Pharmacological targeting of histamine H4 receptor in periodontal disease.

OBJECTIVE: The objective of this study was to investigate whether histamine H4 receptor (H4 R) antagonists could prevent experimental periodontitis (EP)-induced histological, functional and inflammatory alterations in submandibular gland (SMG), periodontal bone and gingiva. METHODS: Bilateral EP was induced for 2 weeks in anaesthetized male rats. The effect of systemic and local administration of H4 R antagonists (JNJ7777120, JNJ10191584) on histopathology and functionality of SMG, bone loss and gingival inflammation was evaluated. RESULTS: The subcutaneous administration of JNJ7777120 prevented periodontitis-induced SMG histological injury, reducing vacuolization and apoptosis and additionally reversed the increased prostaglandin E2 (PGE2) levels in SMG while it partially reversed the methacholine-induced salivation reduction produced by periodontitis. JNJ7777120 attenuated bone loss and the increased PGE2 levels and inflammatory infiltration in gingival tissue of rats with periodontitis. Finally, local administration of JNJ7777120 and JNJ10191584 was also beneficial for improving periodontal parameters. CONCLUSIONS: H4 receptor antagonists are able to ameliorate periodontitis-induced injury on SMG, gingival tissue and bone structure, suggesting that pharmacological targeting of H4 R could be an attractive strategy to improve periodontal health.

Mitochondrial dysfunction as a mediator of hippocampal apoptosis in a model of hepatic encephalopathy.

In this study, we describe the presence of apoptosis, associated with a mitochondrial dysfunction in the hippocampus of animals in an experimental model defined as minimal hepatic encephalopathy (MHE). This experimental model was studied after 10 days of induced portal vein calibrated stricture, leading to portal hypertension and to a moderate hyperammonemia, without the presence of other evident central nervous system changes. The molecular mechanisms here proposed indicate the presence of apoptotic intrinsic pathways that point to hippocampal mitochondria as an important mediator of apoptosis in this experimental model. In this model of MHE, the presence of DNA fragmentation is documented by 2.3-times increased number of TUNEL-positive cells. These findings together with a higher ratio of the Bcl-2 family members Bax/Bcl-xL in the outer mitochondrial membrane of the MHE animals together with 11% of cytochrome c release indicate the presence of apoptosis in this experimental model. A detailed analysis of the hippocampal mitochondrial physiology was performed after mitochondrial isolation. The determination of the respiratory rate in the presence of malate plus glutamate and ADP showed a 45% decrease in respiratory control in MHE animals as compared with the sham group. A marked decrease of cytochrome oxidase (complex IV of the electron transport chain) was also observed, showing 46% less activity in hippocampal mitochondria from MHE animals. In addition, mitochondria from these animals showed less ability to maintain membrane potential (ΔΨ (m)) which was 13% lower than the sham group. Light scattering experiments showed that mitochondria from MHE animals were more sensitive to swell in the presence of increased calcium concentrations as compared with the sham group. In addition, in vitro studies performed in mitochondria from sham animals showed that mitochondrial permeability transition (MPT) could be a mitochondrial mediator of the apoptotic signaling in the presence of NH(4) (+) and calcium.

Atrial natriuretic peptide reduces inflammation and enhances apoptosis in rat acute pancreatitis.

AIM: We previously reported that atrial natriuretic peptide (ANP) reduces serum amylase and intrapancreatic trypsinogen activation in the onset of acute pancreatitis whereas secretin increases them. In the present work, we sought to establish the effect of ANP and secretin on the inflammatory response and cell death in experimental acute pancreatitis. METHODS: The expression and activity of key inflammatory mediators and apoptosis were evaluated in the presence or absence of the atrial peptide, secretin or both in cerulein-induced acute pancreatitis in rats. Also, ultrastructural changes in pancreatic acinar cells were assessed by transmission electron microscopy. RESULTS: ANP significantly reduced NF-κB activation and TNF-α intrapancreatic levels. Furthermore, it decreased inducible nitric oxide synthase and cyclooxygenase 2 expression and activity while it diminished myeloperoxidase activity. ANP also stimulated apoptosis as shown by caspase-3 expression and activation as well as TUNEL assay. These findings correlated well with the ultrastructural changes observed in the exocrine pancreas. Although secretin reduced various inflammatory markers, it also diminished caspase-3 activation and the overall response was the aggravation of the disease as reflected by the ultrastructural alterations of pancreatic acinar cells. In the presence of ANP, various effects evoked by secretin were antagonized. CONCLUSION: Present findings show that ANP significantly attenuated the severity of acute pancreatitis in the rat by inducing apoptosis and reducing the inflammatory response and further suggest that ANP may have eventual therapeutic implications in the disease and/or in medical interventions at risk of its developing like endoscopic retrograde cholangiopancreatography.

Influence of fasting on the effects of dimethylamiloride and oxfenicine on ischaemic-reperfused rat hearts.

To assess whether glycolysis, Na+-H+ exchange and oxidation of fatty acid derived from endogenous lipolysis are involved in the beneficial effects of 24-h fasting on the ischaemic - reperfused heart, it was studied the effects of inhibiting Na+ - H+ exchange using 10 muM dimethylamiloride and fatty acid oxidation using 2 mM oxfenicine, on the functional activity, lactate production and cell viability measured with tetrazolium stain. Since fasting accelerates heart fatty acid oxidation, data were compared to those from fed rats; using Langendorff perfused (glucose 10 mM) hearts of 250-350 g Wistar rats exposed to 25 min ischaemia - 30 min reperfusion. Fasting reduced the ischaemic rise of end diastolic pressure (contracture), improved recovery of contraction and lowered lactate production in comparison with the fed whereas cellular viability was similar in both groups. Dimethylamiloride improved the recovery of contraction (fed control 24 +/- 9%, fed treated 68 +/- 11%, P < 0.05 at the end of reperfusion), attenuated the contracture (fed control 40 +/- 9%, fed treated 24 +/- 11%, P < 0.05 at the beginning of reperfusion) and reduced lactate production in the fed group and increased cellular viability in both groups (fed control 21 +/- 6%, fed treated 69 +/- 7%, P < 0.05, and fasted control 18 +/- 7%, fasted treated 53 +/- 8%, P < 0.05). Oxfenicine reduced the recovery of contraction (fasted control 88 +/- 6%, fasted treated 60 +/- 11%, P < 0.05) and increased lactate production of fasted group and attenuated the contracture in the fed. These data suggest that beneficial effects of fasting owe, at least in part, to a lowered glycolysis probably secondary to the increased fatty acid oxidation and to the accumulation of energy supplying acyl esters. Dimethylamiloride slowing of glycolysis might explain functional improvement, whereas it seems unrelated to the protection on cell viability.

Effects of 5-hydroxydecanoate and ischemic preconditioning on the ischemic-reperfused heart of fed and fasted rats.

This investigation aimed to assess whether the mitochondrial ATP-sensitive potassium channel blocker 5-hydroxydecanoate (5-HD) could abolish the protection conferred by fasting and ischemic preconditioning (IPC) and to ascertain whether these effects are associated with glycogen breakdown and glycolytic activity. Langendorff perfused hearts of fed and 24-h fasted rats were exposed to 25 min ischemia plus 30 min reperfusion. IPC was achieved by a 3 min ischemia plus a 5 min reperfusion cycle. 5-HD (100 microM) perfusion begun 5 min before IPC or 13 min before sustained ischemia in the non preconditioned groups. Fasting improved the reperfusion recovery of contraction, decreased the contracture and the lactate production, increased glycogenolysis and did not affect the percentage of viable tissue. 5-HD abolished the effects of fasting on the contractile recovery but did not affect the contracture. 5-HD decreased the lactate production in the fed group, increased the preischemic glycogen content in both nutritional groups and did not affect the ischemic glycogen fall. IPC improved the contractile function but prevented the contracture only in the fed group, reduced lactate accumulation and glycogenolysis and evoked an increase of the viable tissue. 5-HD abolished the effects of IPC on the contractile recovery and did not affect its effect on the contracture, lactate production, glycogenolysis and viable tissue. These data suggest that the mitocondrial ATP-sensitive potassium channel is involved in the effects of fasting and IPC on the contractile function but the other cardioprotective and metabolic effects appear evoked through other mechanisms. Also suggest that besides the inhibition of the mitochondrial potassium channel, other mechanisms mediate the effects of 5-HD.

Selective cytoprotective effect of histamine on doxorubicin-induced hepatic and cardiac toxicity in animal models.

The aim of the present work was to evaluate the potential protective effect of histamine on Doxorubicin (Dox)-induced hepatic and cardiac toxicity in different rodent species and in a triple-negative breast tumor-bearing mice model. Male Sprague Dawley rats and Balb/c mice were divided into four groups: control (received saline), histamine (5 mg/kg for rats and 1 mg/kg for mice, daily subcutaneous injection starting 24 h before treatment with Dox), Dox (2 mg/kg, intraperitoneally injected three times a week for 2 weeks) and Dox+histamine (received both treatments). Tissue toxicity was evaluated by histopathological studies and oxidative stress and biochemical parameters. The combined effect of histamine and Dox was also investigated in vitro and in vivo in human MDA-MB-231 triple-negative breast cancer model. Heart and liver of Dox-treated animals displayed severe histological damage, loss of tissue weight, increased TBARS levels and DNA damage along with an augment in serum creatine kinase-myocardial band. Pretreatment with histamine prevented Dox-induced tissue events producing a significant preservation of the integrity of both rat and mouse myocardium and liver, through the reduction of Dox-induced oxidative stress and apoptosis. Histamine treatment preserved anti-tumor activity of Dox, exhibiting differential cytotoxicity and increasing the Dox-induced inhibition of breast tumor growth. Findings provide preclinical evidence indicating that histamine could be a promising candidate as a selective cytoprotective agent for the treatment of Dox-induced cardiac and hepatic toxicity, and encourage the translation to clinical practice.

Contribution of phospholipase A2 to the lethal potency of Bothrops alternatus (víbora de la cruz) venom.

Purified phospholipase A2 from Bothrops alternatus venom is one single protein species with a molecular weight of 15,000 and isoelectric point 5.08. When injected i.p. or i.v. at a dose of 0.7 microgram/g body weight it is lethal to mice, eliciting a typical syndrome of dyspnea, tachycardia, arrhythmia and irreversible shock. Post mortem and histopathologic studies have demonstrated that the lungs (massive pulmonary hemorrhage), heart (foci of myocardial and endocardial necrosis with interfibrillar hemorrhage), liver (congestion, hepatocytic microvacuolization with zones of massive necrosis) and kidneys (foci of tubular and glomerular necrosis) were severely injured. Except for the less extensive hemorrhages and the significantly longer survival time, the observed lesions are similar to those observed after the injection of lethal doses of whole venom. The lethal potency of the purified enzyme (LD50 i.p. 0.14 microgram/g body weight) is 46-fold greater than that of the whole venom (LD50 i.p. 6.4 micrograms/g body weight). The contribution of phospholipase A2 to the overall lethal effect of B. alternatus venom is suggested by the decreased lethal potency of a venom sample in which a significant amount of phospholipase A2 has been removed and the full restoration of the lethal potency upon supplementation of the depleted sample with purified enzyme. It is concluded that phospholipase A2 is a major component responsible for lethality of the whole B. alternatus venom, while the contribution of other venom components appears to be significant mainly in reducing the time of survival.

Effect of chemical modification with p-bromophenacyl bromide on the enzymatic and lethal properties of phospholipase A2 from Bothrops alternatus (Víbora de la Cruz) venom.

The effects on lethal potency and enzymatic activity were determined following alkylation, with p-bromophenacyl bromide, of the acidic toxic phospholipase A2 from Bothrops alternatus. The modified B. alternatus enzyme, which lost its enzymatic activity, retained considerable toxicity. Histopathologic studies on mice have demonstrated features similar to those of the native enzyme. However, the distribution of the damage was different and the survival time was longer. It is concluded that the enzyme activity is not important for the lethal action of the enzyme although it influences the distribution of the damage and survival time.

Tyrosine hydroxilase activity in discrete brain regions from prehepatic portal hypertensive rats.

BACKGROUND/AIMS: Portal hypertension in patients and rat models are characterized by splanchnic and systemic hemodynamic alterations. Both the central and autonomic nervous systems are implicated in its pathophysiology. The aim of our research was to study the tyrosine hydroxylase activity and the rate limiting step in the biosynthesis of catecholamines in partial ligated portal hypertensive and in control rat brains. METHODOLOGY: The following seven discrete brain regions were investigated: Subfornical Organ, Organum Vasculosum Lamina Terminalis, Median Eminence, Periventricular Nucleus, Area Postrema, Locus Coeruleus and Nucleus Tractus Solitarius. RESULTS: The enzyme activity showed a significant increment in six nuclei and a decrease in Area Postrema Nucleus when portal hypertensive rats were compared to controls. CONCLUSIONS: These results suggest the participation of some discrete brain regions in the mechanism of hepatic portal hypertension under the present rat model.

Influence of fasting on the effects of diazoxide in the ischemic-reperfused rat heart.

This investigation aimed to assess whether the mitochondrial ATP-sensitive potassium channel opener diazoxide could reproduce the protection conferred by ischemic preconditioning and to ascertain whether its effects are associated with changes in glycogen breakdown and glycolytic activity. Hearts of fed and 24-h fasted rats were perfused with 10 mM glucose containing medium and exposed to 25 min no-flow ischemia plus 30 min reperfusion. Diazoxide (10 microM) perfusion was begun 10 min before ischemia and continued throughout the experiment. Fasting accelerated reperfusion recovery of contraction, reduced the post-ischemic contracture and decreased lactate accumulation during ischemia but had no effects on glycogen levels and cellular viability. Diazoxide, did not affect glycogen catabolism but improved reperfusion recovery of contraction. Furthermore, diazoxide reduced ischemic lactate accumulation and contracture amplitude only in the fed group whereas it improved cell viability in the fed and fasted groups. These data indicate that: 1) reduced lactate production which may attenuate myocyte acidification might explain, at least in part, the beneficial effects of diazoxide on mechanical function, although data obtained with the fasted rat hearts indicate that other mechanisms must be involved as well; 2) the reduction of lactate production occurring in the fed group, does not seem to be related to glycogenolysis; and 3) since diazoxide improved cell viability in the fasted rat group where it did not reduce glycolytic activity, other mechanisms may be responsible for this cytoprotective effect.

Prostanoid production in endothelial and Kupffer liver cells from monocrotaline intoxicated rats.

UNLABELLED: A single dose of monocrotaline, a pyrrolizidine alkaloid, was injected into rats in order to produce 25 (Group I) and 45 (Group II) days later a progressive and so called delayed liver injury. The present study investigated the prostanoid production of Kupffer cells and endothelial cells separated from Monocrotaline and saline (Group III) injected rat livers. Kupffer cells: formation of 6 keto Prostaglandin F1 alpha, the major prostacycline metabolite, gradually decreased in Groups I vs II (P < 0.01) and in both Groups I and II vs Controls (P < 0.01). In addition Prostaglandin F2 alpha showed a significant increase in Groups I and II when compared to Group III, (P < 0.001), and Thromboxane B2 was present in both Groups of Monocrotaline treated animals, while it was not detectable in the control Group III. Endothelial cells: 6 keto Prostaglandin F1 alpha decreased in Groups 1 vs II. This differences was significant when compared, and compared to controls (Group III, P < 0.001). Prostaglandin E2 was detected only in Groups I and II. Prostaglandin F2 alpha and Thromboxane B2 could not be detected in any Group. Ultramicroscopy showed morphological cell damage in nonparenchymal cells in Monocrotaline intoxication in Group II, rats sacrificed 45 days after the injection, while it shows normal features in those treated animals sacrificed 25 days after the injection, as well as in control group. CONCLUSION: A single Monocrotaline injection produces, 25 and 45 days later, severe and progressive alterations in the prostanoid production in Kupffer and Endothelial cells, while ultramicroscopic alterations was only observed 45 days after the injection of Monocrotaline. A decreased production of vasodilators and the presence of vasoconstrictor prostanoids that can participate in the production of the circulatory derangements enhancing liver injury and portal hypertension were also observed.

Variations in rat biochemical parameters after buckshot implant.

Twenty eight albino Wistar rats were implanted with two 100 mg lead spheres: 14 received the implant in the peritoneum (P) and 14 in the thigh (T). Variations in the activity of delta-aminolevulinic dehydratase (ALAD), of urinary delta-aminolevulinic acid (ALAU), of hematoporphyrins (HP) and of lead blood levels (BPb) were then determined at 30, 60 and 90 days with respect to basal values. Parallel determinations were performed by the same schedule in 7 rats implanted with two glass beads and in 8 sham animals receiving surgical incision alone. Techniques employed for ALAD were Berlin and Schaller; for ALAU, Tomokuni and Ogata; for HP, Piomelli; and for BPb, atomic absorption spectrophotometry. As indicators of lead presence, HP and ALAU proved better, both in P and in T rats. The replacement of lead buckshot for small game hunting by other less toxic elements is recommended.

[Functional alterations in central nervous system of prehepatic portal hypertensive rats]. / Cambios funcionales en el sistema nervioso central de ratas hipertensas portales prehepaticas.

Prehepatic Portal Hypertension (PH) leads to morphologic changes in the rat Central Nervous System, including alterations of the blood brain barrier (BBB), and astrogliosis and angiogenesis in CA1 and CA4 hyppocampal fields. The present study investigates functional changes in portal hypertensive rats. Wistar Kyoto rats were used (240 g/bw) and allotted in two groups: GI (n = 8) portal hypertensive rats obtained through a regulated stenosis of the portal vein (Groszmann), and GII (n = 6), sham-operated rats. We have analyzed: BBB integrity with the Trypan Blue diffusion method (TB, Reynolds), protein concentration (PC) in Cerebrospinal Fluid (CSF) and plasma (Bradford method), electroencephalographic activity (EEG), cerebral edema expressed as brain water content (gravidimetric test), and behavior: Animex, righting reflex, pain reflex and Rotarod. TB was positive in GI in peripheral vascular areas in hippocampus, PC in CSF (ug/ml)(mean +/- SED) was GI: 40.6 +/- 6.8 and GII: 16.5 +/- 4.2 (p < 0.005), and the plasma levels were (mg/ml): GI: 108.8 +/- 7.6 and GII: 87.4 +/- 2 (NS). The EEG showed a higher power of the delta band in hypertensive rats (GI: 0.551 +/- 0.033 and GII: 0.342 +/- 0.031, p < 0.008), but water content was not different between GI and GII (water%/per/g/tissue) (GI: 79.21 +/- 0.2, GII: 78.95 +/- 0.18). These results, showing functional changes in the BBB and brain activity without behavioral alterations, suggest the development of a subclinic form of hepatic encephalopathy in our model of PH rats.

[Benzodiazepines metabolism in various models of experimental liver diseases]. / Metabolismo de benzodiacepinas en distintos modelos de hepatopatías experimentales.

The aim of the present paper is to establish possible disturbances in benzodiazepines glucuronidation in two experimental models of liver injury: paracetamol acute intoxication and cholestasis followed by paracetamol acute intoxication. We concluded that, despite the alterations observed in liver microsomal lipid profile, glucuronidation remained similar to controls in paracetamol intoxicated rats. On the contrary, cholestatic animals followed by paracetamol intoxication showed an increment in the glucuronidation of the utilized substrated.

[Development of biochemical and histopathological parameters in paracetamol overdose in experimental cholestasis]. / Evolución de parámetros bioquímicos e histopatológicos en la sobredosis con paracetamol durante la colestasis experimental.

The aim of the present paper is to determine the effect of Paracetamol (P) acute intoxication in cholestatic rats. Four groups of animals were considered: controls, controls intoxicated with P, rats intoxicated with P and cholestatic rats. Hepatic biochemical tests and liver histology were performed in every group. It is concluded that cholestatic rats intoxicated with P showed less Liver damage than in control groups.

[Does TNF-alpha contribute to liver disease physiopathology?]. / Contribuye el TNF-alpha a la fisiopatología de las hepatopatías?

The aim of the present paper is to establish the possible role of serum TNF in the pathophysiology of three experimental models of liver injury: paracetamol intoxication, cholestasis followed by paracetamol intoxication and cholestasis. We concluded that under our experimental conditions the serum TNF-alpha levels were not responsible for the inflammatory phenomena described in our previous paper as apoptosis.

Effects of unilateral ureteric occlusion on renal necrosis occurring in rats fed a methyl-deficient diet.

Wistar male rats were fed from weaning a methyl-deficient diet (groups I and II) or a standard commercial diet (groups III and IV). At the day 3 the left ureter was tied and divided in animals of groups I and III, while those in groups II and IV were sham operated. Rats of all groups were killed on days 8 and 10 to evaluate the incidence and extent of tubular necrosis (day 8) and tubular and cortical necrosis (day 10). The results of this study show that unilateral ureteric obstruction diminishes the incidence, severity and extent of necrosis in the same kidney.