Capillary zone electrophoresis (CZE) was developed to investigate the structure stability of novel camptothecin derivative (L-P) at different pH,the kinetics and thermodynamics of hydrolysis reaction from lactone form to carboxylate form direction at near physiological conditions (pH 7.4,310 K). Uncoated fused-silica capillaries(35 cm×50 μm i. d,with effective length of 26.5 cm) were used. The background electro-lyte( BGE) was 0.025 mol/L sodium phosphate buffer with pH varied at 2.5,4.0,5.0,6.0,7.0,7.4 and 9. 0. The electrophoresis voltage was maintained at 14 kV when the pH of BGE ranged between 2.5 and 5.0,otherwise,the voltage was maintained at 10 kV. The UV detector was set at 260 nm. All samples were introduced using hydrodynamic injection at 5 kPa for 4 s. L-P was found to be lactone form as the solution pH was below 4. 0. As pH increased,the lactone form of L-P would undergo hydrolysis reaction to be carboxylate form. As pH was 9.0,L-P existed almost completely as carboxylate form. The rate constant of the hydrolysis increased as temperature raise. The energy of activation ( Ea) ,the enthalpy ( ΔH) and entropy (ΔS) of the hydrolysis reaction were determined as 72. 6 kJ/mol,10. 5 kJ/mol and 50. 9 J/( mol K) ,respectively. The proposed capillary zone electrophoresis could efficiently separate two pH-dependent structural forms of the novel camptothecin derivative( L-P). The positive enthalpy and entropy values of the L-P hydrolysis indicated that the reaction was endothermic and entropically driven and higher temperature favored.

Objective: To study the relationship between myocardial X-box binding protein 1 (XBP1) expression and myocardial apoptosis in experimental diabetic cardiomyopathy (DCM) rat’s model and to clarify the mechanism of valsartan inhibiting myocardial apoptosis. Methods: A total of 50 Wistar rats were divided into 2 groups: Control group, the rats received intraperitoneal citrate buffer at 65mg/kg,n=10 and Streptozotocin group, the rats received intraperitoneal streptozotocin at 65mg/kg,n=40, all animals were treated for 7 days. DCM model was established in 37 rats (fasting blood glucose ≥ 16.7mmole/L) and they were further divided into 2 groups: DCM group, the rats received intragastric normal saline,n=20 and DCM + valsartan group, the rats received intragastric valsartan at 30mg/kg·day,n=17. The rats were treated for 16 weeks. The body weight, tail blood pressure, glucose and cardiac function were compared among 3 groups. Myocardial apoptosis was detected by TUNEL staining, RNA and protein expressions of myocardial cytochrome C, cleaved caspase 3, glucose regulation protein 78 (GRP78) and XBP1-s were examined by immunolfuorescence, real time RT-PCR and Western blot analysis. Results: Compared with Control group, DCM group showed disordered cardiac structure, more collagen content and myocardial apoptosis,P<0.05; increased RNA and protein expressions of GRP78, XBP1-s, cleaved caspase 3 and cytochrome C,P<0.05. Compared with DCM group, DCM + valsartan group had rather regularly arranged myocardiocytes, less interstitial ifbrosis and myocardial apoptosis,P<0.05; decreased RNA and protein expressions of GRP78, XBP1-s, cleaved caspase 3 and cytochrome C,P<0.05. Conclusion: Valsartan may inhibit myocardial XBP1 activation and therefore, reduce the myocardial apoptosis in experimental DCM rat’s model.

Kv2.1 channel currents in pancreatic beta-cells are thought to contribute to action potential repolarization and thereby modulate insulin secretion. Because of its central role in this important physiological process, Kv2.1 channel is a promising target for the treatment of type 2 diabetes. Jingzhaotoxin-XI (JZTX-XI) is a novel peptide neurotoxin isolated from the venom of the spider Chilobrachys jingzhao. Two-microelectrode voltage clamp experiments had showed that the toxin inhibited Kv2.1 potassium currents expressed in Xenopus Laevis oocytes. In order to investigate the structure-function relationship of JZTX-XI, the natural toxin and a mutant of JZTX-XI in which Arg3 was replaced by Ala, were synthesized by solid-phase chemistry method with Fmoc-protected amino acids on the PS3 automated peptide synthesizer. Reverse-phase high performance liquid chromatography (RP-HPLC) and matrix assisted laser desorption/ ionization time-of-flight mass spectrometry (MALDI-TOF/TOF MS) were used to monitor the oxidative refolding process of synthetic linear peptides to find the optimal renaturation conditions of these toxins. The experiments also proved that the relative molecular masses of refolded peptides were in accordance with their theoretical molecular masses. RP-HPLC chromatogram of co-injected native and refolded JZTX-XI was a single peak. Under the whole-cell patch-clamp mode, JZTX-XI could completely inhibit hKv2.1 and hNav1.5 channels currents expressed in HEK293T cells with IC50 values of 95.8 nmol/L and 437.1 nmol/L respectively. The mutant R3A-JZTX-XI could also inhibit hKv2.1 and hNav1.5 channel currents expressed in HEK293T cells with IC50 values of 1.22 micromol/L and 1.96 micromol/L respectively. However, the prohibitive levels of R3A-JZTX-XI on hKv2.1 and hNav1.5 channels were reduced by about 12.7 times and 4.5 times respectively, indicating that Arg3 was a key amino acid residue relative to the hKv2.1 channel activity of JZTX-XI, but it is also an amino acid residue correlated with the binding activity of JZTX-XI to hNav1.5 channel. Our findings should be helpful to develop JZTX-XI into a molecular probe and drug candidate targeting to Kv2.1 potassium channel in the pancreas.
Animals ; HEK293 Cells ; Humans ; Insulin-Secreting Cells ; metabolism ; Mutant Proteins ; genetics ; pharmacology ; NAV1.5 Voltage-Gated Sodium Channel ; metabolism ; Neurotoxins ; chemical synthesis ; genetics ; pharmacology ; Protein Refolding ; Shab Potassium Channels ; antagonists & inhibitors ; metabolism ; Sodium Channel Blockers ; pharmacology ; Spider Venoms ; genetics ; pharmacology ; Transfection

Objective     To investigate the early safety and efficacy of transapical transcatheter aortic valve implantation (TAVI) for high-risk elderly patients with pure aortic valve insufficiency. Methods     A prospective multicenter clinical study of domestic J-valveTM TAVI for high-risk native non-calcified aortic valve insufficiency was conducted from April 2014 to May 2018, and the early postoperative results were analyzed. A total of 82 patients were enrolled, including 62 patients from West China Hospital, Sichuan University, 16 patients from Zhongshan Hospital, Fudan University, and 4 patients from Beijing Fuwai Hospital, National Center for Cardiovascular Diseases. There were 55 males and 27 females. The age was 61-90 (73.8±6.3) years. The logistic EuroSCORE was 10.0%-44.4% (17.5%±8.1%). All patients underwent TAVI using J-ValveTM system. Clinical evaluation and echocardiography were performed preoperatively and 1 month postoperatively. Multislice spiral CT was reviewed before discharge. Results     Three patients were transferred to thoracotomy for cardiopulmonary bypass operation, and 1 patient had decreased cardiac function due to leakage of the valve 1 week after surgery. The overall technical and procedural success rate was 95.1% and 93.9%, respectively. During hospitalization, 1 patient died of moderate pericyclosis complicated with multiple organ failure, and 1 patient died of pulmonary infection. Six (7.6%) patients received pacemaker implantation due to new onset Ⅲ° atrioventricular block. Echocardiographic follow-up showed paravalvular leak was observed in the few of patients, mild paravalvular leak was in 13 patients on the 30th day. Two patients showed moderate paravalvular leak. Left ventricular end-diastolic volume decreased from 197.7±66.8 mL (pre-TAVI) to 147.2±53.3 mL (30-day post-TAVI) (P<0.05). Mean pressure gradient was 9.5±4.1 mm Hg (30-day post-TAVI). Conclusion     This multicenter study demonstrates that TAVI with the J-Valve system for the treatment of pure aortic regurgitation is associated with sustained clinical and functional cardiovascular benefits in high-risk patients with symptomatic aortic regurgitation early-term follow-up. Our results further support that TAVI with the specific designed J-Valve system is an acceptable alternative therapy for high-risk patients with pure AR. Our result demonstrates good early-term durability and preserved hemodynamic function. The procedure appears to offer an adequate and lasting resolution for selected patients with pure aortic regurgitation.

Ginsenoside Rg1 (Rg1), the major effective component of ginseng, has been shown to have multiple bioactivities, but low oral bioavailability. The aim of this study was to develop a simple, sensitive and rapid high performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) method, which could be used to validate and quantify the concentrations of Rg1 and its metabolites in Sprague-Dawley rat bile, urine, and feces after oral administration (25 mg/kg). Calibration curves offered satisfactory linearity (>0.995) within the determined ranges. Both intra-day and inter-day variances were less than 15%, and the accuracy was within 80-120%. The excretion recoveries of Rg1, ginsenoside Rh1 (Rh1), and protopanaxatriol (Ppt) in bile, urine, and feces combined were all greater than 70%. The fecal excretion recoveries of Rg1, Rh1, and Ppt were 40.11%, 22.19%, and 22.88%, respectively, whereas 6.88% of Rg1 and 0.09% of Rh1 were excreted in bile. Urinary excretion accounted for only 0.04% of Rg1. In conclusion, the observed excretion profiles for Rg1 and its metabolites after oral administration are helpful for understanding the poor oral bioavailability of Rg1 and will aid further investigations of Rg1 as a pharmacologically active component.