ObjectiveThe aim was to construct the recombinant plasmid of pET-28a-G-protein pathway suppressor 2 (GPS2) GPS2,express GPS2 protein in E.coli,and obtain specific polyclonal antiserum of GPS2.MethodsGPS2 gene was obtained and the amplified fragment was then cloned into E.coli expression vector pET-28a to construct recombinant plasmid.The recombinant plasmid was transformed into E.coli expression strain BL21(DE3).IPTG induces the expression protein GPS2 protein,and the induction conditions were optimized.The induced product was purified by Ni2+ affinity chromatography,and the purified product was dialyzed with buffer for refolding.The purified protein can be used as antigen,injected to immunize male New Zealand white rabbit to get polyclonal antiserum.The titer and specificity of the rabbit antiserum were detected by ELISA and Western Blotting.ResultsThe E.coli expression vector pET-28a-GPS2 was constructed successfully and the recombinant protein was efficiently expressed and purified.The purified protein was used to immunize male New Zealand white rabbit to get polyclonal antiserum and the ELISA and Western Blot results showed that the high titer of specific polyclonal antiserum.ConclusionGSP2 could be highly expressed in E.coli.Antiserum of GPS2 protein can be obtained by the purified recombinant to analyze its function.

Objective:To evaluate the clinical effects of Pumpkin Polysaccharide Granules for diabetes Ⅱ.Methods:The clinical effects were evaluated on the governing principle of clinical investigation of diabetes treatment with new Chinese medicinal. In experiment the clinical effects of Pumpkin Polysaccharide Granules (as a treatment group) were compared with that of Xiaoke pill (as a control group).Results:The total effective rate of treatment group was 87.13%, 10 cases of then marked effect, 16 cases effect, 4 cases no improvement. The total effective rate of control group was 70.00%. The treatment group was superior to the control group ( P

Objective: To study the best condition of extracting effective component in Daphne odora Thurrb fresh flower by CO 2 supercritical fluid extraction(SFE). Methods: Uniform design was taken in these 4 factors: extracting pressure, extracting temperature, releasing pressure and releasing temperature, together with the experiments of 7 levels in each factor. Results: The best conditon in SFE: extracting pressure 35MPa, extracting temperature 40 ?C , releasing pressure over 8MPa and releasing temperature 36 ?C . Conclusion: This method is simple, highly selective and efficient in extracting effective component of Daphne odora thurrb fresh flowers.

Objective: To observe the clinical effect of treating remission-stage cerebral infarction with mind-refreshing and orifice-opening needling method. Method: Six hundred cases of cerebral infraction were randomized on the basis of disease phase. The 234 cases in remission stage were randomized into treatment group (116 cases) and control group (118 cases). Besides routine Western therapies, the cases in the treatment group were combined mind-refreshing and orifice-opening needling method and the cases in the control group were combined with conventional needling method. The treatment was done once every day for 4 weeks. The follow-up was done for six months. Result: the baseline material in the two groups has good compatibility (P>0.05) and the treatment group is better than the control group (P<0.05). Conclusion: the mind-refreshing and orifice-opening method is safe and act to improve symptoms of patients during remission stage, reduce disability, prevent disease progression and improve quality of life.

Objective:To evaluate the effect of esketamine on acute kidney injury (AKI) in the rats with sepsis and the role of autophagy.Methods:Forty SPF healthy adult male Sprague-Dawley rats, weighing 200-240 g, were divided into 5 groups ( n=8 each) by a random number table method: control group (Con group), esketamine group (Con+ Ket group), sepsis group (lipopolysaccharide [LPS] group), sepsis plus esketamine group (LPS+ Ket group), and sepsis plus esketamine plus 3-methyladenine (3MA) group (LPS+ Ket+ 3MA group). The model of AKI was established by intraperitoneal injection of LPS in anesthetized rats.Normal saline 10 ml/kg was intraperitoneally injected in Con group.In Con+ Ket group, normal saline 10 ml/kg was intraperitoneally injected, and 30 min later esketamine 10 mg/kg was injected via the tail vein.LPS 10 mg/kg was intraperitoneally injected in LPS group.In LPS+ Ket group, LPS 10 mg/kg was intraperitoneally injected, and 30 min later esketamine 10 mg/kg was injected via the tail vein.In LPS+ Ket+ 3MA group, LPS 10 mg/kg was intraperitoneally injected, and 30 min later esketamine 10 mg/kg and 3-MA 15 mg/kg were injected via the tail vein.The rats were anesthetized at 24 h after intraperitoneal injection of LPS and then sacrificed, and renal tissues were removed for microscopic examination of the pathological changes which were scored and for determination of contents of nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3), apoptosis-associated speck-like protein containing a CARD (ASC), caspase-1, interleukin-1beta (IL-1β) and IL-18 (by enzyme-linked immunosorbent assay) and expression of LC3, P62 and Beclin-1 (by Western blot). Results:Compared with Con group, the score for pathological damage to renal tissues and contents of NLRP3, ASC, caspase-1, IL-1β and IL-18 were significantly increased, LC3-Ⅱ/LC3-Ⅰ ratio was decreased, the expression of Beclin-1 was down-regulated, and the expression of P62 was up-regulated in LPS group ( P<0.05). Compared with LPS group, the score for pathological damage to renal tissues and contents of NLRP3, ASC, caspase-1, IL-1β and IL-18 were significantly decreased, LC3-Ⅱ/LC3-Ⅰ ratio was increased, the expression of Beclin-1 was up-regulated, and the expression of P62 was down-regulated in LPS+ Ket group ( P<0.05). Compared with LPS+ Ket group, the score for pathological damage to renal tissues and contents of NLRP3, ASC, caspase-1, IL-1β and IL-18 were significantly increased, LC3-Ⅱ/LC3-Ⅰ ratio was decreased, the expression of Beclin-1 was down-regulated, and the expression of P62 was up-regulated in LPS+ Ket+ 3MA group ( P<0.05). Conclusion:Esketamine can reduce AKI and autophagy is involved in the process, which is related to inhibiting the activation of NLRP3 inflammasomes and decreasing inflammatory responses in rats with sepsis.