Nitrocobinamide, a new cyanide antidote that can be administered by intramuscular injection.

Currently available cyanide antidotes must be given by intravenous injection over 5-10 min, making them ill-suited for treating many people in the field, as could occur in a major fire, an industrial accident, or a terrorist attack. These scenarios call for a drug that can be given quickly, e.g., by intramuscular injection. We have shown that aquohydroxocobinamide is a potent cyanide antidote in animal models of cyanide poisoning, but it is unstable in solution and poorly absorbed after intramuscular injection. Here we show that adding sodium nitrite to cobinamide yields a stable derivative (referred to as nitrocobinamide) that rescues cyanide-poisoned mice and rabbits when given by intramuscular injection. We also show that the efficacy of nitrocobinamide is markedly enhanced by coadministering sodium thiosulfate (reducing the total injected volume), and we calculate that ∼1.4 mL each of nitrocobinamide and sodium thiosulfate should rescue a human from a lethal cyanide exposure.

Cell cycle regulation of purine synthesis by phosphoribosyl pyrophosphate and inorganic phosphate.

Cells must increase synthesis of purine nucleotides/deoxynucleotides before or during S-phase. We found that rates of purine synthesis via the de novo and salvage pathways increased 5.0- and 3.3-fold respectively, as cells progressed from mid-G1-phase to early S-phase. The increased purine synthesis could be attributed to a 3.2-fold increase in intracellular PRPP (5-phosphoribosyl-α-1-pyrophosphate), a rate-limiting substrate for de novo and salvage purine synthesis. PRPP can be produced by the oxidative and non-oxidative pentose phosphate pathways, and we found a 3.1-fold increase in flow through the non-oxidative pathway, with no change in oxidative pathway activity. Non-oxidative pentose phosphate pathway enzymes showed no change in activity, but PRPP synthetase is regulated by phosphate, and we found that phosphate uptake and total intracellular phosphate concentration increased significantly between mid-G1-phase and early S-phase. Over the same time period, PRPP synthetase activity increased 2.5-fold when assayed in the absence of added phosphate, making enzyme activity dependent on cellular phosphate at the time of extraction. We conclude that purine synthesis increases as cells progress from G1- to S-phase, and that the increase is from heightened PRPP synthetase activity due to increased intracellular phosphate.

Dietary Na+ inhibits the open probability of the epithelial sodium channel in the kidney by enhancing apical P2Y2-receptor tone.

Apical release of ATP and UTP can activate P2Y(2) receptors in the aldosterone-sensitive distal nephron (ASDN) and inhibit the open probability (P(o)) of the epithelial sodium channel (ENaC). Little is known, however, about the regulation and physiological relevance of this system. Patch-clamp studies in freshly isolated ASDN provide evidence that increased dietary Na(+) intake in wild-type mice lowers ENaC P(o), consistent with a contribution to Na(+) homeostasis, and is associated with increased urinary concentrations of UTP and the ATP hydrolytic product, ADP. Genetic deletion of P2Y(2) receptors in mice (P2Y(2)(-/-); littermates to wild-type mice) or inhibition of apical P2Y-receptor activation in wild-type mice prevents dietary Na(+)-induced lowering of ENaC P(o). Although they lack suppression of ENaC P(o) by dietary NaCl, P2Y(2)(-/-) mice do not exhibit NaCl-sensitive blood pressure, perhaps as a consequence of compensatory down-regulation of aldosterone levels. Consistent with this hypothesis, clamping mineralocorticoid activity at high levels unmasks greater ENaC activity and NaCl sensitivity of blood pressure in P2Y(2)(-/-) mice. The studies indicate a key role of the apical ATP/UTP-P2Y(2)-receptor system in the inhibition of ENaC P(o) in the ASDN in response to an increase in Na(+) intake, thereby contributing to NaCl homeostasis and blood pressure regulation.

The phosphatidylinositol 3-kinase/akt cassette regulates purine nucleotide synthesis.

The phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway is highly conserved throughout evolution and regulates cell size and survival and cell cycle progression. It regulates the latter by stimulating procession through G(1) and the G(1)/S phase transition. Entry into S phase requires an abundant supply of purine nucleotides, but the effect of the PI3K/Akt pathway on purine synthesis has not been studied. We now show that the PI3K/Akt cassette regulates both de novo and salvage purine nucleotide synthesis in insulin-responsive mouse mesenchymal cells. We found that serum and insulin stimulated de novo purine synthesis in serum-starved cells largely through PI3K/Akt signaling, and pharmacologic and genetic inhibition of PI3K/Akt reduced de novo synthesis by 75% in logarithmically growing cells. PI3K/Akt regulated early steps of de novo synthesis by modulating phosphoribosylpyrophosphate production by the non-oxidative pentose phosphate pathway and late steps by modulating activity of the bifunctional enzyme aminoimidazole-carboxamide ribonucleotide transformylase IMP cyclohydrolase, an enzyme not previously known to be regulated. The effects of PI3K/Akt on purine nucleotide salvage were likely through regulating phosphoribosylpyrophosphate availability. These studies define a new mechanism whereby the PI3K/Akt cassette functions as a master regulator of cellular metabolism and a key player in oncogenesis.

Hypoglycemia as a predictor of mortality in hospitalized elderly patients.

BACKGROUND: Hypoglycemia during hospitalization occurs in patients with and without diabetes. The aims of this study were to determine the incidence, associated risk factors, and short- and long-term outcome of hypoglycemia among hospitalized elderly patients. METHODS: This is a case-control study conducted at geriatric and medicine departments. All patients 70 years or older with documented hypoglycemia hospitalized within 1 year (n = 281) were compared with a nonhypoglycemic group of 281 elderly, randomly selected patients from the same hospitalized population. RESULTS: Among 5404 patients 70 years or older, 281 (5.2%) had documented hypoglycemia. Compared with the nonhypoglycemic group, we found the following characteristics to be true in the hypoglycemic group: there were more women than men (58% vs 44%, P =.001); sepsis was 10 times more common (P<.001); malignancy was 2.8 times more common (P =.04); the mean serum albumin level was lower (2.8 g/dL vs 3.4 g/dL, P<.001); and the mean serum creatinine and alkaline phosphatase levels were higher (P<.001 for both). Diabetes was known in 42% of the hypoglycemic group and in 31% of the nonhypoglycemic group (P =.03); 70 patients in the hypoglycemic group were taking sulfonylureas or insulin. Multivariate logistic analysis showed that sepsis, albumin level, malignancy, sulfonyurea and insulin treatment, alkaline phosphatase level, female sex, and creatinine level were all independent predictors of developing hypoglycemia. In-hospital mortality and 3-month mortality were about twice as high in the hypoglycemic group (P<.001). Multivariate analysis of mortality found that sepsis, low albumin level, and malignancy were independent predictors, while hypoglycmia was not. CONCLUSIONS: Hypoglycemia was common in elderly hospitalized patients and predicted increased in-hospital 3- and 6-month cumulative mortality. However, in a multivariate analysis, hypoglycemia was not an independent predictor for mortality, implying that it is only a marker.