Laser-induced fluorescence and reflectance spectroscopy for the discrimination of basal cell carcinoma from the surrounding normal skin tissue.

The object of this study was to investigate whether laser-induced skin autofluorescence (LIF) and/or light reflectance spectra could provide a useful contrast between basal cell carcinoma (BCC) tissues and the surrounding healthy skin. Unstained human skin samples, excised from humans undergoing biopsy examination, were irradiated with a nitrogen laser (lambda = 337 nm) for excitation of autofluorescence and a tungsten halogen lamp for the reflectance measurements. The ex vivo spectroscopic results were correlated with the histopathology images to distinguish the areas of BCC from those of the surrounding health skin. A simple spectral analysis technique was also applied for better skin diagnosis. In conclusion, it seems that LIF and reflectance spectra could be used to differentiate neoplastic from normal skin tissue using an appropriate classification model analysis.

Angiogenesis in Hodgkin's lymphoma: a morphometric approach in 286 patients with prognostic implications.

The significance of angiogenesis in Hodgkin's lymphoma (HL) is not well defined. The aim of this study was to evaluate various morphometric characteristics of microvessels in lymph node sections of 286 patients with HL at diagnosis and investigate their relationship with clinicopathologic parameters and prognosis. Microvessel density (MVD), total vascular area (TVA) and several size- and shape-related microvascular parameters were quantitated--after anti-CD34 immunohistochemical staining--in the region of most intense vascularization, using image analysis. An increase in microvessel caliber parameters (area, perimeter, major and minor axis length) and a decrease in MVD were noted with increasing stage. An inverse relationship was recorded between MVD and the number of involved sites (NIS) and LDH. In univariate analysis, overall disease-specific survival was adversely affected by MVD and TVA, whereas inferior failure-free survival (FFS) was associated with the presence of more flattened vessel sections. Multivariate analysis disclosed that the extent of angiogenesis (MVD/TVA), age and the NIS independently affected overall survival. Accordingly, FFS was independently linked to the shape of microvessels and albumin levels or the NIS. In conclusion, our data support the view that angiogenesis in HL provides independent prognostic information, requiring the concomitant evaluation of quantitative and qualitative aspects of microvascular network.

A comparative assessment of proliferating cell nuclear antigen, c-myc p62, and nucleolar organizer region staining in non-Hodgkin's lymphomas: a histochemical and immunohistochemical study of 200 cases.

Proliferating cell nuclear antigen (PCNA) and c-myc p62 oncoprotein are two nuclear proteins expressed in proliferating and transformed cells. They can be recognized immunohistochemically in paraffin sections by the monoclonal antibodies PC-10 and c-myc 1-9E10, respectively. On the other hand, nucleolar organizer regions (NORs) are loops of DNA that carry the r-RNA genes and can be visualized in paraffin sections as black dots (AgNORs) using a silver impregnation method. It has been suggested that the mean number of AgNORs may reflect the cellular kinetics of a tumor. We independently examined 200 cases of non-Hodgkin's lymphomas using the monoclonal antibodies PC-10 and c-myc 1-9E10, as well as the AgNOR method. Our study shows a very significant correlation between PCNA, c-myc expression, and AgNOR count on the one hand and histologic grade on the other (P < .001), although a significant overlap among the three grades exists. PC-10, c-myc 1-9E10, and AgNOR scores are all shown to be linearly related, even though significant discrepancies were observed, and the correlation is stronger between PCNA and AgNORs (PCNA v c-myc p62, r = .551; PCNA v AgNORs, r = .746; c-myc p62 v AgNORs, r = .529; P < .001). A remarkable finding is that the intermediate group of lymphomas is heterogeneous as far as the proliferative rate is concerned: diffuse large cell cleaved/non-cleaved lymphomas (category G of the Working Formulation) are characterized by a significantly higher proliferative index, as evidenced by the elevated PCNA, c-myc p62, and AgNOR scores, in comparison with the other types of intermediate-grade lymphomas (P < .001). However, the proliferative rate is lower than that of the high-grade lymphomas (PCNA, P < .05; c-myc p62, P < .001; AgNORs, P < .005). No significant difference exists between B-cell and T-cell lymphomas except for the higher expression of c-myc p62 in intermediate-grade B-cell lymphomas, obviously due to the higher proliferative rate of diffuse large cell lymphomas. Based on our findings, it appears that the combination of PCNA, c-myc p62, and AgNORs provides an accurate estimate of the proliferative rate of non-Hodgkin's lymphomas in paraffin sections. Clinical studies may show whether this information has prognostic value independent of histologic classification. In addition, our results suggest that category G (diffuse large cell) lymphomas may belong to a malignancy grade higher than the intermediate grade, a suggestion consistent with their more aggressive biologic behavior.

Assessment of proliferating-cell nuclear antigen immunostaining in soft-tissue tumours: relationship to histological grade and mitotic activity.

The proliferative activity in 70 cases of soft-tissue tumours was estimated immunohistochemically using the monoclonal antibody PC-10, which recognizes proliferating-cell nuclear antigen (PCNA) in paraffin sections. The PCNA index (i.e. the percentage of positive neoplastic nuclei) and to a lesser degree the PCNA count (i.e. the number of positive neoplastic nuclei per ten high-power fields) positively correlated with the malignancy grade (PCNA index: P < 0.001; PCNA count: P < 0.01). However, the range of values of PCNA index and PCNA count was similar between benign and grade I tumours. A statistically significant positive correlation was also established between PCNA index and PCNA count on the one hand and mitotic count on the other, but the correlation coefficient was low (r = 0.351, P < 0.01, and r = 0.290, P < 0.05 respectively). These results indicate that PCNA immunostaining may successfully be used as an adjunct to the conventional histopathological parameters in assessing the malignancy grade in soft-tissue tumours although it is of limited value in distinguishing between benign and grade I tumours.

Expression of c-myc p62 Protein in Non-Hodgkin's Lymphomas.

The expression of c-myc oncogene in neoplastic and non-neoplastic lymphoid tissue was studied using the specific monoclonal antibody myc 1-9E10 and the 3 step streptavidin-biotin immunoperoxidase method in paraffin sections. Twelve samples of non-neoplastic reactive lymph nodes of various etiology, 181 of non-Hodgkin's lymphomas (NHL), and five of hairy cell leukemia (HCL) were studied. The staining pattern was mainly nuclear but cytoplasmic reactions were also occasionally observed. The percentage of positive cells per total, cell population was estimated and all cases were classified into four groups: 0.-15%, 16-.30%, 31-45% and more than 45% positive cells. In the reactive lymph nodes, mainly germinal center cells were stained, but the percentage in eleven of the 12 cases was less than 15%. All cases of HCL were negative. In NHL lymphoma, only malignant cells were evaluated. Fifty-six per cent of cases of NHL had more than 15% positive cells. The percentage of positive cells was greater in high grade than in intermediate and low grade NHL (80.6x, 47.6%, and 10.6 respectively had more than 30:< positive cells). This correlation proved to be statistically significant (p < 0.001). The above findings suggest a relation between the expression of the c-myc oncogene and lymphomagenesis and indicate that (evaluation of the expression of c-myc in NHL may be helpful in determining the grade of malignancy.

Expression of C-myc p62 oncoprotein in multiple myeloma: an immunohistochemical study of 180 cases.

The immunohistochemical expression of C-myc p62 oncoprotein was investigated using the immunoperoxidase method and the monoclonal antibody (Mab) MYC 1-9E10 in bone marrow paraffin sections. 180 cases of multiple myeloma (MM) were studied. C-myc expression was found to correlate well with the grade of malignancy, type of myeloma and tumor cell burden (p < 0.001). A relationship was also observed between C-myc p62 and IgA-secreting myelomas. It was found that intermediate grade myelomas are heterogeneous as far as C-myc expression is concerned. Our findings strongly suggest that C-myc might be involved in myelomutagenesis. The evaluation of its expression in MM may be helpful in determining the grade of malignancy and possibly the biological behaviour of this tumor.

Expression of proliferating cell nuclear antigen (PCNA) and nucleolar organizer regions (NORs) in multiple myeloma.

We have studied 72 cases of multiple myeloma (MM) bone marrow trephine biopsies using the monoclonal antibody PC-10 and the colloid silver nitrate method for identification of proliferating cell nuclear antigen (PCNA) and nucleolar organizer regions (NORs), respectively. PC-10 and AgNOR scores were statistically significantly different between low versus intermediate and low versus high grade of malignancy (P < 0.001), whereas their difference was not significant between intermediate and high grade. A strong correlation was identified between these two proliferation-associated indices in each histological grade. There is evidence that the proliferative state of myelomas belonging to the intermediate grade is approximately the same as that observed in high grade (blastic) ones.

Minichromosome maintenance proteins 2 and 5 in non-benign epithelial ovarian tumours: relationship with cell cycle regulators and prognostic implications.

Minichromosome maintenance proteins (MCM) have recently emerged as novel proliferation markers with prognostic implications in several tumour types. This is the first study investigating MCM-2 and MCM-5 immunohistochemical expression in a series of ovarian adenocarcinomas and low malignant potential (LMP) tumours aiming to determine possible associations with clinicopathological parameters, the conventional proliferation index Ki-67, cell cycle regulators (p53, p27(Kip1), p21(WAF1) and pRb) and patients' outcome. Immunohistochemistry was applied in a series of 43 cases of ovarian LMP tumours and 85 cases of adenocarcinomas. Survival analysis was restricted to adenocarcinomas. The median MCM-2 and MCM-5 labelling indices (LIs) were significantly higher in adenocarcinomas compared to LMP tumours (P<0.0001 for both associations). In adenocarcinomas, the levels of MCM-2 and MCM-5 increased significantly with advancing tumour stage (P=0.0052 and P=0.0180, respectively), whereas both MCM-2 and MCM-5 increased significantly with increasing tumour grade (P=0.0002 and P=0.0006, respectively) and the presence of bulky residual disease (P<0.0001 in both relationships). A strong positive correlation was established between MCM-2 or MCM-5 expression level and Ki-67 LI (P<0.0001) as well as p53 protein (P=0.0038 and P=0.0500, respectively). Moreover, MCM-2 LI was inversely correlated with p27(Kip-1) LI (P=0.0068). Finally, both MCM-2 and MCM-5 were associated significantly with adverse patients' outcome in both univariate (> or =20 vs >20%, P=0.0011 and > or =25 vs <25%, P=0.0100, respectively) and multivariate (P=0.0001 and 0.0090, respectively) analysis. An adequately powered independent group of 45 patients was used in order to validate our results in univariate survival analysis. In this group, MCM-2 and MCM-5 expression retained their prognostic significance (P<0.0001 in both relationships). In conclusion, MCM-2 and MCM-5 proteins appear to be promising as prognostic markers in patients with ovarian adenocarcinomas.

The expression of the B-cell marker mb-1 (CD79a) in Hodgkin's disease.

Recent evidence indicates that membrane-bound immunoglobulin on B lymphocytes is associated with a molecule which comprises the products of the mb-1 and B29 genes. This molecule is a highly specific marker for B-cells, presumably because of its central functional role in antigen triggering, and has recently been clustered as CD79a at the 5th Leucocyte Workshop. Recently there has been controversy surrounding reports of B-cell antigen expression by Reed-Sternberg and related cells, and we have therefore studied 108 cases of Hodgkin's disease immunohistochemically using a novel antibody which detects mb-1 protein in paraffin sections. The results were compared with those achieved using antibody L26 to detect CD20. The mb-1 protein was present in the neoplastic cells in all 14 cases of lymphocyte predominance Hodgkin's disease studied, and CD20 immunoreactivity was also found in seven of the eight cases of this subtype studied. Of the non-lymphocyte predominance cases, 20% (19/94) expressed mb-1 and 30% (20/67) CD20 in the Reed-Sternberg cells, but the cells positive for either of these two markers usually constituted only a very small proportion of the neoplastic population. However, in occasional cases (one of 94 for mb-1 and five of 67 for CD20), more than 50% of the neoplastic cells expressed one or both B-cell antigens. These results confirm the B-cell origin of the neoplastic cells in lymphocyte predominance Hodgkin's disease, but they also indicate that, contrary to our previous study, mb-1 expression may occasionally be found in what appears, on histological grounds, to be other types of Hodgkin's disease.

Prognostic relevance of apoptotic cell death in non-Hodgkin's lymphomas: a multivariate survival analysis including Ki67 and p53 oncoprotein expression.

AIMS: To evaluate the independent prognostic value of apoptotic versus proliferative fractions in a series of 92 patients with non-Hodgkin's lymphomas (NHL). METHODS AND RESULTS: Apoptotic fractions were quantified by use of the TdT (terminal deoxynucleotidyl-transferase)-mediated in-situ end-labelling technique (TUNEL), the percentage of positive cells constituting the apoptotic index (AI). Proliferative rate was expressed as percentage of Ki67 positive cells (Ki67 LI). Tissues were also stained for p53 protein with the DO-1 antibody. Patients were followed up until death (n = 33) or for an average of 63 months (n = 56). AI increased with malignancy grade and proliferative activity but was not related to location, cell of origin, clinical stage, bone marrow involvement and p53 expression. In multivariate analysis, overall survival was independently influenced by grade, stage, p53 LI and chemotherapy. The independent predictors of disease-free survival were Ki67 LI location and chemotherapy. AI turned out to be the only independent (negative) predictor of post-relapse survival. On the other hand, a low Ki67 LI increased the risk of relapse (logistic regression analysis) whereas a low p53 LI increased the probability of complete response. CONCLUSIONS: Our results suggest that the combined assessment of apoptotic fraction, proliferative rate and p53 expression may provide important prognostic information independent of other clinicopathological parameters in NHL.