Changes of glycoconjugates in human hepatocellular carcinoma

Summary Receptors of 12 lectins in 25 cases of human hepatocellular carcinomas (HCC) were histochemically investigated by avidin-biotin-peroxidase complex (ABC) methol. Liver tissues of five cirrhotic patients and five normal subjects were used as controls. SJA receptor was absent both in HCC and controls, while LCA and PSA receptors were present in all tissues studied here. Receptors of DBA, PHA, PNA, UEA_I and SBA which did not bind to normal, cirrhotic and pericarcinomatous liver tissues had the positive rates of 4%, 44%, 16%, 4% and 12% in HCC, respectively. Four lectins which strongly bound to the non-cancer liver tissues had their receptors in 96% (ConA, WGA, RCA_I) and 36% (BSA_I) of HCC. The pretreatment of tissue sections with neuraminidase abolished most of WGA receptors and exposed some PNA binding sites. There were many differences in lectin distribution between HCC and noncancer liver tissues. The changes of glycoconjugates in HCC were discussed.     

Changes of glycoconjugates in human hepatocellular carcinoma

Receptors of 12 lectins in 25 cases of human hepatocellular carcinomas (HCC) were histochemically investigated by avidin-biotin-peroxidase complex (ABC) method. Liver tissues of five cirrhotic patients and five normal subjects were used as controls. SJA receptor was absent both in HCC and controls, while LCA and PSA receptors were present in all tissues studied here. Receptors of DBA, PHA, PNA, UEAI and SBA which did not bind to normal, cirrhotic and pericarcinomatous liver tissues had the positive rates of 4%, 44%, 16%, 4% and 12% in HCC, respectively. Four lectins which strongly bound to the non-cancer liver tissues had their receptors in 96% (ConA, WGA, RCAI) and 36% (BSAI) of HCC. The pretreatment of tissue sections with neuraminidase abolished most of WGA receptors and exposed some PNA binding sites. There were many differences in lectin distribution between HCC and noncancer liver tissues. The changes of glycoconjugates in HCC were discussed.     

Identification of fucosylated glycoconjugates in Xenopus laevis testis by lectin histochemistry

Glycoconjugates play roles in many physiological and pathological processes. Previous works have shown important functions mediated by glycans in spermatogenesis, and the carbohydrate composition of testis has been studied by several approaches, including lectin-histochemical methods. However, the testis of Xenopus laevis, an animal model extensively employed in biochemical, cell and developmental research, has not yet been analysed. The aim of this work was to carry out a histochemical study of the fucose (Fuc)-containing glycoconjugates of Xenopus testis by means of lectins, combined with deglycosylation pretreatments. Four Fuc-binding lectins were used: orange peel (Aleuria aurantia) lectin (AAL), gorse seed (Ulex europaeus) agglutinin-I (UEA-I), fresh water eel (Anguilla anguilla) agglutinin (AAA), and asparagus pea (Lotus tetragonolobus) agglutinin (LTA), each recognizing different forms of fucosylated glycans. Labelling with UEA-I, which preferably binds Fucα(1,2) containing oligosaccharides, did not show any appreciable staining. LTA, specific for Fucα(1,3), and AAA, which binds Fucα(1,2), labelled spermatocytes and spermatids, but no labelling was seen when the histochemical procedure was carried out after either β-elimination (which removes O-linked oligosaccharides) or incubation with PNGase F (which removes N-linked oligosaccharides), suggesting that fucosylated glycans are of both N- and O-linked types. AAL, which has its highest affinity to Fucα(1,6), but also recognizes Fucα(1,2) and Fucα(1,3), labelled the whole testis, and the staining remained when the histochemical method was performed after either β-elimination or incubation with PNGase F. Labelling with AAL could be explained by the fact that this lectin could be binding to diverse fucosylated glycans in N- and O-glycans, and even in glycolipids. The importance of these glycans is discussed.     

Lectin histochemistry shows fucosylated glycoconjugates in the primordial germ cells of Xenopus embryos.

Previous works have shown that glycoconjugates with terminal fucose (Fuc) are located in the primordial germ cells (PGCs) of some mammals and might play a role in the migration and adhesion processes during development. The aim of this work was to identify the terminal Fuc moieties of Xenopus PGCs by means of three Fuc-binding lectins: from asparagus pea (LTA), gorse seed (UEA-I), and orange peel fungus (AAA). The histochemical procedures were also carried out after deglycosylation pretreatments: beta-elimination with NaOH to remove O-linked oligosaccharides; incubation with PNGase F to remove N-linked carbohydrate chains; and incubation with alpha(1,2)- and alpha(1,6)-fucosidase. The PGCs were always negative for LTA and UEA-I, two lectins that have the highest affinity for Fuc alpha(1,2)-linked. However, the PGCs were strongly labeled with AAA, which preferentially binds to Fuc with alpha(1,3) or alpha(1,4) linkages and to Fuc alpha(1,6)-linked to the proximal N-acetylglucosamine. There was fainter labeling with AAA when the sections were preincubated with alpha(1,6)-fucosidase, but the labeling remained strong when the sections were pretreated with alpha(1,2)fucosidase. When the beta-elimination procedure was carried out, the PGC labeling with AAA was slight. If the PNGase F incubation was performed, the PGCs remained moderately positive for AAA. These data suggest that the Xenopus PGCs have Fuc moieties in O- and N-linked oligosaccharides, including Fuc alpha(1,6) linked to the innermost GlcNAc, and that the Fuc was not in alpha(1,2)-linkage.     

Cell-surface enrichment of fucosylated glycoconjugates in the 8- to 16-cell mouse embryo

Summary The destination of ³H-fucose was examined in preimplantation mouse embryos at the 8- to 16-cell stage by autoradiography of semi-thin sections. At the end of a 16–18 h incubation in medium containing 200 Ci/ml L-[6-³H] fucose the density of silver grains over the combined surface of cells where they faced their neighbours was 40% greater than that over the outer polar cell surface and 80% greater than that over the cytoplasm. These differences were statistically significant (p<0.01). The outer cell surface also had a grain density above that found over the cytoplasm (p<0.01). There was a rapid decrease of 18–24% in the incorporated radioactivity during the first 1.5 to 3 h in chase medium containing 10 mM cold fucose. Following this the grain density remained relatively stable until it decreased further between 24 and 48 h. By 48 h there was no significant difference between the density of grains over cytoplasm, apposed cell surfaces and outer cell surface: all were reduced compared to the initial values, to 29% in the case of the surfaces and 10% in the case of the cytoplasm. These results indicate that there is an enrichment in the incorporation of ³H-fucose into macromolecules at the cell surfaces where neighbouring cells confront one another relative to the outer polar cell surface, as well as the cytoplasm, at the 8- to 16-cell stage. Conclusions about the relative density of fucoconjugates per unit length of apposed and polar membrane cannot be reached from such a light microscopic study. Fucoconjugates appear to belong to two groups: the first are turned over relatively rapidly (within 3 h after removal from radioactive medium) while the second are more stable, decaying between 24 and 48 h after removal from ³H-fucose.     

Immunohistochemical study on the expression of c-Fos and c-Jun in human skin wounds

An immunohistochemical study on the temporal expression of c-Fos and c-Jun, both of which designate proto-oncogene products, was performed on 60 human skin wounds with different post-infliction intervals. In unwounded skin, c-Fos or c-Jun was immunolocalized at the nuclei of the epidermal cells in the basal layer, hair follicle cells and sweat gland cells. During the early inflammatory phase of wound healing, the nuclei of polymorphonuclear cells (probably neutrophils), mainly infiltrating at the wound site, were labeled with anti-c-Fos or -c-Jun antibody. As the wound age increased, the neutrophils had disappeared at the wound site, and both mononuclear cells (probably macrophages) and spindle-shaped fibroblastic cells, which expressed a c-Fos or c-Jun positive reaction in the nuclei, were mainly observed. Morphometrically, the distribution of the c-Fos-positive ratio was very similar to that of the c-Jun-positive ratio; the positive ratio was considerably increased in wound specimens with a post-infliction interval of 1 day, thus indicating the late inflammatory or proliferative phase. This study showed that c-Fos and c-Jun were closely involved in the inflammatory phase as well as the proliferative phase of the wound healing process.     

A lectin cytochemical study of glycoconjugates in the human retina

Summary The binding to morphologically normal human retina of eleven biotin- or peroxidase-coupled lectins with different carbohydrate specificities was studied. Eight formalin-fixed and paraffin-embedded eyes were examined. Photoreceptor cells bound Lens culinaris (LCA), wheat germ (WGA), peanut (PNA) and Ricinus communis (RCAI) agglutinins, and concanavalin A (ConA). The outer segment region was labeled more strongly than the inner segment region, and PNA labeled only cones. All these lectins except PNA bound to both plexiform layers, and all but PNA and RCAI to the nuclear layers. Pretreatment with neuraminidase to remove sialic acid resulted in increased binding of RCAI and PNA, which now labeled both rods and cones, and in decreased binding of WGA. Bandeiraea simplicifolia (BSAI), Dolichos biflorus (DBA), soybean (SBA), Ulex europaeus (UEAI), and Lotus tetragonolobus (LTA) agglutinins, as well as pokeweed mitogen (PWM) reacted only with retinal vascular endothelial cells, which were also labeled with the other lectins. The results indicate that -mannose, -glucose, -galactose, N-acetyl-d-glucosamine and N-acetylneuraminic acid are present in glycoconjugates of human neuroretina.     

结直肠癌中HSP60和HSP27表达的免疫组织化学研究

目的1观察热休克蛋白60和热休克蛋白27在结直肠癌中的表达及意义。方法：收集结直肠癌80例,其中淋巴结转移40例（转移组）,无淋巴结转移40例（无转移组）;另外,在结直肠癌80例中,有结直肠腺瘤（腺瘤组）以及距肿块15cm以上的正常肠粘膜（对照组）各40例。应用免疫组织化学SP法检测组织中蛋白的表达。结果：HSP60的表达主要定位在癌细胞胞浆,在对照组、腺瘤组、非转移组、转移组中的表达阳性率分别为25％、30％、57．5％、90％,组间比较发现,对照组与转移组、腺瘤组与转移组、转移组与非转移组（x^2=10．912,P〈0．001）的HSP60阳性表达率存在统计学差异;而对照组与腺瘤和非转移组间以及腺瘤与非转移组间无统计学差异。HSP27的表达主要定位在癌细胞的胞浆,在对照组、腺瘤组、非转移组、转移组的表达阳性率分别为5％,35％,50％,90％,组间比较发现,对照组分别与无淋巴结转移组、淋巴结转移组;腺瘤组分别与转移组;非转移组与转移组间存在统计学差异,腺瘤与非转移组间无统计学差异。HSP60和HSP27表达间无统计学相关。结论：HSP27表达可能与结直肠癌发生和转移相关。而HSP60的表达可能在结直肠癌转移中具有重要意义。     

Immunohistochemical analysis for cell proliferation-related protein expression in small cell carcinoma of the esophagus; a comparative study with small cell carcinoma of the lung and squamous cell carcinoma of the esophagus

Small cell carcinoma is a rare neoplasm in the esophagus. To evaluate cell proliferation activity and its underlying mechanisms in this tumor, we examined immunohistochemically 5 cases of small cell carcinoma of the esophagus (SCCE) for expressions of tumor suppressor proteins, oncoproteins and cell proliferation markers including p53, p21WAF1/CIP1, retinoblastoma (Rb) protein, bcl-2, Ki-67 and PCNA, and compared the results with those of 5 cases of small cell carcinoma of the lung (SCCL) and 10 cases of squamous cell carcinoma of the esophagus (SQCE). The prevalence and labeling index of p53-immunoreactivity tended to be higher in SCCE (4/5; 56.6%) and SCCL (4/5; 79.9%) than in SQCE (6/10; 48.8%). Expression of p21WAF1/CIP1 was observed in 2 of 10 cases of SQCE. In contrast, its expression could not be detected in any cases of SCCE and SCCL examined. Expression of Rb protein was observed in 9 out of 10 cases of SQCE, but not in any cases of SCCE and SCCL. SCCE and SCCL showed more frequent and intense immunoreactivity for bcl-2 than SQCE. In expression of cell proliferation markers (Ki-67 and PCNA), no remarkable difference was observed among SCCE, SCCL and SQCE. These results suggest that SCCE and SCCL could share some genetic alternations including mutation of p53, loss of Rb gene and overexpression of bcl-2, and these may be related to the similar biological potentials between the two. Furthermore, SCCE was different from SQCE in expression of Rb protein and bcl-2, and these two types of esophageal carcinoma could arise through different molecular mechanisms.     

Sugar residues of glycoconjugates in the olfactory epithelium of the human fetus: histochemical study using peroxidase-conjugated lectins]

The oligosaccharide content of glycoconjugates was studied at the olfactory epithelium of human fetuses ranging from 8 to 12 weeks of gestation by mean of peroxidase labelled lectins (DBA, PNA, WGA, SBA, ConA, LTA, UEA I). The main results demonstrated that: 1-The olfactory epithelium (olfactory cells, supporting cells and basal cells) was generally characterized by different amount of a-D-mannose, a-D-galactosamine, a-D-glucose, D-galactose-(beta 1-3)-N-acetyl-galactosamine, sialic acid and a-L-fucose. 2-At the 11th-12th week of gestation the largest amount of sugar residues was detected at the olfactory cells and at some basal cells. 3-At the 12th week of gestation, UEA I may be considered a specific marker of the olfactory cells in different stages of development.     

Immunohistochemical study of human pterygium

The purpose of this study has been to evaluate the immunohistochemical characteristics of human pterygial tissues in order to ascertain the possible contribution of an immunological mechanism in the pathogenesis of pterygium and to investigate the presence in the pterygial tissues of some melanoma-associated antigens, in order to evaluate if there may be a small possibility of correlation of the two diseases. Human biopsy specimens of pterygium were obtained by surgery for pterygium excision. Tissue segments were fixed and processed for paraffin embedding. Microtome sections were treated for the immunohistochemical demonstration of IgA, IgM, IgG, CD3, CD20, CD68, HLA-DR, Protein S100, HMB45, and Melan A using the avidin-biotin peroxidase method or the streptavidin biotin-alkaline phosphatase method. The findings suggest that all the effector components of the mucosal immune system are present in the human pterygium and, among the most sensitive markers for melanoma, only S100 shows immunoreactivity. An immunopathogenetic mechanism seems to be responsible for the pathogenesis of pterygium, perhaps being caused by pre-existing conjunctivitis or microtrauma in combination with the patient's predisposition. No correlation between pterygium and melanoma was found.     

The expression of core fucosylated E-cadherin in cancer cells and lung cancer patients： prognostic implications

It is well documented that the glycosylation of E-cadherin is correlated with cancer metastasis, but whether E-cadherin could be core fucosylated remains largely unknown. We found that E-cadherin was core fucosylated in highly metastatic lung cancer cells while absent in lowly metastatic lung cancer cells. Sinceα-1,6 Fucosyltransferase (α-1,6 FucT) is known to catalyze the reaction of core fucosylation, we investigated the biological function of core fucosylation on E-cadherin by α-1,6 FucT targeted RNAi and transfecting α-1,6 FucT expression vector. As a result, calcium dependent cell-cell adhesion mediated by E-cadherin was strengthened with the reduction of core fucosylation on E-cadherin after RNAi and was weakened with the elevated core fucosylation on E-cadherin after α-1,6 FucT over expression. Our data indicated that α-1,6 FucT could regulate E-cadherin mediated cell adhesion and thus play an important role in cancer development and progression. Computermodeling showed that core fucosylation on E-cadherin could significantly impair three-dimensional conformation of N-glycan on E-cadherin and produce conformational asymmetry so as to suppress the function of E-cadherin. Furthermore, the relationship between the expression of core fucosylated E-cadherin and clinicopathological background of lung cancer patients was explored in lung cancer tissue of patients. It turns out to demonstrate that core fucosylated E-cadherin could serve as a promising prognostic indicator for lung cancer patients.     

Aromatase in human lung carcinoma

Lung cancer is the leading cause of cancer mortality in both women and men worldwide but gender differences exist in their clinical and biological manifestations. In particular, among life time non-smoker, female are far more likely to develop lung carcinoma than male. Recent studies demonstrated that estrogens are synthesized in situ in both male and female lung cancers through aromatase, suggesting that sex steroid may contribute to the pathogenesis and development of lung carcinoma. In addition, human lung carcinomas have been recently demonstrated to be frequently associated with expression of estrogen receptors in both male and female patients and a lower expression of aromatase was reported to be associated with better prognosis. Preclinical studies further demonstrated that aromatase inhibitor (AI) suppressed the lung tumor growth both in vitro and in vivo. These findings all suggest a potential role of intratumoral aromatase in biological behavior of non-small cell lung cancer (NSCLC), the most common form of human lung malignancy. Therefore, AIs may become viable therapeutic options for disease management in NSCLC patients but further studies are definitely required to obtain a better understanding of the potential roles of intratumoral aromatase expression as a predictive biomarker for clinical outcome in these NSCLC patients.     

Immunohistochemical demonstration of glycoconjugates bearing the type 2 chain-backbone structure in human fetal,normal and neoplastic gastrointestinal tract

Summary Immunohistochemical distributions of carbohydrate antigens based on the type 2 chain in normal as well as fetal and neoplastic tissues of human gastrointestinal tract were investigated with a monoclonal antibody (MAb) H11 (specific for type 2 chain) alone and in combination with the two MAbs MSG15 (for 26 sialylated type 2 chain) and IB9 (for the 26 sialylated type 2 chain and glycoproteins having NeuAc26GalNAc), and 188C1 (for short- and long-chain Le^x antigens) and FH2 (for the long-chain Le^x antigen). In the pyloric mucosa of secretors, the type 2 chain is oncodevelopmentally expressed, but in non-secretors it is detected in surface mucous cells of normal gastric mucosa. The 26 sialylation, which is confined to endocrine cells of normal pyloric mucosa, occurs in fetal and carcinoma tissues. Irrespective of the secretor status, the short- and the long-chain Le^x antigens can be detected in mature and immature glandular mucous cells of normal gastric mucosa, respectively; both antigens are also expressed in fetal and carcinoma tissues. In the colon, the type 2 chain and its 26 sialylated counterpart are expressed in an oncodevelopmental manner. The short- and the long-chain Le^x antigens are significantly enhanced in colonic carcinoma. The glycoproteins with NeuAc26GalNAc residues appear in gastric and colonic carcinoma as well as intestinalized gastric mucosa and transitional mucosa. Thus, some of these antigens were distinctively expressed in certain epithelial cells lining the normal gastrointestinal tract depending on maturation and patients' secretor status, and some were oncodevelopmental or carcinoma-associated antigens of the human gastrointestinal tract.Abbreviations Fuc fucose - Gal galactose - GalNAc N-acetylgalactosamine - Glc glucose - GlcNAc N-acetylglucosamine - MAb monoclonal antibody - NeuAc N-acetylneuraminic acid - PBS phosphate-buffered saline     

Immunohistochemical study on oxytocin-like substance in the human placenta

Our previous studies have demonstrated that a large quantity of oxytocin (OT)-like substance exists in human placental tissue. In the present study, the localization of the OT-like substance in the human placenta was investigated by the PAP (peroxidase and antiperoxidase complex) immunohistochemical method. The results demonstrated that the site of syncytiotrophoblast was positively stained by specific antiserum to OT, whereas the tissue was not significantly stained by normal rabbit serum (NRS). These results suggest that the OT-like substance might be localized in syncytiotrophoblast of the placental tissue.     

Immunohistochemical study of the expression of trophoblastic beta-1-globulin in the gastric epithelium of man in stomach cancer]

The expression of specific-pregnancy beta 1-globulin (SP1) was studied in gastric mucosa with chronic gastritis. The expression of SP1 was revealed in focus of intestinal metaplasia, dysplasia as well as in cover epithelial cells.     

Immunohistochemical demonstration of glycoconjugates bearing the type 2 chain-backbone structure in human fetal, normal and neoplastic gastrointestinal tract.

Immunohistochemical distributions of carbohydrate antigens based on the type 2 chain in normal as well as fetal and neoplastic tissues of human gastrointestinal tract were investigated with a monoclonal antibody (MAb) H11 (specific for type 2 chain) alone and in combination with the two MAbs MSG15 (for alpha 2----6 sialylated type 2 chain) and IB9 (for the alpha 2----6 sialylated type 2 chain and glycoproteins having NeuAc alpha 2----6Gal-NAc), and 188C1 (for short- and long-chain Lex antigens) and FH2 (for the long-chain Lex antigen). In the pyloric mucosa of secretors, the type 2 chain is oncodevelopmentally expressed, but in non-secretors it is detected in surface mucous cells of normal gastric mucosa. The alpha 2----6 sialylation, which is confined to endocrine cells of normal pyloric mucosa, occurs in fetal and carcinoma tissues. Irrespective of the secretor status, the short- and the long-chain Lex antigens can be detected in mature and immature glandular mucous cells of normal gastric mucosa, respectively; both antigens are also expressed in fetal and carcinoma tissues. In the colon, the type 2 chain and its alpha 2----6 sialylated counterpart are expressed in an oncodevelopmental manner. The short- and the long-chain Lex antigens are significantly enhanced in colonic carcinoma. The glycoproteins with NeuAc alpha 2----6GalNAc residues appear in gastric and colonic carcinoma as well as intestinalized gastric mucosa and transitional mucosa. Thus, some of these antigens were distinctively expressed in certain epithelial cells lining the normal gastrointestinal tract depending on maturation and patients' secretor status, and some were oncodevelopmental or carcinoma-associated antigens of the human gastrointestinal tract.     

Immunohistochemical evaluation of HER-2/neu expression in infiltrating breast carcinoma: a study of reproducibility

OBJECTIVE: To determine interobserver and intraobserver reproducibility in the assessment of the HercepTest- and TAB250-immunostained slides. STUDY DESIGN: Three independent expert pathologists (two with and one without training in HercepTest assessment) evaluated the HercepTest and TAB250-immunostained slides of 108 infiltrating breast carcinomas with a triple-blind method. The evaluation was repeated, with the same method and sequence of view, after 60 days. RESULTS: Expert pathologists, after adequate training in HercepTest evaluation, could reach excellent interobserver (K=.911, P<.001) and intraobserver reproducibility (K of .863-.926; P <.001 for all). The percentage of disagreement in intraobserver reproducibility ranged from 0.9% to 3.7%. Interobserver and intraobserver reproducibility in the evaluation of TAB250-immunostained slides was good (K = .658, P < .001) and from good to excellent (K of .600-.895, P < .001 for all), respectively. CONCLUSION: Optimization of the level of accuracy in HercepTest evaluation is mandatory because the decision to initiate therapy with Herceptin depends on the result. Moreover, considering that the percentage of disagreement in intraobserver reproducibility ranges from 0.9% to 3.7%, it is advisable that two expert pathologists evaluate all HercepTest slides with a double-blind method. If there are discordant results, they must be discussed by the same pathologists.     

S-100 protein in human lung neuroendocrine neoplasms. Immunohistochemical study of 14 cases and review of the literature

A group of lung neuroendocrine (NE) neoplasms are investigated in view of the possible presence of S-100 protein immunoreactivity in their cells. The selected tumours were classified according to Gould et al. (1983a) and Mosca et al. (1985). They comprise 5 carcinoids, 3 neuroendocrine carcinomas of the well-differentiated type, or peripheral carcinoids, 5 neuroendocrine carcinomas of the intermediate cell type, or intermediate-cell, poorly differentiated carcinomas, 3 neuroendocrine carcinomas of the microcytoma type, or small cell carcinomas-SCC and a nodal metastasis of microcytoma. All but 2 tumours were immunoreactive for neuron specific enolase (NSE). Few S-100 immunoreactive cells were detected in 4 out of 5 carcinoids, in 1 out of 3 peripheral carcinoids, in 4 out of 5 poorly differentiated carcinomas and in the 3 microcytomas examined. No S-100 positive cells were found in the SCC's nodal metastasis. The S-100 immunolabelled cells can be interpreted as dendritic reticulum cells migrating through the tumours. However, in one case of typical carcinoid, abundant S-100 positive cells were detected: their stellate morphology and their intimate relation with neoplastic cells suggest that they are part of the neoplasia as a sort of satellite cell.