Domenico Britti

A role for superoxide in gentamicin-mediated nephropathy in rats

Gentamicin is an antibiotic effective against Gram-negative infection, whose clinical use is limited by its nephrotoxicity. Oxygen free radicals are considered to be important mediators of gentamicin-mediated nephrotoxicity, but the exact nature of the radical in question is not known with certainty. We have investigated the potential role of superoxide in gentamicin-induced renal toxicity by using M40403, a low molecular weight synthetic manganese containing superoxide dismutase mimetic, which selectively removes superoxide. Administration of gentamicin at 100 mg/kg, s.c. for 5 days to rats induced a marked renal failure, characterised by a significant decrease in creatinine clearance and increased plasma creatinine levels, fractional excretion of sodium, lithium, urine gamma glutamyl transferase (gamma GT) and daily urine volume. A significant increase in kidney myeloperoxidase activity and lipid peroxidation was also observed in gentamicin-treated rats. M40403 (10 mg/kg, i.p. for 5 days) attenuated all these parameters of damage. Immunohistochemical localisation demonstrated nitrotyrosine formation and poly(ADP-ribose) synthetase (PARS) activation in the proximal tubule of gentamicin-treated rats. Renal histology examination confirmed tubular necrosis. M40403 significantly prevented gentamicin-induced nitrotyrosine formation, poly(ADP-ribose) synthetase activation and tubular necrosis. These results confirm our hypothesis that superoxide anions play an important role in gentamicin-mediated nephropathy and support the possible clinical use of low molecular weight synthetic superoxide dismutase mimetics in those conditions that are associated with over production of superoxide.

Pyrrolidine dithiocarbamate attenuates the development of acute and chronic inflammation

The nuclear factor‐κB (NF‐κB) is a transcription factor which plays a pivotal role in the induction of genes involved in physiological processes as well as in the response to injury and inflammation. Dithiocarbamates are antioxidants which are potent inhibitors of NF‐κB. We postulated that pyrrolidine dithiocarbamate (PDTC) would attenuate inflammation. In the present study we investigate the effects of PDTC in animal models of acute and chronic inflammation (carrageenan‐induced pleurisy and collagen‐induced arthritis). We report here for the first time that PDTC (given at 100, 30 or 10 mg kg−1 i.p. in the pleurisy model or at 10 mg kg−1 i.p. every 48 h in the arthritis model) exerts potent anti‐inflammatory effects (e.g. significant reduction of (A) pleural exudate formation, (B) polymorphonuclear cell infiltration, (C) lipid peroxidation, (D) inducible nitric oxide synthase (iNOS) activity and nitric oxide production (E) plasma and pleural exudates levels of interleukin‐1β and tumour necrosis factor‐α, (F) histological injury and (G) delayed development of clinical indicators). Furthermore, PDTC reduced immunohistochemical evidence of (A) formation of nitrotyrosine, (B) activation of poly (ADP‐ribose) polymerase (PARP), (C) expression of iNOS and (D) expression of cyclo‐oxygenase‐2 (COX‐2) in the lungs of carrageenan‐treated mice and in the joints from collagen‐treated mice. Additionally, Western blotting and immunohistochemical analysis of lung tissue revealed that PDTC prevented degradation of IKB‐α and translocation of NF‐κB from the cytoplasm into the nucleus. Taken together, our results clearly demonstrate that prevention of the activation of NF‐κB by PDTC reduces the development of acute and chronic inflammation. Therefore, inhibition of NF‐κB may represent a novel approach for the therapy of inflammation.

Effect of N-acetylcysteine on gentamicin-mediated nephropathy in rats.

Studies were performed on the mechanisms of the protective effects of free-radical scavengers against gentamicin-mediated nephropathy. Administration of gentamicin, 100 mg/kg s.c., for 5 days to rats induced marked renal failure, characterised by a significantly decreased creatinine clearance and increased blood creatinine levels, fractional excretion of sodium Na(+), lithium Li(+), urine gamma glutamyl transferase and daily urine volume. A significant increase in kidney myeloperoxidase activity and lipid peroxidation was observed in gentamicin-treated rats. Immunohistochemical localisation demonstrated nitrotyrosine formation and poly(ADP-ribose)synthase activation in the proximal tubule from gentamicin-treated rats. Renal histology examination confirmed the tubular necrosis. N-acetylcysteine (10 mg/kg i.p. for 5 days) caused normalisation of the above biochemical parameters. In addition, N-acetylcysteine treatment significantly prevents the gentamicin-induced tubular necrosis. These results suggest that (1) N-acetylcysteine has protective effects on gentamicin-mediated nephropathy, and (2) the mechanisms of the protective effects can be, at least in part, related to interference with peroxynitrite-related pathways.

The role of the peroxisome proliferator-activated receptor-α (PPAR-α) in the regulation of acute inflammation

The peroxisome proliferator‐activated receptor‐α (PPAR‐α) is a member of the nuclear receptor superfamily of ligand‐dependent transcription factors related to retinoid, steroid, and thyroid hormone receptors. The aim of the present study was to evaluate the role of the PPAR‐α receptor on the development of acute inflammation. To address this question, we used two animal models of acute inflammation (carrageenan‐induced paw edema and carrageenan‐induced pleurisy). We report here that when compared with PPAR‐α wild‐type mice, PPAR‐α knockout mice (PPAR‐αKO) mice experienced a higher rate of the extent and severity when subjected to carrageenan injection in the paw edema model or to carrageenan administration in the pleurisy model. In particular, the absence of a functional PPAR‐α gene in PPAR‐αKO mice resulted in a significant augmentation of various inflammatory parameters (e.g., enhancement of paw edema, pleural exudate formation, mononuclear cell infiltration, and histological injury) in vivo. Furthermore, the absence of a functional PPAR‐α gene enhanced the staining (immunohistochemistry) for FAS ligand in the paw and in the lung and the expression of tumor necrosis factor α and interleukin‐1β in the lungs of carrageenan‐treated mice. In conclusion, the increased inflammatory response observed in PPAR‐αΚΟ mice strongly suggests that a PPAR‐α pathway modulates the degree of acute inflammation in the mice.

Effect of Aminoguanidine in Ligature-induced Periodontitis in Rats

The role of nitric oxide and reactive oxygen species is well-demonstrated in inflammation. In this study, we evaluated the effect of aminoguanidine, a nitric oxide synthase inhibitor, in a rat model of periodontitis. We induced periodontitis in rats by placing a piece of 2/0 braided silk around the lower left 1st molar. At day 8, the gingivomucosal tissue encircling the mandibular 1st molar was removed for biochemical and histological analysis. Ligation significantly increased inducible nitric oxide synthase activity and expression, and damaged tissue revealed increased neutrophil infiltration, lipid peroxidation, and positive staining for nitrotyrosine formation and poly (ADP-ribose) polymerase activation. Ligation significantly increased Evans blue extravasation in gingivomucosal tissue and alveolar bone destruction. Aminoguanidine (100 mg/kg i.p., daily for 8 days) treatment significantly reduced all these inflammatory parameters, indicating that it protects against the tissue damage associated with periodontitis by reducing nitric oxide production and oxidative stress.

High density lipoprotein (HDL) reduces renal ischemia/reperfusion injury

High-density lipoproteins (HDL) have been shown to reduce organ injury and mortality in animal models of shock via modulation of the expression of adhesion molecules and pro-inflammatory enzymes. As renal inflammation plays an important role in the development of ischemia/reperfusion (I/R) injury of the kidney, the aim of this study was to investigate the ability of HDL to alleviate renal dysfunction and injury in a rat model of renal I/R. HDL (80 mg/kg, intravenous) was administered to male Wistar rats 30 min before bilateral renal ischemia for 45 min followed by reperfusion for up to 48 h. After 6-h reperfusion, HDL significantly reduced (1) renal and tubular dysfunction, (2) tubular and reperfusion-injury, and (3) histologic evidence of renal injury. HDL also improved renal function (after 24-h and 48-h reperfusion) and reduced histologic signs of renal injury (after 48-h reperfusion). Administration of HDL significantly reduced the numbers of polymorphonuclear leukocytes (PMN) infiltrating into renal tissues during reperfusion, which was reflected by an attenuation of the increase in renal myeloperoxidase activity caused by I/R. Furthermore, HDL markedly reduced expression of the adhesion molecules, intercellular adhesion molecule-1 (ICAM-1), and P-selectin during reperfusion. The increase in renal malondialdehyde levels caused by renal I/R was also significantly reduced by HDL, suggesting attenuation of lipid peroxidation subsequent to oxidative stress. These results demonstrate that HDL significantly reduces renal I/R injury and severity of ischemic acute renal failure. It is proposed that the mechanism of protection involves reduction of the expression of adhesion molecules, resulting in reduction of PMN infiltration and oxidative stress.

Anticancer activity of liposomal bergamot essential oil (BEO) on human neuroblastoma cells.

Citrus extracts, particularly bergamot essential oil (BEO) and its fractions, have been found to exhibit anticancer efficacy. However, the poor water solubility, low stability and limited bioavailability have prevented the use of BEO in cancer therapy. To overcome such drawbacks, we formulated BEO liposomes that improved the water solubility of the phytocomponents and increased their anticancer activity in vitro against human SH-SY5Y neuroblastoma cells. The results warrant further investigation of BEO liposomes for in vivo applications.

Inducible nitric oxide synthase-deficient mice exhibit resistance to the acute pancreatitis induced by cerulein.

Oxidative stress plays an important role in the early stage of acute pancreatitis as well as the associated multiple organ injury. Here we compare the degree of pancreatitis caused by cerulein in mice lacking the inducible (or type 2) nitric oxide synthase (iNOS) and in the corresponding wild-type mice. Intraperitoneal injection of cerulein resulted in wild-type mice in a severe, acute pancreatitis, which was characterized by edema, neutrophil infiltration, tissue hemorrhage and cell necrosis as well as increases in the serum levels of amylase and/or lipase. The infiltration of the pancreatic tissue of these animals with neutrophils (measured as increase in myeloperoxidase activity) was associated with up-regulation/expression of the adhesion molecules ICAM-1 and P-selectin as well as signs of enhanced lipid peroxidation (e.g., increased tissue levels of malondialdehyde). Immunohistochemical examination demonstrated a marked increase in the staining (immunoreactivity) for nitrotyrosine and poly (ADP-ribose) synthetase (PARS) in the pancreas of cerulein-treated iNOS wild-type mice. In contrast, the degree of pancreatic inflammation and tissue injury (histological score), upregulation/expression of P-selectin and ICAM-1, the staining for nitrotyrosine and PARS, and lipid peroxidation was markedly reduced in pancreatic tissue sections obtained from cerulein-treated iNOS-deficient mice. These findings support the view that iNOS plays an important, pro-inflammatory role in the acute pancreatitis caused by cerulein in mice.

Effects of etanercept, a tumour necrosis factor‐α antagonist, in an experimental model of periodontitis in rats

Etanercept is a tumour necrosis factor antagonist with anti‐inflammatory effects. The aim of our study was to evaluate, for the first time, the therapeutic efficacy of in vivo inhibition of TNF‐α in an experimental model of periodontitis.

Calpain Inhibitor I Reduces the Development of Acute and Chronic Inflammation

There is limited evidence that inhibition of the activity of the protease calpain I reduces inflammation. Here we investigate the effects of calpain inhibitor I in animal models of acute and chronic inflammation (carrageenan-induced pleurisy and collagen-induced arthritis). We report here for the first time that calpain inhibitor I (given at 5, 10, or 20 mg/kg i.p. in the pleurisy model or at 5 mg/kg i.p every 48 hours in the arthritis model) exerts potent anti-inflammatory effects (eg, inhibition of pleural exudate formation, mononuclear cell infiltration, delayed the development of the clinical signs and histological injury) in vivo. Furthermore, calpain inhibitor I reduced (1) the staining for nitrotyrosine and poly (ADP-ribose) polymerase (immunohistochemistry) and (2) the expression of inducible nitric oxide synthase and cyclooxygenase-2 in the lungs of carrageenan-treated rats and in joints from collagen-treated rats. Thus, prevention of the activation of calpain I reduces the development of acute and chronic inflammation. Inhibition of calpain I activity may represent a novel therapeutic approach for the therapy of inflammation.