Induction of heme oxygenase-1 expression inhibits platelet-dependent thrombosis

Heme oxygenase-1 (HO-1) plays a key role in protecting tissue from oxidative stress. Although some studies implicate HO-1 in modulating thrombosis after vascular injury, the impact of HO-1 on the rate of clot formation in vivo is poorly defined. This study examined the potential function of HO-1 in regulating platelet-dependent arterial thrombosis. Platelet-rich thrombi were induced in C57BL/6J mice by applying 10% ferric chloride to the exposed carotid artery. Mean occlusion time of wild-type mice (n = 10) was 14.6 +/- 1.0 min versus 12.9 +/- 0.6 min for HO-1-/- mice (n = 11, p = 0.17). However, after challenge with hemin, mean occlusion time was significantly longer in wild-type mice (16.3 +/- 1.2 min, n = 15) than HO-1-/- mice (12.0 +/- 1.0 min, n = 9; p = 0.021). Hemin administration induced an approximately twofold increase in oxidative stress, measured as plasma thiobarbituric acid reactive substances. Immunohistochemical analysis revealed that hemin induced a robust increase in HO-1 expression within the carotid arterial wall. Ex vivo blood clotting within a collagen-coated perfusion chamber was studied to determine whether the accelerated thrombosis observed in HO-1-/- mice was contributed to by effects on the blood itself. Under basal conditions, mean clot formation during perfusion of blood over collagen did not differ between wild-type mice and HO-1-/- mice. However, after hemin challenge, mean clot formation was significantly increased in HO-1-/- mice compared with wild-type controls. These results suggest that, under basal conditions, HO-1 does not exert a significant effect on platelet-dependent clot formation in vivo. However, under conditions that stimulate HO-1 production, platelet-dependent thrombus formation is inhibited by HO-1. Enhanced HO-1 expression in response to oxidative stress may represent an adaptive response mechanism to down-regulate platelet activation under prothrombotic conditions.     

A comparative study of the JAM test and 51Cr-release assay to assess the cytotoxicity of dendritic cells on hematopoietic tumor cells

Dendritic cells (DCs) are potent antigen presenting cells and possess a direct anti-tumor cytotoxic ability. Nevertheless, the mechanism of anti-tumor cytotoxicity by DCs and the methods for its evaluation are not fully elucidated. In order to clarify this mechanism of cytotoxicity, we examined the ability of DCs 1) to suppress [³H] thymidine (³H-TdR) uptake by tumor cells; 2) to induce cytolysis on ⁵¹Cr-labeled tumor cells; 3) and to induce DNA fragmentation on ³H-TdR labeled tumor cells (JAM test). Cytolysis and DNA fragmentation are markers of necrotic and apoptotic mechanisms of cytotoxicity in vitro, respectively. DCs inhibited approximately 38.6% to 54.8% of the growth of B4D6, NB4, U937, and Daudi cells as evaluated by the uptake of ³H-TdR. However no cytolysis was verified by ⁵¹Cr-release assay. On the other hand, cytotoxicity rates found using the JAM test ranged from 3 to 81% depending on the cell line and the effector to target cell ratio. The discrepancy of cytotoxicity between ⁵¹Cr-release assay and the JAM test may be due to the phagocytosis of apoptotic tumor cells or the absorption of released ⁵¹Cr by DCs surrounding the target cells. In conclusion, the JAM test was more sensitive than the 4-h and the 10-h ⁵¹Cr-release assay to investigate cytotoxicity mediated by DCs toward hematopoietic tumor cell lines in vitro.     

Utilization of the 2017 diagnostic criteria for hEDS by the Toronto GoodHope Ehlers–Danlos syndrome clinic: A retrospective review

The new 2017 diagnostic criteria for hypermobile Ehlers–Danlos Syndrome (hEDS) provide a framework for diagnosing hEDS but are more stringent than the previous Villefranche criteria. Our clinical experience at the GoodHope EDS clinic was that the 2017 criteria left many highly symptomatic patients without a diagnosis of hEDS. We conducted a retrospective cohort study to confirm our clinic experience and assess the accuracy of the 2017 diagnostic criteria for hEDS in patients who had a previous hEDS diagnosis based on the Villefranche criteria. Our study found that 15% (n = 20 of 131) of patients with a prior diagnosis of hEDS met the 2017 diagnostic criteria, and many of the traits used to distinguish hEDS were not significantly more frequent in patients who met 2017 criteria versus those who did not. In both groups objective systemic manifestations were found less frequently than subjective systemic manifestations. Beighton score (BS) as assessed by primary care practitioner was found to be higher than assessment by EDS practitioner in 81% (n = 74 of 91) of cases. Generalized joint hypermobility was confirmed in only 46% (n = 51 of 111) of patients who had a previous diagnosis of hEDS. Higher BS did not correlate with increased number of systemic manifestations in our cohort. Common comorbidities of hEDS were found with similar frequency in those who met 2017 criteria and those who did not. Based on our cohort, the 2017 hEDS diagnostic criteria require refinement to improve its diagnostic accuracy.     

动物实验模型中多极导管心室除颤系统电除颤对心脏功能的影响

评价新型的双极和三极导管自动心室除颤系统电除颤对左心室收缩和舒张功能的影响。动物麻醉后,在X光机指导下,分别在10只犬心脏内装置双极导管自动除颤系统（组Ⅰ）;在10只猪心脏内装置三极导管自动除颤系统（组Ⅱ）;并行电除颤试验。使用食管超声心动图在电除颤前后记录二维、M型和多谱勒超声图像。组I动物接受4次电除颤,电量为64J;组Ⅱ接受平均8次电除颤,电量为210J。结果显示：左室收缩面积分数、左室等容舒张时间和二尖瓣血流E波与A波速度比值以及时间-流速积分比值等反映左室舒缩功能的指标在两组动物除颤后均无显著改变。研究表明：两种经静脉导管自动心室除颤系统中反复低能量心内膜电除颤对左室舒缩功能无明显损伤作用;研究结果为经静脉多极导管自动心室除颤系统在临床的应用和电生理研究提供了可靠的实验数据。     

Peptides derived from IgE heavy chain constant region induce profound IgE isotype-specific tolerance

(BALB/c × SJL)F1 mice, perinatally injected with peptide-N-glyconase F-treated, deglycosylated IgE heavy chain or recombinant IgE heavy chain (CH?2-CH?4), were profoundly inhibited in antigen-specific IgE production. There exist minimally two tolerogenic IgE peptides, residing in the CH?2 and CH?4 domains. Peptide I, generated by V8 protease, comprises 39 amino acids within CH?2, beginning at amino acid 103. Peptide E begins at amino acid 312 of the CH?4 domain and extends through the CH?4 domain. The total lack of antigen-specific IgE responses in IgE peptide-treated mice was not due to overproduction of interferon-γ, nor lack of interleukin (IL)-4, as predicted by the Th2/IL-4 paradigm for IgE production. IgE-tolerant mice exhibited comparable levels of circulating anti-IgE antibodies to those of PBS-treated control mice. IgG obtained from sera of both sources failed to inhibit IgE responses in vitro. Moreover, IgE responses of spleen cells from IgE peptides-treated mice were restored by CD4⁺ T cells from PBS-treated control mice. We hypothesize that regulation of antigen-specific IgE responses is mediated by CD4⁺ T cells which normally recognize IgE peptides on IgE precursor B cells, and can be rendered tolerant by perinatal IgE peptide treatment.     

高效凝胶色谱一步法制备级纯化单克隆抗体

作者采用ＰｈａｒｍａｃｉａＳｅｐｈａｃｒｙｌＳ—３００凝胶色谱柱，建立了ＩｇＧ类ＭｃＡｂ的一步法制各级纯化方法。该法是将ＭｃＡｂ腹水直接上样，用ｐＨ７．４１０ｍｍｏｌ／ＬＰＢＳ洗脱（流速０．５ｍｌ／ｍｉｎ），即得到纯化的ＭｃＡｂ。一次上样量４０～５０ｍｌ腹水，回收率为８５％－９０％．整个纯化周期４ｈ。纯化的ＭｃＡｂ经ＳＤＳ—ＰＡＧＥ测定，纯度＞９０％，免疫组化ＡＢＣ法测定活性为１：８００００（７．８×１０－１１ｍｏｌ／Ｌ）。该法操作简单、快速，只要有一台核酸／蛋白检测仪，便可进行制备级水平的纯化。     

采用IL－6cDNA控针检测人IL－6基因表达

采用缺口平移技术对ＩＬ—６ｃＤＮＡ进行了生物素和同位素标记，证明两种标记的ＩＬ－６ｃＤＮＡ（３０ｎｇ／ｍｌ）均可检出低至５ｐｇ的同源ＤＮＡ，而模拟探针（５００μｇ／ｍｌ）对高达１００ｎｇ的同源序列无此反应。应用ＩＬ－６ｃＤＮＡ对富含ＩＬ－６ｍＲＮＡ的标本进行斑点杂交或Ｎｏｒｔｈｅｒｎ杂交，证明人参三醇皂甙促进了淋巴细胞ＩＬ—６基因表达（４倍），人胎脾单个核细胞、人胎脑组织匀浆和人胎胰岛细胞内均含有高浓度ＩＬ—６ｍＲＮＡ。     

Epidemiology of community-acquired Staphylococcus aureus bacteremia.

Methicillin-resistant Staphylococcus aureus (MRSA) is an important nosocomial pathogen which has been isolated with increasing frequency in recent decades. Community-acquired MRSA (CA-MRSA) infections have also become increasingly important in recent years. This study retrospectively analyzed the risk factors, duration of hospitalization, yearly trend and seasonal variation in prevalence, and antibiotic susceptibility of isolates of community-acquired S. aureus (CASA) bacteremia and CA-MRSA bacteremia from patients treated in a teaching hospital in northern Taiwan. A total of 104 clinical isolates of CASA bacteremia were collected between January 1999 and December 2001. Among these, 35 (33.7%) were identified as MRSA. After multivariate analysis, the independent risk factors for developing CA-MRSA bacteremia were diabetes mellitus (p=0.028), chronic obstructive lung disease (p=0.037), and renal insufficiency (p=0.041). Only 6 (17.1%) patients in the MRSA group had no identified risk factors. Most of the isolates of CA-MRSA had a high degree of resistance to most antibiotics, including clindamycin (71.4%), trimethoprim-sulfamethoxazole (65.7%), and chloramphenicol (41.2%). No major trend or seasonal variation in the prevalence was found during the study period. No difference in mortality related to resistance pattern was found. Although CA-MRSA is not the major pathogen in community-acquired bacteremia, it should be included in the differential diagnosis of Gram-positive bacterial bloodstream infection, especially in those patients with risk factors. Early empiric therapy with glycopeptides in these patients may reduce morbidity and mortality.     

Interobserver variability in determining MIB-1 labeling indices in oligodendrogliomas

Several studies have shown that MIB-1 labeling indices correlate well with tumor grade and prognosis in a variety of tumor types. Several factors are responsible for some degree of variability in the determination of labeling indices. Interobserver variability is one of the factors often cited as responsible for this variability. A slide from each of 30 oligodendrogliomas, stained with MIB-1 antibody, was distributed to six pathologists. The same set of slides was reviewed by each individual. Each pathologist was instructed to determine a MIB-1 labeling index by evaluating 1,000 tumor cell nuclei from the area of the slide with the most staining. The labeling index record reflected a percentage of positive-staining tumor cells. Interobserver agreement was compared. MIB-1 labeling indices ranged from 0 to 45.7. Overall agreement was good (> or =0.75) with a concordance coefficient of 0.832 (confidence interval, 0.700 to 0.909). Variability was greater among tumors with higher labeling indices as compared with tumors with labeling indices closer to 0. The overall agreement of MIB-1 labeling indices, while not perfect, was good. The generally minor variability among observers may be related to differences in the area of the slide evaluated and in differing lower thresholds for interpreting positivity. Further improvement of concordance may theoretically be attainable by further training and discussion among observers.