Role of antibody to S100 protein in diagnostic pathology

Normal tissues and various tumors were examined for S100 protein, using anti-S100 protein antiserum, in an immunoperoxidase reaction. Among normal tissues, in addition to the previously reported presence of S100 protein in some neurons, glial, and Schwann cells of the nervous system, melanocytes and Langerhans cells of the skin, interdigitating reticulum cells of lymph nodes, and chondrocytes, we demonstrated it in myoepithelial cells and ducts of sweat glands, salivary glands, and the breast, serous glands of the lung, fetal neuroblasts, and sustentacular cells of the adrenal medulla. Among neoplasms, S100 protein previously has been reported in neurogenic tumors, melanomas, and neuroblastomas; we have demonstrated it in mixed sweat gland tumors, histiocytosis X, pleomorphic adenomas of the salivary gland, medullary carcinomas of the breast, bronchioloalveolar carcinomas of the lung, sustentacular cells of pheochromocytomas, teratomas of the ovary, and tumors of cartilage (enchondromas, osteochondromas, and chondrosarcomas). With S100 protein producing tumors, a normal progenitor cell was identified, indicating that demonstration of S100 protein in tumors confirmed their origin.     

Accuracy of the determination of S100 protein expression in malignant melanoma using a polyclonal antibody directed against S100 and monoclonal antibodies specific for S100 alpha and beta

Abstract

S100 proteins are present in a variety of tissues and perform regulatory functions in numerous metabolic processes. They have an important role in many human cancers, including malignant melanoma. Both polyclonal and monoclonal antibodies have been used to investigate S100 expression in melanoma tissue sections. This study aimed to determine the accuracy and sensitivity of these two types of antibodies in detecting S100 proteins in paraffin processed tissue cases of malignant melanoma. The study compared routinely used rabbit polyclonal anti-S100 antibody raised against both anti-S100A and B isoforms (Dako, Glostrup, Denmark), as per studies by Timar, and compared and contrasted findings with mouse monoclonal anti-S100A and anti-S100B antibodies (Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA). The study involved the assessment of formalin-fixed paraffin-embedded tissue blocks from 56 cases of malignant melanoma, consisting of 23 superficial spreading, nine nodular, eight lentigo maligna, five acral lentigenous forms, five metastatic melanomas (two sentinel lymph node positive cases and three cases of nodal involvement from cases of elective nodal groin dissections), and six cases of desmoplastic malignant melanoma (DMM). The slides were stained by immunohistochemical methods on an automated platform (BenchMark XT; Roche, USA) and employing the iView detection system. All slides were examined by routine light microscopy by two independent assessors. The best results for both intensity of staining and percentage of positive tumor cells were achieved with polyclonal anti-S100 antibody and monoclonal anti-S100B antibody. Anti-S100A antibody yielded weaker staining intensity (with mean intensity of 1·8, compared to 2·8 for both anti-S100B antibody and polyclonal anti-S100 antibody), and a lower percentage of positive melanoma cells (an average of 74% for anti-S100A, compared to 95% for both anti-S100B antibody and polyclonal anti-S100 antibody). This result was statistically significant (P<0·01). Staining in cases of DMM gave the same results (P<0·01). The conclusion from this study is that polyclonal anti-S100 antibody and monoclonal anti-S100B antibody are more suitable than monoclonal anti-S100A antibody for diagnostic investigations of malignant melanoma, irrespective of the histological type of melanoma.     

Monoclonal antibody to desmosomal glycoprotein 1--a new epithelial marker for diagnostic pathology

Desmosomes are intercellular adhesive junctions that occur in almost all epithelia and should therefore be useful as epithelial markers in tumour diagnosis. Here, we describe a monoclonal antibody, 32-2B, to a major desmosomal glycoprotein (dgl) which reacts with human tissues in paraffin sections. This antibody was tested for its ability to stain epithelia and tumours. It reacted with all epithelia tested and with every specimen of a wide range of carcinomas. It also stained meningiomas, another desmosome-containing tumour. It did not stain other types of tumours including lymphomas, melanomas, and various sarcomas, or normal tissues which lack desmosomes. These characteristics demonstrate that 32-2B is a reliable epithelial marker that may have a useful role in diagnostic histopathology.     

Proinflammatory properties of the human S100 protein S100A12

S100 proteins represent a new class of chemoattractants. Here we extend earlier evidence for the proinflammatory properties of human S100A12. A12 induced migration of monocytoid cells, with optimal activity at 10(-10) M and potency of >10(-9) M C5a. Neutrophils were poorly responsive, and lymphocyte migration was not affected. Actin polymerization in monocytoid cells was accompanied by a sustained [Ca(2+)]i flux of a magnitude comparable with C5a. A12 elicited a transient infiltration of neutrophils (4-8 h) and more delayed recruitment of monocytes (8-24 h) in vivo. A12 (approximately 70 nM) was present in synovial fluid (SF) from rheumatoid arthritis patients, and synovium contained A12-positive neutrophils in the sublining and interstitial region, often surrounding the perivasculature but rarely in the synovial lining layer, although some macrophages were positive. The A12 gene was transiently up-regulated in monocytes by tumor necrosis factor alpha (6 h); induction by lipopolysaccharide (LPS) was sustained (12-48 h). A12 may contribute to leukocyte migration in chronic inflammatory responses.     

Role of 100S ribosomes in bacterial decay period

Ribosomal proteins S10 and S2 were each fused with GFP to track the fates of these proteins in the stationary growth phase and the following decay period in Escherichia coli. The fused proteins localized mainly in the cytoplasm, and their amounts were proportional to the colony‐forming unit. S10‐GFP strains that lacked genes responsible for regulating 100S ribosomes and S2‐GFP strain that was unable to form 100S both showed shortened stationary phases. This result indicates that these strains exhibit earlier death in the absence of 100S formation (S2‐GFP, S10‐GFP?rmf and S10‐GFP?hpf) and breakdown (S10‐GFP?yfiA). Therefore, in addition to the mere presence of 100S, the correct timing of 100S formation and breakdown is required to maintain viability. We propose a model in which 100S acts as a tentative repository of ribosomes that are protected from degradation and provide a source of amino acids in later growth period.     

Establishment of S100A8 Transgenic Rats to Understand Innate Property of S100A8 and Its Immunological Role

The innate properties of S100A8 as a regulator in acute inflammation have not yet been elucidated in detail. Our aims are to newly establish S100A8 transgenic rats (Tg-S100A8) and to elucidate the immunological functions of S100A8. Following the treatment with 5% dextran sulfate sodium for 1 week, the body weight in Tg-S100A8 weakly decreased after the start; however, that in Japanese Wistar rats (WT) significantly decreased in the end. The serum level of CRP in Tg-S100A8 was significantly lower than that in WT, although the concentration of CRP apparently increased in both Tg-S100A8 and WT. The dynamic mobility of S100A8 and S100A9 in macrophages was microscopically observed using fluorescent immunological staining, in which the S100A9 was dominantly expressed in many macrophages in the rectal tissue of WT. As determined by PCR and real-time PCR, the levels of S100A8 messenger RNA (mRNA) in several organ tissues of the Tg-S100A8, such as heart and small intestine, were apparently higher than those of WT, respectively. The expression of IL-6 and TNF-α mRNAs was negatively regulated in main organ tissues of the large colon of Tg-S100A8 followed by down-regulation of IL-6 protein. An important result was that the expression of S100A8 mRNA was strongly induced in many macrophages of Tg-S100A8, whereas that of some inflammatory cytokine mRNAs described above were significantly reduced. Tg-S100A8 has potential as a useful experimental model rat not only for investigating the innate properties of S100A8 as a regulator, but also for clarifying its functional role in immune cells from a myeloid origin, particularly macrophages.     

Immunohistochemical localization of S100 protein in skin tumors

We applied a peroxidase-antiperoxidase technique for S100 protein to 73 tumors of skin and skin adnexa. These included 15 eccrine tumors, 11 apocrine tumors, 18 tumors with differentiation toward hair, two sebaceous adenomas, one mixed tumor of the scalp, ten dermatofibromas, ten basal cell carcinomas, five squamous cell carcinomas, and one clear cell acanthoma. Consistent results were obtained. Occasional cells in eccrine tumors showed strong positive staining, as did the Langerhans' cells in the squamous cell carcinomas and the clear cell acanthoma. The cells of the apocrine tumors showed moderate to weak staining, and the tumors with differentiation toward hair, the sebaceous adenomas, and the mixed tumor of the scalp showed uniform negative staining, as did basal cell carcinomas and dermatofibromas.     

Congenital protein S deficiencies; diagnostic difficulties

Protein S is a physiologic inhibitor of coagulation acting as a cofactor of activated protein C (APC) that inhibits factor Va and VIII. Approximately 60% of PS is bound to C4bBP, a protein of the complement system and only the free PS has a cofactor PCa role. Congenital PS deficiencies are diagnosed by immunologic dosage of free and total PS and functional assay evaluating APC cofactor activity. However, it has been demonstrated a direct anticoagulant activity of free PS, non-dependant of APC on the cascade coagulation and even PS bound to C4bBP seems to have anticoagulant properties. So, it appears that functional assays available estimate only a part of PS anticoagulant activities and, in addition, many interferences are reported with these tests (lupus anticoagulant, factor V Leiden, factor VIII excess...). Immunologic dosages are more reliable in spite of rare qualitative PS deficiencies that could be non-diagnosed. PS deficiencies are often difficult to diagnose because of an overlapping between normal and pathological values. Familial studies are necessary to prove the hereditary origin because there are several causes of acquired and sometimes persistent PS deficiencies (liver insufficiency, vitamin K absence, hormonal therapy in women, PS auto immune deficiency). About 200 different mutations were retrieved and, therefore, molecular studies are not of current practice. It is recommended currently to measure in first intention the free PS, if possible in association with PCa cofactor activity.     

人S100蛋白的表达及其抗血清的制备和脑脊液中S100含量测定方法的建立

目的：建立定量检测脑脊液（CSF）及血清中S100蛋白的方法，探讨S100蛋白的检测在辅助诊断克－雅病（CJD）中的应用。方法：利用脑cDNA文库，经PCR获得了S100基因并克隆至原核表达载体pGEX-2T上，在大肠埃希菌中表达了谷胱甘肽－S－转移酶（GST）－S100融合蛋白；融合蛋白经亲和纯化后，免疫家兔，制备抗体；抗体经纯化后，用生物素（BNHS）标记，建立了可定量检测S100蛋白的生物素－亲和素系统ELISA方法，并初步用于临床脑脊液的检测中。结果：所表达的GST－S100蛋白相对分子质量约为35000，以其为抗原制备的S100特异性抗血清具有良好的免疫反应性。建立了定量检测脑脊液中S100蛋白的双抗体夹心ELISA方法，对3例“可能性的CJD”患者（14－3－3蛋白阳性）和15例无痴呆症状患者脑脊液进行检测，结果显示，3例CJD患者脑脊液S100含量均超过2．900μg/L,而在无痴呆症状患者组中14例患者脑脊液S100含量都低于0．180μg/L。对正常人和CJD患者血清进行检测，显示S100蛋白含量个体间差异很大。结论：所建立的方法可用于脑脊液中S100蛋白的检测，进一步扩大标本量有助于明确脑脊液中S100蛋白的检测在辅助诊断CJD中的价值。     

Applications of electron microscopy to diagnostic pulmonary pathology

Viruses and other possible causative agents should be sought light and electron microscopically in all cases of ill-defined diseases including "sarcoid." Ideally, tissue should be prepared for electron microscopic examination as soon as a specimen is obtained; however, when this has not been done, tissue preserved in formalin solution can be used. Viruses, some bacteria, and other agents suspected on the basis of light microscopic findings can be verified electron microscopically by reprocessing paraffin-embedded tissue from areas that show smudge cells, focal necrosis with atypical cellular proliferation, and nuclear inclusions. Electron microscopically, all dying cells show swelling and rupture of cellular organelles and membranes; reactive changes include proliferation of branching tubules and paracrystalline and other types of proteinaceous precipitates (inclusions) in both the nucleus and cytoplasm. Qualitative and quantitative changes of cellular organelles, fibrils, microvilli, and intercellular junctions reflect hyperplasia, metaplasia, or dysplasia of the cell and may enable identification of the diseases, e.g., desquamative interstitial pneumonia. In various conditions, basal laminae become irregular, disruptive, or reduplicated following epithelial necrosis and regeneration. Electron microscopic evidence of immunologic damage to basal lamina and cells and immuno-electron-microscopic features of the lung in general require further studies. Electron microscopic features of transbronchial biopsy specimens may be diagnostic in cases of alveolar proteinosis, histiocytosis X, and amyloidosis. Ultrastructural abnormalities of cilia are common; primary ciliary defects are rare. Finally, light microscopic, scanning electron microscopic, and x-ray energy-dispersive spectrometric examinations of paraffin-embedded sections appear most practical for the pathologic evaluation of cases of pneumoconiosis.     

DNA hybridization in diagnostic pathology

DNA hybridization is becoming an important new adjunct to conventional methods for the diagnosis of infectious diseases, inherited conditions, and neoplasia. Applications of this technology require very small quantities of tissue or body fluids because the DNA probes used in the hybridization assays detect minute amounts of homologous DNA sequence in the test material. Under the proper conditions, these DNA probes are absolutely specific for the pathogen or gene being examined, and hybridization with them usually yields objective answers that require little interpretation. The relatively minor inconveniences currently associated with DNA hybridization are related to the use of radioactivity as a detection signal and the time and labor required to obtain diagnostic data. In the future, technical improvements currently being developed and the preparation of new probes for additional human and microbial genes are likely to create an increasingly larger role for DNA hybridization in diagnostic pathology.     

Urinary cytology in diagnostic pathology

In summary, the place of urinary cytology in diagnostic urologic pathology is now established. Recent emphasis on topical chemotherapy for superficial bladder neoplasms has made cytologic examination of urinary specimens essential for good patient care. Renewed interest in the method has resulted in improved preparations and diagnostic criteria. Important pitfalls in cellular analysis have been identified. New technologies, advances in therapy, and changes in classification will require continual adjustments in our approach to urinary cytology and the diagnostic cytopathologist should remain flexible while acquiring expertise in this area.     

DNA probes in diagnostic pathology

Molecular pathology has become firmly established as a distinctive discipline in medicine. It has introduced radical changes in concepts of disease causation and in classification of disease states affecting humans and other organisms. In addition, molecular pathology represents a "new" diagnostic technology with many potentials that have been heretofore untapped. This overview provides a discussion of the use of DNA probes in the study of human diseases. The role of detectable genetic abnormalities in pathogenesis will be considered, as well as their possible impact on nosology and disease classification.     

胎儿胰腺S100蛋白表达的发育

目的：探索人胚胎发育过程中胰腺S100蛋白表达的发生和分布。方法：采用免疫组织化学EnVision法,选用S100蛋白抗体对30例人胚胎胰腺进行标记。结果：S100蛋白免疫反应性纤维沿腺泡、导管和血管分布。胎龄14周胰腺内开始出现S100蛋白免疫反应性纤维样结构,并随胎龄逐渐增加,至胚胎发育后期（胎龄24-28周）达到高峰,至35周后开始减少。结论：胰腺神经系统的发育有明显的阶段性。     

诊断病理专业理论课教学的几点体会

诊断病理学是一门实践性很强的学科,是介于基础医学与临床医学之间的桥梁学科.病理诊断是运用病理学的基本概念,根据疾病的病变特点,结合临床提供一手材料,由病理科医生所做的定诊性结论.在各种影像诊断和其它先进的诊断方法高度发展的今天,许多疾病的最后确诊还要取决于病理诊断,故病理诊断在疾病诊断中被誉为“金标准”.因病理诊断具有复杂性、独特性和定性性,故病理医生必须有高超的病理诊断水平,病理诊断理论课学习尤其重要.作者在病理学理论课教学中不断进行探讨,将几点体会报道如下.