Objective To determine diazepam, nitrazepam, oxazepam, estazolam, and al-prazolam simultaneously in human plasma by reversed phase high-performance liquid chromatography (RP-HPLC). Methods Ten microliter carbamazepine (50 mg/L)as the internal standard was added into 1 mL sample, which contained the 5 mixed sedative hypnotics as standard substance and human plasma as ground substance. They were extracted with acetoacetate from plasma samples, and then were dissolved by 100 μL mobile phase. The blood drug levels were analyzed by high perform-ance liquid chromatograph with 20 μL sample injection on a chromatographic column C 18 (4.6 mm×250 mm)at 30℃. The mobile phase consisted of methanol and water (65:35) , and the flow rate was 1.0 mL/min. The ultraviolet detection wavelength was 230 nm. Results The linearity range of the 5 drugs was 5～1 200 μg/L(r≥0.9966, P<0.05). The recovery rate was 95.5%～105.6%. The extraction recovery rate was more than 75%. The relative standard deviation (RSD) of intra-day and inter-day was less than 10% (n=5). Conclusion RP-HPLC method is convert-ient, accurate and sensitive for simultaneous determination of the concentration of diazepam, nitraze-pam, oxazepam, estazolam, and alprazolam in human plasma.

Objective To construct the recombinant plasmid that highly expressed human subcellular PTEN,in order to provide a basis for further study on its anti-tumor effect. Methods PTEN cDNA was amplified by RT-PCR based on mRNA of placenta.The PCR product was ligated into T-vector,and transfected into E.coli;the obtained T-PTEN plasmid was identified with restrictive digestion and sequencing.PCR was used to incorporte nuclear signal of localization(NSL) into PTEN when T-PTEN was used as template.Then the PCR product was ligated into T-vector,and transfected into E.coli,and T-NSL-PTEN plasmid was obtained.pcDNA3.1 and T-NSL-PTEN were ligated after digested with EcoRⅠand BamHⅠ,and transfected into E.coli,the recombinant vector pcDNA3.1-NSL-PTEN was obtained,and identified with digestion and sequcncing.Results The recombinant expression vector DUM-PTEN and PUM-NSL PTEN were identified by restrictive digestion and DNA sequencing.As expected,by EcoRⅠ and BamHⅠ digestion,it showed the band of 1 200 bp.The sequencing result showed the NSL was incorporated successfully.The recombinant pcDNA3.1-PTEN was obtained with 1 200 bp,the sequencing result showed that its sequence was same as target gene;the recombinant pcDNA3.1-NSL-PTEN was comfirmed by restrictive digestion and sequencing,and the NSL was incorporated successfully. Conclusion The recombinant expression plasmid pcDNA3.1-NSL-PTEN is constructed successfully which can highly express human subcellular PTEN.

Objective To investigate the infection status of infectious diseases for 2 521 patients before blood transfusion in a Hospital in Changsha.Methods A total of 2 521 patients who would be transfused were selected,and six kind of serum hepatitis B virus indicators,hepatitis C virus antibody (antiHCV),human immunodeficiency virus antibody (anti-HIV/1 + 2),and treponema pallidum antibody (antiTP) of nine common infectious disease targets were detected with enzyme-linked immunosorbent assay (ELISA).Results Among 2 521 patients,HBsAg-positive cases were 8.33％,anti-HCV positive were 0.59％,anti-HIV positive [confirmed by the Provincial Center for Disease Control (CDC)] was 6 cases,and TP-positive were 2.301％.A total of 289 patients were tested positively,with a total positive rate of 11.46％.Conclusions Detection before transfusion may reduce infection risk and decrease the risk of occupational exposure,strengthen medical staff self-protection,and reduce medical malpractice caused by blood transfusion.

Objective:With proteomic approach,to find the differentially expressed proteins spots in the peripheral blood mononuclear cells(PBMCs) of healthy people and intracerebral hemorrhage patients with liver yang forming wind syndrome(LYFWS) treated by Zhenganxifeng decoction(ZGXFD) to investigate the possible proteins of LYFWS and the cure mechanism of ZGXFD.Methods: 10 intracerebral hemorrhage patients with LYFWS were treated by ZGXFD for 7-10 days.10 healthy people were selected taken as healthy controls.10ml venous blood was drawed from every subjects and PBMCs were isolated from blood by using lymphocytes separation medium.The total proteins was extracted from PBMCs.The total proteins from either intracerebral hemorrhage patients or healthy controls were prepared by means of immobilized pH gradient based on two-dimensional gel electrophoresis.After Coomassie brilliant blue G250 staining,gel-image analysis was performed by PDQuest.The differentially expressed proteins spots were identified by matrix-assisted laser desorption ionization-time-of-flight mass spectrometry(MALD I-TOF-MS).Results: Gel-image analysis revealed that there were 22 proteins spots expressed differentially.20 proteins spots were expressed differentially in the intracerebral hemorrhage patients with LYFWS as compared with healthy control,of which 8 proteins were down-regulated,7 proteins were up-regulated and 5 proteins were lost.After treated by ZGXFD,4 proteins were down-regulated,5 proteins were up-regulated and 2 proteins were lost,9 proteins weren’t expressed di erentially as compared with healthy control.8 of 20 di erentially expressed proteins spots in the intracerebral hemorrhage patients with LYFWS identi ed by MALDI-TOF-MS as compared with healthy control.These proteins are related to cell metabolism,signal transduction and so forth,of which 6 proteins were down-regulated,1 proteins were up-regulated and 1 proteins were lost.After treated by ZGXFD,2 proteins were down-regulated,1 proteins were lost,5 proteins weren’t expressed di erentially as compared with healthy control.Conclusion: 2-DE pro les of intracerebral hemorrhage patients with LYFWS in PBMCs had been established.The proteins of intracerebral hemorrhage patients with LYFWS had been identi ed,which were related to cell metabolism,signal transduction and so forth.This research presented that ZGXFD could adjust multiple di erentially expressed proteins of intracerebral hemorrhage patients with LYFWS.It was also indicated that the cure mechanism of ZGXFD was to regulate multitarget proteins.

OBJECTIVE To invesigate the prevalence of SHV-producing Enterobacteriaceae isolates in Hunan Province and identify the subtype of SHV encoding gene and the epidemiological aspect of SHV-producing isolates. METHODS Isolates were collected and identified as well as subjected to ESBLs detection.PCR and DNA sequencing were used to determine the genotype of SHV enzymes.The homology of SHV-producing strains were detected by RAPD. RESULTS Twenty-six of 171 Enterobacteriaceae isolates were confirmed to produce blaSHV genes,and 5 subtypes of SHV-type ?-lactamases were determined,including 9 strains of SHV-28,7 strains of SHV-12,7 strains of SHV-1,2 strains of SHV-11 and 1 strain of SHV-5.There were 6 RAPD types in 19 isolates of SHV-producing Klebsiella pneumoniae,5 RAPD types in 5 isolates of SHV-producing Enterobacter cloacae. CONCLUSIONS SHV-12 is the predominant genotype of SHV ESBLs producing Enterobacteriaceae in Hunan Province,and clone spread has played a certain role in SHV-type ?-lactamase producing K.pneumoniae.SHV ESBLs are not found in Escherichia coli.

Objective:To observe the temporal and spatial expression and function of mcpr1 gene during murine tooth germ development.Methods:The expression of MCPR1 at different stages of mouse tooth germ were detected by immunohistochemical staining.Results:MCPR1 expression was detected at all stages of tooth germ, but the distribution patterns at various stages were different. It indicated that the temporal and spatial expression pattern of MCPR1 during murine tooth germ development was specific.Conclusion:mcpr1 might play an important role in modulating the differentiation and mature of enamel organ.

Objective To examine the causes of high frequency sensorineural hearing loss caused by piston technique of stapes surgery.Methods The retrospective analysis involved in thirty cases(49 ears)with stapedectomy from February 2004 to 2009 at our hospital.Pure tone audiometry was performed for each patient who underwent stapedectomy both during the two days preoperation and 1,3,6 months postoperation.Results One month after operation at 8000 Hz,the air conduction hearing threshold was 8 dB less than that before,4 000 Hz average bone conduction hearing threshold was 10 dB less than that before(P < 0.01).The average high-frequency hearingthreshold of ≥40 years old group was noticeably declined than that of 40 years old group(P < 0.01).There was no association between the tinnitus and high frequency hearing loss.(P >0.05).Conclusion Transient high-frequency sensorineural hearing losses occurred to most patients under going stapedectomy,but showed significant improvement,3 months after the surgery.The average high-frequency hearing threshold of ≥40 years old group was seriously declined than that of 40 years old group.

OBJECTIVE: To explore the different and common features between adults and children with otitis media with effusion (OME) on course of disease, clinical characteristics and audiometric changes. METHOD: Pure tone audiometry, tympanogram and clinical information were analyzed and compared between 34 cases of adults and 42 cases of children with OME. RESULT: The course of disease was relatively short in children. The adenoid vegetation inflammation was more frequently found in children, while the OME occurred more often in adults. The air conduction and bone conduction threshold increased both in children and adults, particularly at high frequencies of 2 and 4 kHz. The bone conduction threshold at 4 kHz in adults was significantly higher than that in children (P < 0.05). In the cases with C type tympanogram, the negative pressure in adults was even lower than that in children (P < 0.05). CONCLUSION The OME in children is often relatively shorter in course of disease, which is mainly caused by adenoid vegetation. Sensorineural deafness can occur both in children and adults, especially at high frequencies. OME in children is less severe, however, it may lead to negative effects in learning and cognition, which should be diagnosed and managed in time.
Acoustic Impedance Tests ; Adolescent ; Adult ; Audiometry, Pure-Tone ; Bone Conduction ; Child ; Child, Preschool ; Female ; Humans ; Inflammation ; pathology ; Male ; Otitis Media with Effusion ; physiopathology ; Young Adult

OBJECTIVE: To explore the correlation between childhood obstructive sleep apnea-hypopnea syndrome and tympanogram. METHOD: A prospective clinical study was carried out. Polysomnography and tympanogram were analyzed in 72 cases. RESULT: Among the 144 ears 37 ears had type C or B tympanogram. There was no correlation between tympanogram and LSaO2. It was found correlation between tympanogram and AHI (P < 0.05). CONCLUSION Hypoxemia may affect middle-ear function. It was necessary to do the examination of acoustic impedance routinely in children with moderate and severe OSAHS.
Acoustic Impedance Tests ; Child ; Child, Preschool ; Ear, Middle ; physiopathology ; Female ; Humans ; Male ; Prospective Studies ; Sleep Apnea, Obstructive ; physiopathology

Objective To investigate the prevalence of thyroid nodules(TN)among centenarians in Hainan province and explore the association between thyroid nodules and major chronic diseases.Methods A mixed cross-sectional study of questionnaire survey,medical examination including ultrasonography and laboratory examination were conducted in elderly who resided in Hainan province and aged 100 and over,the subjects who have signed consent and complete data in both basic information and medical examination were included in this study.Eight hundred and four centenarians were finally enrolled and data of them was analyzed to investigate the prevalence of thyroid nodules and to explore its association with common chronic diseases by comparing the prevalence of chronic conditions between groups with and without thyroid nodules.Results The overall prevalence of TN was 73.5%(591、804)and the prevalence of TN was significantly higher in female when comparing with male[75.1%(505、672)vs 65.2%(86、132)](P <0.001).No significant correlation was found between the prevalence of TN and major chronic diseases (P>0.05),however,the factors associated with dyslipidemia and anemia such as TC[(4.6±1.2)mmol、L vs(4.3±1.5)mmol、L],LDL-C[(2.8±0.8)mmol、L vs(2.7±0.8)mmol、L],and Hb[(110.6±22.5)g、L vs(105.5±31.7)g、L]were significant higher in TN group than those in Non-TN group(P <0.05). Conclusions The thyroid nodule is common in centenarian population and its prevalence was higher in female than male.The associations between TN and biomarkers of dyslipidemia and anemia are found in the present study.