A fast and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the determination of prodrug of paclitaxel (Pro-PTX) and paclitaxel (PTX) in rat plasma was developed. The plasma samples were subjected to protein precipitation with acetonitrile (0.1% formic acid), and then separated by LC with an Ultimate AQ-C18 column (50 mm × 3.0 mm, 3 μm) and acetonitrile-1 mmol·L^-1 ammonium formate (containing 0.1% formic acid) as the mobile phase. Multiple reaction monitoring (MRM) scanning mode was used to detect the ion responses m/z 983.4→415.2 (Pro-PTX), m/z 854.4→286.1 (PTX) and m/z 808.3→527.2 (Docetaxel, internal standard) by using a triple quadrupole tandem mass spectrometer with electrospray ionization source and positive ion mode. The method validation results show that the linear ranges of Pro-PTX and PTX in plasma were 2.00-500 ng·mL^-1 (r > 0.99) and 4.00-1 000 ng·mL^-1 (r > 0.99), the lower limit of quantification was 2.00 ng·mL^-1 and 4.00 ng·mL^-1, respectively; the quality control samples with low, medium and high concentrations of Pro-PTX and PTX were within the batch, the relative standard deviation (RSD) between batches were all less than 9%; the relative deviation (RE) was within the range of ± 9%; In the stability test, both Pro-PTX and PTX in plasma were stable under various storage conditions. The method was sensitive, specific, and reproducible, and was suitable for the pharmacokinetic study of Pro-PTX in rats. Animal experiments were approved by the Ethics Committee of Laboratory Animal Management and Animal Welfare, Institute of Materia Medica, Chinese Academy of Medical Sciences (No.: 00003552).

Objective To assess the clinical safety and feasibility for ultrasound guided paravertebral block anesthesia of percutaneous nephrolithotomy.Methods Between December 2015 to June 2016,180 patients with renal or ureteral calculi were enrolled and evaluated with uhrasonography and CT scan.Of all the 180 patients,108 males and 82 females.Their mean age was 39 years (23-71 years).The clinical characteristics of the patients in each group,such as age,gender,BMI index,ASA status,mean arterial pressure and disease type had no significant differences (P ＞ 0.05).These patients were randomized into general anesthesia group (G group),combined spinal epidural anesthesia group (C group) and paravertebral nerve block anesthesia group (P group).G group:35 males and 25 females.Their mean age was (40.1 ± 11.8) years and BMI was (25.1 ± 3.8) kg/m2;Renal calculi 52 cases,ureteral calculi 8 cases,Average maximum stone diameter was (2.6 ± 0.8)cm.C group:38 males and 22 females.Their mean age was (39.7 ± 12.4) years and BMI was (24.6 ± 4.1) kg/m2;Renal calculi 54 cases,ureteral calculi 6 cases,Average maximum stone diameter was (2.4 ± 0.8) cm.P group:35 males and 25 females.Their mean age was (38.9 ± 12.7) years and BMI was (25.4 ± 4.0) kg/m2;Renal calculi 51 cases,ureteral calculi 9 cases,Average maximum stone diameter was (2.5 ± 0.7) cm.Periprocedural Vital signs,complications,the times of anal discharging gas and postoperative feeding,hospitalized day and expense in these three groups were evaluated.Results Major intraoperative or postoperative complications did not occur in all of the patients.Mean arterial pressure decreased during preoperative changing positions was observed in group G (mean decreased 8.8 mmHg)and group C (mean decreased 1.9 mmHg),with significant difference in intra-group (P ＜ 0.05).Postoperative nausea and vomiting was observed in 8 and 2 patients of group G and group P,respectively (P ＜ 0.05).Postoperative pain was observed in 2 and 7 patients of group C and group P,respectively (P ＞ 0.05).In addition,group P had early post operation feeding time [(6.4 ± 2.4) h],shorter hospitalized day [(4.5 ± 1.1) d] and lower hospitalized expense compared with other groups (P ＜ 0.05).Conclusions Ultrasound guided paravertebral block can provide safe and reliable surgical anesthesia for percutaneous nephrolithotomy.

Objective Microenvironment plays important roles in the proliferation , viability, and apoptosis of tumor cells. This study was to investigate the effects of different functional groups on the biological characteristics of the osteosarcoma Saos -2 cell line in vitro. Methods Using self-assembled monolayers of alkanethiols on gold , we prepared different terminal chemical groups , including methyl (-CH3 ) , amino (-NH2 ) , hydroxyl (-OH) , and carboxyl (-COOH ) .We determined the similar density of different functional groups by contact angle measurement and x-ray photoelectron spectroscopy , and observed the effects of different functional groups on the adhesion , proliferation, viability, and apoptosis of the osteosarcoma Saos-2 cells by fluorescence microscopy , CCK-8 as-say, flow cytometry, and scan electron microscopy (SEM). Results The surface of -COOH and -NH2 promoted the adhesion and proliferation of the of the Saos-2 cells, with a good compatibility , while that of -CH3 was unfavorable for their adhesion and proliferation and even increased their apoptosis . The promoting effects of the functional groups on the adhesion and proliferation of the cells were listed in the following order: -COOH ≥ -NH2 >-OH -CH3 , while their toxicity and apoptosis-increasing effect ranked as -CH3 -OH >-NH2 >-COOH. Conclusion The-CH3 group inhibits the adhesion and proliferation and promotes the apoptosis of Saos-2 cells, which has provided some evidence for the surface design of biomaterials.

In this study, we explored the mechanism of Huganning tablet (HGNP) in the treatment of nonalcoholic fatty liver disease (NAFLD) based on network pharmacology and computer-aided drug design. Firstly, the potential ingredients and targets of HGNP were identified from TCMSP database, Swiss Target Prediction database, Chinese pharmacopoeia (2015) and literatures, and then the targets of HGNP intersected with NAFLD disease targets that obtained in GeneCards database to acquired potential targets. The bioconductor bioinformatics package of R software was used for gene ontology (GO) enrichment and Kyoto encyclopedia of genes and genomes (KEGG) enrichment analysis. The network of “potential ingredient-key target-pathway” was formed in Cytoscape software to study the interactions between potential ingredients of HGNP, key targets, pathways and NAFLD. Based on the results of network pharmacology, the molecular docking analysis of the key targets and potential active ingredients in HGNP tablets with top degree in the network was conducted using Discovery Studio 2020 software, followed by molecular dynamics simulations, binding free energy calculation, drug-likeness properties analysis and ADMET (absorption, distribution, metabolism, excretion and toxicity) properties prediction. In vitro, HepG2 cells were used to establish steatosis model, and the effects of five key compounds on hepatocyte steatosis were analyzed by oil red O staining and triglyceride (TG) content determination. The results showed that 141 ingredients and 151 potential targets were obtained. A total of 2 526 items and 151 pathways were identified by GO and KEGG enrichment analysis. The molecular docking suggested that five components, isorhamnetin, salvianolic acid B, emodin, resveratrol and rhein, exhibited strong binding ability with key targets [retinoic acid receptor RXR-alpha (RXRA), tumor necrosis factor (TNF), glycogen synthase kinase-3 beta (GSK3B), serine/threonine-protein kinase 1 (AKT1)]. It was further verified that isorhamnetin and salvianolic acid B bind to key targets with good structural stability and binding affinity based on molecular dynamics simulations and binding free energy calculations. The drug-likeness properties, pharmacokinetic properties and toxicity of five key compounds were more comprehensively analyzed through drug-likeness properties analysis and ADMET properties prediction. In vitro, all five compounds, isorhamnetin, salvianolic acid B, emodin, resveratrol, and rhein, improved hepatocyte steatosis of HepG2 cells, confirming the reliability of the present study. In conclusion, based on network pharmacology, computer-aided drug design and in vitro validation, this study investigated the mechanism of HGNP for the treatment of NAFLD at multiple levels and provided a basis for its clinical application.

OBJECTIVETo evaluate the value of a new platelet function test PFA P2Y (PFA-200) in monitoring clopidogrel treatment for cardiovascular disease in elderly patients.

METHODSFifty-six elderly patients receiving clopidogrel therapy in the Department of Cardiology of General Hospital of PLA from March to August in 2016 and 85 healthy volunteers were recruited for analysis. All the subjects underwent PFA P2Y, LTA (light transmittance aggregometry) and TEG (Thromboelastograph) tests, and Spearman correlation coefficients were used to test the associations between test results. The agreement among the 3 platelet function test methods was assessed using Cohen's kappa coefficient.

RESULTSCorrelation coefficient (r) was -0.701 (P<0.001) between PFA P2Y and LTA, and 0.475 (P<0.001) between PFA P2Y and TEG. The agreement was 75% between PFA P2Y and LTA and 67.9% between PFA P2Y and TEG. The κ value was 0.434 (P=0.001) between PFA P2Y and LTA and 0.242 (P=0.046) between PFA P2Y and TEG. With ADP-induced maximum platelet aggregation rate of LTA >50% as the laboratory clopidogrel resistance, the cut-off value of PFA P2Y was 119 s (AUC=0.733) with a sensitivity of 75.6% and a specificity of 73.3%.

CONCLUSIONPFA P2Y has a moderate correlation and agreement with LTA, but has a poor correlation and agreement with TEG. PFA P2Y can be useful for assessing the effects of clopidogrel therapy and the association of the cut-off value (119 s) with the long-term clinical ischemic events needs be confirmed in further study.

Biological Assay ; Blood Coagulation Tests ; Blood Platelets ; Cardiovascular Diseases ; drug therapy ; Humans ; Platelet Aggregation ; Platelet Aggregation Inhibitors ; therapeutic use ; Platelet Function Tests ; Sensitivity and Specificity ; Ticlopidine ; analogs & derivatives ; therapeutic use

Adult ; Female ; Humans ; Kidney Neoplasms ; diagnostic imaging ; metabolism ; Neuroectodermal Tumors ; diagnostic imaging ; metabolism ; Radiography ; Young Adult

OBJECTIVETo investigate the expression of KIAA0101 protein in gastric carcinoma cells, and to explore the effects of its down-regulation on the cell proliferation, cell cycle and invasion.

METHODSWestern blot was used to detect KIAA0101 protein expression in three gastric carcinoma cell lines including MKN-28, SGC-7901 and MKN-45. KIAA0101 siRNA and control siRNA were utilized to transfect MKN-45 cells, respectively. CCK-8 was used to analyze the changes of cell proliferation, and flow cytometry to examine the changes of cell cycle distribution. Finally, Boyden chamber was used to detect the ability of cell invasion.

RESULTSRelative level of KIAA0101 protein in MKN-45 cells was significantly higher than those in MKN-28 and SGC-7901 cells, and there was significant difference among the three cell lines (P < 0.05). The result of CCK-8 study demonstrated that, compared with untreated group and control siRNA group, the proliferation of MKN-45 cells in KIAA0101 siRNA group was significantly inhibited (P < 0.05). Additionally, the result of cell cycle analysis revealed that the percentage of cell number in G(0)/G(1) phase in KIAA0101 siRNA group [(61.47 ± 0.89)%] was significantly higher than those in untreated group [(47.43 ± 0.85)%] and control siRNA group [(48.43 ± 0.73)%; F = 271.653, P = 0.000]. Further, Boyden chamber assay showed that the cell numbers migrated to Matrigel in KIAA0101 siRNA group (61.51 ± 4.76) were significantly lower than those in untreated group (138.74 ± 10.16) and control siRNA group (132.93 ± 11.25; F = 65.949, P = 0.000).

CONCLUSIONSDown-regulation of KIAA0101 expression leads to an inhibition of cell proliferation, cell cycle and cell invasion. It may provide a novel target for the treatment of patients with gastric carcinoma.

Carrier Proteins ; genetics ; metabolism ; Cell Cycle ; Cell Line, Tumor ; Cell Movement ; Cell Proliferation ; Down-Regulation ; Gene Expression Regulation, Neoplastic ; Humans ; Neoplasm Invasiveness ; RNA Interference ; RNA, Small Interfering ; genetics ; Stomach Neoplasms ; metabolism ; pathology ; Transfection

OBJECTIVETo determine whether serum homocysteine and folate levels are correlated with the occurrence of neonatal asphyxia and to study the effects of gender and gestational age on serum homocysteine and folate levels.

METHODSThirty-five neonates with mild asphyxia (19 males and 16 females) and 40 normal neonates (control group,18 males and 22 females) were enrolled in this study. The asphyxia and the control groups consisted of 10 and 11 cases of preterm infants respectively. Serum homocysteine levels were measured using ELASA. Serum folate levels were measured using radioimmunity assay.

RESULTSSerum homocysteine level (14.66+/-2.61 micromol/L vs 7.55+/-0.50 mumol/L; P<0.05) was significantly higher and serum folate level (2.47+/-0.24 ng/mL vs 3.28+/-0.28 ng/mL; P<0.05) was significantly lower in the asphyxia group than that in the control group. There were no significant differences in serum levels of homocysteine and folate between males and females either in the asphyxia group or the control group. The asphyxiated neonates born at premature showed increased serum homocysteine level compared with the full-term neonates with asphyxia (21.25+/-5.01 micromol/L vs 12.34+/-2. 01 micromol/L; P<0.05).

CONCLUSIONSThe increased serum homocysteine level and decreased serum folate level are correlated with the occurrence of neonatal asphyxia. Serum homocysteine and folate levels are not associated with the gender. A more significantly increased serum homocysteine level may be found in asphyxiated neonates born at premature.

Asphyxia Neonatorum ; blood ; Female ; Folic Acid ; blood ; Homocysteine ; blood ; Humans ; Infant, Newborn ; Infant, Premature ; Male ; Sex Characteristics

OBJECTIVETo investigate the rebound depolarization of substantia gelatinosa (SG) neurons in rat spinal dorsal horn and explore its modulatory mechanisms to provide better insights into rebound depolarization-related diseases.

METHODSParasagittal slices of the spinal cord were prepared from 3- to 5-week-old Sprague-Dawley rats. The electrophysiologic characteristics and responses to hyperpolarization stimulation were recorded using whole-cell patch-clamp technique. The effects of hyperpolarization-activated cyclic nucleotide gated cation (HCN) channel blockers and T-type calcium channel blockers on rebound depolarization of the neurons were studied.

RESULTSA total of 63 SG neurons were recorded. Among them, 23 neurons showed no rebound depolarization, 19 neurons showed rebound depolarization without spikes, and 21 neurons showed rebound depolarization with spikes. The action potential thresholds of the neurons without rebound depolarization were significantly higher than those of the neurons with rebound depolarization and spikes (-28.7∓1.6 mV vs -36.0∓2.0 mV, P<0.05). The two HCN channel blockers CsCl and ZD7288 significantly delayed the latency of rebound depolarization with spike from 45.9∓11.6 ms to 121.6∓51.3 ms (P<0.05) and from 36.2∓10.3 ms to 73.6∓13.6 ms (P<0.05), respectively. ZD7288 also significantly prolonged the latency of rebound depolarization without spike from 71.9∓35.1 ms to 267.0∓68.8 ms (P<0.05). The T-type calcium channel blockers NiCl2 and mibefradil strongly decreased the amplitude of rebound depolarization with spike from 19.9∓6.3 mV to 9.5∓4.5 mV (P<0.05) and from 26.1∓9.4 mV to 15.5∓5.0 mV (P<0.05), respectively. Mibefradil also significantly decreased the amplitude of rebound depolarization without spike from 14.3∓3.0 mV to 7.9∓2.0 mV (P<0.05).

CONCLUSIONNearly two-thirds of the SG neurons have rebound depolarizations modulated by HCN channel and T-type calcium channel.

Action Potentials ; Animals ; Calcium Channel Blockers ; pharmacology ; Calcium Channels, T-Type ; Cell Polarity ; Cesium ; pharmacology ; Chlorides ; pharmacology ; Cyclic Nucleotide-Gated Cation Channels ; antagonists & inhibitors ; Neurons ; cytology ; Patch-Clamp Techniques ; Pyrimidines ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Spinal Cord Dorsal Horn ; cytology ; Substantia Gelatinosa ; cytology