A. R. Botticelli

Vagus nerve mediates the protective effects of melanocortins against cerebral and systemic damage after ischemic stroke.

A vagus nerve-mediated, efferent cholinergic protective pathway activated by melanocortins is operative in circulatory shock and myocardial ischemia. Moreover, melanocortins have neuroprotective effects against brain damage after ischemic stroke. Here we investigated cerebral and systemic pathophysiologic reactions to focal cerebral ischemia in rats induced by intrastriatal microinjection of endothelin-1, and the possible protective role of the melanocortin-activated vagal cholinergic pathway. In the striatum and liver of saline-treated control rats, the activation of extracellular signal-regulated kinases, c-jun N-terminal kinases, and caspase-3, the increase in tumor necrosis factor-α (TNF-α) concentration and DNA fragmentation, as well as the increase in TNF-α plasma levels, occurred 10 and 20 h after the ischemic insult suggesting an activation of inflammatory and apoptotic responses. Treatment with [Nle4, D-Phe7]α-melanocyte-stimulating hormone (NDP-α-MSH; 3 or 9 h after stroke) suppressed the inflammatory and apoptotic cascades at central and peripheral level. Bilateral vagotomy and pharmacologic blockade of peripheral nicotinic acetylcholine receptors blunted the protective effect of NDP-α-MSH. The present results show that focal brain ischemia in rats causes significant effects not only in the brain, but also in the liver. Moreover, our data support the hypothesis that a protective, melanocortin-activated, vagal cholinergic pathway is likely operative in conditions of ischemic stroke.

Selective melanocortin MC4 receptor agonists reverse haemorrhagic shock and prevent multiple organ damage

In circulatory shock, melanocortins have life‐saving effects likely to be mediated by MC4 receptors. To gain direct insight into the role of melanocortin MC4 receptors in haemorrhagic shock, we investigated the effects of two novel selective MC4 receptor agonists.

Effect of γ-hydroxybutyrate in two rat models of focal cerebral damage

Abstract γ-Hydroxybutyrate (GHB) and its lactone, γ-butyrolactone (GBL) have been previously shown to produce a protective effect in animal models of cerebral ischaemia/hypoxia, as well as in human conditions of head injury-induced coma. The aim of the present research was to study the effect of GHB in experimental conditions of focal cerebral damage, either induced by ischaemia or excitotoxicity. Under general anaesthesia, rats were injected into the right striatum with either endothelin-1 (ET-1, 0.43 nmol) or kainic acid (7.5 nmol) in a volume of 1 μl. Sham-lesioned rats received 1 μl of the solvent. Both ET-1- and kainic acid-lesioned rats were randomly assigned to one of the following intraperitoneal (i.p.) treatments: (i) and (ii) GHB, 100 or 300 mg kg −1 2 h after the lesion, followed by 50 or 100 mg kg −1 , respectively, every 12 h; (iii) saline, 2 ml kg −1 , same schedule. Sham animals were treated with saline, 2 ml kg −1 , same schedule. Treatments lasted for 10 days. The higher dose of GHB produced a significant protection against the ET-1-induced impairments in sensory-motor orientation and coordinated limb use (evaluated 24 and 42 days after the lesion) and in place learning and memory (Morris test, performed 19 and 39 days after the lesion). The same dose regimen reduced the circling behaviour induced by apomorphine in kainate-lesioned rats (10 days after the lesion), and limited or prevented at all the histological damage produced either by ET-1 or by kainic acid (evaluated 43 or 10 days after the lesion, respectively). These results show that GHB limits both histological and functional consequences of a focal ischaemic or excitotoxic insult of the brain, in rats, even if the treatment is started 2 h after the lesion.

Morphological, Histochemical, and Immunocytochemical Study of CO2 and Er:YAG Laser Effect on Oral Soft Tissues

OBJECTIVE The purpose of this study was to investigate the morphological, histochemical, and immunocytochemical changes of the oral mucosa after CO(2) or Er:YAG laser irradiation. BACKGROUND DATA There have been no comparative reports on CO(2) and Er:YAG laser effects on human oral soft tissues. MATERIALS AND METHODS Tissue preservation was studied in 40 oral biopsies of young patients obtained by CO(2) and Er:YAG laser surgery. Hematoxylin-eosin and Giemsa stains, PAS/diastase treatment, AE1 and AE2 cytokeratins, MiB1/Ki67, and bcl-2 immunoreactions were performed on the laser cut edges on formalin fixed, paraffin embedded biopsies. RESULTS CO(2) laser biopsies show blisters, clefts and erosions of the epithelium. Intracellular edema and lengthened nuclei were also seen. The glycogen content results decreased in CO(2) laser biopsies. Good expression for cytokeratins and cell-cycle proliferation markers were found in Er:YAG biopsies, on the contrary the apoptosis marker was better expressed in CO(2) laser biopsies. CONCLUSION The results suggest that Er:YAG laser may be routinely used in surgery, because of its minimal damage of the epithelial tissue, its low inflammatory reaction, its quicker healing process and its lower risk of scarring.

Neuroprotective effect of γ-hydroxybutyrate in transient global cerebral ischemia in the rat

Abstract The effect of γ-hydroxybutyrate on the histological and behavioral consequences of transient brain ischemia was studied in the four vessel occlusion rat model. In saline-treated animals, 30 min ischemia caused a massive loss of neurons in the hippocampal CA1 subfield (normal neurons: 14%, 5%, 23% and 30% on the 3rd, 10th, 15th and 65th day after ischemia, respectively). γ-Hydroxybutyrate — 300 mg/kg intraperitoneally (i.p.) 30 min before or 10 min after arteries occlusion, followed by 100 mg/kg i.p. twice daily for the following 10 days — afforded a highly significant protection (normal neurons on the 3rd, 10th, 15th and 65th day after ischemia: 88% and 91%, 80% and 80%, 91% and 90%, 72% and 71% in rats receiving the first dose before or after arteries occlusion, respectively). The ischemia-induced sensory–motor impairment was significantly attenuated in rats receiving the first dose of γ-hydroxybutyrate before arteries occlusion. Finally, the ischemia-induced impairment in spatial learning and memory, evaluated starting 27 days after the ischemic episode, was significantly attenuated by γ-hydroxybutyrate, either injected first at 30 min before or 10 min after arteries occlusion. Lower doses of γ-hydroxybutyrate had no significant effect. In conclusion, these results indicate that γ-hydroxybutyrate provides significant protection against both histological and behavioral consequences of transient global cerebral ischemia in rats.

Melanocortins protect against progression of Alzheimer's disease in triple- transgenic mice by targeting multiple pathophysiological pathways

Besides specific triggering causes, Alzheimers disease (AD) involves pathophysiological pathways that are common to acute and chronic neurodegenerative disorders. Melanocortins induce neuroprotection in experimental acute neurodegenerative conditions, and low melanocortin levels have been found in occasional studies performed in AD-type dementia patients. Here we investigated the possible neuroprotective role of melanocortins in a chronic neurodegenerative disorder, AD, by using 12-week-old (at the start of the study) triple-transgenic (3xTg-AD) mice harboring human transgenes APPSwe, PS1M146V, and tauP301L. Treatment of 3xTg-AD mice, once daily until the end of the study (30 weeks of age), with the melanocortin analog [Nle(4),D-Phe(7)]-α-melanocyte-stimulating hormone (NDP-α-MSH) reduced cerebral cortex/hippocampus phosphorylation/level of all AD-related biomarkers investigated (mediators of amyloid/tau cascade, oxidative/nitrosative stress, inflammation, apoptosis), decreased neuronal loss, induced over-expression of the synaptic activity-dependent gene Zif268, and improved cognitive functions, relative to saline-treated 3xTg-AD mice. Pharmacological blockade of melanocortin MC4 receptors prevented all neuroprotective effects of NDP-α-MSH. Our study identifies, for the first time, a class of drugs, MC4 receptor-stimulating melanocortins, that are able to counteract the progression of experimental AD by targeting pathophysiological mechanisms up- and down-stream of β-amyloid and tau. These data could have important clinical implications.

Functional recovery after delayed treatment of ischemic stroke with melanocortins is associated with overexpression of the activity-dependent gene Zif268

Melanocortin peptides afford strong neuroprotection and improve functional recovery in experimental ischemic stroke; they also have established neurotrophic actions. The expression of the immediate early gene Zif268 is dependent on synaptic activity and is involved in injury repair and memory formation. Here, we investigated the role of Zif268 in learning and memory recovery after delayed treatment of ischemic stroke with the melanocortin analog [Nle(4), D-Phe(7)]alpha-MSH (NDP-alpha-MSH). A 10-min period of global cerebral ischemia was induced by occluding both common carotid arteries in gerbils. Treatment with a nanomolar dose of NDP-alpha-MSH (every 12h for 11 days) was performed starting 3h or 9h after stroke induction; where indicated, gerbils were pretreated with the melanocortin MC(4) receptor antagonist HS024. Animals were subjected to the Morris water-maze test (four sessions from 4 to 50 days after the ischemic episode). Fifty days after stroke, histological damage and Zif268 expression were investigated in the hippocampus. Treatment with NDP-alpha-MSH significantly reduced hippocampal damage, including neuronal death, and improved learning and memory recovery. This protective effect was long-lasting (50 days, at least) and associated with Zif268 overexpression, with both schedules of NDP-alpha-MSH treatment. Pharmacological blockade of MC(4) receptors prevented these effects. Our data indicate that MC(4) receptor-mediated actions of melanocortins could trigger repair mechanisms able to improve neuronal functionality and synaptic plasticity, and to promote long-lasting functional recovery from ischemic stroke with Zif268 gene involvement.

[Nle4,D-Phe7]α-MSH improves functional recovery in rats subjected to diencephalic hemisection

Rats subjected to diencephalic hemisection were s.c. treated with α-MSH (20 μg/rat daily) or with [Nle4,D-Phe7]α-MSH (10 μg/rat every other day) for two weeks starting on day 3 after lesion. Apomorphine-induced (1 mg/kg s.c.) rotational behavior was studied on days 7, 14 and 21 after lesion, and a sensorimotor test battery was carried out on days 3, 10, 17 and 24 after lesion. [Nle4,D--Phe7] α-MSH greatly reduced rotational behavior and significantly improved sensorimotor performance. Histological studies showed that treatment with α-MSH and, even more markedly, with [Nle4, D-Phe7] α-MSH reduced the size of the lesion and the pseudoinflammatory reaction, and caused a marked proliferation and hypertrophy of astroglia. Binding studies showed that no supersensitivity of striatal dopamine receptors developed on the lesioned side of α-MSH- and [Nle4,D-Phe7]α-MSH-treated rats. The present results seem to further support the trophic role of MSH peptides on nerve tissue.

Cytopathological and chemico-physical analyses of smears of mucosa surrounding oral piercing

OBJECTIVE The aim of this comparative study was to analyze cytopathologically and chemico-physically the mucosa surrounding oral piercing to correlate results with adverse tissue signs. MATERIALS AND METHODS The tongue superficial mucosa of 15 young subjects (control group) and the superficial mucosa surrounding oral piercing of 15 young subjects (test group, TG) were smeared on slides, Papanicolaou stained and analyzed under the optical microscope. Some smears were prepared for (back-scattered) scanning electron microscope (SEM) and X-ray microanalysis to study piercing fragments. RESULTS Smears of TG displayed a variable extent of bacterial cytolysis of epithelial cells, fungi, hyperkeratosis, parakeratosis, granulocyte infiltration, calcium formations and bacterial flora; the four last statistically significant (P < 0.05). Foreign bodies surrounded by keratinocytes were detected under both light and SEM. X-ray microanalyses highlighted piercing alloy aggression, ion release and an inverse gradient of ion concentration inside keratinocytes. CONCLUSIONS The pathological findings in smears correlated with adverse effects of oral piercing. Ion release may be related to direct toxic effects and belated reactions because of metal sensitization. A strict regulation of piercing is warranted.

Antidiuretic and nephrotoxic effects of putrescine in rats

Putrescine, intraperitoneally injected either into intact or into hypophysectomized rats, caused a reduction in urine volume at doses of 200-300 mg/kg. At doses of 100 mg/kg or more, there was also a significant loss of potassium. The highest dose (300 mg/kg) caused haemoglobinuria, proteinuria, increased natriuresis, increased urinary osmolarity, reduced aldosteronaemia, ectasis of glomerular capillaries and tubular damage. The underlying mechanism(s) are probably mostly linked to the strong cationic charge of putrescine and to its binding to fixed anions of tubular-cell membrane.