登革病毒悬液芯片分型检测方法的建立

目的 建立一种基于悬液芯片的登革病毒(dengue virus,DV)检测方法,可对四种血清型登革病毒进行快速检测和鉴定.方法 依据GenBank上4种病毒的基因序列信息,设计并合成相关引物及探针序列.抽提病毒RNA,经反转录后对目的基因进行PCR扩增,产物与核酸探针微球组杂交后于Bio-PlexTM 200系统检测荧光信号值.结果 DV1的悬液芯片检测敏感性约9 DNA拷贝,DV2、DV3、DV4的悬液芯片检测敏感性约90 DNA拷贝.进而将本方法用于检测15份临床标本,其检测结果与分型荧光RT-PCR一致.结论 建立了可同时检测四种血清型登革病毒的悬液芯片检测方法,为快速筛查和鉴定登革病毒提供了新的手段.     

磁珠耳穴贴压治疗偏头痛临床观察

目的比较磁珠耳穴贴压治疗与服用西药治疗偏头痛的疗效差异。方法将偏头痛患者200例随机分为磁珠耳穴贴压组（观察组）和西药组（对照组）,比较其对头部疼痛改善情况。结果观察组总有效率94.6%,明显优于对照组的78.6%（P〈0.01）。结论磁珠耳穴贴压治疗偏头痛疗效肯定,总有效率优于西药治疗。     

流式微球分析技术检测人心肌钙蛋白T方法的建立

目的建立流式微球分析技术（cytoometric bead assay,CBA）检测人肌钙蛋白T（cTnT）方法。方法用鼠抗人的cTnT的单克隆抗体包被在已激活的羧基化聚苯乙烯微球上,然后再用包被好的微球与检测标本进行抗原抗体免疫反应,并加入羊抗人cTnT的多克隆抗体和异硫氰酸荧光素（FITC）标记的驴抗羊的IgG,室温避光免疫反应一定时间后洗涤一次,上流式细胞仪检测FITC的荧光强度,以此测定标本中cTnT含量。结果通过正交试验,在100μl反应体系中,微球含量大约为1.25×106,选择10μg的鼠抗人cTnT单克隆抗体为最佳加入量,加入标本和抗体后,选择避光反应时间为120 min,洗涤选择一次,选择8μg/ml羊抗人cTnT多克隆抗体和20μg/ml FITC标记驴抗羊IgG为最佳工作浓度;方法检测灵敏度为16.0 pg/mL,线性范围是16-5 000 pg/mL,批内重复性变异系数为5.2-11.3%,回收率是94.7-111.5%,高浓度的胆红素、胆固醇和甘油三脂无明显干扰,与Roche Elecsys的电化学发光法有较好的相关性。结论自建CBA检测cTnT技术性能较好,可在临床工作中推广使用。     

Maternal country of birth and previous pregnancies are associated with breast milk characteristics

Populations in high infectious exposure countries are at low risk of some immune-mediated diseases such as Crohn's disease and allergy. This low risk is maintained upon immigration to an industrialized country, but the offspring of such immigrants have a higher immune-mediated disease risk than the indigenous population. We hypothesize that early life exposures in a developing country shape the maternal immune system, which could have implications for the offspring born in a developed country with a low infectious load. The aim of this study was to investigate if exposures in childhood (indicated by country of origin) and subsequent exposures influence immunologic characteristics relevant to stimulation of offspring. Breast milk components among 64 mothers resident in Sweden, 32 of whom immigrated from a developing country, were examined using the ELISA and Cytometric Bead Array methods. Immigrants from a developing country had statistically significantly higher levels of breast milk interleukin-6 (IL-6), IL-8 and transforming growth factor-β1. A larger number of previous pregnancies were associated with down-regulation of several substances, statistically significant for soluble CD14 and IL-8. The results suggest that maternal country of birth may influence adult immune characteristics, potentially relevant to disease risk in offspring. Such a mechanism may explain the higher immune-mediated disease risk among children of migrants from a developing to developed country. Older siblings may influence disease risk through the action of previous pregnancies on maternal immune characteristics.     

Calcium gluconate as cross-linker improves survival and shelf life of encapsulated and dried Metarhizium brunneum and Saccharomyces cerevisiae for the application as biological control agents

Calcium chloride (CC) is the most common cross-linker for the encapsulation of biocontrol microorganisms in alginate beads. The aim of this study was to evaluate if calcium gluconate (CG) can replace CC as cross-linker and at the same time improve viability after drying and rehydration, hygroscopic properties, shelf life and nutrient supply. Hence, the biocontrol fungi Metarhizium brunneum and Saccharomyces cerevisiae were encapsulated in Ca-alginate beads supplemented with starch. Beads were dried and maximum survival was found in beads cross-linked with CG. Beads prepared with CG showed lower hygroscopic properties, but a higher shelf life for encapsulated fungi. Moreover, we demonstrated that gluconate has a nutritive effect on encapsulated fungi, leading to increased mycelium growth of M. brunneum and to enhanced CO₂ release from beads containing Saccharomyces cerevisiae. The application of CG as cross-linker will pave the way towards increasing drying survival and shelf life of various, especially drying-sensitive microbes.     

Serum biomarkers of vascular cognitive impairment evaluated by bead-based proteomic technology

To investigate the biomarkers of vascular cognitive impairment (VCI), we analyzed 30 VCI patients and 30 healthy controls using proteome technology. Peptides in sera were purified using magnetic beads, and analyzed by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and ClinProTool software. Twenty-four peptides were significantly differently expressed between VCI patients and controls. There were seven peptides selected for our diagnostic model. The sensitivity of the model measured by internal and external validation was 95% and 80% respectively, while the specificity was 100% for both. A peptide peak at 4963 m/z was chosen for identification by nano-liquid chromatography–electrospray ionization–tandem mass spectrometry (nano-LC–ESI–MS/MS). The amino acid sequence was consistent with the fragment of the trace-amine associated receptor 6 (TAAR6) in the human international protein index database. In conclusion, the specific fragment of TAAR6 may be important for clinical biomarkers in the diagnosis of VCI. The high specificity and sensitivity of the model may provide an appropriate method for screening VCI patients.     

两种纯化大鼠毛囊干细胞方法的比较

目的:显微分离技术分别与流式细胞仪分选和免疫磁珠分选两种方法联合使用,比较不同方法的优缺点,以进一步探"索毛囊干细胞的体外分选培养的条件.方法:显微分离鼠毛囊隆突部,消化成单细胞悬液.分别利用流式细胞仪和免疫磁珠分选毛囊干细胞.观察细胞形态并检测分选前后细胞的纯度,活性,回收率,进行免疫荧光鉴定.结果:两种方法均能分选出实验所需的毛囊干细胞.流式细胞仪分选细胞纯度高,活性稍弱;免疫磁珠法分选细胞纯度稍低但活性强.结论:流式细胞仪分选和免疫磁珠分选均能有效获取毛囊干细胞.显微分离技术可根据实验需求选择不同的分选方法搭配.     

Deoxynivalenol exacerbates viral bronchopneumonia induced by respiratory reovirus infection.

The trichothecene mycotoxin deoxynivalenol (DON), a frequent contaminant of cereal grains, is known to dysregulate mucosal and systemic immunity. In this study, we tested the hypothesis that DON interferes with the murine immune response to viral respiratory infection. Female Balb/c mice (5 weeks old) were orally gavaged with DON (10 mg/kg body weight [bw]) or saline vehicle and then intranasally instilled with 10(7) plaque-forming units of reovirus serotype 1, strain Lang (T1/L). At 10-day postinstillation (PI), both viral titers and reovirus L(2) gene expression were 10-fold higher in lungs of DON-treated mice than in saline controls. The lowest observed effective DON dose that impaired viral clearance was 2 mg/kg bw. Although DON amplified reovirus-induced interferon (IFN)-beta and IFN-gamma mRNA responses in lung, the toxin suppressed mRNA expression for IFN-alpha, IFN-alphabeta receptor (IFNAR), and IFN-gamma receptor (IFNGR). DON also impaired induction of two type 1 IFN-dependent antiviral genes, double-stranded RNA activated protein kinase R (PKR) and oligoadenylate synthase 2 (OAS2). Respiratory reovirus infection caused a mild bronchopneumonia in mice which was markedly exacerbated by DON as evidenced by severe inflammatory cell infiltration, marked alveolar damage, and a higher volume density of intraepithelial mucosubstances in pulmonary airways. At 3- and 7-day PI, elevations in total protein, MCP-1, TNF-alpha, total cells, macrophages, neutrophils, and lymphocytes were observed in bronchoalveolar lavage fluid (BALF) of control mice infected with reovirus. DON markedly enhanced viral-induced elevations of protein, MCP-1, TNF-alpha, and inflammatory cells in the BALF at 3-day PI. DON exposure also upregulated induction of reovirus-specific immunoglobulin A (IgA) in BALF, fecal pellets, and serum. DON's effect on BALF IgA was preceded by elevated IL-6 expression and secretion in the lung. Taken together, the results suggest that DON compromised resistance to respiratory viral infection. Reduced expression of IFNAR and type 1 IFN-mediated genes in the lung might contribute to DON impairment of pulmonary reovirus clearance, whereas exacerbation of bronchopneumonia and IgA responses corresponded to increased MCP-1, TNF-alpha, and IL-6 expression.     

头颈部肿瘤术后即刻的功能及外形修复

目的探讨组织瓣移植在头颈部肿瘤术后即可修复中的可行性和应用价值。方法观察48例头颈肿瘤术后组织瓣移植,分析所采用皮瓣的类型受区血管,皮瓣成活情况及并发症的发生情况,并分析有可能影响皮瓣成活的各种因素。结果前臂皮瓣、胸大肌皮瓣、腓骨肌皮瓣、股外侧皮瓣为最常用的4种组织瓣,占全部皮瓣的95%,组织瓣成活的临床成功率为96%,术后血栓的发生率为6%,受区和供区总的并发症发生率为10%,但大多数并发症均不严重,没有造成严重的后果。结论头颈部肿瘤术后组织瓣移植是安全可靠的,整形与修复的开展能扩大手术的切除范围,提高生存率,减少局部复发率。