B型流感病毒B/Yamagata/16/88反向遗传操作平台的搭建及BALB/c小鼠感染模型的建立

目的 利用基因工程技术全基因合成B型流感病毒B/Yamagata/16/88的8个基因节段,并利用反向遗传技术从体外拯救B型流感病毒B/Yamagata/16/88,同时建立BALB/c小鼠感染模型,为下一步研究B型流感病毒致病机制、传播机制以及开发新型疫苗奠定基础。方法 通过基因合成和反向遗传技术体外拯救B型流感病毒B/Yamagata/16/88。全基因组测序验证拯救病毒基因组序列与Genbank序列的一致性。将拯救病毒以10⁵EID₅₀的攻毒剂量人工感染BALB/c小鼠,通过体重变化、生存率、肺脏病毒复制等方面进行致病性分析,建立小鼠感染模型。结果 成功从体外拯救出B型流感病毒B/Yamagata/16/88,命名为B-S9。全基因组测序结果表明,B-S9基因组序列与Genbank公布序列一致。B-S9能够人工感染BALB/c小鼠,但不致死,对BALB/c小鼠呈现低致病性;攻毒后第3天,B-S9感染小鼠体重出现下降,攻毒后第8天,小鼠体重开始回升;攻毒后第3天和第6天,B-S9感染小鼠的肺脏内均能检测到病毒复制,且攻毒后第3天的小鼠肺脏病毒滴度比攻毒后第6天的小鼠肺脏滴度高132倍。结论 成功搭建B型流感病毒B/Yamagata/16/88反向遗传操作平台,并建立BALB/c小鼠感染模型。目前国内外对B型流感病毒的研究比较少,该反向遗传操作平台的建立为B型流感病毒致病机制和传播机制的研究奠定了基础,同时也为包括B型流感病毒减毒活疫苗在内的新型疫苗的研制开辟了新途径。     

白附子混悬液对H22荷瘤小鼠抗肿瘤作用的免疫调节机制研究

[目的]研究白附子混悬液对荷瘤小鼠免疫功能的调节作用,探讨其抗肿瘤作用的免疫学机制。[方法]采用H₂₂荷瘤小鼠模型为研究对象,用酶联免疫吸附实验(ELISA)法检测小鼠血清中白介素-2(IL-2)和白介素-4(IL-4)水平;用流式细胞术(FACS)法分析对CD₄⁺,CD₈⁺T细胞的调节作用。[结果]白附子混悬液能够上调荷瘤小鼠血清IL-2细胞因子的表达,降低IL-4水平,可以使CD₄⁺T细胞表达升高,CD₈⁺T细胞表达降低。[结论]白附子混悬液能够通过调节荷瘤机体异常免疫功能状态而发挥抗肿瘤作用。     

p53半剂量缺失纠正1，25（OH）2D3缺乏小鼠的骨质疏松

目的：探索p53半剂量缺失杂合子小鼠能否通过增强抗氧化能力纠正活性维生素D（1,25（OH）₂D₃）缺乏引起的骨质疏松。方法：取10周龄高钙高磷饮食喂养的同窝野生型（wild type,WT）小鼠、p53半剂量缺失杂合子（p53^+/-）小鼠、1α-羟化酶基因敲除［1α（OH）ase^-/-］小鼠及p53半剂量缺失的1α羟化酶基因敲除［1α（OH）ase^-/-p53^+/-］小鼠的长骨,利用X线、micro-CT、组织病理学和分子生物学等方法,比较各组小鼠血清学、长骨骨矿化、骨形成、骨吸收以及氧化应激等表达变化。结果：与WT小鼠相比,p53^+/-小鼠血清钙、磷、甲状旁腺素（parathyroid hormone,PTH）和1,25（OH）₂D₃水平差异无统计学意义,骨密度、总胶原 （total collagen,T-col）阳性面积、成骨细胞数量、碱性磷酸酶（alkaline phosphatase,ALP）和Ⅰ型胶原（collagen typeⅠ,Col-Ⅰ）阳性面积均有所增加,活性氧水平降低,抗氧化酶SOD1表达增加。与1α（OH）ase^-/-小鼠相比,1α（OH）ase^-/-p53^+/-小鼠血清钙、磷和PTH差异无统计学意义,血清中检测不到1,25（OH）₂D₃,骨密度、T-col阳性面积、成骨细胞数量、ALP和Col-Ⅰ阳性面积均明显增加,破骨细胞数量减少,活性氧水平降低,抗氧化酶SOD1表达增加。结论：p53半剂量缺失可通过增强抗氧化能力纠正 1,25（OH）₂D₃缺乏小鼠的骨质疏松。     

人参皂苷Rb1对小鼠T淋巴细胞体外活化、增殖及凋亡的影响

目的 研究人参皂苷Rb₁对小鼠T淋巴细胞体外活化及增殖的影响,并对其免疫调节作用进行初步探讨。方法 分离制备小鼠淋巴结细胞悬液;以双色荧光抗体染色技术结合流式细胞术检测在刀豆蛋白A（ConA）的刺激下,人参皂苷Rb₁对早期活化标志CD3/CD69和调节T淋巴细胞标志CD4/CD25表达的影响;用羧基荧光素乙酰乙酸（CFDA-SE）染色结合流式细胞术和MTT法检测在ConA的刺激下,人参皂苷Rb₁对T淋巴细胞增殖的影响;DIOC/PI双染技术检测在H₂O₂作用下,人参皂苷Rb₁对淋巴细胞凋亡进程的影响。结果 终浓度为5、10、20 μmol/L的人参皂苷Rb₁对ConA刺激的调节T淋巴细胞CD4^＋/CD25^＋的表达具有促进作用,而CD3^＋/CD69^＋的表达明显下调（P＜0.01）;对ConA刺激的T淋巴细胞增殖具有明显的抑制作用（P＜0.05）,且呈剂量依赖性;人参皂苷Rb₁显著抑制H₂O₂诱导的淋巴细胞的凋亡进程（P＜0.05）。结论 人参皂苷Rb₁对小鼠淋巴细胞的体外活化及增殖均具有明显的抑制作用,是一种潜在的免疫抑制剂。     

肝脏脂肪变对胰高血糖素受体的表达影响

目的 观察胰高血糖素受体（GCGR）在ob/ob小鼠肝组织以及肝癌细胞株HepG₂细胞脂肪变后表达变化。方法 分别检测10周龄的雄性ob/ob小鼠和同窝野生对照小鼠的体重、肝功、血糖、血脂以及胰岛素等指标,用荧光定量PCR方法、Western blot法和免疫组化分别检测两组鼠肝脏中gcgr基因、蛋白表达的差异以及定位情况。不同浓度油酸处理HepG₂细胞,油红O染色观察HepG₂细胞脂肪病变程度,QPCR法和Western blot法检测细胞gcgr mRNA和蛋白的表达变化。结果 GCGR广泛表达于多种组织,以肝脏表达较多,ob/ob小鼠呈现明显的脂肪肝性病变,且免疫组化结果发现,ob/ob小鼠与正常对照小鼠相比,肝脏中GCGR表达减少;mRNA表达水平分别为0.709±0.174 vs 1.000±0.000,与对照组比较,差异有统计学意义（P<0.05）;蛋白表达水平分别为0.761±0.211 vs 1.200±0.346,与对照组比较,差异有统计学意义（P<0.05）。不同浓度梯度油酸处理HepG₂细胞,其GCGR mRNA和蛋白水平表达随油酸浓度增加而逐渐降低,差异有统计学意义（P<0.05）。结论 肝脏脂肪性病变降低了肝脏GCGR表达水平,从而可能影响肝脏的糖脂代谢。     

他克莫司联合中药热敷及激光照射改善白癜风患者的疗效及机制

观察在他克莫司软膏治疗的基础上配合中药热敷与308nm准分子激光照射对白癜风的治疗效果的影响,并探讨其机制。结果显示中药热敷与308nm准分子激光照射能改善他克莫司对白癜风的治疗效果,并且可提高患者血清CD₄⁺,CD₂₅⁺,Treg,CD₄⁺CD₂₅⁺FOXP₃⁺Treg浓度。表明他克莫司联合中药热敷、308nm准分子激光照射治疗白癜风可显著提高临床疗效,其机制可能通过调控调节性T细胞CD₄⁺CD₂₅⁺FOXP₃⁺改善机体免疫机能。     

不同品系小型猪对高致病性猪繁殖与呼吸综合征病毒的敏感性筛选

目的 筛选对高致病性猪繁殖与呼吸综合征病毒(HP-PRRSV)敏感的小型猪品系, 以用于HP-PRRS活疫苗的评价。方法 本研究选择蓝塘猪、蕨麻小型猪、巴马小型猪、五指山小型猪(白系)接种PRRSV NVDC-JXA1株强毒, 并以本地二元杂猪作为对照。攻毒后每天观察动物精神、食欲、死亡等情况, 死亡动物进行剖检, 并采取肺组织做RT-PCR检测病毒, 连续观察5周, 试验结束时采取存活动物血清分别检测PRRSV抗体。结果 攻毒后各组都有动物出现精神沉郁、食欲下降、死亡等现象, 死亡动物RT-PCR检测都显示为阳性, 存活动物PRRSV抗体检测阳性。从死亡率看, 巴马小型猪和五指山小型猪白系对PRRSV NVDC-JXA1株强毒的敏感性最高, 并且敏感性超过了对照组二元杂猪。结论 巴马小型猪和五指山小型猪白系对HP-PRRSV敏感, 可用于HP-PRRS活疫苗的检验。     

清开灵和双黄连口服液体内抗禽流感病毒作用

目的 评价清开灵和双黄连口服液体内抗禽流感病毒药效,探讨其对感染流感病毒小鼠免疫功能的影响。方法 建立H9N2亚型禽流感病毒鼠肺适应株人工感染BALB/c小鼠模型,以肺指数抑制率、生命保护率和肺病毒滴度为主要评价指标,研究清开灵和双黄连口服液对感染H9N2亚型禽流感病毒小鼠的防治效果;以脾指数、胸腺指数和T细胞亚群比率（CD4⁺/CD8⁺）为主要评价指标,探讨清开灵和双黄连口服液对感染流感病毒小鼠免疫功能的影响。结果 清开灵和双黄连口服液均能显著抑制H9N2亚型禽流感病毒引起的小鼠肺炎实变,攻毒后第4天小鼠肺指数抑制率分别为34.1%、26.3%。清开灵和双黄连口服液对感染小鼠有显著的生命保护作用,存活率分别为70.0%、60.0%,显著高于病毒对照组的存活率（30.0%）,鼠肺病毒滴度显著低于病毒组（P＜0.01）。清开灵和双黄连口服液对感染病毒后小鼠脾脏和胸腺萎缩具有显著的抑制作用,并能提升感染小鼠脾脏中CD4⁺/CD8⁺值。结论 清开灵和双黄连口服液具有显著的抗禽流感病毒作用,对病毒复制的抑制及对病毒感染导致的小鼠免疫功能下降的调节作用是其发挥抗病毒功效的重要机制。     

新型三唑类化合物Ⅱ3的镇痛作用及对环氧酶活性的影响

目的： 研究化合物Ⅱ₃的镇痛作用及其对环氧酶-1(COX-1)和环氧酶-2(COX-2)活性的影响。方法： 采用小鼠热板实验和扭体实验研究化合物Ⅱ₃的镇痛作用;利用放免法测定巨噬细胞中PGE_{2的含量,考察化合物Ⅱ3对COX-2活性的影响;测定内皮细胞中6-酮-前列腺素F1α(6-keto-PGF1α)含量,考察化合物Ⅱ3对COX-1活性的影响。结果： 化合物Ⅱ3高、中、低3个剂量组均可以延长小鼠的热痛阈值,延长造模后小鼠第一次出现扭体反应的时间并减少15 min内扭体反应出现的次数(P<0.05,P<0.01);在1×10^-5,1×10^-6,1×10^-7 mol/L浓度时均能抑制小鼠腹腔巨噬细胞PGE2的生成, 1×10^-6 mol/L浓度显著地抑制生成后的COX-2活性,在相同浓度下对新生小牛胸主动脉内皮细胞6-keto-PGF1α生成的抑制作用相对较弱。结论： 化合物Ⅱ3具有一定的镇痛作用,而且对COX-2具有选择性抑制作用。}     

PM2.5对大鼠子宫组织形态及氧化应激效应的影响

目的 探究PM_2.5对大鼠子宫组织生理及相关生化应激效应的影响。方法 将30只SD雌鼠随机分为3个不同PM_2.5暴露组（生理盐水对照组、1.5 mg/kg PM_2.5低剂量组和37.5 mg/kg PM_2.5高剂量组）。PM_2.5暴露10 d后,将雌鼠处死,HE染色观察子宫组织病理变化,并检测子宫组织中SOD、GSH、MDA和LDH含量。结果 与对照组相比,PM_2.5暴露组雌鼠子宫组织结构异常,内膜上皮细胞变薄,排列混乱;固有层基质细胞和血管减少。氧化应激指标检测结果显示,高剂量组MDA和LDH含量分别为（6.53±1.24） nmol/mg prot和（265.62±24.65） U/g prot,明显高于对照组（P<0.05）;高剂量组和低剂量组SOD和GSH含量均明显低于对照组（P<0.05）。结论 PM_2.5可对大鼠子宫组织形态造成破坏,并诱发子宫相关的氧化应激反应。     

Value of D-Dimers in patients with acute aortic dissection

Objective: To evaluatel the value of D-dimers in patients with acute aortic dissection (AAD). Methods: This study consisted of 16 patients with AAD and 27 non-AAD patients. Serum D-dimets were measured by Sta-Liatest D-DI immunoturbidimetric assay. Results: D-dimer level was higher (P ＜ 0.001) in patients with AAD(7.91 ± 5.52 μg/ml) than that in non- AAD group(1.57±1.24 μg/ml). D-dimer was positive (＞0.4 μg/ml) in all patients with AAD and in 10 control group patients (37%). Among patients with acute AAD, D-dimers tended to be higher in Stanford A than in Stanford B (8.67 ± 4.31 μg/ml vs. 3.24±1.27 μg/ml, P ＜0.01). D-dimer values tended to be higher in more extended disease(3.84 ± 1.65 μg/ml, 8.57 ± 3.58 μg/ml and 11.87 ± 5.69 μg/ml in thoracic aorta, thoracic and abdominal aorta, thoracic and abdominal aorta and iliacal arteries, respectively, P ＜ 0.05 for both 8.57 ± 3.58 and 11.87 ± 5.69 vs. 3.84 ± 1.65 ). Including the control group into the analysis, we found a sensitivity of 100%, a negative predictive value of 100%, and a specificity of 66% and a positive predictive value of 64% for D-dimer in diagnosis of AAD in our patients with suspected AAD. Conclusion: D-dimer was elevated in patients with AAD. A negative D-dimer test result could be useful in excluding AAD.     

Research of combined liver-kidney transplantation model in rats

Objective： To set up a simple and reliable rat model of combined liver-kidney transplantation. Methods： SD rats served as both donors and recipients. 4℃ sodium lactate Ringer＇s was infused from portal veins to donated livers,and from abdominal aorta to donated kidneys, respectively. Anastomosis of the portal vein and the inferior vena cava （IVC） inferior to the right kidney between the graft and the recipient was performed by a double cuff method, then the superior hepatic vena cava with suture. A patch of donated renal artery was anastomosed to the recipient abdominal aorta. The urethra and bile duct were reconstructed with a simple inside bracket. Results： Among 65 cases of combined liver-kidney transplantation, the success rate in the late 40 cases was 77.5%. The function of the grafted liver and kidney remained normal. Conclusion： This rat model of combined liver-kidney transplantation can be established in common laboratory conditions with high success rate and meet the needs of renal transplantation experiment.     

BLOOD PRESSURE CHANGE WITH AGE IN SALT-SENSITIVE TEENAGERS

Objective To observe blood pressure change with age in salt-sensitive teenagers whose salt sensitivity were determined by repeated testing.Methods Salt sensitivity was determined through intravenous infusion of normal saline combined with volume-depletion by oral diuretic furosemide in 55 teenagers. After five years, salt sensitivity was re-examined and subject blood pressure was followed up. Blood pressure changes in salt-sensitive teenagers were compared to that of non-salt sensitive teenagers over five years.Results After 5 years, the repetition rate of salt sensitivity determined by intravenous saline loading is 92.7%. In teenagers with salt sensitivity on the baseline, both the systolic blood pressure increments and increment rates were much higher than non-salt sensitive teenagers (12.7±12.1 mmHg vs. 2.8±5.2 mmHg, P＜ 0.01; 12.2%± 12.0% vs. 2.5% ±4.4%, P＜ 0.001,respectively). There was a similar trend for diastolic blood pressure (8.4 ± 6.4 mmHg vs. 3.7 ± 6.4 mmHg, P = 0.052; 13.2% ±10.6 % vs. 6.8%± 10.1%, P = 0.053, respectively).Conclusions Salt sensitivity determined by intravenous saline loading showed good reproducibility. Blood pressure increments with age were much higher in salt-sensitive teenagers than non-salt sensitive teenagers, especially in terms of systolic blood pressure.     

安心颗粒对急诊经皮冠状动脉介入术后生活质量影响的研究

目的：评价使用安心颗粒对急诊经皮冠状动脉介入术（PPCI）术后生活质量的影响.方法：将160例接受PPCI的急性ST段抬高型心肌梗死患者随机分为安心颗粒组（术前顿服安心颗粒8.8g,术后安心颗粒4.4 g/次,每日2次）和对照组（仅接受基础药物治疗）.所有患者均服用阿司匹林、氯吡格雷和阿托伐他汀.分别在入院时、出院前1d、出院后180 d时,应用心肌梗死多维度量表（MIDAS）、中文版SF-36评价量表对患者生活质量评分.并观察术后30 d以内的出血并发症、血小板减少症发生情况.结果：入院时和出院前1d,两组患者的心肌梗死MIDAS、SF-36量表评分比较无差异（P＞0.05）;出院后180 d时,与对照组比较,安心颗粒组MIDAS、SF-36评分明显减低（P＜0.05）;组内与入院时比较,两组出院前1d、出院后180 d时,MIDAS、SF-36评分均降低（P＜0.05）.两组患者在随访期间均无大量出血、少量出血、重度和极重度血小板减少症发生,安心颗粒组有4例、对照组有7例发生不明显出血（P＞0.05）.两组发生轻度血小板减少症的患者数比较无差异（P＞0.05）.结论：PPCI使用安心颗粒,能改善急性ST段抬高型心肌梗死患者的生活质量,且不增加出血风险.     

The comparative studies of the influences of Urapidil and Nicardipine on sino-atrial node function, atrio-ventricular node function and hemodynamics

Objective:To investigate the influences of urapidil and nicardipine on rabbit sinus function,atrio-ventricular node function and hemodynamics.Methods:Thirty-two Angora's rabbits were selected and randomly divided into four groups.U1 group:urapidil 0.25 mg/kg;U2 group:urapidil 0.5 mg/kg;N1 group:nicardipine 10 μg/kg;N2 group:nicardipine 20 μg/kg.All these medicine were administrated within 30 seconds.Measurements were taken before and after the administration of urapidil or nicardipine for the following data:mean blood pressure(MAP),heart rate(HR),sino-atrial conduction time(SACT),maximal sinoatrial recovery time(SNRTmax)corrected sinus node recovery time(CSNRT),index of sinus node recovery time(SNRTI),Wenckebach A-V conduction frequency (WB),and P-R interval.Results:Significant MAP and HR changes were identified in all of the four groups before and after administration of both urapidil and nicardipine.No significant changes could be found in the rest of the parameters.Intergroup analysis showed that SACT and CSNRT of N1 and N2 groups were shorter than those of the U2 group(P＜0.01);the MAP decreased(P＜0.01)and the HR increased drastically(P＜0.01).Conclusions:Neither urapidil(0.25 mg/kg,0.5 mg/kg)nor nicardipine(10μg/kg,20μg/kg)has any significant influence on rabbit sinus function or rabbit atrio-ventricular node function.Nicardipine could be a better choice than urapidil for parafunctional sinus node patients.     

Expression of OPGmRNA and ODFmRNA in the patients of hip fracture due to osteoporosis

Objective：To investigate the gene expression of osteoprotegerin（OPG） and osteoclast differentiation factor（ODF） in the bone tissue of patients with hip fracture due to osteoporosis. Methods：OPGmRNA and ODFmRNA in the bone tissue in 50 cases of osteoporosis sufferers（over 50 years old） with hip fracture（Observer Group） and 30 cases of hip facture sufferers with no osteoporosis（Control group） were analyzed with the Semi-Quantitative RT-PCR method. Results：The mRNA expressed of ODF, OPG were both high in the patients with hip fracture. In the control group, the expression of OPG mRNA was observed, while the expression of ODF mRNA was very slight. Conclusion：Aged patients contained all signals including OPG, ODF that are essential for inducing osteoclastogenesis and promoting bone resorption.     

The clinicopathological and immuohistochemical analysis of solid-pseudopapillary tumor of the pancreas： report of 9 cases

Objective：To investigate the clinical features, pathological characteristics and immunophenotype of solid-pseudopapillary tumor of the pancreas（SPTP）. Methods：Nine surgically treated cases of SPTP were retrospectively reviewed. Hematoxylin and Eosin（HE） staining and immunohistochemical staining were used to analyze all cases, and the general clinical data was collected. Results：Six patients were asymptomatic except for a palpable mass. Two patients complained of vague-epigastric pain. One patient appeared jaundice. The tumor was encapsulated and solid tissues alternately with cystic tissues. Histologically, the histological structure of solid portion was pseudopapillary with a fibrovascular core. Tumor cells were uniform and medium-sized which were arranged in sheets ets or nests or pseudopapillary patterns. Immunohistochemical studies demonstrated that SPTP proved positive in vimentin（9/9 cases）, AAT（9/9 cases）, NSE（9/9 cases）, ACT（7/9 cases）, CK20（2/9 cases）, CgA（1/9 cases）, S-100（3/gcases）, PR（4/gcases）, Syn（3/9 cases） and CD56（5/9cases）, negative in CEA and ER. Conclusion：SPTP is a tumor predominantly occurring in young women frequently without special symptoms. This tumor has various characteristical histological patterns with different immunophenotype.     

Scientific principles and rigorous processes should be followed in developing clinical guidelines for therapeutic interventions of integrative medicine

A clinical guideline for the therapeutic interventions of integrative medicine may be defined as a written document which states a series of recommendations on therapeutic interventions of integrative medicine for a special disease or condition. The guideline may provide assistance to medical professionals in making clinical decisions aimed at improving the clinical outcome of patients and reducing the costs of medical care（~＇4~. Recommendations issued by a guideline should be based on the best available evidence in both Western and Chinese medicine. For fulfilling this purpose, the development of clinical guidelines for therapeutic interventions in the field of integrative medicine should follow scientific principles and undergo a rigorous processes.     

Dynamic Changes in Serum Estradiol and Progesterone levels in Patients of Premenstrual Syndrome with Adverse Flow of Liver-qi

Objective:To probe into the influence of changes of ovarian hormones on the pathogenesis of the specific sub-type premenstrual syndrome(PMS)and reveal partial microcosmic mechanisms of adverse flow of liver-qi.Methods:Estradiol(E2)and progesterone(P)levels in serum were determined at different phases of menstrual cycle by radioimmunoassay.Results:In the group of PMS with adverse flow of liver-qi.the secretive peak value Of E2 and P at the follicular phase significantly decreased,and the secretive peak value at the luteal phase did not come into being.Conclusions:Low E2 and P secretive peak at the follicular phase and absence of secretive peak at the luteal phase is one of the microcosmic mechanisms of PMS with adverse flow of liver-qi.One of the pathophysiologic mechanisms of specific sub-type PMS is probably the continuous low level of E2and P.     

Real-time Three-dimensional Echocardiography in Assessment of Left Ventricular and Right Ventricular Volumes

Real-time three-dimensional echocardiography (RT3DE)is a new ultrasound technique that enables dynamic threedimensional visualization and quantification of the heart in real time. Investigation of feasibility and methodology of RT3DE in determining left ventricular (LV) and right ventricular (RV) volumes, RT3DE was performed in 35 normal adults using Philips SONOS 7500 system with a 2-4 MHz matrix array transducer. The 60°×60° "pyramid" volume database was obtained and analyzed on a TomTec echo workstation. Both LV and RV volumes were calculated with four 3DE methods (i.e. apical 2, 4, 8, and 16-plane) through manually tracing ventricular endocardial borders in end diastole and end systole. Stroke volumes were then calculated. LV volume was also measured by 2DE Simpson's rule using GE VIVID 7 ultrasound machine.