Summary The effect of alimentary carbohydrate overload on hoof temperature was investigated to determine the state of the sublamellar vasculature preceding the onset of equine laminitis. Hoof, core and ambient temperatures and heart rate were logged continuously in 21 mature Standardbred horses kept in an environmental chamber set at 10°C. Recording hoof temperature was a successful, noninvasive, method to measure indirectly, shifts in digital blood flow against a background of cold induced, physiological, vasoconstriction. High hoof temperatures were assumed to indicate digital vasodilation and low hoof temperatures digital vasoconstriction. Seven horses were either untreated or sham treated controls. A slurry of ground wheat flour (17.5 g/kg) was administered via nasogastric tube to 13 horses all of which were humanely killed 48 h later. Histological sections of the lamellar tissues were examined for evidence of laminitis. Analysis of mean hoof temperature graphs showed that horses judged laminitis positive had experienced a period of prolonged digital vasodilation 16–40 h after carbohydrate overload. Laminitis negative horses experienced no such period of vasodilation and never had hoof temperatures significantly (except once, at 28 h) above that of controls. The only parameter which significantly differentiated the laminitis positive from laminitis negative horses, between 12 and 32 h after carbohydrate overload, was foot temperature, which was significantly higher in laminitis positive horses (P<0.05). Therefore, a period of sublamellar vasodilation, 12 to 40 h after alimentary carbohydrate overload precedes the onset of laminitis. If the digital circulation sustains vasoconstriction during this period then laminitis does not occur. We propose that the period of increased digital blood flow in laminitis positive horses, concomitant with the severe metabolic crisis brought on by the alimentary carbohydrate overload, may expose the lamellar tissues to a concentration of blood borne factors sufficient to trigger lamellar separation.

Summary.-A series of 300 patients presenting consecutively with primary operable breast cancer has been studied. A significant correlation was found between oestrogen-receptor (ER) content and histological grade: the better-differentiated tumours rarely lacked receptor. This correlation was significant only in women defined as post-menopausal. Data on early recurrence of disease indicate a worse prognosis for women in whom primary tumours are ER-.

No abstract

ABSTRACT:CYP2C9 is distinguished by a preference for substrates bearing a negative charge at physiological pH. Previous studies have suggested that CYP2C9 residues R97 and K72 may play roles in determining preference for anionic substrates by interaction at the active site or in the access channel. The aim of the present study was to assess the role of these two residues in determining substrate selectivity. R97 and K72 were substituted with negative, uncharged polar and hydrophobic residues using a degenerate polymerase chain reaction-directed strategy. Wild-type and mutant enzymes were expressed in bicistronic format with human cytochrome P450 reductase in Escherichia coli. Mutation of R97 led to a loss of holoenzyme expression for R97A, R97V, R97L, R97T, and R97E mutants. Low levels of hemoprotein were detected for R97Q, R97K, R97I, and R97P mutants. Significant apoenzyme was observed, suggesting that heme insertion or protein stability was compromised in R97 mutants. These observations are consistent with a structural role for R97 in addition to any role in substrate binding. By contrast, all K72 mutants examined (K72E, K72Q, K72V, and K72L) could be expressed as hemoprotein at levels comparable to wild-type. Type I binding spectra were obtained with wildtype and K72 mutants using diclofenac and ibuprofen. Mutation of K72 had little or no effect on the interaction with these substrates, arguing against a critical role in determining substrate specificity. Thus, neither residue appears to play a role in determining substrate specificity, but a structural role for R97 can be proposed consistent with recently published crystallographic data for CYP2C9 and CYP2C5.The cytochrome P450 (P450 1 ) group of enzymes are the predominant catalysts of phase I xenobiotic metabolism in humans and other mammals, catalyzing a variety of monooxygenation reactions including, among others, aliphatic and aromatic hydroxylation, epoxidation, N-and O-dealkylation, and N-and S-oxidation. In humans, approximately 15 enzymes from the CYP1-3 families are responsible for the metabolic clearance of most lipophilic chemicals. Among this group, forms from the same subfamily show discrete but often overlapping substrate specificities.CYP2C9, one of the most important forms in overall drug metabolism, is distinguished from other human CYP2C forms by a preference for substrates bearing a negative charge at physiological pH (Smith and Jones, 1992;Mancy et al., 1995); however, neutral or positively charged substrates may also be substrates. Various models have been developed to explain the preference for anionic substrates.Mansuy and colleagues originally proposed a pharmacophore model based on examination of tienilic acid derivatives and other CYP2C9 substrates (Mancy et al., 1995). In this model, a positively charged residue bordering the substrate binding site was proposed to interact electrostatically with the negative center on the substrate.Homology modeling of the CYP2C9 protein based on crystal structures available for other P450 enzyme...

No abstract