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Dipeptidyl peptidase 4 (DPP4) is recognised as an attractive anti-diabetic drug target, and several DPP4 inhibitors are already on the market. As members of the same gene family, dipeptidyl peptidase 8 (DPP8) and dipeptidyl peptidase 9 (DPP9) share high sequence and structural homology as well as functional activity with DPP4. However, the inhibition of their activities was reported to cause severe toxicities. Thus, the development of DPP4 inhibitors that do not have DPP8 and DPP9 inhibitory activity is critical for safe anti-diabetic therapy. To achieve this goal, we established a selective evaluation method for DPP4 inhibitors based on recombinant human DPP8 and DPP9 proteins expressed by Rosetta cells. In this method, we used purified recombinant 120 kDa DPP8 or DPP9 protein from the Rosetta expression system. The optimum concentrations of the recombinant DPP8 and DPP9 proteins were 30 ng/mL and 20 ng/mL, respectively, and the corresponding concentrations of their substrates were both 0.2 mmol/L. This method was highly reproducible and reliable for the evaluation of the DPP8 and DPP9 selectivity for DPP4 inhibitor candidates, which would provide valuable guidance in the development of safe DPP4 inhibitors.
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Objective To evaluate the lethal effect of adenovirus-mediated Ad-hTERTp-HSV-TK/GCV suicide gene system in combination with oxaliplatin (L-OHP) on human hepato carcinoma cell line HepG2 in vitro.Methods It was used that the replication-defective adenovirus carring the HSV-TK gene under control of the hTERT promoter to transfer the HepG2 cells in vitro. Human hepato carcinoma cell line HepG2 was transfected with MOI=100. It was studied that the grow inhibitory effct with Ad-hTERTp-HSV-TK/GCV therapy, oxaliplatin, Ad-hTERTp-HSV-TK/GCV therapy in combination with oxaliplatin through different drug concentration on human hepato carcinoma cell line HepG2. The growth inhibition rate of HepG2 cells was determined by trypan blue exclusion assay and MTT.Results The growth of HepG2 cells Ad-hTERTp-HSV-TK/GCV, oxaliplatin, and combination of Ad-hTERTp-HSV-TK/GCV and oxaliplatin was significantly slower.The more concentration the greater inhibition of cell growth. The growth inhibition rate of combined Ad-hTERTp-HSV-TK/GCV with oxaliplatin was 86.63 %.The growth inhibition rate of Ad-hTERTp-HSV-TK/GCV was 72.12 % compared to the combined therapy (P =0.023). The growth inhibition rate of oxaliplatin was 59.41% compared to the combined therapy (P =0.019). Combination of Ad-hTERTp-HSV-TK/GCV and oxaliplatin resulted in greater inhibition of cell growth compared with TK gene and L-OHP (P ＜0.05).Conclusion HSV-TK/GCV in combination with L-OHP can enhance thelethal effect of suicide gene therapy against HepG2 cells.It is more targetable than the function of single drug therapy.Also, it could reduce drug level and plays an important role on the future's clinical medication.
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Objective To observe the effect of Ad-bTERTp-HSV-TK/GCV system on malignant ascites of mice and probe into its mechanism of action.Methods The SX1 inbred strain mice were injected with H22 cell line of liver cancer and were divided into 4 groups at random.The mice in each group were given corresponding treatment after 48 hours.The production of ascites and survival period were evaluated. The apoptosis rates of tumor cells were detected by FCM.Morphological changes of tumor cells were studied by electromicroscope.Results Compared with other groups.Ad-hTERTp-HSV-TK/GCV Can obviously inhibit the production of ascites(P<0.01),prolong the survival period (P<0.01),and apoptosis rate in this group (27.12±2.12)% was significantly higher than that in other groups.No obvious side effect Was found during the treatment.Conclusion Ad-hTERTp-HSV-TK/GCV system Can inhibit production of ascites and prolong the survical period of mice by inducing apoptosis of hepatoma cells,which is a safe and feasible treatment for hepatoma therapy.
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@#ObjectiveTo investigate the effects of intramuscular Botulinum toxin type A (BTX-A) injection on spasticity of lower limb, walking and activities of daily living (ADL) after stroke. Methods13 patients with spasticity on lower limb were injected intramuscularly with BTX-A, and assessed with Composite Spasticity Scale (CSS), step length and gait velocity, and Barthel index (BI) before and 1, 4, 8 and 12 weeks after treatment. ResultsThe scores of CSS and BI significantly improved after treatment (P<0.05). There was some improvement in step length and gait velocity, but not significant (P>0.05). All the improvement appeared 1 week after injection and lasted to the 12th week. ConclusionLocal intramuscular BTX-A injection can reduce the spasticity on lower limb and improve ADL.
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@# Objective To investigate the effect of hyperbaric oxygen (HBO) on basic activities of daily living (B-ADL) of patients with unilateral spatial neglect (USN) after stroke.Methods 72 patients with USN after stroke were divided into the experimental group (group A1, 36 cases), who received the training for USN and HBO, and observation group (group A2, 36 cases), who received the training for USN alone. Other 90 stroke inpatients without USN at the same time were as control (group B). All of them received routine rehabilitation, and were assessed with Modified Barthel Index (MBI) before and after treatment. Results Before treatment, the scores of MBI in the patients with USN (groups A1 and A2) were significantly lower than those in the control group (B) (P<0.05). The scores of MBI in all groups improved 6 weeks after treatments (P<0.001). While, the scores in the group A2 were lower than that of the group A1, but both were not significantly different from control (P>0.05). Conclusion USN would significantly impair the ADL of stroke patients, and the rehabilitation for USN may improve their ADL. HBO may improve the outcome.
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Objective To construct the recombinant adenovirus vector with hTERT-HSV-TK and observe the killing effect of Ad-hTERTp-HSV-TK/GCV system on hepatocellular carcinoma cells. Methods A recombinant replication defective adenoviral vector of Ad-hTERTp-HSV-TK was constructed via homologous recombination which both shuttle plasmid pSU-Tp-TK and adenovirus backbone plasmid pBHGE3 transfected into the HEK293 packaging cells. Then the Ad-hTERTp-HSV-TK was amplified and purified through PCR. The activity of the HepG2 cells and the L-02 cells were tested by methyl thiazolyl terazolium (MTT) after they were transfected by the recombinant adenovirus of different multiplicities of infection (MOI) and then were added GCV of different conc.entration. Results The recombinant replication defective adenoviral vector of Ad-hTERTp-HSV-TK were identified by PCR successfully. The viral titer was 1.5×1010 pfu/ml after amplification and purification. The HepG2 cells were targetedly suppressed by Ad-hTERTp-HSV-TK/GCV system. The survival rate of cells decreased gradually along with the increase of the MOI and the GCV' s concentration. Conclusion The recombinant replication defective adenoviral vector of Ad-hTERTp-HSV-TK can inhibit the HepG2 cells significantly, but has not influence on the L-02 cells.
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Telomerase, as a factor of tumour's happening and development, is a kind of cell element not existing or with low activity in normal cells. Moreover, the activity of telomerase is a common channel for cell's immortalization and deterioration, and is also the main factor in cloning tumour and developing malignant turnour. Therefore, the tumour gene therapy focusing on telomerase has become the new target of present study of tumor.This therapy has greater prospects on clinical application than the past tumour gene therapy simply directing at one cancer gene.
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After two years' study on the application of PP333 to one-year-grown Angelica sinensis (Oliv. )Diels.,it was found that spraying 300mg/L PP333 on the seedlings markedly increased the average fresh and dry weight of its root,as well as the ash content especialy its acid-soluble ash and alcohol-soluble substance. Thus,this method can inerease not only the yield but also improve its quality,which is a low-investment and high-yield way for the production of A. sinensis (Oliv. )Inels