IMMUNOELECTRON MICROSCOPIC LOCALIZATION OF THYROGLOBULIN (TG) IN THE THYROID GLAND IN BASEDOW'S DISEASE

An electron microscopic immunohistochemical localization of thyroglobulin (TG) using PAP methods from Epok-812 embedded tissues has been made in human thyroids obtained from 15 patients with treated Basedow's disease aged 13 to 43 years. Follicular cells vary considerably in shape and content of cytoplasmic organelles depending mainly upon the size of follicles. Reaction product for TG coincided well with previous investigations studied by electron microscopic autoradiography and is strongly positive in most follicular lumina having no relation to their size, reabsorbed colloid droplets or small subapical vesicles. The reabsorbed colloid droplets are rarely observed in cells surrounding distended follicles and more frequently appeared in small follicles. The reabsorbed colloid droplets after fusion with lysosomes show a variable amount of reaction products from intense deposition to a complete absence of product indicating hydrolysis of TG by lysosomal enzymes. There is no reaction product in rough endoplasmic reticulum or Golgi apparatus in spite of positive reaction product in the subapical vesicles. These subapical vesicles are composed of both endocytotic and exocytotic vesicles.     

大鼠心肌细胞心钠素的免疫电镜定位研究

心钠素是一种由心脏分泌的激素,具有利钠、利尿等多种重要的生理功能。本文应用先进的免疫金技术,在电镜水平上对心钠素在大鼠心肌细胞中的定位和分布进行了探讨。观察结果表明,心钠素分泌颗粒呈圆形,有单位膜包绕,直径约在200-500nm之间,分布于心房肌细胞的肌浆内,尤以核旁最为多见。其次,在肌原纤维间及肌膜下亦可见到。大鼠心钠素与α-人心钠素有完全交叉反应。本文观察结果为今后利用大鼠作为动物模型,深入开展心钠素的研究提供了参考材料。     

AN IMMUNO-ELECTRON MICROSCOPIC STUDY ON INTRACELLULAR LOCALIZATION OF THYROGLOBULIN (TG) IN THE THYROID GLAND IN GRAVE'S DISEASE

Intracellular localization of thyroglobulin (TG) using a pre-embedding diffusion technique and an indirect localization sequence has been made in human thyroid obtained from 20 patients with treated Grave's disease. Both antibodies, anti-human TG-rabbit IgG F(ab')₂ and anti-rabbit IgG F(ab')₂-goat IgG F(ab')₂ fragments easily penetrated the cytoplasm of follicular cells which were dissociated by RPMI-1640 solution containing collagenase, dispase, and deoxyribonuclease. With light microscopic observation of semithin sections positive immuno-reaction for TG was demonstrated as fine granular deposits in the cytoplasm of the dissociated cells. In electron microscopic studies, intracellular antigen was well circumscribed within certain cell organelles in all cases with the positive immuno-reaction for TG being observed in perinuclear space, rough endoplasmic reticulum, Golgi complexes, secretory granules, and reabsorbed colloid droplets. Content of positive immuno-reaction product differed somewhat from one case to another and from one follicle to another even in the same case. There was no immuno-reaction product in nuclei, mitochondria, lysosomes, and lipofuscin-like granules. ACTA PATHOL. JPN. 35 : 591–603, 1985.     

LOCALIZATION OF FERRITIN IN HUMAN LIVER DISEASES STUDIED BY IMMUNO-HISTOCHEMICAL AND IMMUNO-ELECTRON MICROSCOPIC PROCEDURES

Localization of ferritin using a pre-embedding diffusion technique and an indirect localization sequence has been made in 34 cases of human liver under normal and several pathological conditions. With light microscopic observation, positive immuno-staining for ferritin was demonstrated as diffuse deposits in the hepatocytes and Kupffer cells. Intensity of the positive immuno-staining for ferritin in these cells appeared to roughly coincide with serum ferritin levels of each patient, but showed no disease specificity, although hepatoma cells contained weak deposits or were negative from immuno-staining for ferritin. With electron microscopic studies, intracellular antigen was well preserved in the hepatocytes and Kupffer cells in most cases with the positive immuno-staining for ferritin being observed in cytosol and a few cisternae of rough endoplasmic reticulum. Content of the positive immuno-staining for ferritin differed considerably from one case to another and one cell to another even in the same case. There was no immuno-staining for ferritin in hemosiderin pigment, lysosome, most of rough endoplasmic reticulum, Golgi complexes, and nucleus in both cells.     

神经垂体内5-羟色胺的免疫电镜研究

本文用免疫电镜方法研究了大鼠神经垂体内5-HT的超微结构分布。结果表明:神经垂体内的5-HT免疫反应产物定位于神经末梢的大颗粒囊泡内、小透亮囊泡表面和线粒体外膜。有时,垂体细胞胞浆也呈5-HT免疫反应阳性。5-HT免疫反应阳性的神经末梢分布于毛细血管周围和垂体细胞附迈。本文首次报道含5-HT的神经末梢可与非5-HT神经末梢形成对称性轴-轴突触。其中,5-HT末梢为突触前成分。本文结果提示:神经垂体内的5-HT不仅可通过非突触方式而且可通过突触方式释放,参与神经内分泌功能的调节。     

ENDOCYTOSIS OF HUMAN THYROGLOBULIN (TG) IN ADENOMATOUS GOITER STUDIED BY AN IMMUNO ELECTRON MICROSCOPIC PROCEDURE

Five cases of adenomatous goiter have been studied by an electron microscope using an immuno-reaction for thyroglobulin (TG) and focusing on the mechanism of endocytosis. Positive stain for TG was demonstrated in follicular lumina, large reabsorbed colloid droplets and small subapical vesicles. Endocytotic vesicles ranging from 320 nm to 1600 nm in diameter were observed in the cytoplasm as pits in the apical plasma membrane. Some of them showed direct connection with the positive stain for TG in the follicular lumen and the others were completely ingested in the cytoplasm. With statistic analysis, a majority of the vesicles showing the positive stain for TG in the cytoplasm distributed in the range of 200 nm to 1200 nm in diameter with the peak in 300 nm to 399 nm and was situated within an extent of the diameter measured from the endocytotic vesicles. Engulfment of colloid by pseudopods and fusion of the reabsorbed colloid droplets were encountered as extremely rare findings and appeared to play no major role for formation of large colloid droplets in adenomatous goiter.     

IMMUNOELECTRON MICROSCOPIC STUDY OF THYROID MICROSOMAL ANTIGEN

Using human IgG F(ab')₂ prepared from pooled sera containing a high titer of thyroid microsomal antibody (TMAb), localization of thyroid microsomal antigen (TMAg) was studied electron microscopically in dissociated follicular cells obtained from 25 thyroids of patients with Basedow's disease. Twenty out of the 25 cases were positive for serum-TMAb. In 18 of these cases, the reaction product for spontaneous membrane-bound IgG was demonstrated only on the apical plasma membrane of a small number of follicular cells, but was not observed anywhere in 5 cases without serum-TMAb. After masking the spontaneous membrane-bound IgG with anti-human IgG-goat IgG Fab, deposits of the reaction product disappeared or were significantly decreased. After the masking, all cases incubated in the above-mentioned TMAb-positive IgG F(ab')₂ showed an intense reaction product located linearly on the apical plasma membrane of a large number of follicular cells, but not on the lateral and basal plasma membrane and in the cytoplasm. On the other hand, after the masking, the control group incubated in human IgG F(ab')₂ from a normal subject revealed the same result as that in the group with masked spontaneous membrane-bound IgG. These findings indicate that TMAg is located only on the apical plasma membrane of follicular cells. ACTA PATHOL JPN 38 : 1141 -1148, 1988.     

IMMUNOELECTRON MICROSCOPIC STUDY ON THE CROSS-REACTIVITY OF ANTIBODIES TO ADULT T-CELL LEUKEMIA-ASSOCIATED ANTIGENS IN HUMAN and MONKEY SERA

The cross-reactivity of antibodies to adult T-cell leukemia (ATL)-associated antigens (ATLA) in human and monkey sera was investigated by indirect immunoperoxidase and immunoferritin methods using a human cell line (MT-2) carrying a type C virus (HTLV), two monkey cell lines (Si-1 and Si-3) carrying HTLV, and a monkey cell line (Si-2) carrying a type C virus isolated from an anti-ATLA-positive monkey. Anti-ATLA-positive but not-negative human and monkey sera gave positive immunoperoxidase reaction with all four virus-positive cell lines when studied by light microscopy. Electron microscopic findings revealed ferritin or peroxidase labeling of virus particles and plasma membranes of these four cell lines with antibody-positive but not-negative human and monkey sera. These results clearly indicate the cross-reactivity of anti-ATLA antibodies in human and monkey sera at light and electron microscopic levels, and the presence of antigenic determinants common to the surface of type C virus particles of human and monkey origin. ACTA PATHOL. JPN. 35: 863–870, 1985     

AN IMMUNO-ELECTRON MICROSCOPIC STUDY ON INTRA and EXTRACELLULAR LOCALIZATION OF ALPHA-FETOPROTEIN IN HUMAN HEPATOCELLULAR CARCINOMA

Intra and extracellular localization of alpha-fetoprotein (AFP) has been studied by an indirect peroxidase labeled antibody method using 12 cases of human hepatocellular carcinoma (HCC). With light microscopic observation, positive immuno-staining for AFP was observed in 6 out of 12 cases and demonstrated as granular or diffuse deposits in the cytoplasm of neoplastic hepatocytes. In electron microscopic studies, 8 cases showed the positive immuno-staining for AFP in the neoplastic hepatocytes. Intracellular antigen was well circumscribed within certain cell organelles with the positive immuno-staining for AFP being observed in perinuclear space, cisternae of rough endoplasmic reticulum (RER), Golgi complexes, and secretory vesicles. In addition, the positive immuno-staining for AFP was observed in bile canaliculus-like space in most cases with increased levels of serum AFP and in some cases which showed normal levels of serum AFP. Furthermore, the positive immuno-staining for AFP was also observed in intercellular, Disse's-like and sinusoid-like spaces, and micropinocytotic and lysosome-like vesicles in the endothelial cells in a few cases which showed excessively high value of serum AFP. ACTA PATHOL. JPN. 37:915–928, 1987.     

大鼠神经垂体内后叶加压素神经分泌末梢的GABA能神经支配的免疫电镜证实

为了探讨神经垂体内后叶加压素（ＶＰ）释放的γ－氨基丁酸（ＧＡＢＡ）能神经调控，本研究用包埋前ＡＢＣ法结合免疫电镜双标技术，研究了大鼠神经垂体内ＶＰ能神经和ＧＡＢＡ能神经的超微结构分布及其相互联系。先用ＤＡＢ法显示ＧＡＢＡ免疫反应，然后用钼酸铵－ＴＭＢ法显示ＶＰ免疫反应，再用ＤＡＢ－氯化钴稳定后作免疫电镜包埋。电镜观察发现：在神经垂体内ＧＡＢＡ样免疫反应产物呈电子密度高的颗粒状沉淀，定位于神经末梢内的小清亮羹泡周围和线粒体膜上；ＶＰ样免疫反应产物呈电子密度高的不规则形块状或针状散在于神经分泌末梢内。ＧＡＢＡ样神经末梢分布于神经分泌末梢之间或紧贴毛细血管和垂体细胞，可与神经分泌末梢紧密接触甚至形成突触。ＶＰ样神经分泌末梢内含小透亮囊泡和大颗粒囊泡，并可与ＧＡＢＡ样轴突末梢形成突触。在这种情况下，ＧＡＢＡ样轴突为突触前成分，突触前、后膜呈对称性，突触间隙宽度小于２０ｎｍ，内含电子致密物质。以上结果证实，在大鼠神经垂体内ＶＰ释放受到ＧＡＢＡ神经的直接突触调控。     

人甲状腺球蛋白的纯化和鉴定

新鲜甲状腺组织经硫酸铵沉淀和两次ＳｅｐｈａｄｅｘＧ－２００凝胶层析后，可获得较高纯度的甲状腺蛋白。经ＳＤＳ－ＰＡＧＥ鉴定，显示了３条区带，主带为１９Ｓ，另两条为２７Ｓ和１２Ｓ。经双向琼脂扩散和免疫电泳鉴定，提纯的甲状腺球蛋白未见血清蛋白质的污染。     

大鼠神经垂体催产素能神经元内神经分泌颗粒的免疫电镜观察

目的　观察大鼠神经垂体催产素（ＯＴ）能神经元轴突及终末内颗粒囊泡形态学及免疫细胞化学特点。方法　应用电镜包埋后免疫细胞化学方法,对大鼠神经垂体进行ＯＴ免疫染色。结果　在ＯＴ能神经元轴突终末内,可见ＯＴ免疫反应性的致密核芯大颗粒囊泡和无ＯＴ免疫反应性的清亮小泡存在。部分囊泡与轴膜靠近,并可见颗粒囊泡与轴膜融合形成的胞吐现象;在轴突非终末部分,也可见ＯＴ免疫反应性的颗粒囊泡,散在分布轴浆内,还可见无ＯＴ免疫反应性的颗粒囊泡,呈散在或聚集分布,两种囊泡均可位于微管之间。结论　位于轴突和轴突终末内的囊泡,在形态学和ＯＴ免疫染色方面有明显差异,推测它们在功能上也有不同。     

大鼠最后区内神经降压肽电镜免疫细胞化学定位

采用电镜免疫细胞化学技术观察了大鼠最后区内神经降压肽样免疫反应神经成分的超微结构。免疫反应产物要见于核周质，树突，轴突及其终末内。神经降压肽样免疫反应轴突终末与末标记树突形成轴－树突触。未标记轴突终末与神经降压肽样免疫反应树突呆构成轴－树突触。     

ELECTRON MICROSCOPIC STUDY ON MITOCHONDRIA IN HÜRTHLE CELL ADENOMA OF THYROID

In this investigation Hürthle cell adenoma of the thyroid was observed by electron microscopy. The cytoplasm of the cells consisted chiefly of an accumulation of numerous irregularly shaped mitochondria. Very few other organellae were observed. In addition electron microscopy revealed various degrees of electron density in the matrix of the mitochondria. When 3 dimensional models of the mitochondria were constructed, they revealed the mitochondria to have 3 dimentional forms which sometimes looked as if they might be dividing into two or three parts. The authors felt that this indicated a process of vigorous division of mitochondria and an extension of their life span. However, it was not possible to identify the cause of this division of mitochondria.     

人胎盘脱氢表雄酮的免疫组织化学研究

目的　观察脱氢表雄酮 (DHEA)在人胎盘的定位及其含量的周龄变化。　方法　用免疫组织化学及定量方法。　结果　人胎盘两层滋养层细胞和基质细胞均呈 DHEA免疫反应性 ,反应物质分布于胞质 ,胞核无免疫反应。在人胎盘绒毛的不同发育阶段中 ,DHEA的相对含量随周龄的增加而增加。　结论　 DHEA对胚胎及胎盘的发育可能具有重要的调节功能。     

血栓调节蛋白在成人及胎肺组织中的表达

目的 研究血栓调节蛋白在成人肺组织及胎肺组织中的表达意义。方法 以发育１８周的人胎肺组织、正常成人肺组织为研究对象,应用免疫组织化学ＳＰ法检测血栓调节蛋白的存在。结果 在１８周人胎肺组织中,血栓调节蛋白表达于围绕原如肺泡上皮细胞团周围的血和内皮细胞中,在原始肺泡上皮细胞及原始支气管上皮细胞、软骨均呈阴性表达。正常成人肺组织内的血管内皮细胞呈阳性表达。结论 血栓调节蛋白在人胎中期肺组织中未见表达。     

CRYSTALLINE STRUCTURES IN HUMAN PANCREATIC BETA CELL ADENOMA

An electron microscopic observation on a pancreatic tumor removed from a 34-year-old woman revealed the fine structural morphology of a functional beta cell adenoma. Characteristic PAS positive crystalline structures were frequently observed in the cytoplasm of the tumor cells. They were not bounded by a membrane and had a rectangular or irregular hexagonal shape. Highly regular patterns were seen as such as lattice or honeycomb and parallel ripple structures. They are similar to the Reinke's crystal or crystalline structures reported in human hepatocytes suffering from several different diseases and considered as a protein-carbohydrate complex. Occasionally, small paracrystalline structures appeared to indicate an immature type of these structures in the opaque fine fibrillar mass. Crystalline or paracrystalline structures were not detected in the normal pancreatic tissue removed with the tumor from the patient.