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PubMed2000

Fine-needle aspiration cytology of hemangiopericytoma: report of two cases.

The fine-needle aspiration biopsy (FNAB) findings in two cases of hemangiopericytoma (HP), arising in the parotid gland and on the inner chest wall, respectively, are reported. Smear preparations in each case showed cytologic features of an undifferentiated spindle-cell neoplasm, whereas a core needle biopsy specimen of the chest wall mass showed a spindle-cell tumor with a "staghorn-like" arrangement of endothelium-lined vascular channels. Immunostains performed on this core biopsy, and on the surgical resection specimens in both cases, showed positive staining of tumor cells for vimentin and CD34, with negative staining for a variety of smooth muscle, epithelial, neural, and neuroendocrine markers. Electron microscopy performed in one case further supported the diagnosis of HP. With adequate sampling and appropriate use of ancillary studies, a diagnosis of HP can be reliably suggested on the basis of FNAB and core biopsy of a soft-tissue mass.

Aged↗

PubMed1988

Flow cytometric detection of herpes simplex virus type 2 infected and transformed cells by immunoenzymatic and by indirect immunofluorescence staining.

The glucose oxidase antiglucose oxidase (GAG) immunoenzymatic staining procedure has been used to detect herpes simplex virus (HSV) antigens microscopically. In this study, the GAG procedure was adapted to cells in suspension, and its potential usefulness in flow cytometry was examined. HSV-2 infected monkey kidney and HSV-2 transformed mouse cells were stained using antisera to HSV-2 or to an HSV-2 specific protein with a molecular weight of 38 Kd, respectively, with the GAG procedure. Flow cytometric analysis of the GAG stained cells was then performed by the measurement of scattered light intensity in the angular intervals 1 degree-2 degrees, 2.5 degrees-19 degrees, and 3 degrees-6 degrees. The greatest scattered light intensity decrement caused by staining occurred in the 3 degrees-6 degrees angular interval, as predicted by previous work. In infected cells, which stain intensely by immunofluorescence, the difference between positively and negatively stained cells was adequate for detecting infected cells using the GAG method; however, this was not the case for the lightly staining transformed cells. The indirect immunofluorescence method of analysis of the same populations was superior to the scattered light method of analysis of the GAG stained infected and transformed cells.

Animals↗

PubMed2006

Fine-needle aspiration of a Xp11.2 translocation/TFE3 fusion renal cell carcinoma metastatic to the lung: report of a case and review of the literature.

A 57-yr-old woman presented to the National Cancer Institute (NCI) with a history of nephrectomy for a clear cell renal cell carcinoma (RCC), Fuhrman grade 3 of 4 diagnosed 1 yr prior to admission to the NCI. A CT scan done upon admission revealed multiple bilateral lung masses. A CT-guided fine-needle aspiration (FNA) of one of the lung masses revealed a cellular specimen composed primarily of follicular structures surrounding dense hyalinized central cores. The cells in the follicular structures displayed bland nuclei and had granular to vacuolated cytoplasm. Papillary structures were also appreciated. Immunocytochemical studies showed tumor cells that were strongly vimentin and TFE3 positive. Focal staining for AE1/AE3 and CD10 was observed, as was negative staining for EMA. A surgical biopsy specimen reflected the FNA findings and demonstrated a similar immunoprofile. These findings correspond to the recently described Xp11.2 translocation/TFE3 fusion renal cell carcinoma. To our knowledge, this is the first report describing the cytologic features of an Xp11.2 translocation/TFE3 fusion RCC.

Basic Helix-Loop-Helix Leucine Zipper Transcriptio↗

PubMed1990

Improved detection of lymphocyte membrane proteins in purified form and as a crude mixture using native and denaturing polyacrylamide gel electrophoresis by optimisation of coomassie brilliant blue and silver staining.

Optimised silver staining protocols were devised for the detection of membrane proteins in purified form and as a crude mixture. These were adduced in both sodium dodecyl sulphate (SDS) and native polyacrylamide gel electrophoresis and consisted of ethanol-acetic acid-formaldehyde fixation, Coomassie Brilliant Blue prestaining, Rapidfix pretreatment, formaldehyde enhancement and finally ammoniacal silver staining. With these modifications, numerous staining problems of membrane proteins were overcome. These included reduction in background staining, enhanced detection sensitivity in native gels, elimination of negative staining and the avoidance of metallic silver deposition on the gel surface. In overcoming these problems, some factors determining the colour and stainability of membrane proteins in their native state were determined. Both the anionic Coomassie Brilliant Blue dye and SDS detergent improved the sensitivity of silver staining in native gels, and ammoniacal silver was more sensitive than neutral silver, suggesting silver staining to be a charge dependent process.

Ammonia↗

PubMed1989

Dynamic G- and R-banding of human chromosomes for electron microscopy.

Synchronized human lymphocytes were exposed to 5-bromo-2'-deoxyuridine (BrdUrd) for incorporation in either G- or R-bands. The substituted bands were revealed by monoclonal anti-BrdUrd antibodies disclosed with either gold-labeled antibodies or with the protein A-gold complex. Sharp G- or R-banding, specific for electron microscopy (EM), was obtained. These banding patterns, referred to as GB-AAu (G-bands by BrdUrd using Antibodies and gold [Au]) and RB-AAu (R-bands by BrdUrd using Antibodies and gold [Au]), resemble dynamic band patterns (GBG and RBG) much more than they do morphologic band patterns (GTG and RHG). The G- and R-band patterns allow accurate chromosome identification and karyotyping. An actual karyotype of human GB-AAu-banded chromosomes at the 750 band level, photographed in the EM, is presented. The method produces excellent band separation and band contrast. Variations in band staining intensities were noted and correlated with BrdUrd enrichment. The C-band regions were positively stained after GB-AAu banding while they were negatively stained after RB-AAu banding. Telomeres appeared heterogeneous after GB-AAu banding suggesting that part of the telomeric bands might be late replicating.

Antibodies, Monoclonal↗

PubMed1985

Glutaraldehyde-induced changes in the axially projected fine structure of collagen fibrils.

The fine structure of the collagen fibril, as seen in axial projection, is changed by treatment with glutaraldehyde. The changes are detectable in electron-optical staining patterns and in the intensities of the low-angle meridional X-ray diffraction maxima. Current knowledge of the amino acid sequence of collagen and of the axial arrangement of molecules in fibrils permits interpretation in terms of specific alterations to the axial distribution of electron density along the fibril. Analysis of fibril staining patterns from glutaraldehyde-treated calf skin collagen shows that uptake of staining ions in positive staining patterns is inhibited at residues known to interact with glutaraldehyde (lysyl, hydroxylysyl and probably histidyl side-chains) and on other charged residues in the immediate neighbourhood of the glutaraldehyde-reactive residues. This can be seen as a "stain-exclusion effect" due to the presence of bulky polymeric complexes of glutaraldehyde molecules at cross-linking sites. Such stain exclusion accounts for the drastic changes in the negative staining pattern following treatment with glutaraldehyde. The intensity changes observed in the low-angle meridional X-ray reflections from rat tail tendon, similarly treated, also can be explained by the presence of these bulky complexes. Existing data have been used to predict a model of the altered electron density profile indicating the axial distribution of glutaraldehyde along a D-period of moist tendon collagen.

Aldehydes↗

PubMed1999

Diagnostic criteria for sarcoidosis of the sinuses.

BACKGROUND: Although newer technologies facilitate its diagnosis and treatment, little is known about sarcoidosis of the paranasal sinuses. OBJECTIVES: We sought to better characterize sarcoidosis of the sinuses and establish criteria for diagnosis. METHODS: Case-finding criteria were established, and over 50 reports of sarcoidosis of the sinuses in the medical literature were reviewed. Nine case reports of patients fulfilling the case-finding criteria were identified, as were 6 additional patients from our clinics. RESULTS: Nasal obstruction and chronic sinusitis were the usual initial complaints from patients and were associated with mucosal crusting, studding, plaque-like changes, or polyps in the nose in 5 of 6 of our patients. The most consistent finding in nose and sinuses was an erythematous, edematous, friable, hypertrophied mucosa. Like 5 of 6 of our patients, most patients had extra-pulmonary sarcoidosis involving multiple organs, but some had isolated upper respiratory disease. Radiologic studies showed extensive and often complete opacification of the sinuses and nose similar to that seen in diffuse polyposis associated with chronic bacterial and fungal sinusitis. No specific histopathologic findings distinguished sinus disease from those reported with pulmonary involvement. Pharyngeal involvement was present in 2 case reports and caused the apparent asphyxiation of 1 of our patients. CONCLUSION: Sarcoidosis of the sinuses should be considered in the differential diagnosis of sinusitis, especially in association with nasal polyposis, even when the sarcoidosis has not been otherwise diagnosed. On the basis of this experience, we propose diagnostic criteria for sarcoidosis of the sinuses. These include (1) radiologic evidence of sinusitis, (2) histopathologic confirmation of noncaseating granuloma in the sinus tissue supported by negative stains for fungus and acid-fast bacilli, (3) negative serologic test results for syphilis and antineutrophil cytoplasmic antibodies, and (4) no clinical evidence of other disease processes associated with granulomatous nasal and sinus inflammation. These criteria will provide the basis for further studies to assess both the natural history and the effectiveness of treatment in sarcoidosis of the sinuses.

Adult↗

PubMed1973

Immunoferritin localization of intracellular antigens: the use of ultracryotomy to obtain ultrathin sections suitable for direct immunoferritin staining.

A general method for the ultrastructural localization of intracellular proteins and antigens by immunoferritin techniques has been developed. The method involves direct staining of ultrathin sections of mildly glutaraldehyde-fixed and frozen tissues cut by means of a cryo-ultramicrotome. Bovine pancreatic sections were cut, mounted on grids, and stained with ferritin-rabbit antibovine RNase conjugates. After negative staining with 0.2% phosphotungstic acid, electron micrographs revealed specific labeling of all of the zymogen granules and the cisternae of the rough endoplasmic reticulum. No significant labeling was seen in the nucleus, mitochondria, or cell sap regions. The observation that no significant labeling was found in any region of rat pancreatic sections was consistent with the fact that rat RNase is immunologically non-crossreactive with bovine RNase. In addition, the labeling seen in bovine pancreas was completely absent if the sections were first incubated with free antibody. The method used here avoids prolonged fixation, dehydration, and other harsh chemical or physical treatments, and should extend the usefulness of immunoferritin techniques to the intracellular localization of many protein antigens beyond previously available methods.

Animals↗

PubMed1984

Immunohistochemical staining for keratin and carcinoembryonic antigen in the diagnosis of malignant mesothelioma.

Using formalin-fixed, paraffin-embedded tissue and commercial antisera, we evaluated the usefulness of immunohistochemical staining for carcinoembryonic antigen (CEA) and keratin in the diagnosis of malignant mesothelioma. All 18 adenocarcinomas of lung examined stained for CEA, usually strongly, while only eight of 22 mesotheliomas stained for CEA and the staining was generally weak. Staining for keratin was observed in 10 of 22 mesotheliomas and 12 of 18 adenocarcinomas; there were no differences in intensity of staining between the groups. We conclude that strong diffuse staining for CEA favors a diagnosis of carcinoma, and negative staining for CEA is against a diagnosis of carcinoma, but these are relative and not absolute criteria. We find that staining for keratin is of no use in distinguishing these types of tumors.

Adenocarcinoma↗

PubMed2006

p16INK4A is a robust in vivo biomarker of cellular aging in human skin.

The cell-cycle regulating gene, p16INK4A, encoding an inhibitor of cyclin-dependent kinases 4 and 6, is considered to play an important role in cellular aging and in premature senescence. Although there is an age-dependent increase of p16INK4A expression in human fibroblast senescence in vitro, no data are available regarding the age dependency of p16INK4A in vivo. To determine whether p16INK4A expression in human skin correlates with donor age, p16INK4A expression was analyzed by immunohistochemistry as well as the expression of the p16INK4A repressor BMI1. Samples from the age groups 0-20, 21-70, and 71-95 years were selected from a bank of healthy human skin. We show that the number of p16INK4A positive cells is significantly higher in elderly individuals compared to the younger age groups. The number of p16INK4A positive cells was found to be increased in both epidermis and dermis, compartments with strictly different proliferative activities. BMI1 gene expression was significantly down-regulated with increasing donor age, whereas no striking age differences were observed for Ki67. In immunofluorescence co-expression studies, Ki67-positive cells were negative for p16INK4A and BMI1-expressing cells also stained negatively for Ki67. In conclusion, we provide for the first time evidence that p16INK4A expression directly correlates with chronological aging of human skin in vivo. p16INK4A therefore is a biomarker for human aging in vivo. The data reported here suggest a model for changes in regulatory gene expression that drive aging in human skin.

Age Distribution↗

PubMed1975

Examination of fimbriation of some gram-negative rods with and without twitching and gliding motility.

Negatively stained preparations of 30 different strains of gram-negative rods representing 20 different taxa were examined in the electron microscope. Thirteen of the strains studied exhibited twitching and six of the strains exhibited motility. Additionally, non-twitching substrains of two of the twitching strains and a non-gliding substrain of one of the gliding strains were examined. A variety of cultural media, preparations for negative straining and negative strains were used. It was found that all strains with twitching motility possessed fimbriae, the diameter of which was approximately 50 A in all but one strain; the fimbriae of this strain had a diameter of approximately 40 A. The fimbriae were judged to be of polar origin in all cases where the origin could be determined with certainty. On none of the strains without twitching motility could fimbriae be demonstrated. Only one of the six strains with gliding motility possessed fimbriae.

Alcaligenes↗

PubMed1992

Cell-, age- and stage-dependent distribution of connexin43 gap junctions in testes.

Immunocytochemical data demonstrate that the distribution of gap junction connexin43 (Cx43) in rodent testes is dependent on cell type, testis maturation, and stage of the mature seminiferous epithelium. Western blotting and indirect immunofluorescence microscopy using anti-peptide antisera to Cx43 revealed abundant Cx43 in rat and mouse testes and mouse TM3 and TM4 cells. Cx43 mRNA was detected in rat testes and mouse TM4 cells by Northern blot analysis. Cx43 was localized by immunogold electron microscopy to gap junctions on Sertoli cells and Leydig cells. A punctate distribution of Cx43 was observed on peritubular cell surfaces following indirect immunofluorescence of detergent-permeabilized tubule segments. In cryosections from testes of immature (to 30 days) rats, and mature rats and mice, Leydig cells showed a punctate surface distribution of Cx43 following indirect immunofluorescence. A diffuse cytoplasmic fluorescence was also seen in spermatocytes and spermatogonia. Cx43 staining associated with Sertoli cells was age- and stage-dependent. Over 90% of the tubules in immature tests (22-30 days) contained Cx43 in the region of Sertoli-Sertoli occluding junctions and in the adluminal compartment. In mature rat testes, however, Cx43 immunostaining was detected in only 60% of 1195 tubule sections where it was abundant proximal to the Sertoli cell occluding junctions. All strongly stained tubules were from stages I-VIII, while negatively stained tubules were at stages IX-XIV. Cx43 immunostaining in mature mouse testes was also stage-dependent with all positive tubules at stages VI-VIII. In contrast to Cx43, Cx26 and Cx32 were detected by immunofluorescence only in the apical regions of the seminiferous epithelia in 90% of tubules from mature rats. Consistent with the observed Cx43 immunostaining, octanol-sensitive in situ dye-coupling was observed between Leydig cells, between peritubular cells and between Sertoli cells, suggesting the occurrence of functional gap junctions in these cell types. These observations provide evidence for extensive gap junction-mediated communication between a variety of testis cell types important to the support of spermatogenesis.

Animals↗

PubMedDate not supplied

RT-PCR amplification of RNA extracted from formalin-fixed, paraffin-embedded oral cancer sections: analysis of p53 pathway.

We present a new approach towards the detection of the mRNAs in formalin-fixed, paraffin-embedded samples using a reverse transcriptase (RT)-polymerase chain reaction (PCR). The total RNAs were extracted from 10-micron-thick sections and were reverse-transcribed, then the RT-products were subjected to PCR amplification of GAPDH mRNA for screening the mRNA degradation. Next, nested PCR was performed for examining the expression of p53-related genes, p21WAF1, MDM2, p33ING1 and p14ARF. GAPDH mRNA expression was detectable in 12 out of 21 oral squamous cell carcinoma (SCC) samples. p21WAF1 mRNA expression was detectable in 5 out of 12 SCC samples, MDM2 mRNA expression was detectable in 5 our of 12 SCC samples and p33ING1 mRNA expression was detectable in 6 out of 12 SCC samples. However, the expression of p14ARF mRNA was not detectable in any of the samples. Seven out of 12 oral SCC samples showed abnormal nuclear accumulation of p53 protein by immunohistochemical staining, whereas 5 out of 12 oral SCCs showed negative staining for p53 protein. Of of p33ING1 mRNA. One of these was a verrucous carcinoma in which the p53 gene products might be inactivated by the oncoprotein E6 of human papilloma virus. Thus, the p53 tumor suppressor pathway was disrupted in most oral SCCs at the cellular levels, due to either an abnormality in p53 itself or loss of expression of p53 regulatory factors. This method would assist in making diagnosis, determining therapeutic strategy and predicting the prognosis of various cancers including oral SCCs.

Animals↗

PubMed2006

[Identification of human testicular spermatogenic cells at different stages].

OBJECTIVE: To establish the method to identify human testicular spermatogenic cells. METHODS: Cells were dispersed by mechanic disintegration on testicular biopsy samples of obstructive azoospermic patients. Diff-Quik staining was applied to mixed cell smears. Live cells were observed under inverted microscope equipped with Hoffman modulation contrast optics, and classified according to their morphology. Chromogenic in situ hybridization (CISH) using chromosome 17 centromere probe and anti-c-kit immunocytochemistry staining were applied to classified cell smears. RESULTS: Sertoli cell, primary pachytene spermatocyte (PPS), spermatogonia, round spermatid were the main four round cell groups that can be classified under Hoffman optics. In CISH, Sertoli cell, PPS and spermatogonia displayed 2 centromere signals, while round spermatid and elongated spermatid/sperm displayed 1 centromere signal. In immunocytochemistry staining, PPS and spermatogonia displayed positive staining, while Sertoli cell, round spermatid and sperm displayed negative staining. CONCLUSION: Dispersed human testicular cells displayed different characteristics in live/staining morphology, ploidy analysis, cell surface membrane antigen expression. All of these methods can be chosen to identify human testicular spermatogenic cells at different stages, while cell morphology classifying under Hoffman optics is a simple and effective method for live cell identification.

Adult↗

PubMed1988

B-cell lymphoma of probable thymic origin: case report.

A case of B-cell lymphoma of probable thymic origin is reported. A 34-year-old woman was found to have an anterior mediastinal tumor in November 1986. The surface lymph nodes were not palpable. A total resection of the tumor mass was performed. The tumor invaded the right pleura, the right lung and the pericardium. Histologically, normal thymus was found at the margin of the tumor tissue. The neoplasm was predominantly composed of large lymphoid cells, separated by rather thick fibrous bands of nodular fashion in some areas. Immunohistochemical staining demonstrated monoclonal cytoplasmic IgG and kappa chains in a small portion of the neoplastic cells in fixed tissue. The cells showed positive staining with cluster of differentiation (CD) 20 (B1) but negative staining with antibodies reactive with T-cells in unfixed tissue. "Malignant lymphoma, diffuse, large cell type (B)" was the diagnosis. The arrangements of immunoglobulin (Ig) and T cell receptor (TCR) beta genes were studied. Clonal rearrangement bands of IgH and Ig kappa genes were observed in the same sizes in both the tumor and the peripheral blood before chemotherapy. The patient received chemotherapy until September 1987, and is in complete remission at present (January 1988). The peripheral blood showed germ line patterns of IgH and Ig kappa genes in complete remission. No rearrangement bands of TCR beta genes were detected throughout. The B-cell lineage was proved both from gene arrangement analysis and with immunohistochemistry.

Adult↗

PubMedDate not supplied

Extended lymphadenectomy and the quality of life in rectal cancer patients.

For a better quality of life in rectal cancer patients, high dose radiotherapy following abdominoperineal resection of the rectum with a pelvic partition is another surgical option replacing extended abdominoperineal resection. In addition to pelvic partition with polyglycolic acid mesh, the tissue expander was inserted into the pelvic cavity to support the intestine upward and the bladder forward. The mean total radiation dosage was 5040 cGy. Between 1989 and 1991, 10 patients were treated according to this method. Out of 10 patients 9 were free of recurrence, and only one had hepatic metastasis. In addition, postoperatively, the average residual urine by this method was 39.1 ml and was statistically different compared to a figure of 200 ml in conventional abdominoperineal resections (p < 0.001, "t"-test). In order to individualize the operative procedures among a variety of surgical options, the molecular biological technique was utilized. In p53 stain analysis of 114 colorectal cancer patients, patients with p53 positive staining reached a higher stage than those with p53 negative staining (p < 0.05, chi 2 analysis). Therefore, we surmised that the positivity of the p53 stain could be one of the factors gauged as an indication of postoperative high-dose radiation. In conclusion, high-dose postoperative radiotherapy was thought to be one of the treatment modalities to improve the survival and quality of life of advanced rectal cancer in selected cases.

Adult↗

PubMed1994

Amyloid fibril formation in gelsolin-derived amyloidosis. Definition of the amyloidogenic region and evidence of accelerated amyloid formation of mutant Asn-187 and Tyr-187 gelsolin peptides.

BACKGROUND: We have recently shown that the actin-modulating cytoskeletal and plasma protein gelsolin is involved in the pathogenesis of familial amyloidosis of Finnish type. To define the amyloidogenic region(s) in gelsolin and clarify the mechanisms involved in amyloid formation, we tested the amyloidogenicity of synthetic gelsolin peptide analogues. EXPERIMENTAL DESIGN: The in vitro amyloid fibril formation was studied using 22 synthetic peptides 7 to 30 residues long having sequence homology with wild-type or mutant gelsolins. Amyloid formation was monitored by Congo-red staining and polarization microscopy of the peptide aggregates, by negative staining electron microscopy, and by quantitative fluorometry with thioflavine T. RESULTS: Ultrastructurally, amyloid-like fibrils were formed from the mutant Asn-187 and Tyr-187 gelsolin peptides corresponding to the naturally occurring missense mutations found in familial gelsolin amyloidosis syndromes, as well as from a gelsolin peptide having a Val-187 substitution. The shortest peptide tested that was capable of forming amyloid-like fibrils was 9-residue mutant Asn-187 peptide. The corresponding wild-type peptide did not form amyloid. Quantitative fluorometry at the emission maximum 482 nm revealed highly accelerated amyloid fibril formation of the mutant Asn-187, Tyr-187 and Val-187 peptides as compared with the corresponding wild-type peptides. CONCLUSIONS: We have defined the amyloidogenic region of gelsolin to a 9-residue sequence in the highly conserved repetitive motif B and showed that residue 187 represents a critical site where a substitution of an amino acid with a charged side chain (Asp) with an amino acid with an uncharged (Asn) or hydrophobic side chain (Tyr, Val) creates a conformation that is highly amyloidogenic thus providing an explanation for the amyloidogenicity of the Asn-187 and Tyr-187 gelsolin variants.

Amino Acid Sequence↗

PubMed1998

Selective expression of inducible nitric oxide synthase in human prostate carcinoma.

BACKGROUND: Nitric oxide (NO) is synthesized by inducible nitric oxide synthase (iNOS) and plays an important role in tumor growth and angiogenesis. NO generation by iNOS also influences the cytotoxicity of macrophages and tumor-induced immunosuppression. Before now, the expression of iNOS in prostate carcinoma tissue had not been determined. METHODS: In this study, tissue sections from 16 patients with prostate carcinoma were studied immunohistochemically and compared with tissue specimens from 10 patients with benign hyperplasia. RESULTS: Positive iNOS immunostaining was detected in all sections from patients with prostate carcinoma. The malignant epithelial cells were highly positive. The antibody against iNOS also marked round cells, which had the same cell shape as that observed for macrophages. These cells were located in stroma and epithelium adjacent to tumor islets. However, round cells in benign tissue stained negative for iNOS. None of the benign hyperplasia specimens stained positive for iNOS immunohistochemically. CONCLUSIONS: Prostate carcinoma tissue had a high iNOS content, whereas benign tissue did not. The authors suggest that epithelial iNOS expression can be used as a specific immunohistochemical marker for prostate carcinoma. NO generation by iNOS may play multiple roles in the development of this disease.

Adenocarcinoma↗