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The recently described telomerase reverse transcriptase (TERT) promoter mutations are recurrent in cutaneous melanoma. Several authors have described an association between these molecular alterations, some histological parameters, and patient survival. BRAF mutations are very frequent in melanoma, but their actual role in the evolution of the disease is still unclear. Here, we investigated the relationship of TERT promoter mutations and BRAF mutations with the most relevant clinicopathological parameters, individually and coexisting, in order to evaluate their role as independent prognostic markers and to determine the effect of their coexistence. A TERT promoter alteration was found in 20 of 53 cases (38 %), significantly associated with histological type, increasing tumor thickness and mitotic rate, more advanced pathologic tumor (pT) stage, and absence of regression. A BRAF mutation was found in 21 of 53 cases (40 %), significantly associated with tumor thickness and presence of metastases in the sentinel lymph node. Coexistence of a TERT promoter and BRAF mutation was detected in 11 of 53 cases (21 %). This was associated with increasing thickness, high mitotic rate, lymph node metastasis, presence of ulceration, and absence of regression. Coexistence of a mutation in the TERT promoter and in the BRAF gene correlated with more prognostically relevant factors than either mutation alone. Our data lead us to hypothesize that TERT promoter and BRAF mutations cooperate in cutaneous melanoma. Further studies in larger cohorts of patients are needed to investigate how this synergistic effect is involved in the evolution of the disease.

Aims and Background Thickness and level of invasion are the main morphological elements for an approximate but not sufficiently sensitive prognostic evaluation of cutaneous melanomas. By using immunohistochemical methods it is possible to detect biological markers related to prognosis. We have studied p53, PCNA, Bcl-2 and P-gp expression in 49 primary cutaneous melanomas. Materials We used the immunophosphatase APAAP immunohistochemical method. The percentage of labeled cells (according to four classes of positivity: 50%) and the localization of immunoreactivity were expressed for each marker. Statistical analysis was performed to determine the correlations between markers and level or thickness of melanomas. Results We found a good correlation between p53 expression and melanoma thickness (P Conclusions Our data seem to indicate an unfavorable prognostic role of higher nuclear p53 expression. However, we believe that our results need to be integrated with patients’ clinical follow-up and with the study of the expression of these markers in benign melanocytic lesions to gain more accurate information about their prognostic significance.

Plants of Boea hygroscopica F. Muell were subjected to dehydration for 25 days until the relative water content was 23%. The rate of water loss was very slow during the first 12 days of dehydration while it dramatically increased during the last 3 days of the treatment. On day 12 of dehydration total glutathione content was reduced to 24% of the control level and was mainly present in the oxidized form. At that time an oxidation of the sulfhydryl groups of soluble proteins was observed. A protection of glutathione against oxidation of the sulfhydryl groups of soluble proteins was established only after 22 days of desiccation, at the same time as glutathione began to accumulate. During the whole desiccation period enzyme activities related to glutathione utilization and regeneration and the activity of NADP + -dependent glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.12), which contains essential sulfhydryl groups, were maintained. The results of the present experiment suggest that during dehydration of Boea hygroscopica glutathione has, above all, a primary role in the protection of the sulfhydryl groups of the thylakoid proteins, which were maintained in the reduced form during the whole dehvdration period.

In leaves of Boea hygroscopica subjected to either rapid or slow dehydration and rehydration, the response to H 2 O 2 production was studied by monitoring the changes in the amounts of ascorbic and dehydroascorbic acids as well as the amounts of reduced and oxidized glutathione and related enzyme activities. Boea hygroscopica is a resurrection plant in which drought tolerance depends on the rate of desiccation. For this reason, excised leaves of B. hygroscopica were subjected either to rapid or slow dehydration, by a 144-hr exposure to 0% or 80% relative humidity, respectively. Slowly dried leaves, rehydrated for 36 hr, were able to resume their activities completely, while rapidly dried leaves were not able to revive. After dehydration, H 2 O 2 production decreased both in slowly and in rapidly dried leaves, whereas the levels of total ascorbate and glutathione became two and 50 times as high, respectively, as in the control. During slow drying, reduced glutathione was oxidized, since its content in slowly dried leaves was 50% lower than in rapidly dried leaves. The levels of soluble proteins were found to be, respectively, five and six times as high in rapidly and slowly dried leaves as in the control and rehydrated leaves. The electrophoretic patterns of soluble proteins were similar in both dehydrated leaves, but different from control and rehydrated leaves, which also presented similar patterns. The specific activities of dehydroascorbate reductase (EC 1.8.5.1), glutathione reductase (EC 1.6.4.2) and ascorbate peroxidase (EC 1.11.1.11) also had the same values in both slowly and rapidly dried leaves. Following rehydration, oxidative processes were intensified: H 2 O 2 levels and the oxidation of ascorbate and glutathione mostly increased during this period in which the functioning of the ascorbate/glutathione cycle was induced.

Adenocarcinoma is becoming the most common histologic type of lung cancer in both sex. Although most cases are seen in smokers, it develops more frequently than other histologic types in individuals who have never smoked. This evidence suggests that other putative etiologic factors, such as sex hormones, need to be investigated. Several subtypes of lung adenocarcinoma have been recently described with distinct clinicopathologic features and prognostic implications. The purpose of this study is to investigate the role of estrogen receptor beta in lung adenocarcinoma, with particular attention paid to its different histologic subtypes. Nuclear estrogen receptor beta expression was evaluated by immunohistochemistry in 112 lung adenocarcinomas, including both "single subtype" and "mixed subtype" samples. Using a 2-level (high/low) score system, estrogen receptor beta expression was high in most (75%) adenocarcinomas and turned out to be strongly related to the histologic subtypes. In fact, estrogen receptor beta expression was low or negative in 68.2% of solid subtypes, whereas it was high in 76.5% of nonmucinous bronchioloalveolar, in 69.4% of acinar, and in 61.2% of papillary patterns (P = .00004). Furthermore, a strong association between estrogen receptor beta expression and tumor histologic grade was observed: estrogen receptor beta was highly expressed predominantly in well- and moderately differentiated tumors (P = .0014). In conclusion, estrogen receptor beta expression has distinct patterns in lung adenocarcinoma, suggesting a specific role for estrogen receptor beta in the pathogenesis of different histologic subtypes of this type of cancer. Moreover, loss of estrogen receptor beta expression in poorly differentiated (G3) tumors could represent a crucial step in the dedifferentiation process of lung adenocarcinoma.

Summary Angiogenesis is a central process in the growth of solid tumors. The purpose of our study was to analyze the angiogenic pattern in squamous and basal cell carcinomas and to point out differences in microvessel density that could explain their different biological behaviour. Thirty-nine skin tumors (26 basal and 13 squamous cell carcinomas) were analyzed. In all samples, the microvessels density (MVD) and the levels of vascular endothelial growth factor mRNA (VEGFmRNA) were analyzed, together with the inter-relationship between these two variables. Using the median value of the entire series (33 vessels per 2.22 mm2), tumors with low and high MVD were identified. The majority of cancers with high vascularization belonged to the squamous histotype (12 of 39), while 19 of the 26 basal cell carcinomas showed a lower number of microvessels than the median value (p = 0.0001). The median value of VEGFcDNA quantitation allowed us to distinguish tumors with high VEGF expression (>470 molecules cDNA) from those with low (≦470 molecules) VEGF expression: 20 of the 26 basal cell carcinomas showed low VEGF expression, while 11 of the 13 squamous cell carcinomas showed high VEGFcDNA levels (p = 0.0003). Moreover, a significant association between a high microvessel density and high VEGFmRNA levels (p = 0.006) was found. Furthermore, when studying VEGF expression by immunohistochemistry, we obtained similar results and noted a correlation with VEGFmRNA expression (p The association between high vascularization, high VEGF levels, and squamous cell histotype suggests the possible role of neoangiogenesis in determining the more aggressive biological behaviour of this type ofcancer.

Changes in the amounts of ascorbic acid, dehydroascorbic acid and glutathione and in the levels of related enzyme activities were studied in Sporobolus stapfianus leaves, a desiccation-tolerant or resurrection plant, in response to desiccation and rehydration. Detached leaves of S. stapfianus were rapidly dried for 120 hr at 0% rel. hum. A 24 hr desiccation period was sufficient to achieve extremely low water contents (0.2 %). Dried leaves were then rehydrated for 24 hr. The soluble-protein content decreased after drying but did not increase after rehydration without death ensuing. The enzymes related to the glutathione-ascorbate cycle reacted differently to the rapid water loss: dehydroascorbate reductase (EC 1.8.5.1) and glutathione reductase (EC 1.6.4.2) activities increased while ascorbate peroxidase (EC 1.11.1.11) activity decreased. All these enzymes resumed their activities after rehydration. Oxidative processes were intensified following desiccation and even more after rehydration: i.e. the H 2 O 2 level and the dehydroascorbate/ ascorbate molar ratio increased in desiccated leaves and even more in rehydrated leaves, while the oxidized glutathione/reduced glutathione molar ratio, which decreased in desiccated leaves, reached the control value with rehydration.

Telomerase, a ribonucleoprotein, is capable of adding telomeric sequences (TTAGGG hexameric repeats) to the ends of chromosomes and, thereby, halting the erosion of chromosome at each cell division. Whereas most normal somatic cells contain minimal or no detectable telomerase activity, most immortal and tumour cells exhibit significant levels of telomerase activity and show no net loss of telomere length during proliferation. The evaluation of telomerase has been proposed for diagnostic and therapeutic purposes in human cancer. Skin cancer is the most common cancer in humans; the precise molecular events in skin carcinogenesis are numerous and complicated and not yet completely clarified. In this study, we evaluated telomerase in 35 basal cell carcinomas and in 14 squamous cell carcinomas in order to determine if activation of the telomerase enzyme was a pivotal step in the development of skin cancer and whether telomerase activity levels were different between the two histotypes. A higher enzymatic level was shown to be associated with squamous cell carcinomas, while low levels were mainly detected in the basal cell histotype (chi2 test; p=0.02). Telomerase complex activity is dependent on its catalytic subunit, telomerase reverse transcriptase hTERT. By reverse transcription-PCR, using primers within the reverse transcriptase domain of hTERT, we observed a significant correlation between hTERT expression and telomerase activity in our skin tumour samples (p=0.0003). We detected the presence of multiple, alternately spliced transcripts, corresponding to full-length messages as well as spliced messages with critical reverse transcriptase motifs deleted. A higher telomerase messenger level was shown to be associated with squamous cell carcinomas (chi2 test; p