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Biomedical subjects

A Baroni

Publications and source records attributed to A Baroni.

At least 19 recordsLinked to original sources
PubMed2005

Immunomodulatory effects of a set of amygdalin analogues on human keratinocyte cells.

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-beta, alpha-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-beta and alpha-v integrin than the native peptide. TNF-alpha 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.

Amygdalin↗
PubMed2004

Pemphigus serum and captopril induce heat shock protein 70 and inducible nitric oxide synthase overexpression, triggering apoptosis in human keratinocytes.

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.

Angiotensin-Converting Enzyme Inhibitors↗
PubMed2004

3-O-methylfunicone, a secondary metabolite produced by Penicillium pinophilum, induces growth arrest and apoptosis in HeLa cells.

3-O-Methylfunicone (OMF) is a secondary metabolite produced by the soil fungus Penicillium pinophilum which has cytostatic properties. The aim of this study was to investigate the mechanisms by which such properties are exerted, with special reference to any anti-proliferative and apoptotic potential, on HeLa cells. OMF treatment caused about 44% inhibition of cell growth after 24 h, and modifications in the tubulin fibre organization. In addition, a significant increase in p21 mRNA expression and a decrease in cyclin D1 and Cdk4 mRNA expression resulted at the same time. Apoptosis induction was demonstrated by the annexin V assay, cytofluorimetric analysis of the DNA content of the sub-G1 fraction and DNA laddering. Taken together, our data showed that the compound inhibits proliferation of HeLa cells by several mechanisms, such as disruption of tubulin fibres, cell cycle arrest and apoptosis induction. The capacity of the compound to affect the cell cycle and to modulate apoptosis is indicative of a potential for the development of a new agent for cancer chemotherapy.

Annexin A5↗
PubMed2003

Early vitronectin receptor downregulation in a melanoma cell line during all-trans retinoic acid-induced apoptosis.

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 micromol 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 micromol 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 micromol 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-alpha(v)beta(3) or anti-alpha(v)beta(5) VnR monoclonal antibodies. Functionally, the treatment of melanoma cells with 10 micromol 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 alpha(v)beta(3) and alpha(v)beta(5) VnR levels were reduced upon exposure to 10 micromol 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.

Antineoplastic Agents↗
PubMed2003

Granuloma annulare in a site of healed herpes zoster: Wolf's isotopic response.

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.

Antiviral Agents↗
PubMed2002

Immunomodulatory effects of peptide T on human keratinocyte cells.

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)-beta and heat shock protein (HSP)-70 in human keratinocyte cells treated with PT. The HSP-70 increase is modulated by TGF-beta. In fact, we demonstrated that anti-TGF-beta antibodies reduce HSP-70 overexpression. In addition, we show a modulation of alphav 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.

Adult↗
PubMed2002

Prevention and treatment of keloids with intralesional verapamil.

BACKGROUND: The current treatment of keloids is based on surgery, medical therapy and physical treatment. The optimal treatment remains undefined. OBJECTIVE: This clinical study determines the effectiveness of verapamil hydrochloride on the prevention and treatment of keloids. METHODS: Two groups of patients with keloids were divided according to lesion site and age. The first group was treated by perilesional surgical excision of keloids and topical silicone, combined with adjuvant treatment of intralesional 2.5 mg/ml verapamil hydrochloride injections administered at timed intervals. The second group served as control, receiving the same treatment except the verapamil hydrochloride. RESULTS: A periodical examination to check specific clinical parameters and an 18-month follow-up determined that keloids were cured in 54% of the cases in the first group, compared to 18% in the second group. Moreover, in the remaining 36% of patients in the first group in whom keloids recurred, there was an improvement in size and above all in consistence. CONCLUSION: These findings suggest that verapamil hydrochloride is an effective tool for the prevention and therapy of keloids.

Administration, Topical↗
PubMed2001

UVA irradiation induces L-isoaspartyl formation in melanoma cell proteins.

It has been reported that UVA effects are partly mediated by production of reactive oxygen species. Moreover, oxidative stress increases protein damage, involving the occurrence of isoaspartyl residues, a product of protein deamidation/isomerization reactions. This work was undertaken in order to study the effects of UVA irradiation, mediated by oxidation, on sensitive protein targets. Melanoma cells exposed to UVA rays have been chosen as a model for monitoring the occurrence of L-isoaspartyl sites. A dramatic increase of these abnormal residues, specifically recognized and methylated by the enzyme L-isoaspartate(D-aspartate) O-methyltransferase (PCMT; EC 2.1.1.77), can be detected after exposure of M14 cells to raising doses of UVA. The effect of UVA on NO and TBARS accumulation, as well as on DNA fragmentation, has also been investigated. NO formation parallels the increase in isoaspartyl formation, while lipid peroxidation occurs only at the highest UVA doses. No DNA fragmentation has been detected under the employed experimental conditions. These results, as a whole, indicate that protein damages are one of the early events on UVA-induced cell injury. The endogenous activity of PCMT remains remarkably stable under UVA treatment, suggesting that this enzyme might play a crucial role in the repair and/or disposal of damaged proteins in UVA-irradiated cells.

Animals↗
PubMed2001

Uptake of Malassezia furfur by human dermal fibroblasts: effect of ketoconazole and cytoskeleton inhibitors.

We showed the ability of human dermal fibroblasts to take up Malassezia furfur and the effect of ketoconazole and cytoskeleton inhibitors, including cytochalasin D and colchicine, on invasivity. Engulfment was evaluated by May Grunwald Giemsa stain and confirmed by acridine orange staining and electron microscopy. Both revealed the different steps of engulfment, including a fusion event between lysosomes and phagosomes containing M. furfur. Subinhibitory concentrations of ketoconazole (5 microg/ml) reduced the invasive capacity compared to controls (52.0+/-6.3 vs 10.0+/-1.2). M. furfur induced changes in the cytoskeleton of human dermal fibroblasts, with signs of disaggregation of actin fibres. We also studied the effect of the cytoskeleton inhibitors, cytochalasin D (1 microg/ml) and colchicine (1 microg/ml), on engulfment. Cytochalasin D, an inhibitor of actin polymers, inhibited the uptake of M. furfur by human dermal fibroblasts. Colchicine, a microtubule inhibitor, reduced the uptake of M. furfur less markedly. This suggests that the process of engulfment is F-actin-dependent, but the integrity of microtubules is also important in "non-professional" phagocytic cells such as dermal fibroblasts.

Antifungal Agents↗
PubMed2001

Malassezia furfur invasiveness in a keratinocyte cell line (HaCat): effects on cytoskeleton and on adhesion molecule and cytokine expression.

The lipophilic yeast Malassezia furfur is a member of the cutaneous microbiota, also associated with several chronic diseases such as pityriasis versicolor, folliculitis, seborrhoeic dermatitis, and some forms of atopic dermatitis, psoriasis and confluent and reticulate papillomatosis. In this study we determined the immunomodulatory and invasive capacity of M. furfur in a human keratinocyte cell culture, HaCat. At a yeast cell to HaCat ratio of 30:1, M. furfur penetration was only 30% with poor phagolysosome fusion and with cytoskeleton modification. Transglutaminase I gene expression was also inhibited, supporting the hypothesis that M. furfur causes an initial break in the barrier function of the epidermis. Moreover, we demonstrated that M. furfur modulates proinflammatory and immunomodulatory cytokine synthesis by downregulating IL-1alpha and by inhibiting IL-6 and TNF-alpha and by upregulating IL-10 and TGF-beta1. The suppressed inflammatory response induced by M. furfur may play a role in chronic disease.

Cell Division↗
PubMed2001

Peripheral blood T lymphocytes from systemic sclerosis patients show both Th1 and Th2 activation.

Our objective was to investigate the phenotype of helper T cells in the peripheral blood of patients with systemic sclerosis (SSc). PBMC from 15 patients with SSc and 15 sex- and age-matched controls were investigated for lymphocyte subpopulations (CD3, CD4, CD8, CD19, CD16/CD56, CD3-DR); IL-2, IL-4, and IFN-gamma mRNAs; and the relative cytokines in their cytoplasm. The last assay was carried out both in unstimulated and in PMA-activated PBMC. SSc patients presented a higher percentage of activated T cells, CD3+ DR+ (19.7 +/- 9.9 vs 5.1 +/- 2.5%; P < 0.0001); 12 of them presented IFN-gamma mRNA-positive cells; and none IL-2 or IL-4 mRNAs. Under basal conditions, PBMC from six SSc patients contained IL-2, IL-4, and IFN-gamma (i.e., they showed both Th1 and Th2 activation), and 1 IFN-gamma only. PMA-stimulated PBMC of patients differed from those of controls only in the increased percentage of IFN-gamma positive cells (52 +/- 12 vs 37 +/- 11%; P < 0.01). Our study demonstrates that Thl activation occurs in the peripheral blood of SSc patients. This evidence must be faced with from both a pathogenetic and a therapeutical point of view.

Adult↗
PubMed2001

Porins from Salmonella typhimurium accelerate human blood coagulation in vitro by selective stimulation of thrombin activity: implications in septic shock DIC pathogenesis.

The effect of porins, major hydrophobic outer membrane proteins purified from Salmonella typhimurium, on human blood coagulation was investigated. It was found that micromolar concentrations of porins accelerated markedly human blood coagulation in vitro. Using appropriate experiments, data were obtained showing that the main target of the porin-induced procoagulant effect was thrombin. A possible binding of porins with thrombin has been suggested to be the basis of this effect. The implications of this finding in the pathogenesis of the disseminated intravascular coagulation syndrome (DIC) occurring during the Gram-negative septic shock is discussed.

Antithrombin III↗
PubMed2001

Histopathological features and modulation of type IV collagen expression induced by Pseudomonas aeruginosa lipopolysaccharide (LPS) and porins on mouse skin.

BACKGROUND: Pseudomonas aeruginosa (P. aeruginosa) is responsible for serious infections in the immunocompromised host. Many skin lesions induced by P. aeruginosa have been described. Few investigations have been performed on the local action of P. aeruginosa components. OBJECTIVES: To shed light on the "in vivo" activity of lipopolysaccharide (LPS) and porins extracted from P. aeruginosa, by verifying their effects after inoculation in mouse skin through the observation of histological changes and immunohistochemical expression of collagen IV. RESULTS: Both substances were able to induce a similar inflammatory process and a characteristic reversible change in collagen IV distribution. Interestingly, a fibroblast increase was observed at 24 h in the skin treated with porins, while it appeared later in the skin treated with LPS. Besides these changes, porins particularly increased collagen edema, together with disgregation of hypodermal structures. Moreover "in vitro", porins were able to stimulate fibloblasts 3T3 to convert 72 kDa type IV collagenase into the activated 62 kDa form and to release the 92 kDa collagenase. CONCLUSION: LPS and porins, released by gram-negative bacteria during cell growth and lysis, interact with the host at target cells, such as keratinocytes, fibroblasts and immunocompetent cells, thus contributing significantly to the pathogenesis of P aeruginosa skin infections.

3T3 Cells↗
PubMed2000

Hyperinsulinaemia in offspring of Type 2 diabetic patients: impaired response of carbohydrate metabolism, but preserved cardiovascular response.

AIMS: To investigate the effects of endogenous insulin on haemodynamics in nine offspring of Type 2 diabetic patients (P), compared with 18 subjects without family history of diabetes (C), all with normal glucose tolerance. METHODS: All subjects underwent a 180-min oral glucose tolerance test with continuous blood pressure and ECG recording. Low-to-high frequency ratio (LF/HF), an index of the sympatho-vagal balance, was calculated by heart rate spectral analysis. RESULTS: At baseline, LF/HF correlated with fasting plasma insulin (r = 0.44, P < 0.03) and with insulin/glucose ratio (r = 0.46, P < 0.03). Plasma insulin, basally similar in the two groups, was significantly increased in P (342 +/- 34.2) when compared to C (177.6 +/- 25.2 pmol/l), P < 0.005 from time 30min onward. Blood glucose, also similar at baseline, remained not significantly different in P (5.74 +/- 0.25) vs. C (5.08 +/- 0.27 mmol/l), throughout the study. Diastolic blood pressure significantly decreased in P, but not in C during the first hour of the study. Finally, LF/HF ratio significantly increased in P (2.5 +/- 0.4 vs. C, 1.7 +/- 0.2) during the first hour. CONCLUSIONS: In conclusion, the offspring of Type 2 diabetic patients with normal glucose tolerance display an increased insulin secretion; however, they are not resistant to the haemodynamic effects of insulin, as suggested by the reduction of diastolic blood pressure. This, in turn, may determine a chronic sympathetic activation, which could be involved in the pathogenesis of Type 2 diabetes mellitus.

Adult↗
PubMed2000

Lactoferrin levels in term and preterm milk.

OBJECTIVE: Even though there is no doubt that human milk is the best nourishment for the neonate, there is still controversy regarding its suitability for preterm infants. The aim of this study was to contribute to the knowledge of the anti-infective properties of preterm milk, measuring lactoferrin levels, which are a non-specific protective factor. METHODS: Samples from 26 preterm and 20 term mothers (mean gestational age +/- standard deviation, 30.9+/-2.6 and 39.5+/-1.1 weeks, respectively) were collected during the first month post-partum. Milk samples were obtained by total expression of one breast between 10 a.m. and noon. An aliquot was kept at -20 degrees C until analyzed by SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel electrophoresis). Each sample was run in duplicate. RESULTS: Lactoferrin levels (X +/- SD) in colostrum and mature milk varied from 575.0+/-218.2 mg/dL to 459.4+/-190.7 mg/dL in preterm samples and from 970.6+/-288.6 mg/dL to 292.0+/-167.4 mg/dL in term samples. No significant differences were observed between preterm and term groups, in spite of the trend observed in colostrum, where term milk tended to show higher levels than preterm milk. Decreasing values were observed in both groups along time (ANOVA, p<0.05). However, in the preterm group, lactoferrin levels seemed to maintain rather constant values from the eighth post-partum day onwards. CONCLUSIONS: The trend to higher levels of lactoferrin in preterm mature milk would allow maintenance of the protective effect of human milk in preterm infants in spite of the small volumes ingested by these neonates. These findings support the practice of feeding premature infants with their own mothers' milk at a time when their immune systems have not completely developed.

Colostrum↗
PubMed2000

New strategies in dandruff treatment: growth control of Malassezia ovalis.

BACKGROUND: Cutaneous infections induced by Malassezia ovalis (Pityrosporum ovale) represent a therapeutic problem due to the high rate of recurrence. OBJECTIVE: We studied feasible strategies to control the growth of M. ovalis, compatible with topical use in cosmetic formulations. Studies were performed on the effects of pH, ionic strength, cinnamic acid and related compounds on mycotic growth. METHODS: M. ovalis was cultivated in modified Sabouraud agar. The effects of pH, ionic strength and cinnamic acid and related compounds on mycotic growth were studied by the membrane filter method. RESULTS: In vitro growth of M. ovalis is strongly affected by pH and ionic strength. pH 4.5 induced a growth inhibition of about 95% and 1 M NaCl, at the optimal growth pH, reduced cell growth by over 90%. Cinnamic acid showed an inhibitory effect of 50% at 0.005 g/dl; 30 min incubation with cinnamic acid 0.5 g/dl had a mycocidic effect. CONCLUSION: These results suggest the use of cosmetic compositions containing cinnamic acid or buffered acidic lotions and shampoos in the treatment of M. ovalis infections of the scalp, eventually in addition or alternative to antimycotic drugs or in maintenance therapy. Cosmetic formulations with high ionic strength or skin irritant derivatives such as cinnamaldehyde cannot be proposed for practical use.

Acrolein↗
PubMed1999

Ketoconazole inhibits lipopolysaccharide-induced activation of the nitric oxide synthase gene in the murine macrophage cell line J774.

The aim of this study was to determine whether ketoconazole can affect the expression of the nitric oxide (NO) synthase gene in the murine macrophage cell line J774. The inducible enzyme (i-NOS) is activated in murine macrophages by LPS and cytokines. Exposure of the J774 cell line to ketoconazole for 24 h did not induce any NO release. Cells preincubated with ketoconazole and treated with LPS showed a significant decrease in nitrite levels in the culture medium, compared with controls (cells treated with LPS alone). The addition of 1 mM N-monomethyl-L-arginine (L-NMMA), a structural analogue of arginine, reduced nitrite levels by about 88+/-9.2% in cells treated with LPS alone, whereas in those treated with ketoconazole + LPS, the levels were comparable to the baseline values detected in control cells. Northern blotting, used to assess i-NOS mRNA expression in the J774 cells, showed that ketoconazole reduced the LPS-induced increase in i-NOS mRNA activation by about 50%. These results support another mechanism for the antiinflammatory effect of ketoconazole (i.e. reduction in i-NOS gene expression and consequently inhibition of reactive radical NO production), that may explain the antierythema and antiedema action of this compound, besides its antimycotic effects.

Animals↗