聚合酶链反应（PCR）对单纯疱疹病毒性脑炎的临床诊断价值

应用聚合酶链反应（ＰＣＲ）检测技术对１１８例颅内感染性疾病患者及３７例无神神经系统疾病患者脑脊液（ＣＳＦ）中的单纯疱疹病毒ＤＮＡ（ＨＳＶ－ＤＮＡ）进行了检测及分型。结果提示：“散发性脑炎”组的阳性率为３８．４６％（２０／５２），细菌、真菌性脑膜炎、其它病毒性脑炎组以及无神经系统疾病组均为阴性。作者认为：①本检测是目前单纯疹病毒性脑炎（ＨＳＥ）较为简便而准确的早期诊断方法之一；②ＨＳＶ分型检测，对病原诊断更具有全面性；③两型单纯疱疹病毒均可引起ＨＳＥ。     

Biochemical analysis of plasma-soluble invariant chains and their complex formation with soluble HLA-DR

The invariant chain (CD74) is preferentially localized in the cytoplasm and regulates the loading of exogenous derived peptides into HLA class II heterodimers. In addition, a small proportion of CD74:class II complexes is also expressed on the cell surface. We identified and quantified soluble CD74 (sCD74) molecules in the plasma and sCD74:sHLA-DR complexes by ELISA. EDTA plasma samples from 86 healthy probands were analyzed. sCD74 could be detected in all samples with a mean concentration of 1.14 relative units±1.04 SD (range 0.17-4.31). Approximately 10% of the samples had increased amounts of sCD74 (3.0 relative units). Complexes of sCD74 and sHLA-DR were detected in all samples and their quantities were positively correlated (r=0.83, p0.001) with the sCD74 concentrations. SDS-PAGE analysis of plasma samples with high sCD74 concentrations (3.0 relative units) revealed four isoforms of sCD74 with molecular weights of 45, 43, 35, 31 kDa corresponding to known sizes of intracellular CD74. However, only molecular weights of the 45 and 43 kDa isoforms of sCD74 are found complexed with sHLA-DR. Our data demonstrate, that CD74 molecules are present in their soluble form in the plasma of healthy probands and form complexes with soluble HLA-DR molecules.     

Activation of guinea pig eosinophil respiratory burst by leukotriene B4: role of protein kinase C

Leukotriene B4 (LTB4) and the protein kinase C activator, 4-beta-phorbol dibutyrate (PDBu), both induced a pronounced and concentration-dependent stimulation of hydrogen peroxide (H2O2) generation by purified guinea pig peritoneal eosinophils in the concentration range 1 nM-1 microM. The LTB4 response was inhibited competitively by the specific LTB4 receptor antagonist, U-75302, with a KB of 25 nM, while the concentration-response curves for both stimuli were shifted rightwards (3.8-fold and 2.8-fold for LTB4 and PDBu, respectively) by the competitive protein kinase C inhibitor, 1-O-hexadecyl-2-O-methylglycerol at a concentration of 300 microM. LTB4 appears, therefore, to induce respiratory burst in eosinophils via a receptor-mediated mechanism involving protein kinase C.     

Extracorporeal membrane oxygenation (ECMO) as lung or heart assist

Extracorporeal membrane oxygenation (ECMO) may serve as extracorporeal lung assist (ECLA) in patients with acute respiratory failure (ARF) or as extracorporeal heart assist (ECHA) in patients with low output syndrome (LOS) after open heart surgery. From 1988 to 1992 seven patients underwent ECMO in our hospital; four suffered from ARF and three from LOS. Various bypass techniques were employed. Two ARF patients, aged 58 and 18 years, had veno-venous bypass; in the latter, ECMO was reinstituted as a veno-arterial bypass one week after weaning. In a three-year-old boy, the ECMO outflow tubing was primarily connected to the pulmonary artery, and shortly afterwards relocated to the common carotid artery. In a 31-year-old man with ARF, and three LOS patients, a 56-year-old woman, and two men aged 68 and 70 years, ECMO was veno-arterial with direct access to the ascending aorta. A heparin-coated system was used, and all but one patient, who was treated with warfarin, received a daily low dose of heparin, which was withdrawn after from one to nine days.
Six patients were weaned off ECMO after 4.5 to 21 days. Three ARF patients recovered completely; the child died. In one LOS patient, ECMO was withdrawn due to a poor general condition. Two others were weaned off ECMO and the intra-aortic balloon pump, and the inotropic support was significantly reduced, but both died of multiple system organ failure. Although no firm conclusions can be drawn from these few case reports, the heparin-coated system used as ECLA appears promising, whereas ECHA seems to imply a poor prognosis in patients who are not candidates for cardiac transplantation.     

High level production of soluble single chain antibodies in small-scale Escherichia coli cultures

We have investigated the effect of growth and induction conditions on the production of soluble single-chain Fv antibody fragments in Escherichia coli under the control of wt lac promoter. The scFv was directed into the periplasmic space by a pelB leader sequence. Addition of sucrose to the medium gave a 15–25-fold increase in the yield of soluble scFv-phOx (3.0 mg/l) for bacterial shake-tube cultures and an increase of 80–150-fold (16.5 mg/l) for shake-flask cultures. Using flask culture in the presence of 0.4 M sucrose, a significant amount of scFv was released into the medium. We found that the scFv could be made to accumulate in the periplasm or be secreted into the medium by simply changing the incubation conditions and the concentration of the inducer. The ratio between soluble antibody fragments and insoluble scFv aggregates proved to be dependent on the strength of the promoter. Lowering the incubation temperature below 20°C had no effect on the yield of soluble antibody fragments in the periplasm, but they were no longer secreted into the medium. An example of high level production in shake-flask cultures and one-step purification by immobilized metal affinity chromatography (IMAC) is described for a soluble scFv specific for the T cell surface antigen CD3. The biological activity of the purified anti-CD3 scFv was demonstrated by flow cytometry. This method should be especially useful for the functional screening of a large number of clones in small-scale cultures.     

一种直接高效克隆PCR产物的载体制备

Ｂｌｕｅｓｃｒｉｐｔ经ＥｃｏＲ Ｖ酶切后，在Ｔａｑ酶作用下加入ｄＴＴＰ使其３’端加上碱基“Ｔ”而成为３’端突出的粘性末端载体。用此载体直接与ＰＣＲ产物连接进行克隆。克隆效率比采用平端载体克隆的效率提高约５０倍。     

Factors affecting the neonatal response to artificial surfactant

Objective : To define the individual neonatal response to the artificial surfactant, Exosurf, and factors that may influence the response.
Methodology : Eighty-two consecutive, preterm neonates with respiratory distress syndrome, who received Exosurf at <12h of age were studied. Their response was categorized from the graphical change in the oxygenation index with postnatal age, for 12h after each of two doses of surfactant and assessed independently by two observers. Clinical factors were analysed for their effect on the four pre-defined categories of response, namely: none; mild; good; relapsed; and good: sustained.
Results : Within the first 12h, 11% of the neonates showed no response, 5% a mild response and 84% a good response, but 34% relapsed. By 24h, 6% still showed no response (all died), 11% showed a mild response and 83% a good response, of whom half relapsed. At 24h, no response was significantly associated with low gestational age and asphyxia mild response with less severe lung disease. According to the response there was a gradation in the risk of death during the first week.
Conclusions : The response to Exosurf can be individually and reproducibly categorized and demonstrated that 83% of neonates had a good response but half relapsed. No response was associated with extreme prematurity and asphyxia     

Electrode Configurations for a Respiratory Impedance Measurement Suitable for Rate Responsive Pacing

A rate responsive cardiac pacemaker will benefit the patient only if the input parameter is a reliable indicator of exercise, Part of the reliability is dependent upon selection of the best physiological parameter and part is dependent upon the measurement technique. Twenty-six patients have been studied in order to ascertain the best measurement configuration for an intravascular impedance respiratory monitor. Four electrode configurations have been examined at rest and during exercise. The results have been compared using measures of sensitivity and specificity. The ideal configuration requires that a current field be established between the right ventricular blood and the pacemaker case. And, that voltage sensing be done between an electrode in the SVC and the pacemaker case.     

成牙本质细胞中上游刺激因子1对骨桥素表达调控的研究

目的检测成牙本质细胞中上游刺激因子1（upstream stimulatory factor 1，USF1）对骨桥素表达的影响。方法培养成牙本质细胞MDPC-23，稳定转染PCMV-USF1和酸性-USF（A-USF）质粒，提取总RNA，半定量反转录聚合酶链反应检测骨桥素的表达水平，并对结果进行统计学分析。结果筛选出稳定转染USF1和A-USF的抗性克隆，USF1、A-USF转染组和对照组骨桥素相对灰度比值分别为：60．33％±4．51％、229．33％±7．09％、110．00％±15．62％，转染组与对照组差异有统计学意义（P〈0．01）。结论USF1可调控成牙本质细胞骨桥素mRNA转录，该作用被A-USF部分阻断。