Michela Corrias

Activated Notch1 expression is associated with angiogenesis in cutaneous melanoma.

An early event in melanocytic tumor growth is the upregulation of Notch signaling. When an active form of Notch1 is overexpressed in primary human melanocytes, it increases cell growth, survival and invasive properties, promoting melanoma progression. Recent evidence suggested that tumor initiation and growth are driven by a subset of tumor-initiating cells termed cancer stem cells. Notch1 plays a predominant role in the maintenance of melanoblasts, including melanocyte stem cells, by preventing initiation of apoptosis. Moreover, the importance of Notch1 in the regulation of tumor angiogenesis is supported by growing evidence in various cancers. Nestin has been widely used as a marker for melanocyte stem cells as well as an angiogenic marker to evaluate neovascularity of endothelial cells in tumors. To gain an insight into the impact of Notch1 activation on the maintenance of melanocyte stem cells and angiogenesis in melanoma, the expression levels of activated Notch1 and nestin were analyzed by immunohistochemistry in 114 primary cutaneous melanomas and 35 lymph node metastases. Activated Notch1 and nestin expression was also evaluated in four dysplastic melanocytic nevi. This study provides evidence that activated Notch1 is overexpressed in cutaneous melanoma, in tumor cells as well as in microvessel endothelium, and that it can promote tumor angiogenesis. Indeed, the overexpression of activated Notch1 in both tumor and vascular endothelial cells was significantly associated with microvascular density in melanoma samples. Thus, activated Notch1 inhibitors may provide a therapeutic strategy in the treatment of melanoma by blocking tumor-associated vascularization.

Nestin and vimentin colocalization affects the subcellular location of glucocorticoid receptor in cutaneous melanoma

Aims:  Nestin (a neuronal stem cell/progenitor cell marker of central nervous system development), vimentin (which is ubiquitously expressed in mesenchymal cells), and the glucocorticoid receptor (GR, which is involved in the immune response, cell proliferation, and apoptosis) have been shown to interact in embryonic and undifferentiated tissues in modulating cell proliferation. The aim of this study was to analyse nestin, vimentin and GR expression in tumour tissue (melanoma), and their association with clinicopathological variables, to evaluate any effect on tumour progression.

Association between the ACE insertion/deletion polymorphism and pterygium in Sardinian patients: a population based case–control study

Objective The purpose of the study was to examine whether the insertion (I) and/or deletion (D) polymorphism of ACE confers susceptibility to primary pterygium in Sardinian patients in a case–control study. Methods and results Polymorphism genotyping was performed by nested PCR using genomic DNA extracted from the whole peripheral blood of participants with (n=251) and without (n=260) pterygium. DD, ID and II genotype frequencies were: 48%, 39% and 13%, respectively, for patients with pterygium, and 15%, 40% and 44%, respectively, for the control group. A statistically significant difference was found between the pterygium and control groups for the ACE I/D polymorphism (p<0.001). Moreover, a statistically significant difference was found between the DD and II groups (p<0.01; OR=10.49; 95% CI 6.18 to 17.79), DD+ID versus II group (p<0.01; OR=5.23; 95% CI 3.37 to 8.13) and DD versus ID groups (p<0.01; OR=3.21; 95% CI 2.04 to 5.04). Conclusions Statistical analysis showed that the DD genotype is associated with an increased risk of developing pterygium, and with a good chance that the D allele may play an important role in the development of disease.

Angiotensin II: immunohistochemical study in Sardinian pterygium

The Angiotensin II (Ang II) is the principal effector peptide of the RAS system. It has a pleiotropic effect and, beside its physiological role, it has the property to stimulate angiogenesis and activate multiple signalling pathways related to cell proliferation. The purpose of the study was to determinate the Ang II expression and localization in Sardinian pterygium and normal conjunctiva by immunohistochemistry, and its possible involvement in the development and progression of the disease. Twenty-three pterygiums and eleven normal conjunctiva specimens obtained from Sardinian patients, were processed for paraffin embedding and assessed for the immunohistochemi-cal revelation of Ang II. Significant Ang II expression was identified in pterygium and conjunctiva. Particularly, thirteen pterygium specimens (n=13) displayed exclusively moderate to strong nuclear staining; some specimens (n=5) showed exclusively a moderate cytoplasmic immunoreactivity, and few specimens (n=2) displayed moderate to strong immunoreactivity in both cytoplasm and nucleus. Only 3 specimens were negative. Statistical significance difference in respect of nuclear and cytoplasmic localization was observed between normal conjunctiva and pterygium (P=0.020). The results showed a predominant intranuclear localization of Ang II in pterygium epithelial cells, in spite of conjunctiva that mainly showed cytoplasmic localization. These findings suggest a possible role for Ang II in the development and/or progression of pterygium mediated by the activation of local RAS system.

Association between the angiotensin-converting enzyme (ACE) insertion/deletion (I/D) polymorphism and pterygium in Sardinian patients

Pterygium is a common ocular surface disorder characterized by proliferation, inflammatory infiltrates, fibrosis, angiogenesis and extracellular matrix breakdown. Epidemiological studies indicate exposure chronic to UVB light as the most important risk factor for the development of pterygium. The Angiotensin Converting Enzyme (ACE) is the major component of the Renin-angiotensin system (RAS). It converts the inactive decapeptide Angiotensin I (Ang I) to the active octapeptide Angiotensin II (Ang II). Ang II is the most potent vasoconstrictor and stimulant of the aldosterone release. Recent discoveries have demonstrated that Ang II is also involved in cell proliferation, apoptosis, angiogenesis and tissue fibrosis. Moreover, it acts as growth factor and participates in inflammatory responses. The gene encoding ACE is mapped on chromosome 17q23; it contains 25 introns and 26 exons and shows a polymorphism characterized by the presence (insertion, I) or absence (deletion, D) of a 287-bp Alu sequence of DNA in intron 16. The presence or absence of Alu sequence in the ACE gene leads to the D/D, I/D and I/I genotypes. Novel studies have reported that the absence or presence of specific ACE I/D polymorphisms within ACE gene in several illnesses, such as cardiovascular diseases and breast cancer, can confer increased risk to develop the pathologies. Due to these evidences and the pterygium features, the aim of our study is to evaluate the ACE I/D gene polymorphism type in a group of Sardinia pterygium patients and to establish the possible correlation between ACE-type polymorphism and the development of pterygium in a case-control study.

Vitamin D and vitamin D receptor in patients with ophthalmic pterygium

Pterygium, an ultraviolet radiation (UV)-related disease, is a relatively benign process, but since it displays tumor-like features, it has been proposed to be a neoplastic- like growth disorder. Vitamin D performs a number of functions in addition to calcium homeostasis, as inhibition of cell proliferation, activation of apoptotic pathways, and inhibition of angiogenesis. Since the antitumor actions of vitamin D are mediated primarily through the nuclear vitamin D receptor (VDR), the aim of the present study was to investigate vitamin D status in patients with pterygium and in control subjects, and VDR immunohistochemical expression in samples of pterygium and normal conjunctiva in order to evaluate a possible role of vitamin D pathway in the pathogenesis of the disease. Serum vitamin D concentration was measured among 41 patients with pterygium and 47 volunteers by an automated chemiluminescence immunoassay. Moreover, 23 formalin- fixed and paraffin-embedded pterygium biopsy samples and 24 conjunctiva specimens were treated for the immunohistochemical demonstration of VDR using the streptavidin-biotin alkaline phosphatase method. No differences were observed about vitamin D level between patient with pterygium and control group, but significant differences between VDR immunolocalization in pterygium and normal conjunctiva were observed (P=0.00001). In conjunctiva, the immunoreactivity, localized mainly in cytoplasm of epithelial cells, may probably demonstrate VDR regulation of cell growth, differentiation, and apoptosis, while in pterygium VDR co-localization in the nucleus and cytoplasm of epithelial cells may indicate alternative nuclear pathways by which vitamin D might exert its antiinflammatory and anti-proliferative effects by the regulation of gene expression.

Immunohistochemical detection of Vitamin D receptor in pterygium

Pterygium is a chronic condition characterized by the encroachment of altered conjunctiva into the normal cornea. Several factors have been proposed as causative agents in the development of pterygium, however the exact mechanism of its pathogenesis is still unclear, even if several investigators consider pterygium an ultraviolet radiation (UV)-related disease. It is a relatively benign process, but since it displays tumor-like features, it has been proposed to be a neoplastic-like growth disorder (1). Vitamin D performs a number of functions outside of calcium homeostasis. Vitamin D status has been inversely associated with risk for various cancer, since Vitamin D inhibits cell proliferation, activates apoptotic pathways, inhibits angiogenesis, and exerts prodifferentiative effects in a wide variety of cancers (2). Since the antitumor actions of vitamin D are mediated primarily through Vitamin D receptor (VDR), and little is known about this subject in pterygium, the knowledge of VDR status may be important in understanding the pathogenesis of the disease. Keeping in mind all pathogenetic pterygium features, such as excessive cell proliferation, inflammation, fibrosis, extracellular matrix remodelling and intense neovascularization, we might suppose an alteration in the Vitamin D pathway. Therefore, the purpose of the study was to demonstrate, by immunohistochemistry on formalin-fixed and paraffin embedded sections of primary pterygium, the presence of VDR in pterygium, and to correlate it with the serum level of Vitamin D, in order to evaluate a possible role of Vitamin D pathway in the pathogenesis of the disease. The results will be discussed.

Immunohistochemical detection of progenitor cells in pterygium

Pterygium is a degenerative and hyperplastic ocular surface disorder characterized by excessive cell proliferation, inflammation, fibrosis, extracellular matrix remodelling and angiogenesis. Several factors have been thought to be involved in the development of pterygium, however the exact mechanism of its pathogenesis is still unclear. Nestin is almost an acronym for “neuroepithelial stem cell protein”. It is an intermediate filament (IF) protein expressed in proliferating cells during the developmental stages in a variety of embryonic and fetal tissues. It is also expressed in some adult stem/progenitor cell populations, such as newborn vascular endothelial cells and it is reactivated in response to injuries or other pathological conditions. CD34 is a hematopoietic progenitor antigen expressed on hematopoietic progenitors as well as on small vessels endothelial cells and embryonic fibroblasts. CD34 is thought to function as an adhesion molecule for early hematopoietic progenitors mediating the attachment of stem cells to extracellular matrix or stromal cells. Some authors have already demonstrated the presence of some stemness markers in pterygial tissues, suggesting the involvement of bone marrow originated stem cells in the pathogenesis of pterygium. The aim of the present study was to evaluate, by immunohistochemistry on formalin-fixed and paraffin embedded sections of primary pterygium, the presence of some more stem cells markers, such as nestin and CD34 in order to support the classification of pterygium as proliferative disorder. The results will be discussed.

Glucorticoid receptor in human cutaneous melanoma: immunohistochemical and immunofluorescence study

GR is a nuclear receptor which, when activated by its specific ligand, can act as a transcription factor that binds to glucocorticoid response elements (GRE) or negative GRE. It affects inflammatory responses, differentiation and cell proliferation. The ligand activated glucocorticoid receptor induces a G1 cell cycle arrest or apoptosis in immature thymocytes and impairs proliferation of fibroblasts of undifferentiated mammary epithelial cells. It impairs proliferation and differentiation of neural progenitor cells in vivo and in vitro. Glucocorticoids are widely used in cancer therapy and have cell type-specific pro- or antiapoptotic effects. In melanoma, however, the antitumor activity of glucocorticoids remains an open question. A recent report demonstrated that in mouse embryo tissue and in human undifferentiated cells, cytoplasmic accumulation of GR is determined by nestin in conjunction with vimentin, copolymerised into an intermediate filament system, and that this anchoring of GR to the nestin/vimentin etheromeric complex is related to the maintenance of a high proliferation rate. The aim of this study was to analyse the expression of subcellular GR in cutaneous melanoma by immunofluorescence, immunohistochemistry and laser scanning confocal microscopy and to evaluate any effect in melanoma progression. The results will be discussed.