AIMTo simultaneously determine three unconjugated neurosteroids, dehydroepiandrosterone (DHEA) , pregnenolone (PREG), allopregnenolone (AP), from several brain regions of the rat.

METHODSNeurosteroids were isolated separately in a two steps procedure by using ethyl acetate-n-hexane (90:10) as the first step to extract the unconjugated steroids, then the steroid fractions were further purified by SPE. All steroids were derivatized with 2-nitro-4-trifluoromethylphenylhydrazine (2-NFPH) and analyzed by HPLC-MS ( APCI) using selected-ion monitoring. Methyltestosterone was chosen as the internal standard. Results The linear calibration curve of DHEA was obtained in the concentration range of 0.030-2.00 microg x L(-1). The linear calibration curves of PREG and AP were obtained in the concentration range of 0.025-2.00 microg x L(-1). The concentrations of DHEA, PREG and AP in male rat brain regions were (0.70 +/- 0.23), (4.8 +/- 1.9), (1.1 +/- 0.6) ng x g(-1) for frontal cortex, (0.57 +/- 0.28), (6 +/- 3), (0.5 +/- 0.3) ng x g(-1) for hippocampus, (1.5 +/- 1.0), (9 +/- 5), (1.4 +/- 0.9) ng x g(-1) for amygdale, (0.52 +/- 0.14), (7.7 +/- 2.8), (0.5 +/- 0.6) ng x g(-1) for striatum, (2.9 +/- 1.6), (18 +/- 9), (1.6 +/- 1.3) ng x g(-1) for nucleus accumbens, (4.0 +/- 2.0), (27 +/- 12), (0.8 +/- 0.5) ng x g(-1) for pituitary gland, (1.7 +/- 1.2), ( 16 +/- 10), and (0. 8 +/- 0.7) ng x g(-1) for hypothalamus, respectively.

CONCLUSIONGood linearity and accuracy were observed for each steroid. The procedure was suitable for measuring concentrations of the unconjugated steroids in rat brain simultaneously.

Amygdala ; chemistry ; Animals ; Brain Chemistry ; Chromatography, High Pressure Liquid ; Corpus Striatum ; chemistry ; Dehydroepiandrosterone ; analysis ; Hippocampus ; chemistry ; Hypothalamus ; chemistry ; Male ; Mass Spectrometry ; methods ; Nucleus Accumbens ; chemistry ; Prefrontal Cortex ; chemistry ; Pregnenolone ; analogs & derivatives ; analysis ; Rats ; Rats, Sprague-Dawley ; Sensitivity and Specificity

Objective To optimize the short tandem repeats(STR)which link closely to survival motor neuron(SMN)and have redundant polymorphism information contents,and to use these STR in the prenatal diagnosis of spinal muscular atrophy(SMA).Methods Eleven STR loci(D5S435,D5F153, DSF151,D5S637,D5S1413,D5S125,D5S464,D5S1556,DSF149,D5S351,MAP1B-5')were amplified by PCR.Then the PCR products were detected by polyacrylamide gel electrophoresis(PAGE)and analyzed by silver staining.STR loci were evaluated and optimized by their PIC values.PCR-PAGE and gene scan were combined to make genetic link analysis for SMA families based on the optimized STR.Results Three STR loci(D5S435,DSF149 and D5S351)were selected with 8,19 and 18 polymorphic fragments detected respectively in 100 normal individuals.Their PIC values were 0.84,0.91 and 0.92 respectively.Four carriers and 2 normal individuals were detected from 6 SMA families with linkage analysis by using the 3 STR.Conclusion This genetic diagnosis system based on the 3 STR loci can provide rapid prenatal diagnosis for SMA families,can eliminate maternal blood contamination,and also can discriminate carriers from normal individuals in the fetuses,which makes the prenatal diagnosis system of SMA perfect.

Objective To explore the relationships of serum leptin(LP),tumor necrosis factor-?(TNF-?)and insulin resistance in simple obesity children.Methods Forty-seven children with simple obesity were divided into two groups: high insulin group(HIG) and normal insulin group(NIG) according to the levels of fasting state of serum insulin and glucose.The fasting state of serum insulin(FINS),LP,TNF-? and fasting blood glucose(FBG) were measured.The relationships of LP,TNF-? and insulin resistance index(HOMA-IR) were also analyzed.Results 1.The BMI,waist,hip,waist hip ratio(WHR),LP,TNF-?,HOMA-IR were obviously higher in the HIG than those in the NIG and NCG groups(Pa0.05).3.In the HIG group,the FINS was positively correlated to LP,TNF-? and HOMA-IR(r=0.560,0.413,0.864 P

OBJECTIVETo investigate the role of Stat3 (one of the signal transductant and activator) signal transduction pathway in proliferation of vascular smooth muscle cells (VSMCs).

METHODSVSMC was transfected with Stat3 antisense oligonucleotide. ELISA analysis was performed on Stat3 expression in cultured rat thoracic VSMC. Cell proliferation and toxicity assay by Methyl Thiazole (MTT) was used to measure the cell proliferation. The expression of Stat3, p-Stat3, Cyclin D1 and Bcl-x(L) were measured by Western blot.

RESULTSStat3 protein expression was positive in VSMC. Targeting of Stat3 using antisense oligonucleotide directed against the translation site, resulted in significant growth inhibition and downregulation of Stat3, p-Stat3 and Cyclin D1.

CONCLUSIONSThe increase of Stat3 correlates significantly with cell proliferation of VSMC. Cyclin D1 may be the critical target gene in mediating proliferation. Blocking Stat3 signal transduction pathway may inhibit the growth of VSMCs.

Animals ; Cell Line ; Cell Proliferation ; Cyclin D1 ; metabolism ; Male ; Muscle, Smooth, Vascular ; metabolism ; Myocytes, Smooth Muscle ; metabolism ; Rats ; Rats, Sprague-Dawley ; STAT3 Transcription Factor ; genetics ; metabolism ; Signal Transduction ; Transfection

Child, Preschool ; Humans ; Male ; Myiasis ; complications ; diagnosis ; therapy

BACKGROUNDEndothelial dysfunction is a key event in the onset and progression of atherosclerosis in diabetic patients. Apoptosis may lead to endothelial dysfunction and contribute to vascular complications. However, no study has addressed apoptosis in human umbilical vein endothelial cells (HUVECs) induced by an intermittent high-glucose media and its association with adiponectin receptor 1 (adipoR1), adipoR2, or adenosine monophosphate (AMP)-activated protein kinase (AMPK).

METHODSHUVECs were cultured in continuous normal glucose (5.5 mmol/L), continuous high glucose (25 mmol/L), alternating normal and high glucose and mannitol. In the alternating normal and high-glucose media, HUVECs were treated under different conditions. First, cells were transfected with the adipoR1-specific small-interfering RNA (siRNA) and then stimulated with globular adiponectin (gAD). Second, cells were cultured in both gAD and the AMPK activator 5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside (AICAR). Third, cells were cultured in the AMPK inhibitor adenine-9-β-D-arabino-furanoside (araA), gAD, and in AICAR.

RESULTSHUVEC apoptosis increased more significantly in an intermittent high-glucose medium than in a constant high-glucose medium. HUVEC apoptosis induced by an intermittent high-glucose medium was inhibited when the cells were pretreated with 3 µg/ml gAD, which rapidly activated AMPK and adipoR1 in HUVECs. However, adipoR2 was not activated.

CONCLUSIONSWe found that adipoR1, not adipoR2, is involved in mediating intermittent high-concentration glucose-evoked apoptosis in endothelial cells. gAD activated AMPK through adipoR1, leads to the partial inhibition of HUVEC apoptosis. A fluctuating glucose medium is more harmful than a constant high-glucose medium to endothelial cells.

AMP-Activated Protein Kinases ; antagonists & inhibitors ; genetics ; metabolism ; Adiponectin ; pharmacology ; Aminoimidazole Carboxamide ; analogs & derivatives ; pharmacology ; Apoptosis ; drug effects ; Glucose ; pharmacology ; Human Umbilical Vein Endothelial Cells ; cytology ; drug effects ; metabolism ; Humans ; RNA, Small Interfering ; Receptors, Adiponectin ; genetics ; metabolism ; Ribonucleotides ; pharmacology ; Signal Transduction ; drug effects ; genetics

OBJECTIVETo explore the clinical value of "triple-rule-out" protocols using dual-source computed tomography for aortic dissection (AD) assessment.

METHODSTotally 25 patients suspecting of suffering from AD were examined on a dual-source computed tomography scanner. Two-dimensional and three-dimensional reconstruction was performed in all patients by means of multiplanar reconstruction, curved planar reformation, maximum intensity projection, and volume rendering. All images were read by two experienced radiologists in consensus. All patients were divided into AD group (n=12) and NO AD group (n=13) , The average Hounsfield unit values of true and false lumen were compared between superior of the aortic around the first endoleak and inferior of the aortic around renal artery.

RESULTSIn AD group, there were 6 patients with DeBakey type 1, 2 patients with DeBakey type 2, and 4 patients with DeBakey type 3. The image quality was rated on a 3-point scale as excellent in 10 patients (83.3%) and good in 2 patients (16.7%) . All cases was fully evaluable in NO AD group. The average Hounsfield unit values of true lumen between superior of the aortic around the first endoleak and inferior of the aortic around renal artery showed no significant difference between AD and NO AD group.

CONCLUSIONDual-source computed tomography offers a non-invasive, accurate, and rapid way to evaluate AD.

Adult ; Aged ; Aged, 80 and over ; Aneurysm, Dissecting ; diagnostic imaging ; Angiography ; methods ; Aortic Aneurysm ; diagnostic imaging ; Female ; Humans ; Male ; Middle Aged ; Tomography, X-Ray Computed ; methods ; Young Adult

This study was aimed to investigate the effects of ouabain at low concentrations on growth regulation in various leukemia cell lines and to determine the therapeutic potential of ouabain in leukemia. By using MTT, flow cytometry (FCM), the changes in cell growth and cell cycle of leukemia cell lines were observed after treating with ouabain at low concentrations ( Cell Line, Tumor ; Cell Proliferation ; drug effects ; Dose-Response Relationship, Drug ; Humans ; Leukemia ; pathology ; Leukemia, Megakaryoblastic, Acute ; pathology ; Ouabain ; pharmacology ; Sodium-Potassium-Exchanging ATPase ; metabolism

OBJECTIVETo construct a method for detecting the copy number of survival of motor neuron 1 gene (SMN1) with single copy difference based on real-time fluorescence quantitative PCR, and to make practical use of the method for acquiring the data on SMN1 copy number in Chinese as well as for screening the carriers of spinal muscular atrophy (SMA) from healthy individuals and SMA families.

METHODSExon 7 and flanking area of SMN1 gene were amplified by real-time fluorescence quantitative PCR in 264 healthy individuals, in 1 standard sample having 2 SMN1 but having no SMN2, and in 88 parents of SMA patients. The samples for detecting were diluted to 30 ng/microL and the standard sample was diluted to 15 ng/microL, 30 ng/microL, 45 ng/microL, 60 ng/microL; the unknown samples and 4 standard samples with different concentrations were amplified at the same time, a standard curve could be drawn out according to the results of the 4 standard samples, then the copy number of samples could be calculated.

RESULTSOf 88 parents' samples, 84 samples each had 1 copy of SMN1, and the rest 4 each had 2 copies of SMN1. Of 264 healthy individuals' samples, 5 samples each had only 1 copy of SMN1 (an indicator of definite gene carriers), 232 samples each had 2 copies of SMN1, 25 samples each had 3 copies of SMN1, and 2 samples each had 4 copies of SMN1. Of the samples of 32 members of SMA families, 2 samples each had only 1 copy of SMN1 indicating definite gene carriers, 25 samples each had 2 copies of SMN1, and 5 samples each had 3 copies of SMN1.

CONCLUSIONSMN1 copy number could be detected precisely by real-time fluorescence quantitative PCR; the screening of gene carriers could provide essential data for genetic counseling.

Exons ; Family Health ; Female ; Fluorescence ; Gene Dosage ; Humans ; Male ; Muscular Atrophy, Spinal ; genetics ; Polymerase Chain Reaction ; methods ; Survival of Motor Neuron 1 Protein ; genetics

OBJECTIVETo retrospectively compare the clinical outcome in patients with portal hypertension treated with transjugular intrahepatic portosystemic shunt (TIPS) using Fluency stent-graft (PTFE-covered stents) or bare stents.

METHODSApproval of study and treatment protocol and waiver of informed consent for the retrospective study were obtained from institutional review board. Informed consent was obtained from each patient before procedure. Sixty consecutive patients with portal hypertension treated with TIPS from April 2007 to April 2009 were included. TIPS creation was performed with Fluency stent-graft in 30 patients (group A) and with bare stents in 30 patients (group B). Liver function, TIPS patency and clinical outcome were evaluated every 3 months.

RESULTSDuring hospitalization, there was no hepatic encephalopathy and recurrency of variceal bleeding.Acute shunt occlusion was observed in one patient with group A and another patient with group B.Follow-up was performed with average time of (6.2 +/- 3.9) months in group A and (8.3 +/- 4.4) months in group B. The rates of recurrent bleeding, acute shunt occlusion, hepatic encephalopathy and death were 3.3% and 20.0%, 0 and 30.0%, 16.7% and 20.0%, 0 and 13.3% in group A and B. The rates of recurrent bleeding, acute shunt occlusion and death in group A was lower than those in group B. There was no difference of hepatic encephalopathy between group A and B. The decrease of portal pressure and portosystemic pressure gradient, and the increase of portal flow and shunt flow in group A were higher than those in group B. There were no difference of liver function, ammonia and MELD between group A and B.

CONCLUSIONSFluency stent-graft is safe and effective in TIPS creation, with high patency rate. Covered-stent can improve the clinical outcome of portal hypertension.

Adult ; Aged ; Female ; Follow-Up Studies ; Humans ; Hypertension, Portal ; surgery ; Male ; Middle Aged ; Portasystemic Shunt, Transjugular Intrahepatic ; instrumentation ; Retrospective Studies ; Stents ; Treatment Outcome ; Young Adult