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Notes: Summary Background Recent evidence assigns the vitronectin receptors (VnRs) an important role in regulating tumour cell invasion and dissemination. In vivo and in vitro studies document that all trans-retinoid acids (ATRAs) inhibit growth-inducing apoptosis in melanomas. Objectives We have analysed the effects of ATRA treatment on melanoma cell adhesion and motility. Methods Human M14 melanoma cells were treated with 10 µmol L−1 ATRA for different times and stained with rhodamine–phalloidin to analyse the effect of treatment on cytoskeleton organization. Cell adhesion and cell migration assays were performed to analyse the role of VnRs in the ATRA-induced early stages of apoptosis. VnR expression was evaluated by Western blot, immunoprecipitation and immunocytochemistry assays. Results First, using an annexin V assay, we found that apoptosis was triggered by 48 h with 10 µmol L−1 ATRA exposure. At this time point, decrease in the F-actin polymerization as well as inhibition of cell adhesive ability to vitronectin (Vn) was exerted by ATRA treatment. In the presence of serum, exposure to 10 µmol L−1 ATRA for 48 h produced a dramatic inhibition of the cell adhesion ability that was comparable with that exerted by untreated cells preincubated with anti-αvβ3 or anti-αvβ5 VnR monoclonal antibodies. Functionally, the treatment of melanoma cells with 10 µmol L−1 ATRA for 48 h causes an inhibition of directional cell migration towards Vn-coated filters. Therefore, we analysed the effect of ATRA on the VnR expression. Both αvβ3 and αvβ5 VnR levels were reduced upon exposure to 10 µmol L−1 ATRA for 48 h as shown by Western blot, immunoprecipitation and immunocytochemistry assays. Conclusions Altogether, our data indicate that treatment of M14 melanoma cells with ATRA downregulates VnR expression and that this reduction is closely correlated with the ATRA-dependent inhibition of actin-fibre organization, cell adhesion and migration. Although the mechanism by which ATRA regulates the expression of VnR in M14 melanoma cells needs further elucidation, this system may represent a model for understanding the molecular basis of ATRA therapy in melanoma.

Notes: Background  Captopril is an angiotensin-converting enzyme inhibitor with sulphydryl groups in its chemical structure. It is commonly used as an antihypertensive drug. The occurrence of pemphigus vulgaris has repeatedly been reported in patients receiving captopril. The capacity of captopril and pemphigus serum to induce acantholysis, in vivo or in vitro, has been demonstrated experimentally.Objectives  To show that captopril and pemphigus serum, acting by a biochemical and immunological mechanism, respectively, trigger apoptosis.Methods  Human keratinocyte cells were treated with 15 mmol L−1 captopril or with pemphigus serum. DNA was extracted and the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labelling method was used to detect apoptosis.Results  DNA fragmentation occurred after 72 h of treatment. Increased expression of p53, c-myc and inducible nitric oxide (NO) synthase (iNOS) mRNA were observed by polymerase chain reaction (PCR) in the treated cells compared with the untreated ones. The increase in iNOS gene expression was associated with overproduction of NO. Moreover, the addition of 1 mmol L−1N-monomethyl-L-arginine, a structural analogue of arginine, reduced nitrite levels by about 70% in cells treated with captopril or pemphigus serum. Western blot analysis revealed an overexpression of heat shock protein 70 (hsp70) in cells treated with captopril or pemphigus serum. Finally, total inhibition of the keratinocyte transglutaminase gene was shown by PCR analysis in the same samples, compared with control cells.Conclusions  These data demonstrate the involvement of apoptosis in keratinocytes treated with captopril or pemphigus serum, with induction of the iNOS gene and hsp70 in the cascade of events leading to programmed cell death.

Notes: A case of granuloma annulare that developed in a site of healed herpes zoster is reported. Polymerase chain reaction failed to detect VZV DNA in a biopsy specimen. The occurrence of different types of cutaneous reactions in a body area previously affected by herpes virus infection is known as Wolf's isotopic response. Pathogenesis may be due to a local neuroimmune dysregulation set off by herpesvirus-induced lesions of dermal sensory nerve fibres.

Notes: Summary Background Peptide T (PT) is an octapeptide shown to resolve psoriatic lesions. PT is from the V2 region of HIV-1 gp120, an exterior envelope glycoprotein that is a target for host immune responses. The anti-inflammatory mechanisms of PT are not well understood. Objectives We studied the immunomodulatory effects of PT on the human keratinocyte cells. Methods Cultured human keratinocytes were treated with PT, proteins extracted and analysed by Western blotting and reverse transcriptase polymerase chain reaction. Results Our findings show reduced expression of intercellular adhesion molecule 1 and an increase in transforming growth factor (TGF)-β and heat shock protein (HSP)-70 in human keratinocyte cells treated with PT. The HSP-70 increase is modulated by TGF-β. In fact, we demonstrated that anti-TGF-β antibodies reduce HSP-70 overexpression. In addition, we show a modulation of αv integrins after 4 hours of treatment with PT. These receptors favour keratinocyte migration and epidermal regeneration. It has been reported that overexpression of HSP results in dramatic changes to intermediate filaments. These proteins act on keratin intermediate filaments and determine their retraction. The consequence is cell–cell contact detachment and inhibition of cellular hyperproliferation. Conclusions Our results support the beneficial effect of PT found in vivo, suggesting, moreover, the primary role of keratinocytes upon which PT acts directly by stimulating the anti-inflammatory function and favouring the regeneration of tissue.

Notes: Pemphigus is an autoimmune disease where both endogenous (genetic) and exogenous (environmental) factors play a part. Viral infections, in particular herpesvirus infections, have been identified as a possible triggering factor for pemphigus. In this study, using the polymerase chain reaction, we studied peripheral blood mononuclear cells (PBMC) and skin biopsies from patients with pemphigus, and in some of these were able to demonstrate the presence of DNA sequences of herpes simplex virus 1/2 (50% in PBMC and 71% in skin biopsies), Epstein–Barr virus (15% in PBMC and 5% in skin biopsies) and human herpesvirus 6 (20% in PBMC only). However, the inability to detect herpesvirus DNA consistently in these cases suggests that viral infection may only be an occasional factor triggering the outbreak or exacerbation of the disease. The possible role of interferons and interleukins in the pathogenesis of virus-induced pemphigus is discussed.

Notes: Abstract:  Peptide T (PT) is an octapeptide shown to resolve psoriatic lesions. Our previous investigations suggest that keratinocytes play an important role in conditioning the therapeutic effects of the PT in psoriasis. However, peptides are not good therapeutic agents, because they exhibit poor absorption, are easily metabolized and are immunogenic. Using computational methods, the natural product amygdalin was identified as peptidomimetic of PT. However, amygdalin exhibits a toxic profile due to its cyanide group. To overcome this deleterious effect, we synthesized analogues lacking the cyanide group. Human keratinocytes were treated with PT or with three different peptidomimetics of PT. To study its effects on the expression of HSP-70, TGF-β, α-v integrin, ICAM-1 and cytokines, we analysed the protein levels by Western blot and ELISA. Our results show that the different peptidomimetics of PT tested exhibit a similar biological behaviour in regard to the overexpression of HSP-70, TGF-β and α-v integrin than the native peptide. TNF-α is overexpressed by PT and SVT-03018; between the other two analogs, SVT-03016 do not produce any significant change in regard to the control, while SVT-03017 shows only a moderate increase in regard to control. SVT-03018 provokes a remarkable upregulation of IL-10, stronger than SVT-03016, SVT-03017 and PT. All the other three analogues reduce comparably to the PT, the expression of ICAM-1 and do not increase the release of proinflammatory cytokines. The results highlighted that the three analogues of amygdalin with the cyanide group removed exhibit the same biological effects of PT. Therefore, they can be considered peptidomimetics, suggesting their possible use in the treatment of psoriasis.

Notes: Abstract A total of 41 faeces and 5 vomits of the Weddell seal Leptonychotes weddelli was collected at Harmony Point, Nelson Island, South Shetland Islands, from 14 January to 1 February 1996. The diet indicated by the remains in the samples was diverse and comprised both pelagic and benthic-demersal species. Fish were the most frequent (95.7%) and numerous prey (46.2%), but molluscs were the most important by mass (65.8%). Octopods, mainly Pareledone charcoti, constituted the bulk of the diet (63.1% by mass), but the importance of the remaining molluscs was negligible. Otoliths represented 510 fish of which 491 were identified as belonging to 5 species: Gymnoscopelus nicholsi, Electrona antarctica, Lepidonotothen nudifrons, Gobionotothen gibberifrons and Nototheniops nybelini. The myctophid Gymnoscopelus nicholsi was the most important fish prey, and the contribution of benthic-demersal species was low. However, the importance of that fish was over-estimated since 96% of the specimens were obtained from the five vomits analysed. The biases associated with the faecal analysis technique are discussed.