Combined application of RT-PCR and immunohistochemistry on paraffin embedded sentinel lymph nodes of prostate cancer patients

The detection of tumor cells in the sentinel lymph node (SLN) is of great importance for the prognosis of cancer patients. At present, immunohistochemistry and RT-PCR for tumor marker expression are the most sensitive techniques available for this analysis. However, so far, most RT-PCR-based analyses of SLNs have been performed on fresh material, excluding a direct comparison with the (immuno)histologic results. In our view, this does not entirely aid routine diagnosis. We established an efficient method for RNA extraction and RT-PCR from paraffin sections of SLNs from prostate cancer patients and compared the results with the (immuno)histologic data of adjacent sections. Amplifiable RNA was obtained from 133 SLNs of 68 prostate cancer patients. Correlation of PSA-specific RT-PCR with (immuno)histologic findings showed a positive and negative predictive value of 83% and 100%, respectively, for the prostate cancer patients investigated. Four of 12 patients with biochemical relapse, but without (immuno)histologically detectable tumor cells were RT-PCR-positive for PSA. We found that single sections of paraffin-embedded SLNs are suitable for routinely performed RT-PCR. Combined with (immuno)histology, PSA-specific RT-PCR is a revealing supplementary technique for the detection of tumor cells in SLNs of prostate cancer patients.     

Evidence of macrophage and lymphocyte, but not dendritic cell, infiltration in posterior uveal melanomas, whilst cultured uveal melanomas demonstrate pluripotency by expressing CD68 and CD163

Although variation in the level of macrophage infiltration has been reported in uveal melanoma, little is known about the expression of other leucocyte markers. An immuno- histochemistry study of the levels of expression of macrophage and other leucocyte markers, in a series of 10 primary choroidal melanoma biopsies, was undertaken. Biopsies were either fixed immediately in formalin and embedded in paraffin wax or established as short-term cultures. Using single- and double-labelling immunohistochemistry, cultured cells and paraffin sections were analysed for a range of melanoma (HMB45, Melan A, S100 and tyrosinase) and immune cell (CD68, CD163, CD45 and CD1a) markers. All samples expressed at least two known melanoma markers. Infiltrating macrophages were present in the majority of sections. When cultured specimens were studied by double-labelling immunofluorescence, uveal melanoma cells were seen to express macrophage markers or have cross-reactivity with related proteins. Expression of the leucocyte antigen CD45 was observed in three tumours but was not present in any cultured cells, whilst the expression of the dendritic cell marker CD1a was absent from all samples.     

Prognostic value of RT-PCR tyrosinase detection in peripheral blood of melanoma patients

Malignant melanoma (MM) prognosis has been related to tumour thickness and clinical stage and metastasis risk has been associated with presence of tumour cells in peripheral blood. The aim of this study was to determine the relationship between presence of tyrosinase in peripheral blood of MM patients and their clinical prognosis. Blood samples from 58 MM patients (stage I-IV) were analysed, using RT-PCR assay to detect tyrosinase mRNA. The results showed that positive RT-PCR assay for tyrosinase were significantly associated with clinical status and tumour thickness. After a median follow-up of 24 months, RT-PCR results were found to be significant correlated with recurrence (p<0.05) and clinical stage III (p<0.05). Separate analysis of stage III tumours to determine the prognostic value of tyrosinase presence in peripheral blood showed an overall 24-month survival rate of 70% in the RT-PCR negative group versus 10% in the positive group (p<0.02). These results suggest that detection of circulating melanoma cells may be especially relevant in stage III patients, in whom RT-PCR positivity defines a subpopulation at high risk of recurrence.     

Micromorphometric features of positive sentinel lymph nodes predict involvement of nonsentinel nodes in patients with melanoma

The aim of the present study was to determine whether micromorphometric features of positive sentinel lymph nodes (SLNs) from patients with melanoma are useful for predicting further nodal involvement in completion lymph node dissection (CLND) specimens. Of 986 patients with melanoma undergoing SLN biopsy between March 1992 and February 2001, 175 (17.7%) had at least 1 positive SLN and 140 had subsequent CLND specimens available for review. Further nodal involvement in CLND specimens was present in 24 (17.1%) of 140 patients. Of 8 micromorphometric features of the SLNs that were assessed, the presence of metastases in CLND specimens was correlated significantly with a tumor penetrative depth (maximum distance of melanoma cells from the inner margin of the SLN capsule) of more than 2 mm (P < .05), a deposit size of more than 10 mm2 (P < .01), the presence of melanoma cells in perinodal lymphatic vessels (P < .01), and the effacement of nodal architecture by metastatic melanoma cells (P < .05). Our results indicate that some morphologic features of melanoma metastases in SLNs predict the likelihood of further nodal involvement in CLND specimens.     

Nodal melanocytic nevi in sentinel lymph nodes. Correlation with melanoma-associated cutaneous nevi

Melanocytic nevi occurring in lymph nodes create diagnostic difficulty by mimicking metastases. Few studies describe nodal nevi in sentinel lymph nodes (SLNs) excised for melanoma. We evaluated 72 cases in which patients had undergone SLN biopsy for melanoma. Lymph nodes and cutaneous melanomas were evaluated according to a standard protocol. Nodal nevi were identified in 8 patients (11%). Of these, 6 (75%) had an associated cutaneous nevus (P = .006). Of 21 patients with an associated nevus, 4 (19%) with nodal nevi had a cutaneous nevus with congenital features (P = .01). The incidence of nodal nevus correlated with a Breslow thickness greater than 2.5 mm (P = .02). Nevi were not seen in non-SLNs. Nodal nevi appear more frequently in patients with melanoma-associated cutaneous nevi, particularly if congenital features are present. The increased frequency of nodal nevi in SLNs relative to non-SLNs suggests an etiology of mechanical transport of nevus cells.     

应用地高辛标记的cRNA探针原位杂交检测胰岛素mRNA

成年雄性Ｗｉｔａｒ大鼠，用地高辛标记的ｃＲＮＡ探针在胰腺切片上进行原位杂交，以检测胰岛Ｂ细胞胰岛素基因表达产物ｍＲＮＡ；同时并用免疫组织化学法显示相邻切片胰岛素免疫反应阳性细胞，进行比较，胰腺经４％多聚甲醋灌注固定，经Ｂｏｕｉｎ液再固定２０ｈ，常石蜡包埋和切片，经胰岛素ｃＲＮＡ探针杂交的胰腺切片中胰岛Ｂ细胞和核仁呈阳性反应，阳性信号为蓝色颗粒，胰岛其他细胞及胰腺泡细胞为阴性，方法照切片中均无阳性信     

Desmoplastic spitz nevus: report of a case and review of the literature

Desmoplastic spitz nevus (DSN) is an uncommon melanocytic lesion. The histologic features of this benign tumour may mimic those of certain benign (dermatofibroma and desmoplastic cellular blue nevus) or malignant (metastatic carcinoma and malignant melanoma) neoplasms. We report the case of a male with a DSN, and also review the clinical characteristics, histologic features and differential diagnosis of this extremely rare lesion. The lesion is identified as an asymptomatic reddish-brown dome-shaped papule on the forearm. Microscopic examination showed a symmetric lesion in the dermis. The neoplastic cells were large and epithelioid-shaped, and were either isolated as individual cells or arranged in small nests in a paucicellular hyalinized stroma. Nuclei were large and vesicular with small nucleoli and no mitoses. Diffuse expression of S100 and the absence of staining with antibodies to melan-A and HMB45 was observed. A diagnosis of DSN with free margins was made. The histologic presentation of this benign lesion mimics both benign and malignant neoplasms. The immunohistochemical profile (S100 positive, HMB-45 negative and cytokeratin negative) may be helpful to differentiate it from other lesions.