Histopathological periodic acid-schiff stains of nail clippings as a second-line diagnostic tool in onychomycosis.

The diagnosis of onychomycosis, using direct microscopy and fungal cultures, is often negative despite the presence of disease. Periodic acid-Schiff (PAS) staining of nail clippings, using histopathological processing, may be positive in these cases. It is not always clear, however, whether the fungal elements detected by PAS staining are pathogenic fungi or some are saprophytes. We aimed to study the efficacy of histopathological PAS staining of nail clippings as a second-line diagnostic tool in onychomycosis. The study included 100 consecutive cases in which direct microscopy and fungal cultures from suspected onychomycosis were negative on one occasion or more. The obtained nail clippings were processed for routine histology, stained with hematoxylin and eosin and PAS, and examined microscopically. Of the 100 cases, 38 (38%) showed positive fungal elements. As a result, 9 patients had sought and received oral antifungal therapy and all achieved complete clinical cure. The histological examination also revealed parakeratosis and globules of plasma, which were statistically significantly more common in the fungal infected nail samples. This may indicate an ongoing inflammatory process associated with onychomycosis. Neutrophils and bacteria were not statistically and significantly more common in the fungal infected nails. We conclude that as a second-line diagnostic tool, PAS stain of nail clippings increases markedly the diagnostic yield of onychomycosis and, consequently, the outcome of therapy.

Acute phosphate nephropathy-an emerging threat.

BACKGROUND: Acute phosphate nephropathy (APN) is a clinicopathological entity causing renal failure, after ingestion of oral sodium phosphate solution (OSPS). Approximately 25 cases have been described, but OSPS is still widely used. This study reports a further 5 cases and discusses the ever-growing significance of APN. METHODS: Five cases of APN were included, 3 retrospectively whereas 2 were diagnosed prospectively. In all, use of OSPS was established, and other causes of nephrocalcinosis were excluded. RESULTS: Average age was 67.4 +/- 7.0 years, with a female preponderance (4:1). All patients had hypertension. Baseline serum creatinine: 0.7 to 1.2 mg/dL (creatinine clearance: 52 to 77 mL/min). Time from colonoscopy to presentation was 56 +/- 36 days. Serum creatinine levels at presentation: 1.4 to 3.6 mg/dL. Time from colonoscopy to renal biopsy was 123 +/- 88 days. Urinalysis showed minimal proteinuria, leucocyturia, and hematuria. One patient had renal glucosuria. All patients were anemic (hemoglobin 8.8-11.4 gr/dL). Serum calcium and phosphate were normal. One required hemodialysis. Mean follow-up was 36 +/- 17 months. Serum creatinine levels at end of follow-up were 1.3 to 3.1 mg/dL. Renal function did not recover completely in any patient. Four required long-term erythropoietin treatment. The prominent histopathological findings were calcium-phosphate tubular depositions (100%), interstitial fibrosis (80%), hypertensive changes (80%), and acute tubular degenerative and regenerative changes (60%). CONCLUSIONS: APN is a serious, irreversible renal complication of OSPS. It is probably under-recognized. Risk factors include female gender, older age, hypertension, and renal failure, although it may occur with preexisting normal renal function.

The applicability of the new WHO-EORTC classification of primary cutaneous lymphomas to a single referral center.

Recent years have witnessed differences between the World Health Organization (WHO) and the European Organization for Research and Treatment of Cancer (EORTC) classification systems of primary cutaneous lymphomas (PCLs). Recently, a joint WHO-EORTC classification system for PCLs has been reached. This study was performed to assess the applicability of this new classification to a single referral center. All new PCL cases, excluding mycosis fungoides and Sezary syndrome, who were referred from 1999 to 2005 were included. The histological, immunohistochemical stainings and molecular studies were reviewed, and additional stains were performed as needed. The cases were then reclassified according to the WHO-EORTC classifications. The clinical files were also studied, and the patients were followed up clinically. There were 43 new non-mycosis fungoides/Sezary syndrome PCLs, including 29 B-cell lymphomas of which 14 were follicle center lymphoma, 10 marginal zone lymphoma, 4 diffuse large-B-cell lymphoma, leg type, and 1 diffuse large-B-cell lymphoma, other. The 14 T-cell lymphomas included 5 cases of lymphomatoid papulosis, 2 CD30+ anaplastic large-cell lymphomas, 1 NK/T-cell lymphoma, and 6 peripheral T-cell lymphomas, unspecified. Of the 6 "unspecified" T-cell lymphomas, 3 were CD4+ small/medium-sized pleomorphic T-cell lymphoma, which is considered currently a provisional entity under the unspecified T-cell category. The remaining 3 cases could not be classified beyond the unspecified T-cell category, of which 2 cases had an aggressive course. The new WHO-EORTC classification is applicable to most non-mycosis fungoides/Sezary syndrome PCL cases, especially the B-cell lymphomas. However, there is still a substantial subset of T-cell PCLs which cannot be classified beyond the unspecified peripheral T-cell category, some of which may have an aggressive course.

Abnormal deposition of collagen around hepatocytes in Wilson's disease is associated with hepatocyte specific expression of lysyl oxidase and lysyl oxidase like protein-2.

BACKGROUND/AIMS: Lysyl-oxidases catalyze the oxidation of lysine residues in collagen and elastin thereby promoting their polymerization. We have studied here the expression of four lysyl-oxidases in normal and diseased human liver. METHODS: The expression of the different lysyl-oxidases in paraffin embedded liver sections was studied using in-situ hybridization and immunohistochemistry. The enzymatic activity of lysyl-oxidase like protein-2 (Loxl2 or LOR-1) using a previously described lysyl-oxidase assay. RESULTS: We have found that the four lysyl-oxidases which we examined are not significantly expressed in the normal liver. By contrast, Wilson's disease and primary biliary cirrhosis (PBC) patients express lysyl-oxidase (Lox) and lysyl-oxidase like protein-2 (Loxl2 or LOR-1) in hepatocytes, and the expression is accompanied by collagen deposition around the hepatocytes. Lysyl-oxidases are also expressed in additional fibrotic liver diseases such as hepatitis B and C but in these diseases the expression is confined to the fibrotic lesions and collagen does not accumulate around hepatocytes. We have found that Loxl2 is able to oxidize lysine residues of collagen, and behaves in that respect similarly to Lox. The copper chelator D-penicillamine inhibits Loxl2 induced oxidation of collagen but the Lox inhibitor beta-aminopropionitrile did not inhibit the oxidation using a BAPN concentration at which Lox activity was completely inhibited. Loxl2 also catalyzed the oxidation of cell surface proteins on HepG2 hepatoblastoma cells and inhibited their proliferation. CONCLUSIONS: Upregulation of Lox and Loxl2 in hepatocytes of Wilson's disease and PBC patients may contribute to liver damage by various mechanisms. The upregulation of Lox and Loxl2 in Wilson's disease could perhaps be utilized for diagnostic purposes since their expression is up-regulated in hepatocytes even before the onset of fibrosis.

Activated status of tumour-infiltrating lymphocytes and apoptosis in testicular seminoma.

Testicular seminoma is characterized by a prominent lymphoid infiltrate and an excellent prognosis. Cytotoxic T-lymphocytes (CTLs) infiltrating seminoma tumour nests constitute a major subset of the lymphoid infiltrate. The objective of this study was to determine whether CTLs express markers of cytotoxic potential and activity and whether the number of activated CTLs correlates with the extent of apoptosis in testicular seminomas, as opposed to non-seminomatous testicular germ cell tumours (NSTGCTs). Twenty cases of pure seminoma as well as 20 cases of NSTGCTs including 16 mixed germ cell tumours (MGCTs) were studied. Immunohistochemistry for the cytotoxic markers TIA-1 (cytotoxic potential) and granzyme B (cytotoxic activity) and the T-cell markers CD3 and CD8 was performed on formalin-fixed, paraffin-embedded sections. The apoptotic index (AI) was determined by the TUNEL method. The number of CD3(+), CD8(+), TIA-1(+), and granzyme B(+) cells in tumour cell nests was markedly increased in testicular seminomas, compared with NSTGCTs (p<0.01). Activated granzyme B(+) cells numbered 25.6+/-5.2 per high power field in seminomas and 8.9+/-3.2, 8.1+/-3.9, and 0.4+/-0.2 for embryonal carcinomas, yolk sac tumours, and immature teratomas, respectively. Double immunohistochemical staining for granzyme B and CD8 revealed that 82.6+/-8.5% of granzyme B-expressing cells were CD8(+). The tumour cell AI was significantly increased in embryonal carcinoma, compared with the seminoma, yolk sac tumour, and immature teratoma subgroups (6.7+/-1.3, 2.3+/-0.3, 3.0+/-1.1, and 2.3+/-1.1, respectively, p<0.001). TUNEL/CD3 double immunostaining revealed that a significant proportion of the apoptotic seminomatous tumour cells were in direct contact with one or more CD3(+) lymphocytes (47.2+/-6.2%). The number of activated granzyme B(+) CTLs showed a strong linear correlation with the AI in the seminoma group (r=0.71, p<0.0001) but not in other subgroups. TUNEL/granzyme B double immunolabelling revealed that a proportion of activated granzyme B(+) lymphocytes (20%) were often seen in close contact with apoptotic tumour cells. The presence of increased numbers of activated cytotoxic lymphocytes in testicular seminomas suggests that apoptotic tumour cell death in this neoplasm may be triggered by cytotoxic granule effectors. This phenomenon may be one of the key host immune mechanisms leading to the excellent prognosis in this tumour.