Objective To observe the effect of fluvastatin on the activity of extracellular signal-regulated kinase(ERK)1/2,expression of connective tissue growth factor(CTGF) in glomerular mesangial cells stimulated by high glucose,and explore the pathogenic mechanism of diabetic nephropathy(DN) and its early prevention and treatment.Methods The study consisted of two parts,the first part was to repeat the existing experienment of stimulatory effect of high glucose on mesangial cells,and validated that high glucose could up-regulated the expression of CTGF through stimulating ERK,and the change was unrelated with hypertonic conditions.There were five groups:①normal control group(NG);②high glucose group(HG);③mannitol group(MN);④NG+ERK inhibitor PD98059 group(NG+PD);⑤HG+ ERK inhibitor PD98059 group(HG+PD).The second part was to observe the changes of the expressions of CTGF protein and mRNA in mesangial cells at high glucose concentration before and after fluvastatin intervention.There were also five groups:①normal control group(NG);②high glucose group(HG);③normal glucose and fluvastatin group(NG+F);④high glucose + fluvastatin(HG+F);⑤high glucose + fluvastatin and mevalonic acid group(HG+F+M).The activity of ERK1/2 and the expressions of CTGF protein and mRNA in cultured glomerular mesangial cells stimulated by high glucose were detected with Western blotting and RT-PCR.The amount of fibronectin(FN) in the supernatant of cells was analyzed by ELISA.Intracellular total protein level was measured by Coomassie brilliant blue method.Results The first part showed that the activity of ERK1/2 and the expression of CTGF protein in HG were more higher than those in NG(P

Objective To construct recombinant plasmids containing HPV18E7 gene ,and explore the optimization condition of its expression in Escherichia coli .Methods The genomic DNA extracted from HeLa cell line which served as a template to the HPV18 E7 gene was amplified using PCR method ;and the amplified product of HPV18E7 gene was connected to the pET-32a(+ ) vector ,which composed the pET-32a(+ )-HPV18E7 recombinant plasmid ;the positive recombinant plasmids were transformed into BL21-DE3-pLysS competent cells and the optimized expression condition was explored in order to obtain a large amount of HPV18E7 oncogenic protein .Results The fragment length of PCR products of HeLa cell genomic DNA was consistent with that of HPV18 E7 gene .In LB medium ,the expression level of the target protein was not high under such conditions as different concentra-tion of IPTG and lactose ,different temperatures and different induction starting amount .Therefore the ZYM-5052 auto-induction medium was tried in this experiment ,and the expression amount of the fusion protein was much higher than that induced with IPTG and lactose .Conclusion The amount of HPV18E7 fusion protein in ZYM-5052 automatic induction medium is much higher than that induced with IPTG and lactose .

Objective To explore the reversibility of early stage tubular interstitial injury as well as the timing of reparation through the pig relief of unilateral ureteral obstruction (R-UUO) model.Methods Eight three-month-old female Guangxi BA-MA mini pigs were selected for the construction of R-UUO models.Five time points were set which were UUO 0 day,UUO 3 days,R-UUO 7 days,R-UUO 14 days,and R-UUO 21 days.Renal function,histological structure,and protein expressions of α-smooth muscle actin (α-SMA),vimentin and E-cadherin were evaluated at different time points.Results After 3 days of UUO,compared with UUO 0 day,serum creatinine levels were increased obviously and the kidney tissues presented varying degrees of damage.The expressions of α-SMA and vimentin were increased and E-cadherin expression was decreased (P ＜ 0.05).Following R-UUO after 3 days of UUO,compared to UUO,serum creatinine levels were significantly decreased.Renal pathological tissue damage was repaired.The expressions of α-SMA and vimentin were decreased and E -cadherin expression was increased (P ＜ 0.05).Conclusions The pig R-UUO animal model may provide a good platform to study the kidney injury and repair.The tubular injury may be fully reversed and repaired when removing the pathogenic factors if the renal tubular injury was at an earlier stage.

In the original publication of the article the keywords are incorrectly online published. The correct keywords should read as α-Conotoxin; Nicotinc acetylcholine receptor; Acetylcholine binding protein; X-ray crystallography".

The α3* nAChRs, which are considered to be promising drug targets for problems such as pain, addiction, cardiovascular function, cognitive disorders etc., are found throughout the central and peripheral nervous system. The α-conotoxin (α-CTx) LvIA has been identified as the most selective inhibitor of α3β2 nAChRs known to date, and it can distinguish the α3β2 nAChR subtype from the α6/α3β2β3 and α3β4 nAChR subtypes. However, the mechanism of its selectivity towards α3β2, α6/α3β2β3, and α3β4 nAChRs remains elusive. Here we report the co-crystal structure of LvIA in complex with Aplysia californica acetylcholine binding protein (Ac-AChBP) at a resolution of 3.4 Å. Based on the structure of this complex, together with homology modeling based on other nAChR subtypes and binding affinity assays, we conclude that Asp-11 of LvIA plays an important role in the selectivity of LvIA towards α3β2 and α3/α6β2β3 nAChRs by making a salt bridge with Lys-155 of the rat α3 subunit. Asn-9 lies within a hydrophobic pocket that is formed by Met-36, Thr-59, and Phe-119 of the rat β2 subunit in the α3β2 nAChR model, revealing the reason for its more potent selectivity towards the α3β2 nAChR subtype. These results provide molecular insights that can be used to design ligands that selectively target α3β2 nAChRs, with significant implications for the design of new therapeutic α-CTxs.
Animals ; Aplysia ; Binding Sites ; Conotoxins ; chemistry ; Crystallography, X-Ray ; Humans ; Protein Structure, Quaternary ; Receptors, Nicotinic ; chemistry