Effects of PINK1 mediated mitophagy on senescence and function of rat bone marrow endothelial progenitor cells / 中国药理学通报

Aim To investigate the effects of PTEN-induced putative kinase1(PINK1)mediated mitophagy on senescence and function of rat bone marrow endothelial progenitor cells(EPCs)by using small interfering RNA(siRNA)technology to knock down the PINK1 gene in rat bone marrow EPCs.Methods EPCs from bone marrow in rats were isolated, cultured and identified.After counting, EPCs were randomlydivided into control group, negative control group(NC siRNA), and Pink1 transfection group(PINK1 siRNA).The expression of PINK1 mRNA and protein in cells in various groups were detected by qRT-PCR and Western blot.At the same time, different time points were chosen to simulate the aging process based on the best knock down time.The senescence of cells was detected by SA-β-galactosidase staining and p16 protein expression.The function of cell proliferation, migration and tubule formation was detected by CCK-8, Transwell chamber and in vitro angiogenesis kit.ROS level was detected by flow cytometry.The expressions of PINK1, Parkin, LC3, and p62 were detected by Western blot.Mitochondria and autophagosomes were observed by transmission electron microscope.Results 48 h after PINK1 siRNA transfected, PINK1 was effectively knocked down.Compared with control group, the positive rate of blue staining and the expression of p16 protein in PINK1 siRNA group increased significantly 48 h and 96 h after transfection.The function of cell proliferation, migration and tubule formation decreased significantly.The level of ROS increased significantly, while the expression of PINK1, Parkin and LC3 protein decreased significantly, and p62 protein expression increased significantly.Under the transmission electron microscope, the mitochondria swelled and denatured, and the number of autophagosomes decreased in the PINK1 siRNA group.Conclusions The down-regulation of PINK1 gene can aggravate the senescence of EPCs, and PINK1 mediated mitophagy may participate in the regulation of senescence and function of EPCs.

Comparison of the Concentrations of Stiripentol and Its Self-nanoemulsifying Drug Delivery System in Plasma and Brain of Rats / 中国药房

OBJECTIVE：To establish a method for de termining stiripentol （STP）concentration in plasma and brain of rats ， and to compare the concentrations of STP and its self-nanoemulsifying drug delivery system （STP-SNEDDS）in plasma and brain. METHODS：Using xanthone as internal standard ，HPLC-fluorescence（HPLC-FLR）method was adopted. The determination was performed on Diamonsil C 18 column with mobile phase consisted of acetonitrile- 25 mmol/L KH 2PO4 solution [ 44 ∶ 56（V/V），pH 2.6] at a flow rate of 1.5 mL/min；the excitation and emission wavelengths were 210 nm and 400 nm，respectively；the column temperature was 30 ℃；the sample size was 10 μL. Totally 36 rats were randomly divided into 2 groups，with 18 rats in each group. They were given STP-SNEDDS and STP suspension （40 mg/kg，by STP ）intragastrically. Blood and brain tissue samples were collected at 0.5，1，2 h after administration （6 rats in each group at different time point ）. After the protein was precipitated by acetonitrile （brain tissue should be homogenized ），the concentrations of STP were determined by the above chromatographic conditions. RESULTS ：The linear ranges of STP concentration in plasma and brain tissue were 0.02-8.00 μg/mL（r were 0.999 6， 0.999 4，respectively）. The limits of quantitation were both 0.02 μg/mL. The inter-day and intra-day RSDs were all less than 5％. The extraction recovery and method recovery were all no less than 90％. Compared with STP suspension group ，the plasma concentration（except for 1 h after administration ）and cerebral concentration （except for 2 h after administration ）of STP in STP-SNEDDS group were all significantly increased （P＜0.05），showing significant linear relationship between them （for STP-SNEDDS）. CONCLUSIONS ：Established HPLC-FLR method presents high accuracy and precision ，and can be used for the distribution of STP and STP-SNEDDS in plasma and brain.The concentration of STP in plasma and brain tissue isincreased after STP is made into SNEDDS.

Preparation of curcumin TPP-PEG-PE nanomicelles with mitochondrial targeting and lysosomal escape functions and its effect on promoting breast cancer cell apoptosis / 中国中药杂志

Orthogonal experiments were used to optimize the process parameters of curcumin TPP-PEG-PCL nanomicelles; the particle size, electric potential and morphology under the electron microscope were systematically detected for the curcumin TPP-PEG-PCL nanomicelles; and the stability and in vitro release of the curcumin TPP-PEG-PCL nanomicelles were investigated. With DID fluorescent dye as the fluorescent probe, flow cytometry was used to study the uptake of nanomicelles by breast cancer cells, and laser confocal microscopy was used to study the mitochondrial targeting and lysosomal escape functions of nanomicelles. Under the same dosage conditions, the effect of curcumin TPP-PEG-PCL nanomicelles on promoting the apoptosis of breast cancer cells was evaluated. The optimal particle size of curcumin TPP-PEG-PCL nanomicelle was(17.3±0.3) nm, and the Zeta potential was(14.6±2.6) mV in orthogonal test. Under such conditions, the micelle appeared as regular spheres under the transmission electron microscope. Fluorescence test results showed that TPP-PEG-PCL nanomicelles can promote drug uptake by tumor cells, escape from lysosomal phagocytosis, and target the mitochondria. The cell survival rate and Hoechst staining positive test results showed that curcumin TPP-PEG-PCL nanomicelles had a good effect on promoting apoptosis of breast cancer cells. The curcumin TPP-PEG-PCL micelles can significantly reduce the mitochondrial membrane potential of breast cancer cells, increase the release of cytochrome C, significantly increase the expression of pro-apoptotic protein Bcl-2 and reduce the expression of anti-apoptotic Bax protein. These test results were significantly better than those of curcumin PEG-PCL nanomicelles and curcumin, with statistically significant differences. The results revealed that curcumin TPP-PEG-PCL nanomicelles can well target breast cancer cell mitochondria and escape from the lysosomal capture, thereby enhancing the drug's role in promoting tumor cell apoptosis.

PreoperativePredictionoftheExpressionofCK19inHepatocellularCarcinomaby Gd-EOB-DTPAEnhancedMRI / 中山大学学报(医学科学版)

@#【Objective】 To investigate the predictive value of preoperative Gd- EOB- DTPA enhanced MRI in the expression of cytokeratin 19（CK19）in hepatocellular carcinoma（HCC）.【Methods】A total of 102 patients，including 94 male and 8 female，with single HCC confirmed by pathology after operation who underwent preoperative enhanced MRI were retrospectively analyzed. A total of 25 were CK19-positive HCC and 77 were CK19-negative HCC. Two radiologists evaluated MR features including tumor size，tumor margin，intratumoral vessels，signal intensity（SI）on arterial phase （AP） ，enhancement pattern ，arterial rim enhancement ，peritumoral enhancement ，internal cystic or necrotic portion，hemorrhage，intratumoral fat，tumor capsule，vascular invasion，lymph node metastasis，intratumoral septum， target sign on diffusion weighted imaging（DWI）or hepatobiliary phase（HBP），peritumor hypointensity，SI on ADC，SI on HBP ，T1 relaxation times and T1 reduction rate between pre- and post- contrast enhancement. The associations between these imaging features and CK19 expression were investigated. 【Results】SI on AP（P = 0.013），arterial rim enhancement（P = 0.018），target sign on DWI（P = 0.001）and target sign on HBP（P = 0.005）were significantly associated with CK19 expression. Delayed enhanced intratumoral septum（P = 0.042）was associated with CK19 expression between HCCs less than 5 cm. Target sign on DWI（P = 0.001，OR = 4.875，95%CI：1.838~12.927）were independent significant factors of CK19- positive HCC.【Conclusion】Preoperative enhanced MRI with Gd- EOB- DTPA is helpful to predict CK19 expression of HCC.

A de novo GJA1 mutation identified by whole-exome sequencing in a patient with oculodentodigital dysplasia / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To explore the genetic basis for a patient with oculodentodigital dysplasia.</p><p><b>METHODS</b>Genomic DNA was extracted from peripheral blood samples from the patient and his parents. Whole-exome sequencing was carried out for the trio family. Suspected mutation was verified by Sanger sequencing.</p><p><b>RESULTS</b>A de novo c.412G>A mutation of the GJA1 gene was identified in the patient, which was validated by Sanger sequencing.</p><p><b>CONCLUSION</b>The c.412G>A mutation of the GJA1 gene probably underlies the disease in the patient.</p>

Effect and cost analysis of two kinds of transparent dressings on the central venous maintenance / 现代临床护理

Objective To compare the effects and medical cost of 2 kinds of transparent dressings on the central venous catheter maintenance. Methods Selecting 75 patients with single lumen central venous catheter in the Geriatrics and ICU of the Tertiary Hospital during April to June 2017,according to the number of puncture, 38 cases (odd numbers)with bordered transparent dressings were selected as the experimental group, and 37 cases (even numbers) with transparent dressings were selected as control group. The dressings were changed weekly,and will be changed if there's rolling edge, dermatitis, seepage and wet. Unplanned tube drawing after 28 days, adverse reaction in the site of puncture, unwell fixed condition of dressings and the related medical costs was evaluated.Results The incidence rate of loosing dressings and dermatitisin the experimental group is significantly lower than those in the control group(P<0.05).The direct cost,indirect cost and total cost in experimental group were significantly lower than those in control group(P<0.05). Conclusion Bordered transparent dressings can promote the effect of tube fixing and decrease nursing manpower cost and medical cost in central venous catheter maintenance,which can be popularized in clinics.

Application of perimembranous ventricular septal defects closure solely by femoral vein approach under transesophageal echocardiography guidance / 中南大学学报(医学版)

Objective:To investigate the feasibility and safety of perimembranous ventricular septal defects (PmVSD) closure solely by femoral vein approach under transesophageal echocardiography (TEE) guidance.Methods:From January 1,2014 to May 31,2016,26 patients with PmVSD in Second Xiangya Hospital were selected,with age at 3.2-6.0 (4.3±0.7) years old and body weight at 15.0-19.5 (16.7±1.4) kg.The diameter of VSD was 3.5-4.8 (4.1±0.3) mm.All patients were treated by percutaneous PmVSD closure solely by femoral vein approach under TEE guidance.The effect of the procedure was evaluated by TEE and transthoracic echocardiography (TTE).The clinical follow-up study was conducted by TTE at 1,3,6 and 12 month (s) after the procedure.Results:Twenty cases were successfully treated with percutaneous PmVSD closure solely by femoral vein approach under TEE guidance,and the success rate was 76.9％.Six patients were converted to perventricular closure under TEE guidance because the guide wire in two cases or catheter in other cases could not pass through PmVSD.The diameter of symmetrical VSD occluder was 6.0-7.0 (6.2±0.4) mm.The procedural time was 12.0-64.0 (26.8±6.3) min.The residence time at ICU was 1.8-2.4 (26.8±6.3) h.The in-hospital time was 4.0-5.0 (4.4±0.5) d.There were 3 patients with immediate post-operative trivial residual shunt and incomplete right bundle branch block (IRBBB).All patients survived with no peripheral vascular injury or complications such as tricuspid regurgitation,pericardial tamponade and pulmonary infection.The residual shunt disappeared in 3 patients and IRBBB became normal rhythm in 3 patients at 1 month follow-up time point.No patients suffered from occluder malposition,residual shunt,pericardial effusion,arrhythmia (atrio-ventricular block),aortic valve regurgitation and tricuspid regurgitation.Conclusion:TEE-guided percutaneous PmVSD closureby femoral vein approach is safe and effective.

Pediatric drugs in China:problems and countermeasures / 国际药学研究杂志

Taking the differences between the pediatric drugs and adult drugs as the starting-point,this article analyzes the main problems of the pediatric drugs in China,such as the shortage of the pediatric drugs. The causes of the problems mainly include high research and development cost of the pediatric drugs,their limited market capacity,difficulty in recruiting trial participants,the less clinical trial institutions in pediatrics,and governments′incentive polices on research and development of pediatric drugs did not really play their roles. The countermeasures to solve these problems mainly include introducing relevant policy to guarantee the re?search and development companies′profit,selecting appropriate varieties of the pediatric drugs to research and develop,promoting the establishment of pediatric clinical trial institutions as soon as possible,and establishing particular passage for pediatric drugs ap?proval,in order to provide references for the government when making policies on pediatric drugs.

Establishment of a NOD/SCID mouse model with human immune reconstitution bearing human triple-negative breast cancer / 南方医科大学学报

<p><b>OBJECTIVE</b>To establish a NOD/SCID mouse model with human immune reconstitution and observe its immune response to human triple-negative breast cancer xenograft.</p><p><b>METHODS</b>Twenty-four NOD/SCID mice without immune leakage were subjected to cyclophosphamide (CTX) treatment 3 days prior to immune reconstitution with human peripheral blood mononuclear cell (PBMC) injection and subcutaneous transplantation of human triple-negative breast cancer MDA-MB-231 cells, CTX treatment and PBMC injection without tumor cell transplantation, MDA-MB-231 cell transplantation only, or no treatments. The tumor growth and immune responses of the mice were observed at regular intervals.</p><p><b>RESULTS</b>Compared with the tumor-bearing mice, the tumor-bearing mice with immune reconstitution showed prolonged incubation period of tumor formation, slower tumor growth rate and increased survival rate. Human IgG and CD3(+) T cells were detected in the peripheral blood of the mice 1 week after human PBMC injection. The percentage of CD3(+) T cells in the spleen cells was 55.3% at 9 weeks in tumor-bearing mice with immune reconstitution and 52.7% in tumor-bearing mice without immune reconstitution. The spleen index of the tumor-bearing mice with immune reconstitution was much higher than that in mice with only immune reconstitution and the control mice (9.64 vs 3.82∓0.31 and 1.51∓0.14 mg/g).</p><p><b>CONCLUSION</b>A stable NOD/SCID mouse model with immune reconstitution has been established successfully, which shows immune responses to triple-negative breast cancer xenografts and allows studies of immunological therapy study of triple-negative breast cancer.</p>

Determination of 10 Residual Organic Solvents in Norvancomycin Hydrochloride Raw Material by HS-GC / 中国药房

OBJECTIVE:To establish the method for determining 10 residual organic solvents in norvancomycin hydrochloride raw material. METHODS:Headspace gas chromatography was performed on the column of nitro modified polyethylene terephthal-ate glycol as stationary phase capillary column;the oven temperature program started at 40 ℃ for 3 min and increased at a rate of 8 ℃/min up to 150 ℃ for 10 min;the temperature was 200 ℃ with carrier gas of high-purity nitrogen gas,the constant flow rate was 5 ml/min with split ratio of 15∶1;the headspace vial equilibrium temperature was 85 ℃ with equilibrium time of 40 min,and the volume was 1 ml. RESULTS:The concentration of n-pentane,acetone,ethanol,benzene,acrylonitrile,toluene,xylene,chlo-robenzene,styrene,divinylbenzene had good linear relationship with its peak area values(r=0.995 7-0.999 9);the RSDs of preci-sion,repeatability tests was ≤6.6%;average recovery was in the range of 94.3%-106.6%(RSD=0.5%-4.5%,n=9). CONCLU-SIONS:The method is fast,sensitive and accurate,and can be used for the determination of residual organic solvents in norvanco-mycin hydrochloride raw material.

Effects of paeoniflorin on cerebral blood flow and the balance of PGI2/TXA2 of rats with focal cerebral ischemia-reperfusion injury / 药学学报

This study is to investigate the effects of paeoniflorin on cerebral blood flow and the balance of PGI2/TXA2 of rats with focal cerebral ischemia-reperfusion injury. A total of 72 SD rats (3) were randomly divided into 6 groups: sham operation group, cerebral ischemia-reperfusion model group (I/R gourp), low (10 mg.kg-1), middle (20 mg.kg-1) and high (40 mg.kg-1) doses of paeoniflorin groups and nimrnodipine group. Focal cerebral ischemia in rats was made by inserting a monofilament suture into internal carotid artery for 90 min and then reperfused for 24 h. The effects of paeoniflorin on neurological deficit scores and the infarction volume of brain were detected. Relative regional cerebral blood flow (rCBF) was continuously monitored over ischemic hemispheres by laser-Doppler flowmetry (LDF). The expression of COX-2 in hippocampal CAl region was estimated by immunohistochemistry and the contents of prostacyclin I2 (PGI2), thromboxane A2 (TXA2), and ratio of PGIJ2/TXA2 in serum were measured by ELISA kits. Paeoniflorin significantly ameliorated neurological scores, reduced the infarction volume, and increased regional cerebral blood flow relative to the I/R group. In addition, paeoniflorin could inhibit COX-2 expression and the release of TXA2 and prevent the downregulation of PGI2 induced by I/R injury. The neuroprotective effects of paeoniflorin against focal cerebral ischemia-reperfusion rats might be attributed to improve the supply of injured hemisphere blood flow and adjust the balance between PGI2/TXA2.

Effects of paeoniflorin on cerebral blood flow and the balance of PGI2/TXA2 of rats with focal cerebral ischemia-reperfusion injury / 药学学报

This study is to investigate the effects of paeoniflorin on cerebral blood flow and the balance of PGI2/TXA2 of rats with focal cerebral ischemia-reperfusion injury. A total of 72 SD rats (3) were randomly divided into 6 groups: sham operation group, cerebral ischemia-reperfusion model group (I/R gourp), low (10 mg.kg-1), middle (20 mg.kg-1) and high (40 mg.kg-1) doses of paeoniflorin groups and nimrnodipine group. Focal cerebral ischemia in rats was made by inserting a monofilament suture into internal carotid artery for 90 min and then reperfused for 24 h. The effects of paeoniflorin on neurological deficit scores and the infarction volume of brain were detected. Relative regional cerebral blood flow (rCBF) was continuously monitored over ischemic hemispheres by laser-Doppler flowmetry (LDF). The expression of COX-2 in hippocampal CAl region was estimated by immunohistochemistry and the contents of prostacyclin I2 (PGI2), thromboxane A2 (TXA2), and ratio of PGIJ2/TXA2 in serum were measured by ELISA kits. Paeoniflorin significantly ameliorated neurological scores, reduced the infarction volume, and increased regional cerebral blood flow relative to the I/R group. In addition, paeoniflorin could inhibit COX-2 expression and the release of TXA2 and prevent the downregulation of PGI2 induced by I/R injury. The neuroprotective effects of paeoniflorin against focal cerebral ischemia-reperfusion rats might be attributed to improve the supply of injured hemisphere blood flow and adjust the balance between PGI2/TXA2.

Expression and gene mutation of phospho-platelet derived growth factor receptor alpha and C-kit in gastrointestinal and extra-gastrointestinal stromal tumors / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To investigate the expression of phospho-platelet derived growth factor receptor alpha (P-PDGFR-alpha) in gastrointestinal stromal tumors (GIST) and extra-gastrointestinal stromal tumors (EGIST) and its clinical significance.</p><p><b>METHODS</b>Expression of P-PDGFR-alpha in 28 samples of positive CD117 and 13 samples of negative CD117 was detected by Envision immunohistochemical staining. Direct PCR sequencing was used to investigate the mutation status of c-kit gene exons 9, 11, 13, 17 and PDGFR-alpha gene exons 12 and 18.</p><p><b>RESULTS</b>The positive rate of P-PDGFR-alpha expression in GISTs with negative CD117 was 69.2%, which was significantly higher than that in GISTs with positive CD117 (7.1%, P<0.05). The positive rates of P-PDGFR-alpha expression in epithelioid GISTs(27.3%) and mixed GIST(63.3%) were both significantly higher than that in fusiform GISTs (9%, P<0.05). The positive rate of CD117 expression in fusiform GISTs (53.6%)was significantly higher than that in epithelioid GISTs (7.1%) and mixed GISTs(39.3%, P<0.05). C-kit gene mutation was found in 19 GIST cases with positive CD117. C-kit gene mutation was found in 19 of 28 GIST patients with positive CD117, among them, mutation of exon 11 occurred in 15 cases and exon 13 in 4 cases. No C-kit gene mutation was seen in 13 GIST patients with negative CD117. PDGFR-alpha gene mutation was found in 4 of 11 GIST cases with positive P-PDGFR-alpha and all occurred in exon 18.</p><p><b>CONCLUSIONS</b>Examination of P-PDGFR-alpha expression may provide reliable evidence for the further improvement of pathological diagnosis,pathological typing and treatment for GISTs with negative CD117. Phosphorylated protein induced by PDGFR-alpha mutation may be associated with the important alternative molecular mechanism and the biological behavior of GIST development.</p>