Cryptomphalus aspersa mollusc eggs extract promotes migration and prevents cutaneous ageing in keratinocytes and dermal fibroblasts in vitro.

BACKGROUND: The search of substances that minimize cutaneous ageing has increased in the last few years. Previous studies have described the regenerative properties of the secretion of the mollusc Cryptomphalus aspersa (C. aspersa) when applied topically. OBJECTIVE: We evaluate the in vitro effects of a new product derived from the eggs of C. aspersa, IFC-CAF, on cell proliferation, migration, distribution of cytoskeletal proteins, production of extracellular components as well as its ability to prevent cutaneous ageing because of intrinsic or extrinsic factors (exposure to UVB) by determination of ageing markers. METHODS: We have used the human keratinocyte cell line (HaCaT cells), primary dermal fibroblasts (HDF) and senescent dermal fibroblasts (SHDF). The effects of the compound on cell proliferation and on the cell cycle were determined by the MTT colorimetric assay, estimation of total protein and/or trypan blue test and by flow cytometry, respectively. We also studied cell migration using the wound-healing migration assay, whereas ELISA assays, Western Blot and immunofluorescence microscopy were carried out to test the expression of proteins related to cytoskeleton, extracellular matrix and with ageing. RESULTS: We have found that IFC-CAF does not promote proliferation but induces migration of HaCaT, HDF and SHDF in a time- and dose-dependent manner; a better organization of cytoskeletal proteins (F-actin and vimentin) and promotes the production of extracellular components (fibronectin, collagen 1 and MMPs) and the adhesion to cell-substrate vinculin protein. IFC-CAF also prevents cutaneous ageing. The treatment decreases the expression of the ageing-related markers b-Gal, p53 and p16INK4 in SDDF cells, and improves cell survival after UVB irradiation and nuclear repair in HaCaT cells. CONCLUSION: IFC-CAF has regenerative properties and protects against ageing factors being, therefore, a potential therapeutic agent for treating or preventing skin ageing.

Lack of correlation between DNA copy number and mRNA expression levels of c-myc in gamma-radiation-induced mouse thymic lymphomas by using quantitative real-time PCR.

BACKGROUND: It is well documented that over-expression of the c-myc proto-oncogene occurs in the vast majority of mouse thymic lymphomas induced by gamma-irradiation, evidencing the importance of this gene in T-cell lymphomagenesis. However, it remains unknown whether elevated levels of c-myc expression are driven by extra c-myc copy numbers. MATERIALS AND METHODS: Here we use a quantitative test on the basis of real-time PCR to determine the cellular copy number of c-myc in a set of 14 g-radiation- induced thymic lymphomas obtained from (C57BL/6J x BALB/cJ) F1 hybrid mice with increased mRNA c-myc expression. RESULTS: Since 5 out of 14 (35.7%) cases had no extra copy numbers of c-myc, gene amplification was obviously not the cause of c-myc over-expression in these tumours. In the remaining 9 tumours, c-myc over-expression was also accompanied with extra DNA copy numbers. Therefore, c-myc amplification might be a consequence of the genomic instability subsequent to the up-regulation of c-myc. However, linear regression analysis showed a lack of correlation between increasing DNA copy numbers and mRNA over expression of c-myc in these tumours (r = 0.029, p = 0.94). CONCLUSION: De-regulation of c-myc does not necessarily imply amplification of this gene in these tumours. This report is, to our knowledge, the first one comparing c-myc amplification with expression in lymphomas of the T-cell lineage.