A triple stain of reticulin, glypican-3, and glutamine synthetase: a useful aid in the diagnosis of liver lesions.

CONTEXT: The correct histologic diagnosis of mass lesions of the liver can be difficult, especially in biopsy samples. Reticulin, glypican-3, and glutamine synthetae are stains that can help distinguish hepatocellular carcinoma, hepatic adenoma, and focal nodular hyperplasia. OBJECTIVE: To evaluate the utility of a triple stain of reticulin, glypican-3, and glutamine synthetae in distinguishing hepatocellular carcinoma, hepatic adenoma, and focal nodular hyperplasia. DESIGN: Whole tissue sections and tissue microarrays were evaluated with a triple stain of reticulin, followed by glutamine synthetae (diaminobenzidine, brown chromogen) and glypican-3 (alkaline phosphatase, red chromogen). The 109 cases evaluated included whole tissue section hepatocellular carcinoma (n = 16), tissue microarray hepatocellular carcinoma (n = 19), whole tissue section hepatic adenoma (n = 15), tissue microarray hepatic adenoma (n = 13), whole tissue section focal nodular hyperplasia (n = 13; 12%), tissue microarray focal nodular hyperplasia (n = 13), as well as nonmalignant liver parenchyma adjacent to hepatocellular carcinoma (n = 20). All cases were scored for reticulin being intact or lost, positive or negative staining for glypican-3, and diffuse, maplike, perivenular, or negative staining for glutamine synthetae. RESULTS: The combination of intact reticulin with either glypican-3 negativity or negative glutamine synthetae was 92% sensitive and 95% specific in the distinction of tissue microarray hepatic adenoma from hepatocellular carcinoma. For the distinction of tissue microarray focal nodular hyperplasia and hepatic adenoma, maplike glutamine synthetae was most useful and was 85% sensitive and 100% specific. CONCLUSIONS: The triple stain of reticulin, glypican-3, and glutamine synthetae is useful in the differentiation of hepatocellular carcinoma, hepatic adenoma, and focal nodular hyperplasia on biopsy specimens. Furthermore, this triple stain is advantageous to single stains and can help when aberrant staining patterns are observed.

Oncocytic adrenocortical tumors: diagnostic algorithm and mitochondrial DNA profile in 27 cases.

The pathologic diagnosis of adrenocortical carcinoma (ACC) relies on microscopic features that are sometimes equivocal in special variants, including oncocytic adrenocortical tumors (OACTs). We report a series of 27 unpublished OACTs (15 pure and 12 mixed or focal) and assess for the first time in OACTs the diagnostic utility of an algorithm recently proposed by our group ("reticulin" algorithm) for conventional ACCs on the basis of a combination of reticulin staining and assessment of only 3 Weiss parameters. Overall, 12 cases were malignant according to the Lin-Weiss-Bisceglia (L-W-B) system for pure tumors and the original Weiss system for mixed or focal tumors; extensive or focal disruption of the reticulin network was found in 16 of 27 OACTs. This was associated with either a high mitotic index, presence of necrosis, and/or vascular invasion in 14 of these, which were thus considered malignant according to our algorithm. From a clinical standpoint, OACTs, at least in the pure form, are "low grade" lesions with a low mean Weiss score, mitotic and Ki-67 indices, and uncommon capsular or vascular invasion. They, including unequivocal morphologically malignant cases, generally pursue an indolent clinical course. In addition, the 4977 bp mitochondrial DNA "common deletion" was detected using real-time polymerase chain reaction in 54% of cases from this study and an additional validation series of 23 OACTs, with a heterogenous (heteroplasmic) intratissue and intracellular distribution (as detected by a modified FISH procedure) and a marked association with the presence of intact reticulin framework.

Apoptotic death induces Abeta production and fibril formation to a much larger extent than necrotic-like death in CGNs.

In this study we report that apoptotic death of primary cultures of cerebellar granule neurons is accompanied by release of thioflavin-binding proteins - indicative of the presence of beta-sheet structures - and fibril formation in the culture medium. When the same neurons are subjected to an excytotoxic death caused by 100 microM glutamate exposure, the amount of thioflavin binding is markedly reduced. Western blot analysis shows that fibrils contain monomers, dimers and trimers of amyloid-beta (Abeta) which, when observed at the electron microscope, have morphologies reminiscent of fibrils of senile plaques. These findings demonstrate that triggering an apoptotic pathway leads to beta-sheet transition and fibril formation of a protein primarily involved in Alzheimer's disease and may be of direct relevance to the possible link between apoptosis and this neuropathology.

Fine needle aspiration cytodiagnosis of liver tumors.

OBJECTIVE: To present our experience with liver fine needle aspiration (FNA) diagnosis and the adjunctive use of cell blocks with reticulin stain. STUDY DESIGN: The authors reviewed the results of cytopathologic diagnosis obtained by FNA biopsy over a 1-year period, from January 2000 to December 2000, in patients who presented primarily with ultrasonographically suspected liver nodules. FNA smears from 936 patients and cell blocks from 796 patients were reviewed. RESULTS: Among the 936 aspirates studied, the most common malignancy was hepatocellular carcinoma (HCC), which was diagnosed in 427 cases (45.6%), followed by metastatic adenocarcinoma, with 52 cases (5.6%). The concurrent cell block was available in 796 cases. Among them, 574 (72.1%) contained sufficient tissue for diagnosis. Combining analysis of cytologic and histologic specimens, the sensitivity of ultrasound-guided FNA for diagnosis of liver tumors was 85.1% and the specificity 98.7%. The results were better than isolated cytologic analysis, which gave a sensitivity of 78.4% and specificity of 97.4%. The lower diagnostic accuracy of cytology resulted mainly from its lower ability to distinguish well-differentiated HCC from benign lesions. In the cell block sections with reticulin stain, all HCCs showed a decreased or absent reticulin pattern, whereas all the benign hepatocellular lesions usually had a normal trabecular reticulin framework. CONCLUSION: FNA cytology assisted by cell block examination can be an accurate and minimally invasive method for the definitive pathologic diagnosis of primary benign and malignant liver masses and for confirmation of tumors metastatic to the liver. In addition, reticulin staining should be part of the routine assessment of cell blocks. It enhances diagnostic accuracy, particularly for well-differentiated HCC.

Persistent reelin-expressing Cajal-Retzius cells in polymicrogyria.

Cajal-Retzius (CR) cells are early-developing cells important in mammalian corticogenesis. Reelin, a protein secreted by CR cells, is essential for completion of neuronal migration and cortical lamination. Lack of reelin causes the 'reeler' phenotype in mice and autosomal recessive lissencephaly with cerebellar hypoplasia in man. Focal increases in reelin and CR cells are associated with thickening and local invaginations of the marginal zone and microgyria in animal studies. It has been suggested that abnormalities of reelin expression may be involved in human polymicrogyria. We have studied CR cells and reelin expression in pathological sections of human polymicrogyria to explore this possibility. Occurrence, distribution, morphology and reelin expression in CR cells were studied in 12 cases of human polymicrogyria, ranging from 21 gestational weeks to 10 years of age. Findings were compared with age-matched controls. Large, reelin-positive CR-like cells were more numerous in the majority of the polymicrogyria cases and persisted for longer than usual, up to 10 years of age. The CR-like cells tended to cluster and were most frequent in fused molecular layers in the polymicrogyria. Reelin-expressing CR-like cells were also found in bridges between the molecular layer and overlying leptomeningeal heterotopia and within the heterotopia itself. Clusters of CR-like cells were also found in adjacent non-polymicrogyric cortex. No clusters were seen in the control subjects. Increased numbers of CR-like cells were seen in both familial and acquired cases. In contrast to previous reports, the findings show that large CR-like cells persisted for longer than usual, up to 10 years of age, and that they may continue to express reelin. Their maximal aggregation in regions of polymicrogyria and overlying leptomeningeal heterotopia suggest an association between the presence of these cells and polymicrogyria, which we interpret in the light of recent findings concerning the roles of reelin and its downstream signalling pathway in neuronal and glial developmental dynamics and post-developmental function.

The str gene cluster for the biosynthesis of 5'-hydroxystreptomycin in Streptomyces glaucescens GLA.0 (ETH 22794): new operons and evidence for pathway-specific regulation by StrR.

Two divergently oriented operons, strXU and strVW, located within the gene cluster for 5'-hydroxystreptomycin (5'-OH-Sm) biosynthesis in Streptomyces glaucescens strain GAL.0 (ETH 22794), were analysed by DNA sequencing and transcription/regulation studies. Three genes, strU and strVW, are conserved in a similar arrangement but in a different location within the str/sts gene cluster of the Sm-producing strain S. griseus N2-3-11. The four putative products resemble NDP-4-ketohexose 3,5-epimerases (StrX, M(r) 20.2 kDa), NAD(P)-dependent oxidoreductases (StrU, 45.6 kDa), and ABC-transporters (StrV, 61.8 kDa; StrW, 63.4 kDa). These genes are apparently involved in the biosynthesis of 5'-OH-Sm because the promoters of both operons are activated in trans by the activator StrR of S. griseus N2-3-11, when cloned in S. lividans 66 TK23. A sequence motif resembling the consensus sequence GTTCGActG(N)11CagTcGAAc for binding of StrR was identified within the intergenic region of strX and strV. Specific binding of StrR to this site was demonstrated by gel retardation assays using purified His*Tag-StrR.

Enzymatic synthesis of aminocyclitol moieties of aminoglycoside antibiotics from inositol by Streptomyces spp.: detection of glutamine-aminocyclitol aminotransferase and diaminocyclitol aminotransferase activities in a spectinomycin producer.

Extracts of stationary-phase mycelia of the spectinomycin producer Streptomyces flavopersicus ATCC 19756 catalyzed inositol dehydrogenase, L-glutamine:inosose aminotransferase, 2-epi-streptamine:inosose aminotransferase, streptamine:inosose aminotransferase, N3-methyl-2-deoxystreptamine:inosose aminotransferase, and aminodeoxy-scyllo-inositol:inosose aminotransferase reactions, as detected with a new rapid assay procedure. These results suggest that one or both amino groups of the N1,N3-dimethyl-2-epi-streptamine moiety of spectinomycin are derived by transamination from the alpha-amino group of L-glutamine. An enzymatic procedure for distinguishing among N1- and N3-monomethyl diaminocyclitol derivatives and their diaminocyclitol biosynthetic precursors is described. A scheme showing key roles of glutamine-aminocyclitol aminotransferases in biosynthesis of major aminoglycoside antibiotics is presented.

Bone marrow changes in patients with hairy cell leukemia treated by recombinant alpha 2-interferon.

Bone marrow specimens from 21 patients with hairy cell leukemia (HCL) who were entered into a program to study the efficacy of treatment with recombinant alpha 2-interferon were evaluated. Patients were treated with the interferon, 2 X 10(6) U/m2 subcutaneously three times weekly, and were scheduled to undergo bone marrow aspiration and biopsy at study entry and after three (21 patients) and six (16 patients) months of treatment. Bone marrow samples after three months of treatment showed an overall decline in cellularity, from an average of 77 +/- 20 to 57 +/- 22 per cent, with a marked decrease in the percentage of neoplastic mass (from 87 +/- 9 to 59 +/- 24 per cent). The bone marrow changes were associated with significant improvement in hematologic values, including hemoglobin levels and granulocyte and platelet counts. The bone marrow changes and improved hematologic values remained stable with continuation of interferon therapy. However complete bone marrow remission did not occur in any of the patients after three or six months of interferon therapy. The HCL cell mass in more than 60 per cent of the patients remained at or above 50 per cent of the marrow cellularity and dropped to less than 25 per cent in 14 per cent of the patients. In all of the patients increased amounts of reticulin fibers were identified in the bone marrow prior to therapy, and 89 per cent of bone marrow aspirations failed (dry tap). The amounts of reticulin fibers remained increased in most of the patients (91 per cent), with a high incidence of dry taps (73 per cent), after therapy. Interferon therapy also changed the tartrate-resistant acid phosphatase(TRAP)-positive HCL cells to TRAP-negative, suggesting inhibition of activity and/or production of TRAP in HCL cells.

Hydroxystreptomycin production and resistance in Streptomyces glaucescens.

The wild-type strain of Streptomyces glaucescens produces hydroxystreptomycin and shows an inherent natural resistance to streptomycin group aminoglycoside antibiotics. Cell free extracts of the wild-type strain were able to inactivate streptomycin, hydroxystreptomycin and dihydrostreptomycin in the presence of ATP. The phosphotransferase did not inactivate other aminoglycoside antibiotics, including spectinomycin. Mutant strains were isolated, which were highly sensitive to streptomycin group aminoglycosides, had no measurable phosphotransferase activity and were unable to form detectable amounts of hydroxystreptomycin. This suggests either a correlation between phosphotransferase activity, streptomycin resistance and hydroxystreptomycin formation or defects in more than one gene in the strS mutant strains tested.

Spontaneous cardiac tumors in eight rats.

Spontaneous cardiac neoplasms were diagnosed in one Sprague-Dawley and seven Fischer 344 rats ranging in age from eight months to two years. All eight neoplasms were diagnosed as neurilemmomas. One neoplasm was composed of dense Antoni type A tissue with Verocay body formation, while seven others were composed principally of loose Antoni type B tissue. No specific site predilection was found.