The adaptation of the plaque reduction assay for measuring specific cytotoxic cells in limiting dilution cultures

The use of the original haemolytic plaque reduction technique to measure cytotoxic T lymphocytes (CTL) has been developed further as a rapid screening assay, particularly suitable for limiting dilution analyses. Using hybridoma cells as targets, the cytotoxicity has been measured by the loss of haemolytic plaque formation and by the reduction of the amount of haemolytic monoclonal antibody secreted from viable target cells into the assay supernatants. The assessment of large numbers of cytotoxic samples has been greatly facilitated by quantitating the amount of haemoglobin released in the assay with an automated microELISA multiscanner and by scoring visually using a modification of the spot test. Using these new techniques, relatively high frequency estimates of cytotoxic cell precursors in an allogeneic response (1 in 462 spleen cells) and an anti-fluorescein response (1 in 3970 spleen cells) were obtained.     

Characterization and cross-reactivity of rabbit antisera to plaque toxins

The effects of heat labile, high molecular weight water-soluble toxins from bacterial plaque on HL60 promyelocytic cells were examined. On gel filtration, four inhibitors of HL60 cell growth and two inhibitors of HeLa cell growth (PT1, PT2) were detected. The first and third HL60 cell inhibitors corresponded to the two HeLa cell inhibitors. The last eluted HL60 cell inhibitor (plaque leukotoxin, PL) did not inhibit HeLa cell growth. Anti-PT2 antibodies reduced the activity of enriched PT2 by 20-50%, but all other antisera tested exhibited no effect. Anti-PL antibodies detected antigens from Actinobacillus actinomycetemcomitans, although anti-A. actinomycetemcomitans and anti-Capnocytophaga sputigena antibodies did not react with plaque extract. These findings suggest that the plaque toxins examined in this study were probably not derived from these two bacteria.     

False results associated with darkground microscopy of subgingival plaque

This study considers false results which may arise due to problems in the preparation or examination of specimens for darkground microscopy of subgingival plaque. Subgingival plaque samples obtained with a sterile curette were placed in 0.1-0.3 ml sterile full or 1/4 strength Ringer's solution: 0.85% saline, 1% gelatin in 0.85% saline, formal saline or pyrogen-free water for injection. Test slides were prepared from the original dispersion, and control slides from the corresponding sterile solution. Optimal dispersion solution, syringe dispersion frequency and the effect on motility of delay in processing samples were tested. Slides were also prepared from dispersions of 11 representative subgingival "periodontopathic" organisms. Problems in sampling included variability in counts between sites with comparable pocket depths, contamination of the sample and reduction of the sample volume after scaling. Problems in dispersion included contamination, uneven distribution of the different morphotypes and destruction of delicate organisms. Problems in slide preparation included slide contamination, limitation in the number of samples that can be assessed by one examiner at a given time without loss of activity of motile cells, and preparation of a cell monolayer. Problems in identification and counting included confusion of Brownian movements with motility, coccoid particles with cocci, spirochetes with campylobacter, flagella with flagella-like structures, size of cocci, counting of fragmented spirochetes and non-motile flagellated organisms and motile cells, and also bias in counting. Problems in morphotype grouping included the observation that many (10 of the 11 representative) periodontitis-related organisms were in the non-motile groups and not all cells of the motile species (Campylobacter, Capnocytophaga) showed motility. The results indicate that each stage of subgingival plaque darkground microscopy, sampling, dispersion, slide preparation, counting, morphotype grouping and interpretation may lead to false results if not representative or reproducible. Procedures are suggested for the minimisation of problems in the preparation and examination of subgingival plaque specimens for darkground microscopy.     

社区呼吸康复治疗在慢性呼吸系统 疾病患者干预管理中的应用*

摘 要： 目的：探究中老年人颈动脉斑块与血清25羟维生素D （25-OH-D） 的相关性。 方法：选取2019年1月—2020 年 12 月自愿参与该研究的上海市浦东新区北蔡社区常住居民 412 人为研究对象,测定及记录其一般临床资料及血清 25-OH-D 等实验室检测结果。依据血管 B 超结果将研究对象分为有斑块组 268 人和无斑块组 144 人,比较两组人群血清 25-OH-D水平差异,用Pearson相关性分析各变量的关系,采用logistic回归分析颈动脉斑块形成的危险因素。结果：有斑块 组血清 25-OH-D 为 （45.18±18.71） nmol/L,无斑块组为 （56.12±19.54） nmol/L,两组差异有统计学意义 （χ2=5.573,P＜ 0.05）。相关性分析显示颈动脉斑块与收缩压、HbA1c、年龄呈正相关 （r值分别为0.388、0.119和0.128,P值均＜0.05）;与 血清 25-OH-D 呈负相关 （r=-0.365,P＜0.01）。血清 25-OH-D 是颈动脉斑块形成的独立相关因素 （OR=0.973,95%CI： 0.960,0.985,P＜0.05）。结论：低水平血清25-OH-D是颈动脉斑块形成的独立危险因素。     

颈动脉粥样斑块与冠状动脉病变

目的 研究颈动脉粥样斑与冠状动脉病变的关系。方法 将82例冠心病待排和冠心病患者按冠状动脉造影结果分成正常组和冠脉粥样病变组,同期超声检测颈动脉粥样硬化程度。结果 颈动脉粥样斑块与冠脉弱粥样硬化之间有较密切的关系。颈动脉粥样斑块对狭窄性冠状动脉病变预测的敏感性为71.7%,特异性为91.9%,阳性和阴性预测值分别为91.7%和73.9%,其准确性为81.7%。结论 颈动脉超声检测可作为预测冠状动脉粥样硬化有价值的指标。     

登革病毒甲基纤维素微量蚀斑技术的建立

目的建立登革病毒甲基纤维素微量蚀斑技术。方法将4株登革病毒接种于C6/36细胞内,5-7d后染色,观察蚀斑情况并计数。将测得值与组织培养半数感染量测定结果比较。结果登革病毒甲基纤维素微量蚀斑技术重复性好,比组织培养半数感染量法敏感。结论甲基纤维素徽量蚀斑技术可靠易行,可用于登革病毒滴定。     

Immunohistological study of senile brains by using a monoclonal antibody recognizing β amyloid precursor protein: significance of granular deposits in relation with senile plaques

Summary Immunochemical analyses revealed that a monclonal antibody Am-3 recognized amyloid precursor protein (APP) in senile plaques extracted from Alzheimer's brain, but did not recognize amyloid protein. Immunohistochemically, however, the staining pattern of Am-3 in frozen section of Alzheimer's brain was almost the same with that of rabbit polyclonal antibody to amyloid peptide which could recognize both amyloid protein and APP. In other words, APP was present in senile plaques of various types, cerebrovascular amyloid and granular deposits. The granular deposits were 5–10 m in size and laminarily distributed in the 1st, 3rd and 4th layers of cerebral cortex. They were especially abundant in 1st and 4th layers where senile plaques were usually fewer in number. Although the distribution in the cerebral cortex was different between the senile plaques and the granular deposits, the number of the granular deposits was well correlated with that of senile plaques. The granular deposits were negative in Congo-red birefringence, but contained amyloid protein as well as APP fragment judging from positive staining by both Am-3 and polyclonal antibody to synthetic amyloid peptide. Thus, they could be regarded as pre-amyloid.     

Amyloid-P-component-like immunoreactivity in β/A4-immunoreactive deposits in Alzheimer-type dementia brains

An immunohistochemical study using the mirror-image technique was performed in order to establish whether amyloid P component is involved in the mechanism of deposition of amyloid fibrils in senile plaques (SPs) in Alzheimer-type dementia (ATD). Ninety percent of /A4 protein-immunoreactive SPs were also stained by the anti-amyloid P component immunchistochemistry, and this applied to all of the diffuse, primitive and classical types of /A4 deposits. These findings may suggest an involvement of amyloid P component in the formation of amyloid fibrils in senile plaques in ATD brains.     

Polyclonal 111In-IgG, 125I-LDL and 125I-endothelin-1 accumulation in experimental arterial wall injury

To test iodine-125 labelled low-density lipoprotein (¹²⁵I-LDL), polyclonal indium-111 labelled immunoglobulin G (¹¹¹In-IgG) and iodine-125 labelled endothelin-1 uptake in metabolically active atheromatous plaques after arterial wall injury, we performed balloon de-endothelialization of carotid arteries or abdominal aortas in 24 New Zealand male rabbits which were fed with a normal diet (n=14) or a hypercholesterolaemic diet (n=10) after surgery. Six weeks later the animals were injected with 200 Ci of (¹²⁵I-LDL), and/or with 100 Ci of ¹¹¹In-IgG or with 9 Ci of ¹²⁵I-endothelin-1. Forty-eight hours later the animals were sacrificed. Carotid arteries and aortas were removed, counted and fixed for autoradiography and light microscopy examination. Contralateral carotid arteries and thoracic aortas served as controls.Significant ¹¹¹In-IgG uptake was observed in the injured arteries at autoradiography, with localization mainly in the healing edges, and at well counting. The percentage of the injected dose per gram (%D.inj/g) was 0.0188±0.06 versus 0.0059±0.003 in controls (P< 0.05). There was no difference in ¹¹¹In-IgG uptake between arteries with injury alone and those with active atheroma formation at the site of the injury. Significant (¹²⁵I-LDL), uptake was observed only when lipid deposition was present at light microscopy (%D.inj/g of 0.0024±0.0005 vs 0.0010±0.0003 in controls, P < 0.05). ¹²⁵I-endothelin-1 accumulation was observed in four of five injured aortas both at autoradiography, with diffuse localization, and at well counting (%D.inj/g of 0.0012±0.0004 in the abdominal aortas vs 0.0008±0.0003 in the thoracic aortas).Polyclonal IgG may accumulate in injured arteries without active atheroma formation. Inflammatory reaction at the site of the injury may cause ¹¹¹In-IgG uptake independently of atheromatous plaque formation. LDL accumulation takes place only with active atheroma formation at the site of the injury. Use of labelled peptides such as endothelin-1 may provide further insight into the mechanisms of atheromatous plaque formation.     

颈动脉斑块与心脑血管阻塞性疾病关系的临床探讨

目的 探讨颈动脉斑块与心脑阻塞性疾病及视网膜中央动脉阻塞(CRAO)的关系。方法 1995年10月至2002年7月采用彩色多普勒超声对我院心脑血管疾病患者149例行颈动脉检查。结果 检出颈动脉斑块者74例，未检出斑块者75例。有斑块者较无斑块者颈动脉内膜中层厚度明显增加(P＜0．01)。结论 颈动脉斑块严重程度的增加，其内膜中层厚度亦呈增厚趋势。前者是造成脏器梗死的重要原因之一。颈动脉斑块的检测和治疗可作为心脑血管及CRAO疾病的重要预防措施。