Reassessment of mitochondrial cyclophilin D as a target for improving cardiac arrest outcomes in the era of therapeutic hypothermia.

Uncertainty exists regarding whether cyclophilin D (CypD), a mitochondrial matrix protein that plays a key role in ischemia-reperfusion injury, can be a pharmacological target for improving outcomes after cardiac arrest (CA), especially when therapeutic hypothermia is used. Using CypD knockout mice (CypD-/-), we investigated the effects of loss of CypD on short-term and medium-term outcomes after CA. CypD-/- mice or their wild-type (WT) littermates underwent either 5 minute CA followed by resuscitation with and/or without hypothermia at 33°C-34°C (targeted temperature reached within minutes after resuscitation), or a sham procedure. Brain and cardiac injury were assessed using echocardiography, neurological scores, MRI and biomarkers. Seven day survival was compared using Kaplan-Meier estimates. The rate of restoration of spontaneous circulation was significantly higher in CypD-/- mice (with shorter cardiac massage duration) than in WT mice (P < 0.05). Loss of CypD significantly attenuated CA-induced release of troponin and S100ß protein, and limited myocardial dysfunction at 150 minutes after CA. Loss of CypD combined with hypothermia led to the best neurological and MRI scores at 24 hours and highest survival rates at 7 days compared to other groups (P < 0.05). In animals successfully resuscitated, loss of CypD had no benefits on day 7 survival while hypothermia was highly protective. Pharmacological inhibition of CypD with cyclosporine A combined with hypothermia provided similar day 7 survival than loss of CypD combined with hypothermia. CypD is a viable target to improve success of cardiopulmonary resuscitation but its inhibition is unlikely to improve long-term outcomes, unless therapeutic hypothermia is associated.

Dose and timing of injections for effective cyclosporine A pretreatment before renal ischemia reperfusion in mice.

BACKGROUND: There is experimental evidence that lethal ischemia-reperfusion injury (IRI) is largely due to mitochondrial permeability transition pore (mPTP) opening, which can be prevented by cyclosporine A (CsA). The aim of our study is to show that a higher dose of CsA (10 mg/kg) injected just before ischemia or a lower dose of CsA (3 mg/kg) injected further in advance of ischemia (1 h) protects the kidneys and improves mitochondrial function. METHODS: All mice underwent a right unilateral nephrectomy followed by 30 min clamping of the left renal artery. Mice in the control group did not receive any pharmacological treatment. Mice in the three groups treated by CsA were injected at different times and with different doses, namely 3 mg/kg 1 h or 10 min before ischemia or 10 mg/kg 10 min before ischemia. After 24 h of reperfusion, the plasma creatinine level were measured, the histological score was assessed and mitochondria were isolated to calculate the calcium retention capacity (CRC) and level of oxidative phosphorylation. RESULTS: Mortality and renal function was significantly higher in the CsA 10 mg/kg-10 min and CsA 3mg/kg-1 h groups than in the CsA 3mg/kg-10 min group. Likewise, the CRC was significantly higher in the former two groups than in the latter, suggesting that the improved renal function was due to a longer delay in the opening of the mPTP. Oxidative phosphorylation levels were also higher 24 h after reperfusion in the protected groups. CONCLUSIONS: Our results suggest that the protection afforded by CsA is likely limited by its availability. The dose and timing of the injections are therefore crucial to ensure that the treatment is effective, but these findings may prove challenging to apply in practice.

Role of hypothalamic melanocortin system in adaptation of food intake to food protein increase in mice.

The hypothalamic melanocortin system--the melanocortin receptor of type 4 (MC4R) and its ligands: α-melanin-stimulating hormone (α-MSH, agonist, inducing hypophagia), and agouti-related protein (AgRP, antagonist, inducing hyperphagia)--is considered to play a central role in the control of food intake. We tested its implication in the mediation of the hunger-curbing effects of protein-enriched diets (PED) in mice. Whereas there was a 20% decrease in food intake in mice fed on the PED, compared to mice fed on an isocaloric starch-enriched diet, there was a paradoxical decrease in expression of the hypothalamic proopiomelanocortin gene, precursor of α-MSH, and increase in expression of the gene encoding AgRP. The hypophagia effect of PED took place in mice with invalidation of either MC4R or POMC, and was even strengthened in mice with ablation of the AgRP-expressing neurons. These data strongly suggest that the hypothalamic melanocortin system does not mediate the hunger-curbing effects induced by changes in the macronutrient composition of food. Rather, the role of this system might be to defend the body against the variations in food intake generated by the nutritional environment.

Deregulation of hepatic insulin sensitivity induced by central lipid infusion in rats is mediated by nitric oxide.

BACKGROUND: Deregulation of hypothalamic fatty acid sensing lead to hepatic insulin-resistance which may partly contribute to further impairment of glucose homeostasis. METHODOLOGY: We investigated here whether hypothalamic nitric oxide (NO) could mediate deleterious peripheral effect of central lipid overload. Thus we infused rats for 24 hours into carotid artery towards brain, either with heparinized triglyceride emulsion (Intralipid, IL) or heparinized saline (control rats). PRINCIPAL FINDINGS: Lipids infusion led to hepatic insulin-resistance partly related to a decreased parasympathetic activity in the liver assessed by an increased acetylcholinesterase activity. Hypothalamic nitric oxide synthases (NOS) activities were significantly increased in IL rats, as the catalytically active neuronal NOS (nNOS) dimers compared to controls. This was related to a decrease in expression of protein inhibitor of nNOS (PIN). Effect of IL infusion on deregulated hepatic insulin-sensitivity was reversed by carotid injection of non selective NOS inhibitor NG-monomethyl-L-arginine (L-NMMA) and also by a selective inhibitor of the nNOS isoform, 7-Nitro-Indazole (7-Ni). In addition, NO donor injection (L-arginine and SNP) within carotid in control rats mimicked lipid effects onto impaired hepatic insulin sensitivity. In parallel we showed that cultured VMH neurons produce NO in response to fatty acid (oleic acid). CONCLUSIONS/SIGNIFICANCE: We conclude that cerebral fatty acid overload induces an enhancement of nNOS activity within hypothalamus which is, at least in part, responsible fatty acid increased hepatic glucose production.

Portal sensing of intestinal gluconeogenesis is a mechanistic link in the diminution of food intake induced by diet protein.

Protein feeding is known to decrease hunger and subsequent food intake in animals and humans. It has also been suggested that glucose appearance into portal vein, as occurring during meal assimilation, may induce comparable effects. Here, we connect these previous observations by reporting that intestinal gluconeogenesis (i.e., de novo synthesis of glucose) is induced during the postabsorptive time (following food digestion) in rats specifically fed on protein-enriched diet. This results in glucose release into portal blood, counterbalancing the lowering of glycemia resulting from intestinal glucose utilization. Comparable infusions into the portal vein of control postabsorptive rats (fed on starch-enriched diet) decrease food consumption and activate the hypothalamic nuclei regulating food intake. Similar hypothalamic activation occurs on protein feeding. All these effects are absent after denervation of the portal vein. Thus, portal sensing of intestinal gluconeogenesis may be a novel mechanism connecting the macronutrient composition of diet to food intake.