To determine the content of isopsoralen in Gushuling Capsule. High performance liquid chromatography was performed on C 18 column. The chromatographic conditions were as follows: methanol-water (48∶52) as mobile phase, flow rate being 1.1?mL/min and the detecting wavelength at 245?nm. A good linearity of isopsoralen was shown in the range of 0.0342～0.2736??g, r=0.9998. The average recovery is 92.11%, RSD=1.89% (n=5). The method is with good reproducibility,RSD=2.05%.[Conclusion]This method can supply evidence for the quality control of Gushuling Capsule.

OBJECTIVETo establish an assay method for detecting the migration of transferred apoptotic cells into the recipient using flow cytometry.

METHODSSpleen lymphocytes were isolated and labeled with an intracellular amine dye, carboxyfluorescein diacetate succinimidyl ester (CFSE), to allow discrimination. The labeled cells were induced with dexamethasone to undergo apoptosis and transferred into recipient mice via tail venous transfusion. Flow cytometry and histological examination of different tissues were performed at different time points. The stability of CFSE labeling for apoptotic cells was also tested.

RESULTSThe CFSE-labeled apoptotic cells were highly fluorescent with a positive labeling rate of (98.0+/-1.9)%. The stability of CFSE-labeling was testified, and the CFSE-labeled apoptotic cells entering different tissues at different time points were detected by flow cytometry and verified by histological examination.

CONCLUSIONFlow cytometry using CFSE labeling is reliable, sensitive, precise and convenient for apoptotic cell tracing in vivo and in vitro.

Adoptive Transfer ; methods ; Animals ; Apoptosis ; Dexamethasone ; pharmacology ; Female ; Flow Cytometry ; methods ; Fluoresceins ; chemistry ; pharmacokinetics ; Fluorescent Dyes ; chemistry ; pharmacokinetics ; Lymphocytes ; chemistry ; cytology ; drug effects ; Mice ; Mice, Inbred BALB C ; Reproducibility of Results ; Spleen ; cytology ; Succinimides ; chemistry ; pharmacokinetics

Objective To observe apelin and its receptor (APJ) expressions in human osteoblasts and evaluate the effect of apelin on osteoblasts.Methods The expressions of apelin and APJ in human osteoblasts were tested by RT-PCR and Western blot.After human osteoblasts were treated with apelin,cell proliferation was measured by [~3H] thymidine incorporation and cell counting.Cell function was measured by alkaline phosphatase (ALP) activity,the secreted osteocalcin level and typeⅠcollagen production .The activation of signaling cascades was tested by Western blot.Small-interfering RNA (siRNA) to blockade APJ was applied to observe effects of apelin on cell proliferation and the activation of signaling cascades.Results Both apelin and APJ were expressed in human osteoblasts.Apelin increased the proliferation and did not show the influences on ALP activity, osteocalcin secretion and type I collagen production in human osteoblasts.Apelin induced activation of phosphatidylinositol-3 kinase (PI3K) downstream effector (Akt),but not mitogen-activated protein kinase (MAPK) such as c-jun N-terminal kinase (JNK),p38 and ERK1/2 in human osteoblasts.Suppression of APJ with siRNA or LY294002 (PI3K inhibitor) abolished the apelin-induced cell proliferation and the activation of Akt.Conclusion Human osteoblasts express apelin and APJ.Apelin stimulates the proliferation of human osteoblast via APJ/PI3K/Akt pathway,but has no effect on osteoblast differentiation.

OBJECTIVE: To investigate the effect of two core binding factors alpha 1 (Cbfa1) isfroms (Cbfa1/P56 and Cbfa1/P57) on the apoptosis of mesenchymal cell line MBA-1. METHODS: The two Cbfal isfroms were transiently transfected into MBA-1 cells, then the changes of apoptosis rate were observed by flow cytometer. The protein expressions of Cbfa1, Bax, Bcl-2, caspase-3, and caspase-9, cytochrome-C and TNF-alpha were determined by Western immunoblot. RESULTS: After the transient transfection with the two isforms of Cbfa1, MBA-1, the cells apoptotic rates increased, and the ratio of Bax/Bcl-2, the expressions of cytochrome-C, caspase-3, caspase-9, and TNF-alpha were significantly increased. CONCLUSION Cbfa1 can promote the apoptosis in mesenchymal cell line MBA-1. Bax/Bcl-2, cytochrome-C, caspase-9, caspase-3, and TNF-alpha are also involved in the apoptosis pathway.
Animals ; Apoptosis ; physiology ; Bone Marrow Cells ; cytology ; Caspase 9 ; Caspases ; metabolism ; Cell Line ; Core Binding Factor alpha Subunits ; pharmacology ; Cytochromes c ; metabolism ; Mesenchymal Stem Cells ; cytology ; Mice ; Protein Isoforms ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; bcl-2-Associated X Protein ; metabolism

BACKGROUNDBoth repaglinide and gliclazide are insulin secretagogues widely used in the treatment of type 2 diabetes. They stimulate insulin secretion through distinct mechanisms and may benefit patients from different aspects. The present study was to evaluate the effects of repaglinide or gliclazide on glycaemic control, insulin secretion, and lipid profiles in type 2 diabetes patients.

METHODSA total of 47 newly diagnosed type 2 diabetes patients were randomized 1:1 to receive a 4-week treatment with repaglinide or gliclazide. The standard mixed meal tolerance test was performed before and after the treatment. Plasma glucose (PG), insulin concentration, and lipid profiles were measured. The area under insulin concentration curve (AUC(ins)) and the early-phase insulin secretion index (ΔI(30)/ΔG(30)) were calculated.

RESULTSAfter the trial, fasting and postprandial PG and postprandial insulin improved significantly in both groups (P < 0.05). The maximum insulin concentration occurred earlier in the repaglinide group than that in the gliclazide group. AUC(ins) increased in both groups (P < 0.05), but no significant difference was found between groups. ΔI(30)/ΔG(30) increased in both groups (P < 0.05), especially in the repaglinide group (P < 0.05). Triglyceride and total cholesterol decreased significantly in the repaglinide group in some time points, while no significant change was observed in the gliclazide group.

CONCLUSIONSRepaglinide and gliclazide had similar effects on glycaemic control and total insulin secretion, while repaglinide had more effects on improvements in β-cell function and lipid metabolism.

Adult ; Blood Glucose ; drug effects ; Carbamates ; therapeutic use ; Cholesterol ; blood ; Diabetes Mellitus, Type 2 ; blood ; drug therapy ; metabolism ; Fasting ; blood ; Female ; Gliclazide ; therapeutic use ; Humans ; Hypoglycemic Agents ; therapeutic use ; Insulin ; secretion ; Male ; Middle Aged ; Piperidines ; therapeutic use ; Postprandial Period ; drug effects ; Treatment Outcome ; Triglycerides ; blood

A simple, sensitive and reliable method was developed for simultaneous determination of ten banned drugs residues including zeranols(ZALs),chloroamphenicol,pentachlorophenol,etc. in swine urine using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The urine samples were pretreated using lyophilization and QuEChERS procedures, respectively. Acetonitrile and ammonium acetate (5 mmol/L) were chosen as mobile phases. Target compounds were separated well in ZorbaxSB-C18by following the optimized gradient elution program and determined by LC-MS/MS in negative electrospray ionization mode. The linearity of the matrix-matched standard curve of ten analytes in two methods was good in the range of the experimental concentration with correlation coefficients more than 0.99. The recoveries of ten drugs were in the range of 80.7%-107.7% and 73.5%-103.3% at the spiked levels of 5,10 and 20 μg/L by lyophilization and QuEChERS methods,respectively. The coefficients of variation were less than 15%. The limits of detection (LOD) and the limits of quantification (LOQ) from lyophilization and QuEChERS method were 0.1 to 2.0 μg/L and 0.2 to 5.0 μg/L,respectively.

Hematopoietic stem cells (HSC) maintain homeostatic hematopoiesis via their multi-lineage differentiation and self-renewal potentials. HSC can be enriched and purified by flow cytometry relying on their cell surface markers and functional characteristics, however, these methods can not meet the need for deep analysis of HSC biological property and function because of the poor purity. Recent studies have successfully purified and tracked HSC using specifically expressed genes, which can enhance the purification efficiency of HSC, thus provide a better tool for the in-vivo study of HSC. This review summarizes the new techniques and discusses their advantages and disadvantages.
Cell Differentiation ; Flow Cytometry ; Gene Expression ; Hematopoiesis ; Hematopoietic Stem Cells

To study the quality control of three traditional Chinese medicines derived from Gleditsia sinensis [Gleditsiae Sinensis Fructus(GSF), Gleditsiae Fructus Abnormalis(GFA), and Gleditsiae Spina(GS)], this paper established a multiple reaction monitoring(MRM) approach based on ultra-high performance liquid chromatography-triple quadrupole-linear ion-trap mass spectrometry(UHPLC-Q-Trap-MS). Using an ACQUITY UPLC BEH C_(18) column(2.1 mm × 100 mm, 1.7 μm), gradient elution was performed at 40 ℃ with water containing 0.1% formic acid-acetonitrile as the mobile phase running at 0.3 mL·min~(-1), and the separation and content determination of ten chemical constituents(e.g., saikachinoside A, locustoside A, orientin, taxifolin, vitexin, isoquercitrin, luteolin, quercitrin, quercetin, and apigenin) in GSF, GFA, and GS were enabled within 31 min. The established method could quickly and efficiently determine the content of ten chemical constituents in GSF, GFA, and GS. All constituents showed good linearity(r>0.995), and the average recovery rate was 94.09%-110.9%. The results showed that, the content of two alkaloids in GSF(2.03-834.75 μg·g~(-1)) was higher than that in GFA(0.03-10.41 μg·g~(-1)) and GS(0.04-13.66 μg·g~(-1)), while the content of eight flavonoids in GS(0.54-2.38 mg·g~(-1)) was higher than that in GSF(0.08-0.29 mg·g~(-1)) and GFA(0.15-0.32 mg·g~(-1)). These results provide references for the quality control of G. sinensis-derived TCMs.
Flavonoids/analysis* ; Alkaloids ; Chromatography, High Pressure Liquid/methods* ; Mass Spectrometry ; Drugs, Chinese Herbal