Objective To investigate the effects of midazolam on GABAA receptor-activated currents in isolated dorsal root ganglion (DRG) neurons in rats.Methods Sprague-Dawley rats of both sexes,weighing 200-250 g,aged 4 weeks,were used in the study.The DRG neurons were isolated and GABAA receptor-activated currents were recorded using the whole-cell patch-clamp technique.GABAA receptor-activated currents were recorded after administration of the mixture of midazolam 3.00 μmol/L (final concentration)and the different final concentrations (0.03,0.10,1.00,10.00,100.00 and 1000.00 μmol/L) of GABA,after different concentrations of midazolam (0.03,0.10,1.00,3.00,10.00 and 100.00 μmol/L) was given,after administration of the mixture of different final concentrations(0.03,0.10,1.00,3.00,10.00 and 100.00 μmol/L) of midazolam and GABA 100.00 μmol/L (final concentration),and after administration of the mixture of midazolam 1.00μmol/L (final concentration) and GABA 100.00 μmol/L (final concentration)at the preset time points of perfusion with different concentrations of midazolam (0,20,40,60 and 120 s of perfusion).The enhancement rate of the currents was calculated.Results No change in the membrane currents was found after midazolam was perfused in the neurons sensitive to GABA.GABAA receptor-activated currents were enhanced after administration of the mixture of different concentrations of GABA and midazolam.GABAA receptor-activated currents were enhanced after different concentrations of midazolam were given compared with that before administration,and the enhancement rate of the GABAA receptoractivated currents was gradually increased with the increase in the concentration of midazolam and reached the peak at the concentration of 3.00 μmol/L.The enhancement rate of the GABAA receptor-activated currents was gradually increased with the prolongation of perfusion time and peaked at 40 s of perfusion.Conclusion Midazolam can enhance the GABAA receptor-activated currents in rat dorsal root ganglion neurons,indicating that midazolam increases the role of GABA through increasing the activity of GABAA receptors and has analgesic effect at the spinal cord level.

To further explore the result of bilingual teaching in pediatrics,we randomly chose 200 undergraduates of 4 class and released students'questionnaires about bilingual teaching with teaching content before and after class to assess students'understanding of bilingual teaching and analysed appraisal result.We found no significant difference of student score between students accepting bilingual teaching and not accepting the bilingual teaching,but there was difference for English tests and expression level.So we think that students can fully accept the bilingual teaching of pediatrics under the premise with selecting appropriate teaching methods and means.

Objective:To express rationally engineered antibodies against EGFR and assess their affinity to EGFR and anti-tumor cell migration effect. Methods:L and V_H genes of humanized antibodies against EGFR were designed and synthesized. Genes encoding V_H and C_H were connected and then cloned into a pIRES based bicistronic expression vector. Gene encoding the corresponding L gene was also cloned into the same vector. 293T cells were transfected with the recombinant plasmid and the antibody expression was confirmed by Western blotting. The antibodies were purified by protein A based affinity chromatography. Binding of the humanized antibody to the EGFR was assessed by Surface Plasmon Renainance with Biacore3000, and the biological activity of the humanized antibody was determined by tumor cell invasion test.Results:Three expression vectors were constructed and the humanized anti-EGFR antibodies were expressed and purified successfully. In reducing SDS-PAGE, the antibodies exhibited two bands of approximately 25×10~3and 50×10~3, respectively. Western blot assay showed that the humanized antibodies had recognition specificity to goat-human IgG antiserum. Biacore assay revealed that the humanized antibody C3 binds to EGFR with high affinity(6.13×10~(-10)M). Cell migration test showed that C2,C3 and C5 could suppress growth and migration of tumor cells.Conclusion:Three anti-EGFR humanized antibodies (C2,C3 and C5) have been constructed and expressed successfully, and the C3 antibody retained high affinity for EGFR and showed improved inhibitory effect on tumor cell growth and migration.

BACKGROUND:Recent studies have found that bone marrow mesenchymal stem cels that culturedin vitro for a long time can naturaly differentiate into neural stem cels, which then differentiate into neurons and glial cels, thereby providing a new therapeutic thinking for Parkinson’s disease, sequela of cerebral infarction, cerebelar atrophy and brain dysplasia.
OBJECTIVE:To discuss the influence of neural stem cel transplantation on neurologic function of rats with cerebral hemorrhage at recovery stage and the relevant mechanism of action.
METHODS: Sixty male Sprague-Dawley rats were randomly divided into normal group (n=18), cerebral hemorrhage group (n=21) and transplantation group (n=21). Cerebral hemorrhage models were established in the latter two groups using VII type colagen enzyme induction method. At 21 days of modeling, rats in the transplantation group were injected neural stem cels via the tail vein, and those in the other two groups received the same volume of normal saline. At 7, 14, 21 days after cel transplantation, modified adhesive removal test (MST) was employed to evaluate the neurologic function of rats, and then the rats were kiled. RT-PCR was used to detect angiopoietin-1 mRNA expression in the bleeding tissues, and western blot assay was employed to measure tyrosine kinase receptor-2 protein expression.
RESULTS AND CONCLUSION:Compared with the normal group, the MST scores in the cerebral hemorrhage group and transplantation group were significantly decreased (P< 0.05). From the 7th day after transplantation, MST scores in the transplantation group were significantly higher than those in the cerebral hemorrhage group (P < 0.05). At 7, 14, 21 days after transplantation, expressions of angiopoietin-1 mRNA and tyrosine kinase receptor-2 protein were ranked as folows: transplantation group > cerebral hemorrhage group > normal group, and there was a significant difference among the three groups (P< 0.05). These findings indicate that neural stem cel transplantation can effectively promote the neurologic recovery of rats with cerebral hemorrhage at recovery stage, and the concrete mechanism may be related to the increase of angiopoietin-1 mRNA and tyrosine kinase receptor-2 protein in the bleeding tissues.

OBJECTIVETo explore the influence of exogenous putrescine and cadaverine on pro-inflammatory factors in the peripheral blood of rabbits.

METHODSForty ordinary adult New Zealand rabbits were divided into saline, necrotic tissue homogenate (NTH), putrescine, and cadaverine groups according to the random number table, with 10 rabbits in each group. Saline, NTH, 10 g/L putrescine, and 10 g/L cadaverine were respectively peritoneally injected into rabbits of corresponding group in the amount of 1 mL/kg. The blood sample in the volume of 2 mL was collected from the central artery of rabbit ears before injection and at 2, 6, 12, 24, 30, 36, 48, 60 hours post injection (PIH). Contents of TNF-α, IL-1, and IL-6 in the serum were determined with enzyme-linked immunosorbent assay. Data were processed with repeated measurement data analysis of variance and Spearman correlation analysis, and cubic model curve was applied in curve fitting for the contents of inflammatory factors.

RESULTS(1) The serum contents of TNF-α, IL-1, and IL-6 were increased in NTH, putrescine, and cadaverine groups in different degrees at most post injection time points. There was no significant change in the concentrations of the three pro-inflammatory factors in saline group, and they were significantly lower than those of the other three groups at most post injection time points (with F values from 3.49 to 13.58, P values all below 0.05). The serum contents of TNF-α, IL-1, and IL-6 in putrescine group began to increase at PIH 2, 6, and 6, which was similar to the trend of NTH group, but the changes were delayed compared with those of cadaverine group(all at PIH 2). The peak values of TNF-α, IL-1, and IL-6 in putrescine group were respectively (339 ± 36), (518 ± 44), and (265.9 ± 33.5) pg/mL, which were significantly lower than those of cadaverine group [ (476 ± 86), (539 ± 22), and (309.4 ± 27.1) pg/mL], with F values respectively 5.11, 1.90, and 5.56, P values all below 0.05. (2) The period of time in which contents of TNF-α, IL-1, and IL-6 began to increase (PIH 3-4) and the peaking time of the three pro-inflammatory cytokines (PIH 18-30) in putrescine group appeared later than those of cadaverine group (PIH 2 and 12-30). The duration of peaking time of the three pro-inflammatory cytokines in putrescine group was shorter than that of cadaverine group (PIH 18-30 vs. PIH 12-30). The increasing period and the duration of peaking time of TNF-α, IL-1, and IL-6 in putrescine group were close to those of NTH group (PIH 3-5 and 18-30). The correlation coefficient test analysis showed that the trends of changes in contents of three pro-inflammatory cytokines in putrescine group were significantly correlated with those of NTH group (r(TNF-α) = 0.933, P < 0.01; r(IL-1) = 0.967, P < 0.01; r(IL-6) = 0.950, P < 0.01). The obvious correlation between cadaverine group and NTH group was only found in the contents of IL-1 and IL-6 (r(IL-1) = 0.913, P < 0.01; r(IL-6) = 0.883, P < 0.05).

CONCLUSIONSBoth exogenous putrescine and cadaverine can cause inflammatory reaction in rabbits. The trend of the inflammatory reaction induced by putrescine was similar with that by NTH, suggesting that putrescine may play a leading role in the inflammatory reaction induced by necrotic tissue decomposition.

Animals ; Cadaverine ; adverse effects ; Inflammation ; blood ; Interleukin-1 ; blood ; Interleukin-6 ; blood ; Necrosis ; blood ; Putrescine ; adverse effects ; Rabbits ; Tumor Necrosis Factor-alpha ; blood

OBJECTIVETo investigate the effects of ropivacaine on Gamma-aminobutyric acid(GABA)-activated currents in dorsal root ganglion (DRG) neurons in rats and discuss the analgesia mechanism of ropivacaine.

METHODSBy means of using whole-cell patch-clamp technique, to investigate the modulatory effects of ropivacaine on GABA-activated currents (I(GABA)) in acutely isolated dorsal root ganglion neurons.

RESULTS(1) In 48 out of 73DRG cells (65.7%, 48/73), to perfusion ropivacaine bromide (0.1 - 1 000 micromol/L) were sensitive. Which produce in 0 to 380 pA current. (2) The majority of the neurons examined (74.5%, 73/98) were sensitive to GABA. Concentration of 1 - 1 000 micromol/L GABA could activate a concentration-dependent inward current, which manifested obvious desensitization, and the inward currents could be blocked byGABA-receptor selective antagonist of bicuculline (100 micromol/L). (3) After the neurons were treated with ropivacaine (0.1 - 1000 micromol/L) prior to the application of GABA (100 micromol/L) 30 s, GABA currents were obviously increased. Ropivacaine could make dose-response curve of the GABA up, EC50 is 23.46 micromol/L. Ropivacaine shifted the GABA dose-response curve upward and increased the maximum response to the contrast about 153%.

CONCLUSIONThe enhancement of ropivacaine to DRG neurons activation of GABA current, can lead to enhancement of pre-synaptic inhibition at the spinal cord level. This may be one of the reasons for the anesthetic effect and analgesia for ropivacaine in epidural anesthesia.

Amides ; pharmacology ; Animals ; Ganglia, Spinal ; cytology ; physiology ; Membrane Potentials ; drug effects ; Neurons ; cytology ; drug effects ; physiology ; Patch-Clamp Techniques ; Rats ; Rats, Sprague-Dawley ; Receptors, GABA-A ; physiology

BACKGROUNDWhile the abnormal appearance of the concealed penis has been well recognized, the effect of buried penis on the structure and function of corpus cavernosum has not been well studied. To explore this issue, we established a rat model and evaluated the effect of buried penis on cavernosum weight, contents and ultrastructure of tissue, and nitric oxide synthase (NOS) activity.

METHODSTwo hundred and ten rats were randomly divided into 3 equal cohorts for 2, 4 and 6 months study (groups A, B and C). Each group was randomly divided into buried group (n = 40), control group (n = 15), and normal group (n = 15), respectively. Intra-purse-string suture of the root of the penis was used to establish the model. Macroscopic development was judged by measuring the weight of the corpus cavernosum. Masson's trichrome staining was performed for observing microstructure while a transmission electron microscope was used for observing ultrastructure. The NOS activity was detected by a NOS activity assay kit.

RESULTSBuried penis had no significant influence on the appearance and weight of the corpus cavernosum. Buried penis resulted in decreased smooth muscle content (P > 0.05 in group A, and P < 0.05 in groups B and C) and increased fibrous connective tissue content (P > 0.05 in groups A and B, and P < 0.05 in group C) compared with the normal and control groups. Ultrastructural abnormalities of corpus cavernosum were observed in the 6-month buried group. Moreover, there was decrease of NOS activity in groups B and C (P < 0.05 in group B and P < 0.01 in group C) when compared with the normal and control groups.

CONCLUSIONBuried penis affects the structure and function of corpus cavernosum in rats and the effect is positively correlated with the buried time, but there is no significant effect on the macroscopic development.

Animals ; Male ; Nitric Oxide Synthase ; metabolism ; Penis ; anatomy & histology ; pathology ; physiopathology ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo evaluate the efficacy and toxicity of the combined chemotherapy with docetaxel, capecitabine and cisplatin (TXP) in the treatment of metastatic nasopharyngeal carcinoma (NPC).

METHODSThis retrospective analysis involved 22 patients with metastatic NPC receiving treatment with the TXP regimen. The patients were given docetaxel at 60 mg/m² on day 1, cisplatin at 20 mg/m² on days 1-3, and capecitabine at 1 250 mg/m² on days 1-14, and the treatment cycle was repeated ever 3 weeks.

RESULTSOf the 22 patients, 14 (63%) achieved partial remission, 2 (9%) had complete remission, and 5 (23%) showed stable disease. The overall clinical response rate of the patients was 72% with a 1-year survival rate of 68%, median progression-free survival of 8 months, and overall survival of 14 months. The main toxicity was myelosuppression; 7 (32%) patients experienced grade 3/4 neutropenia, and 5 (23%) had grade 3/4 anemia. All the other adverse effects were tolerable and reversible.

CONCLUSIONThe TXP regimen is safe and effective for treatment of metastatic NPC, and the results are comparable with those of the reports in recent literatures.

Adult ; Aged ; Antineoplastic Combined Chemotherapy Protocols ; therapeutic use ; Bone Neoplasms ; drug therapy ; secondary ; Capecitabine ; Carcinoma, Squamous Cell ; drug therapy ; pathology ; Cisplatin ; administration & dosage ; Deoxycytidine ; administration & dosage ; analogs & derivatives ; Female ; Fluorouracil ; administration & dosage ; analogs & derivatives ; Humans ; Lung Neoplasms ; drug therapy ; secondary ; Male ; Middle Aged ; Nasopharyngeal Neoplasms ; drug therapy ; pathology ; Retrospective Studies ; Taxoids ; administration & dosage

A fluorescence enhancement probe (ZY8) for the detection of N2H4was designed and synthesized by employing 3-hydroxyflavone as a fluorophore,and its spectral properties had been investigated. The results showed that ZY8 had relatively good selectivity and specificity to N2H4in Tris-HCl-ethanol solution (9:1, V/V, pH 7.40). The fluorescent intensity of ZY8 exhibited good linear relationship with concentration of N2H4in the concentration range of 1.6×10-7mol/L-6.2×10-5mol/L,and its detection limit was estimated to be 1. 6×10-7mol/L. ZY8 itself had weak fluorescence, upon addition of N2H4, an approximate 9-fold fluorescence enhancement was observed, and the color of the solution changed from light grayish green to bright grass-green at UV light of 365 nm. So ZY8 might be used to the visual recognition of N2H4. ZY8 could detect N2H4in near-physiological pH range, and it had fast response and strong anti-interference ability. Moreover,ZY8 could be loaded as test paper for naked-eye detection of N2H4at mmol/L level in water solution,and it was also applied in the determination of N2H4in various water samples by the standard addition recovery experiments, with the recovery ratio ranged from 96.0% to 104.2% %, and RSD of all< 4%. The results of this study demonstrated that ZY8 had potential application to the detection of N2H4in the monitoring of environmental pollution.

OBJECTIVETo investigate the intervention effect of Danggui Shaoyao San on rats with cirrhotic ascites, and discuss the effect of arginine vasopressin (AVP) on cirrhotic ascites.

METHODMale SD rats were randomly divided into the control group, the model group, Danggui Shaoyao San low, middle and high dose groups. The cirrhotic ascites rat model was established by CCl4 combined with phenobarbital. Their urines were collected at 24 h to observe urine excretion of each group. Filter papers were used to determine the amount of ascites. The levels of serum alanine aminotransferasa (ALT) , aspartate aminotransferase (AST) were detected by the automatic biochemistry analyzer. Plasma prothrombin time (PT) was evaluated by the blood coagulation analyzer. The concentration of AVP in plasma was detected by enzyme-linked immunosorbent assay (ELISA). Pathological changes in livers were observed by HE staining.

RESULTCompared with the model group, the Danggui Shaoyao San group showed significant improvement in live indexes, with notable decrease in serum ALT and AST and the time of PT, improvement in liver pathological changes. Simultaneously, the amount of ascites decreased to varying degrees, with notable increase in urine in 24 h and decrease in AVP concentration in plasma.

CONCLUSIONDanggui Shaoyao San can notably improve liver functions of rats with cirrhotic ascites, reduce the generation of ascites and delay the progress of liver pathological changes. Its mechanism may be related to AVP.

Animals ; Arginine Vasopressin ; blood ; Ascites ; blood ; complications ; drug therapy ; physiopathology ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Liver ; drug effects ; enzymology ; metabolism ; physiopathology ; Liver Cirrhosis ; complications ; Male ; Rats ; Rats, Sprague-Dawley