Helper and cytotoxic T lymphocyte responses to influenza vaccination in healthy compared to diabetic elderly

This study was designed to determine the effect of Type II diabetes mellitus in older adults on two measures of the cell-mediated immune response to influenza vaccination. Twenty-two elderly persons with diabetes mellitus were compared to 20 healthy seniors, all of whom were living independently in the community. Peripheral blood mononuclear cells (PBMC) were challenged in vitro with live influenza virus, pre-vaccination and 4 and 12 weeks post-vaccination. PBMC culture supernatants were assayed for IL-2 activity as a measure of the helper T-cell response to vaccination. The cytotoxic T-lymphocyte response was determined using an assay of granzyme B activity in PBMC lysates. Initial analysis of the data showed increased IL-2 production in post-vaccination PBMC cultures from the diabetic group compared to the healthy group. However, when vaccination histories were used in an analysis of variance, it was found that the difference between the two groups was related to vaccination history. Study subjects vaccinated one year prior to participation in this study compared to those who had not been previously vaccinated, demonstrated a significantly suppressed IL-2 response to vaccination. Type II diabetes mellitus had no effect on the IL-2 response to vaccination. The granzyme B response to vaccination was not significantly affected by previous vaccination and results were similar for the healthy and diabetic elderly groups.     

Fate of the excluded abdominal aortic aneurysm sac: long-term follow-up of 831 patients

PURPOSE: Nonresective treatment of the infrarenal abdominal aortic aneurysm by proximal and distal ligation of the aneurysm sac (exclusion) combined with aortic bypass has been previously reported. A 10-year experience with 831 patients undergoing this procedure was reviewed. METHODS: From 1984 to 1994, 831 (761 elective, 70 urgent) of 1103 patients being treated for abdominal aortic aneurysm underwent repair with the retroperitoneal exclusion technique. Perioperative morbidity and mortality, estimated blood loss, transfusion requirements, natural history of the excluded aneurysm sac, and long-term survival were all assessed. RESULTS: The operative mortality rate for patients undergoing exclusion and bypass was 3.4%. The incidence of nonfatal perioperative complications was 5.2%. Colon ischemia requiring resection occurred in 2 (0.2%) of the 831 patients. Estimated blood loss was 638 +/- 557 cc (50 to 330 cc). On follow-up 17 (2%) patients were found to have patent aneurysm sacs as detected by duplex examination. Fourteen patients required surgical intervention. No cases of graft infection or aortoenteric fistula have been noted. CONCLUSION: Retroperitoneal exclusion and bypass is a viable alternative to traditional open endoaneurysmorraphy in surgery for abdominal aortic aneurysm. Most excluded aneurysm sacs have thrombosis without any long- or short-term complications; however, in a small number of patients delayed rupture of patent aneurysm occurs, thus emphasizing the need for diligent follow-up and appropriate intervention.     

侧明151井φ139.7mm套管走向开窗侧钻小井眼技术的研究

侧明151井是中原油田φ139．7mm套管定向开窗侧钻小井眼技术攻关的第一口井。该井斜向器陀螺定向、φ139．7mm套管开窗、修窗一次成功，整个裸眼段φ116mm小井眼轨迹控制良好，完全达到设计要求，为中原油田老井侧钻、特别是小井眼套管定向开窗侧钻小井眼技术的提高打下了坚实的基础。文中详细介绍了该井的施工情况及技术特点。     

Is ataxic gait the predominant presenting manifestation of Creutzfeldt-Jakob disease? Experience of 14 Chinese cases from Taiwan

The Cre recombinase of bacteriophage P1 catalyses site-specific recombination between lox-recombination target sites both in prokaryotic and eukaryotic cells and has thus become a popular tool in genetic research. Stable, Cre-mediated integration of DNA sequences at pre-existing lox sites in the eukaryotic genome is facilitated when a Cre recombinase protein rather than a cre-expression plasmid is used to direct site-specific recombination (Baubonis and Sauer (1993) Nucleic Acids Res., 21, 2025-2029). We bacterially produced a Cre recombinase containing a nuclear localisation signal as a fusion protein with the E. coli maltose binding protein (MBP) and purified the protein by one step affinity chromatography. Subsequent cleavage with the protease factor Xa releases the Cre recombinase including the nuclear localisation signal from the maltose binding protein. Surprisingly, we found that the recombination activity of the uncleaved MBP-Cre fusion protein is virtually identical to that of the native Cre recombinase. This suggests that the MBP portion of the fusion protein behaves as a separate protein domain which does not interfere with Cre activity and can thus be used as an independent molecular tag. Additionally, the fusion protein is very resistant to proteolytic degradation and active over a wide range of temperatures. It efficiently catalyses excision and integration reactions in vitro and in eukaryotic cells. Finally, we could show that, by using MBP-Cre, it is possible to concomitantly excise a lox-flanked DNA sequence from a plasmid and integrate it into a pre-existing lox site in the genome in one transfection experiment. Vector backbone sequences which might have undesirable effects can thereby be excluded. The MBP-Cre fusion protein described here will be a useful tool not only for the catalysis of Cre-mediated recombination reactions in vitro and in vivo but also for the analysis of the mechanism of site-specific recombination.     

Site-directed mutagenesis of the arginine-glycine-aspartic acid in vitronectin abolishes cell adhesion

Vitronectin (VN), a major cell adhesion protein, is found in plasma and in the extracellular matrix. At least three distinct cell surface receptors for vitronectin belonging to the integrin superfamily have been identified in normal and neoplastic cells. Many cell adhesion ligands, including vitronectin, contain an Arg-Gly-Asp (RGD) sequence mediating, in part, the ligand-receptor interaction. These ligands bind different integrins with varying specificity and affinity. The mechanism of receptor specificity remains controversial. To determine the role of the RGD sequence in receptor specificity, we amplified the cDNA for human vitronectin from a liver cDNA library and generated two separate mutants by utilizing site-directed mutagenesis resulting in aspartic acid (Asp47) to glutamic acid (Glu47) substitution and glycine (Gly46) to alanine (Ala46) substitution. The mammalian expression vector, pZEM229R, was used to transfect baby hamster kidney cells which secreted recombinant proteins into the supernatant. All recombinant proteins were isolated by heparin-agarose chromatography and tested for interaction with three known vitronectin receptors, namely, alpha IIIb beta 3 on thrombin-activated platelets, alpha v beta 3 on human umbilical vein endothelial cells and alpha v beta 5 on Panc-1 cells. Recombinant wild-type vitronectin behaved in a fashion similar to plasma-derived vitronectin. Both the RGE-VN and RAD-VN recombinant mutant proteins showed complete loss of cell adhesion activity, regardless of the receptor. These results confirm the essential and central role of the RGD sequence in vitronectin for cell adhesion. This expression system allows further structure/function analysis of vitronectin.     

Early postischemic dantrolene-induced amelioration of poly(ADP-ribose) polymerase-related bioenergetic failure in neonatal rat brain slices

In the infant brain, ischemia-induced ionic and enzyme mechanisms may independently lead to cell death by energy depletion: resequestration of calcium mobilized from intracellular stores consumes ATP, and activated poly(ADP-ribose) polymerase (PARP) uses oxidized nicotinamide adenine dinucleotide to form polyADP-ribosyl nuclear proteins associated with DNA damage. Using 31P nuclear magnetic resonance spectroscopy, we have monitored intracellular pH and cellular energy metabolites in ex vivo neonatal rat cerebral cortex before, during, and after substrate and oxygen deprivation. In an insult that exhibited secondary energy failure and apoptosis we identified a relative 25% augmentation of high-energy phosphates at the end of recovery when the ryanodine-receptor antagonist, dantrolene, was introduced in the early (0- to 40-minute) but not late (40- to 120-minute) stage of recovery (P < 0.05). In contrast to the absence of a late dantrolene-sensitive effect, inhibition of PARP with 3-methoxybenzamide was as effective (P < 0.05) as early dantrolene, even when introduced after a 40-minute delay. The dantrolene and 3-methoxybenzamide effects on high-energy phosphates were not additive, rather the early dantrolene-sensitive effect nullified the potential 3-methoxybenzamide effect. Therefore, in this vascular-independent neonatal preparation, postischemic mobilization of calcium from intracellular stores is associated with PARP-related energy depletion. Inhibition of either of these processes confers improved postischemic bioenergetic recovery in the developing brain.     

Quality assessment of intraoperative blood salvage and autotransfusion

Intraoperative blood salvage and autotransfusion are commonly used to minimize exposure to banked blood. Although this technique has been used widely for years, data vary regarding the quality of autotransfused blood. Salvaged blood may contain plasma, residual heparin, and free hemoglobin released from damaged cells. All of these factors may contribute to the adverse sequelae sometimes seen with autotransfusion. For these reasons, we have monitored autotransfused blood to assess its quality. Intraoperative blood salvage was used during most cardiac procedures and at the discretion of the surgeon in other specialties. Blood was collected through a double lumen catheter that was anticoagulated with heparin, filtered, centrifuged, and washed with saline. A sample of the blood was removed for analysis, which included hematocrit, heparin assay, fibrinogen, and free hemoglobin levels. Over a 6-year period, 1593 patients had intraoperative blood salvage with quality assessment. The majority of patients underwent cardiac operations (941 patients, 59%), whereas 243 had orthopedic (15%) and 208 had vascular (13%) procedures. Additionally, there were 127 pediatric patients (8%) and 74 miscellaneous procedures (5%). The highest average yield of salvaged blood was during vascular procedures (1073 +/- 76 mL), whereas orthopedic cases had the lowest yield (378 +/- 19 mL) and hematocrit (39%). There was minimal residual heparin activity, even in patients requiring systemic anticoagulation (0.3 to 0.5 units/mL). Patients undergoing pediatric procedures had the lowest concentration of free hemoglobin (476 mg/L), whereas all adult patients had higher free hemoglobin levels, especially vascular operations (990 mg/L). Intraoperative salvaged blood has minimal heparin activity, even in procedures requiring systemic anticoagulation. Fibrinogen, a marker of residual plasma, was undetectable in the majority of cases. These data indicate that intraoperative blood salvage generally results in a high-quality product (good hematocrit, low heparin, minimal plasma), although there are significant differences in free hemoglobin levels depending on the operative procedure.     

The IPCS Collaborative Study on Neurobehavioral Screening Methods: V. Results of chemical testing. Steering Group

The goal of the International Programme on Chemical Safety (IPCS) Collaborative Study on Neurobehavioral Screening Methods was to determine the intra- and inter-laboratory reliability of a functional observational battery (FOB) and an automated assessment of motor activity in eight laboratories worldwide. The control data were crucial to the outcome of the studies in terms of sensitivity and reliability of the test measures, which in turn impact on the between-laboratory comparisons of chemical effects. In addition, analyses of control data can aid in determining endpoints that may require modification to improve their sensitivity and reliability. The control data from the eight laboratories were examined in terms of the following parameters: 1) control variability within studies for each laboratory; 2) within-laboratory replicability of control values across studies; 3) within-laboratory stability of control values over the course of testing for a given study; and 4) between-laboratory comparisons of parameters (1), (2), and (3). The analyses indicated considerable differences across endpoints, wherein some measures showed high variability and little replicability, while others were extremely reproducible. Generally, there were similar ranges of variability and replicability of control data across laboratories, although in some cases one or two laboratories were markedly different from the others. The physiological (weight, body temperature) and neuromuscular (grip strength, landing foot splay) endpoints exhibited the least variability, whereas the subjective assessments of reactivity varied the most. These data indicate a reasonable degree of comparability in the data generated in the participating laboratories.