The study aims to establish a quantitative nuclear magnetic resonance (QNMR) method for the determination of the absolute content of bosentan. Proton nuclear magnetic resonance spectroscopy [1H NMR] spectra were obtained in CDCl3 with the internal standard dimethyl terephthalate and zg30 pulse sequence by using a Bruker AVANCE II 400 spectrometer. The content of bosentan is determined with QNMR in comparison with the result obtained by mass balance method. The result is 96.25% by QNMR and 96.54% by mass balance method. A rapid and accurate QNMR method has been established for the quantitative determination of the absolute content of bosentan. The study provides a new way for the quality control and calibration of a new reference standard material, it could be the complementary with the mass balance method for the assay of standard reference.
Calibration ; Magnetic Resonance Spectroscopy ; methods ; Molecular Structure ; Protons ; Quality Control ; Sulfonamides ; chemistry

Objective To investigate the effects of rhBMP2 on the differentiation of human dental pulp stem cells and expression of Delta protein in vitro. provide a theoretical basis for research on human dental pulp stem cells. Methods Monocell suspension was separated from the human adult dental pulp with collagenase Ⅰ and dispase digestion.Clonogenic cells were observed under light microscope and the expressions of surface markers were determined with immunofluorescence. divided into experimental group (containing 50 ?g?L-1rhBMP2 in the culture medium) and negative control group (containing culture medium only).alkaline phosphatase and the expression of Delta proterin of the cells at at passage 5 were detected.Results The human dental pulp stem cells showed colony growth,the nestin and vimentin staining were positive by immunohistochemical staining.STRO-1 was positive in cells by immunofluorescence.Compared with negative control group, the activities of alkaline phosphatase increased after treatment with rhBMP2 for 7,14 and 21d(P

Objective To observe any influence of hyperbaric oxygen (HBO) treatment on the formation of glial scars,and to explore how HBO suppresses the inflammatory reaction to injury.Methods A total of 96 healthy,adult,male,Sprague-Dawley rats were used to model cerebral puncture injury.They were then randomized into a control group and a treatment group,with 48 rats in each group.The treatment group received HBO treatment,while the control group received no special treatment.At 1,3,7,14 and 28 days after the puncture injury,the rats' right brain tissues were harvested and immunohistochemical staining was employed to compare the changes in number of astrocytes and microglial cells around the injury in the two groups.The level of tumor necrosis factor α (TNF-α) and interleukin 1 β (IL-1β) in the cerebral tissue was examined using ELISA.Results Among the control group the average wound areas after 7,14 and 28 days were (2.73 ± 0.05)μm2,(3.42 ± 0.18)μm2 and (2.41 ± 0.09) μm2,a significant reduction after 28 days compared with 7 and 14 days.The corresponding average wound areas of rats in the treatment group were (2.78±0.12)μm2,(2.59 ±0.08)μm2 and (1.20 ±0.06)μm2.There the average wound area had decreased significantly after 14 days,and the further reduction after 28 days was also significant.The numbers of GFAP-positive astrocytes at 14 and 28 days had increased significantly compared with after 7 days in both the control group and the treatment group.The average number of GFAP-positive astrocytes in the control group at 28 days had decreased significantly compared with after 14 days.Compared with the control group at the same time points,the number of GFAP-positivc astrocytes in the treatment group was significantly less.After modeling,the number of ionized calcium-binding adapter molecule Ⅰ (Ibal)-positive microglial cells increased significantly,but there was a significant decrease in both the control and treatment groups by 7 days.The average number of Ibal-positive microglial cells in the treatment group was significantly less than in the control group at all of the time points.Compared with the first day after modeling,the TNF-α concentration of the controls at 3 and 7 days was significantly higher,but by the 7th day it was significantly lower than it had been after 3 days.The average IL-1β concentration in the control group and TNF-α concentration in the treatment group had increased by day 3,but then decreased by day 7.The IL-1β concentration of the treatment group declined gradually.The average TNF-α and IL-1 β concentrations of the treatment group were significantly lower than those of the control group at all of the time points.Conclusion HBO treatment has a relatively good curative effect on cerebral puncture injury.It can accelerate wound healing and reduce the formation of glial scars.Its mechanism could be related to the deactivation of astrocytes and microglia cells and reducing the levels of cell factors that promote inflammation.

OBJECTIVETo investigate the therapeutic effect of IL-6 mAb on experimental autoimmune myocarditis (EAM) in rats, and search the mechanism of the role of IL-6, helper T cells 17 (Th17) and regulative T cells (Treg) in EAM pathogenesis.

METHODSThirty-four Lewis rats were divided into three groups randomly, i.e. control group (n = 6), EAM group (n = 12), and IL-6 mAb intervention group (n = 16). Rats in EAM group and IL-6 mAb intervention group were injected intracutaneously with myosin to establish EAM model. Rats in IL-6 mAb intervention group were injected intraperitoneally with 1 mg IL-6 mAb on 1st, 7th to 20th day after cardiac myosin immune injection. Myocardial inflammation was examined by HE stain, Masson stain, and TdT assay (TUNEL reaction) on 21st and 84th day after IL-6 mAb therapy in order to assess the therapeutic role. Spleen cells were analyzed by flow cytometry to illustrate Th17 and Treg cells? number and function. The serum concentration of IL-6, IL-10, IL-17, and TGF-beta in each group was measured by ELISA, concentration of STAT3, RORgammat, and Foxp3 mRNA in each group was determined with RT-PCR. Spleen cells derived from EAM were stimulated by IL-6 mAb in vitro, and the concentration of IL-10, IL-17 and TGF-beta was measured by ELISA.

RESULTSInflammation score, fibrosis score, and apoptosis index in IL-6 mAb intervention group were significantly decreased as compared with those in EAM group (P < 0.01). The number of Th17 and Treg cells in EAM group on the 21st day (experimental acute peak stage) were increased, and those in intervention group on the 21st day were significantly inhibited (P < 0.01). The concentration of serum IL-6, IL-10, IL-17 and TGF-beta in intervention group on the 21st day was decreased dramatically in comparison with that in EAM group on the same day (P < 0.01). The levels of peripheral blood STAT3, RORgammat, Foxp3 mRNA in intervention group on the 21st day was decreased significantly as compared with that in EAM group (P < 0.01). The expression of IL-10, IL-17 and TGF-beta was increased significantly (P < 0.01) by stimulation of IL-6 mAb on spleen cells derived from EAM in vitro.

CONCLUSIONSIL-6 mAb could neutralize IL-6, and ameliorate myocarditis and reduce heart autoimmune responses. IL-6 mAb has significantly protective effects on EAM by suppressing Th17 and Treg cells.

Animals ; Antibodies, Monoclonal ; therapeutic use ; Autoimmune Diseases ; drug therapy ; immunology ; Disease Models, Animal ; Forkhead Transcription Factors ; metabolism ; Interleukin-10 ; metabolism ; Interleukin-17 ; metabolism ; Interleukin-6 ; immunology ; Male ; Myocarditis ; drug therapy ; immunology ; Nuclear Receptor Subfamily 1, Group F, Member 3 ; metabolism ; Rats ; Rats, Inbred Lew ; STAT3 Transcription Factor ; metabolism ; Th17 Cells ; immunology ; Transforming Growth Factor beta1 ; metabolism

Adolescent ; Alkaline Phosphatase ; metabolism ; Antineoplastic Combined Chemotherapy Protocols ; therapeutic use ; Bleomycin ; therapeutic use ; Cisplatin ; therapeutic use ; Diagnosis, Differential ; Dysgerminoma ; pathology ; Etoposide ; therapeutic use ; Female ; Gonadoblastoma ; drug therapy ; metabolism ; pathology ; surgery ; Humans ; Hysterectomy ; methods ; Inhibins ; metabolism ; Isoenzymes ; metabolism ; Ovarian Neoplasms ; drug therapy ; metabolism ; pathology ; surgery ; Young Adult

Objective To analyze the changes of clinical and pathological features in the patients of IgA nephropathy with anemia.Methods Four hundred and nine patients of IgA nephropathy diagnosed by renal biopsy were classified into two groups:IgA nephropathy with nonanemia (group 1) and IgA nephropathy with anemia (group 2).Changes were studied retrospectively between the groups.Results Serum hemoglobin level was correlated with the clinical parameters of IgA nephropathy.Companed to group 1,changes in group 2 were as followed:serum creatinine increased,eGFR decreased,proteinuria increased; the global sclerosis,segmental sclerosis,crescents and tubulointerstitial lesions worsened.The glomerular and tubulointerstitial lesions were negatively correlated with serum hemoglobin and eGFR,but positively correlated with serum uric acid and proteinuria (P＜0.05).Multivariate Logistic regression analysis revealed that anemia was an independent risk factor for the tubulointerstitial lesion.Conclusion Clinical feature and pathological damages in the patients of IgA nephropathy with anemia are more serious than those with non-anemia.

Objective To evaluate the value of low-density lipoprotein-cholesterol (LDL-C) and non-high-density lipoprotein-cholesterol (non-HDL-C) in differential diagnosis of familial hypertriglyceridemia (FHTG) and familial combined hyperlipidemia (FCHL).Methods We recruited 9 FHTG pedigrees (94 subjects) and 24 FCHL pedigrees (94 subjects) and then divided them into affected groups and non-affected groups according to lipid abnormality.Another 10 normal control pedigrees (57 subjects) served as controls.We compared the routine lipid levels such as triglyceride (TAG),total cholesterol (TC),HDL-C and LDL-C and non-HDL-C between the groups.After stratification based on TAG level,we observed the relationship between LDL-C and non-HDL-C.Last we confirmed and analyzed the cut-off value of differential diagnosis between FHTG and FCHL with receiver operating characteristic (ROC) curve.Results The levels of TAG,TC,and non-HDL-C were significantly higher in the affected group of FHTG than in the non-affected group of FHTG and the normal group (P＜0.01 or P＜0.05).The levels of TAG,TC,HDL-C,LDL-C and non-tHDL-C wcrc significantly higher in the affected group of FCHL than in the non-affected group of FCHL and the normal group (P＜0.01 or P＜0.05).The levels of TAG were significantly higher (P＜0.01) while TC,HDL-C,LDL-C and non-HDL-C levels were significantly lower (P＜ 0.01 or P＜0.05) in the affected group of FHTG than in the affected group of FCHL.The association between LDL-C and non-HDL-C was positive both in FHTG and FCHL,but the relationship became weaker as TAG level increased.The cut-off value of LDL-C and non-HDL-C was 3.575 mmol/L and 4.525 mmol/L,respectively.Conclusion In addition to the routinely used lipid indexes,non-HDL-C may be a new index for differential diagnosis of FHTG and FCHL,and may be superior to LDL-C in this regard.

Objective To explore the effects of endoplasmic reticulum (ER) stress in the hyperoxia-induced lung injury in premature rats. Methods Forty-eight premature Wistar rats were randomized into two groups 12 hours after birth:hyperoxia group (n=24) inhaled 95%oxygen and control group (n=24) inhaled air. Eight rats were sacriifced in each group on day 1, 3, 7 after the treatment and the left lungs were embedded. The pathological changes in the HE stained sections of lung tissues were observed. The expressions of ER related protein ERp57 and c/EBP homologous protein CHOP were detected by immuno histo-chemistry and the apoptosis of lung cells was detected by TUNEL analysis. Results The typical pathological characteristics of acute lung injury were observed in hyperoxia group. The expressions of ERp57 and CHOP were increased with the exposure time in hyperoxia group, and were signiifcantly higher than in control group (P<0.05). The apoptosis rate of lung cells in hyperoxia group was signiifcantly higher than in control group (P<0.01). There was signiifcant positive correlation between cell apoptosis index and expressions of Erp57 and c/EBP homogeneous protein. Conclusions ER stress initiated apoptosis participates and plays an important role in the process of hyperoxia-induced lung injury in premature rats.

Objective To compare the safety and clinical efficiency between minimally invasive percutaneous nephrolithotomy(MPCNL)in supine position and flexible ureteroscopy(FURS)in management of proximal ureteral calculi.Methods From Oct.2010 to May.2012,76 patients with single proximal ureteral calculus between 10-20 mm failed in SWL or other conservative therapy accepted MPCNL (32 cases)or FURS(44 cases).There was no significant difference between the groups in base-line parameters.Stone sizes were(15.6±2.5)mm and(14.9±2.3)mm,P＞0.05.Procedural time,post-operative hospitalization stay,complication rates(Clavien degree Ⅱ or over)and stone free rates were compared.Results In these two groups,procedural time was(49.3± 11.7)and(67.2± 17.3)min,P＜0.05,postoperative hospitalization stay were(4.2±1.1)and(1.8±0.8)days,P＜0.05,complication rates were 12.5％ and 6.8％,P＞0.05 and stone free rates(residual fragments≤3 mm)were 93.7％ and 84.1％,P＞0.05.Conclusions For patients with surgically indicated proximal ureteral calculi,both minimally invasive percutaneous nephrolithotomy in supine position and flexible ureteroscopy are effective and safe therapeutic modalities.Patients treated with flexible ureteroscopy have faster postoperative recovery.

Objective To study any protection against hippocampal neuron damage induced by epilepsy (SE) provided by transcutaneous stimulation (TNS) of the trigeminal nerve and to document any effect of such stimulation on the expression of glutamic acid decarboxylase (GAD) 65/67.Methods Pilocarpine injection was used to induce epilepsy in healthy male Sprague-Dawley rats which were then randomly divided into a treatment group and a model group.Rats which had not received the pilocarpine injection served as normal controls.In the treatment group the rats were given electrostimulation for one month after the first spontaneous seizure following the injection of pilocarpine.In the model group they were given sham TNS for one month.After the month of stimula-tion,immunohistochemistry was used to detect the expression of GAD65/67 in the hippocampus.Terminal deoxy-nucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assays and Nissl staining were applied to deter-mine apoptosis and neuron loss in the hippocampus.Results Significantly less apoptosis was observed in the treatment group than in model group at 24 h,48 h and 72 h post-injection.Compared to the model group,average GAD65/67 expression had increased significantly in the treatment group at 24 h,72 h,1 week,2 weeks and 4 weeks post-stimulation.GAD65 expression reached its peak from 72 h to 1 week post-stimulation,then decreased to the level of the control group by 4 weeks post-stimulation.The expression of GAD67 remained elevated at all the time points employed.Conclusions TNS can significantly protect hippocampal neurons from damage in epilepsy,at least in rats.The underlying anti-epileptic and neuroprotective mechanisms may involve increased inhibitory transmission induced by the stimulation.