single chain Fv(ScFv) antibody specific for gama-seminoprotein.Methods:The nanocomposite probes(FMCNPs-ScFv) were prepared by conjugating fluorescent magnetic nanoparticles with singlegama-chain Fv antibody specific gama-seminoprotein,and were characterized by high resolution transmission electron microscopy,fluorescent spectrum and magnetic spectrum.Nanocomposite probes were incubated with prostate cancer LNCaP cells,and the targeting results of nanocomposite probes were observed by fluorescent microscopy.The cytotoxicity effect of the nanocomposite probes was measured by MTT.Nude mice models of prostate cancer were established and identified by immunohistochemistry method.The nanocomposite probes were injected into nude mice via tail vein.The distribution of nanocomposite probes in the nude mice was observed by Micro-animal imaging system,targeted imaging of the prostate cancer was observed by MR instrument.The nude mice with prostate cancer were irradiated with 100 W magnetic field for 30 min,and the changes of tumor sizes were observed.Results:The FMCNPs-ScFv nanocomposite probes were successfully prepared.Nanocomposite probes entered into the cytoplasm of cancer cells and exhibited low cytotoxicity effect.Nude mice model with prostate cancer were successfully fabricated;the nanocomposite probes distributed quickly in the main organs of mice,and gradually concentrated on the tumor tissues within 24 h.MR images showed that the tumor images were gradually enhanced from 6 h to 24 h after injection of the nanocomposite probe.Four days after magnetic irradiation,the tumors in the nude mice grew slower compared with the control nude mice(P