An improved fixation method for transmission electron microscopy for the histological study of the lens.

An improved fixation technique for transmission electron microscopic observation that enables good fixation of all areas of the rat lens was devised. Immersion fixation with 0.1 M phosphate-buffered 1.25% glutaraldehyde at 4 degrees C for 12 hours followed by postosmication produced good results for all areas of the lens--anterior, equatorial, and posterior zones. The technique was particularly suitable for maintenance of the shape of the lens since practically no irregularity, vacuolar degeneration, or expansion of the intercellular spaces was noted. This technique, which requires only a relatively short time, was also useful for the detection of early ultrastructural changes associated with cataract in spontaneously diabetic WBN/Kob rats. We anticipate that our procedure will be widely applied.     

Trehalose 6,6'-dimycolate (cord factor) of Mycobacterium tuberculosis induces foreign-body- and hypersensitivity-type granulomas in mice

Granulomatous inflammation is characterized morphologically by a compact organized collection of macrophages and their derivatives. It is classified as either a hypersensitivity type or a foreign-body type. Lipid components of the Mycobacterium tuberculosis cell wall participate in the pathogenesis of infection. Strains of M. tuberculosis have cord factor (trehalose 6,6'-dimycolate [TDM]) on their surface. To clarify host responses to TDM, including immunogenicity and pathogenicity, we have analyzed the footpad reaction, histopathology, and cytokine profiles of experimental granulomatous lesions in immunized and unimmunized mice challenged with TDM. In the present study, we have demonstrated for the first time that TDM can induce both foreign-body-type (nonimmune) and hypersensitivity-type (immune) granulomas by acting as a nonspecific irritant and T-cell-dependent antigen. Immunized mice challenged with TDM developed more severe lesions than unimmunized mice. At the active lesion, we found monocyte chemotactic, proinflammatory, and immunoregulatory cytokines. The level was enhanced in immunized mice challenged with TDM. This result implies that both nonimmune and immune mechanisms participate in granulomatous inflammation induced by mycobacterial infection. Taken together with a previous report, this study shows that TDM is a pleiotropic molecule against the host and plays an important role in the pathogenesis of tuberculosis.     

A mouse monoclonal antibody, S2n8, detects a 140 kDa protein on the surface of human endometrial stromal cells and decidual cells

We developed a mouse monoclonal antibody, S2n8, by immunizingmice i.p. with human decidual cells collected in the first trimesterof pregnancy. By indirect immunofluorescence staining of frozensections, S2n8 was found to react with decidual cells and endometrialstromal cells throughout the menstrual cycle, but not with endometrialglandular cells or with the endometrial surface epithelium.Judging from the fluorescence intensity, the antigen expressionon stromal cells was weak in the proliferative phase, and becamestronger in the secretory phase. Decidual cells in the firsttrimester of pregnancy and decidual cells at term showed strongexpression of this antigen. Indirect immunofluorescence stainingof enzymatically dispersed decidual tissue revealed that theS2n8 antigen was expressed on the decidual cell surface. Flowcytometric analysis of 12 freshly prepared stromal cell-enrichedcell suspensions showed that 74.8–94.2% (mean ±SD 86.1 ± 6.6%) of the cells carried the antigen. Theexpression of S2n8 antigen on cultured stromal cells was enhancedby the addition of oestradiol and/or progesterone. The antigenicmolecule was purified by immunoaffinity chromatography fromdecidua collected in the first trimester of pregnancy, and themolecular weight was estimated to be 140 kDa. These findingsindicate that the S2n8 antigen is a useful cell surface markerfor stromal cells/decidual cells and is associated with theirdifferentiation. cell surface antigen/decidual cells/endometrial differentiation/endometrial stromal cells/monoclonal antibody     

IMMUNOHISTOCHEMICAL LOCALIZATION OF TYPE I, II, III, AND IV COLLAGENS IN THE LUNG

Type specific rabbit antibodies to bovine type I, 11, 111, and IV (basement membrane) collagens showing no cross-reaction with other types of collagen were prepared by cross-adsorption and diethylamiuoethyl-cellulose romatography. The antibodies to bovine type I and I11 collagens showed a high cross-reaction with the corresponding human collagens, but those to type I1 and IV collagens did moderate and no cross-reactions with human type I1 and IV collagens, respectively. By using these antibodies, tissue distribution of various types of collagen in normal bovine lung was examined by indirect immunofluorescence microscopy. Both type I and I11 collagens were found to distribute widely in the interstitium of bronchial tree, bronchial
lamina propria and of interlobules as well as alveolar nipples and adventitia of pulmonary arteries. Type I1 collagen was located only in bronchial cartilage. The tissues mainly stained for type 11 collagen were the alveolar interstitium (also stained faintly for type I collagen) and the intima and media of the arteries. Type IV collagen was located in a membranous fashion in alveolar septa and bronchial smooth muscles and subepithelial layers as well as capillaries and the intima and media of arteries. ACTA PATHOL. JPN. 31: 601–610, 1981.     

Changes in the P100 latency of the visual evoked potential and the saccadic reaction time during isometric contraction of the shoulder girdle elevators

We investigated changes in the P100 latency of the visual evoked potential (VEP) and the saccadic reaction time (SRT) in relation to the degree of activity of the shoulder girdle elevators. Muscle force was set in 10% increments from 0% to 50% of the maximal voluntary contraction (MVC). The VEP was derived from a midline occipital electrode with reference electrodes on the ears when the right retina was stimulated through the eyelid by light emitting diodes while the eyes were closed. The P100 latency of the VEP was defined as the time from the stimulus onset to the main positive peak. The SRT was defined as the latency until the beginning of eye movement toward the lateral target, which was moved at random time-intervals. P100 latency was shortened until 30% of the MVC, and which it lengthened. The SRT changed in a pattern similar to that observed for the P100 latency. The ratio of the shortening in P100 latency relative to that of the SRT was approximately 20%. All data is presented as the mean value, plus the standard deviation. We believe that the information processing time in the neural pathway from the retina to the visual cortex was shortened up to a certain muscle force of the shoulder girdle elevators, and then this processing time lengthened. These findings indicate that shortening of information processing time in the neural pathway beyond the visual cortex is included in the shortening of the SRT.     

Influence of blood collection on incidence of 2,7-FAA induced rat leukemia.

The oral administration of N, N'-2,7-fluorenylenebisacetamide induced leukemia, especially mature granulocytic leukemia, in rats. The peripheral blood was examined on various schedules, once or twice a week for a long term. The blood volume lost by a blood collection was little, but it was not negligible in small animals such as rats when the loss was repeated. We investigated the relationship between the volume of blood loss and the incidence of leukemia. The incidence of leukemia rose as the volume of blood loss increased. There was a positive correlation between them. The induction of mature granulocytic leukemia was thought to be increased by the promotion of the granulopoiesis which had been suppressed by 2,7-FAA. It was concluded that the blood loss by repeated blood collection for examination raised the incidence of 2,7-FAA induced leukemia.     

Production of CA125 by human lung cancer cell lines

CA125, which until recently was considered an ovary specific tumor marker, is elevated in the serum of patients with many pathological conditions, including lung cancer. In order to investigate the production of CA125 by human lung cancer cell lines, cell culture and immunochemical staining were performed in three cell lines. Our results showed the cell surface expression of CA125 in both adenocarcinoma and large cell carcinoma cell lines and the production of CA125 in culture medium. This is considered as evidence for in vitro production of CA125 by human lung cancer, and suggests that CA125 elevation is not only the result of ovarian cancer but may be due to other pathological conditions, including lung cancer.