Preoperative blood glucose concentrations and postoperative outcomes after elective non-cardiac surgery: an observational study.

BACKGROUND: The association between preoperative blood glucose (BG) concentration and outcomes after non-cardiac surgery and the impact of the diabetes diagnosis status remain unclear. We tested two hypotheses: that preoperative BG is related to surgical outcomes; and that this relationship depends on the diabetes diagnosis status of the patient. METHODS: We retrospectively analysed data on 61 536 consecutive elective non-cardiac surgery patients treated at our tertiary care facility. Logistic regression models were used to test the hypotheses before and after adjustment for baseline patient characteristics. Our primary outcome was a composite of in-hospital serious complications and mortality. A second primary outcome was 1 yr mortality. RESULTS: The crude incidence of the composite in-hospital outcome was significantly related to preoperative BG (P<0.001), but not after covariable adjustment (P=0.40). This relationship did not significantly differ between patients with and without diagnosed diabetes (P=0.09). One year mortality was significantly related to preoperative BG, both univariably (P<0.001) and after covariable-adjustment (P<0.001). Patients with diagnosed diabetes and preoperative euglycaemia generally had worse 1 yr mortality than those without diabetes at the same BG {e.g. odds ratio (OR) [95% confidence interval (CI)] of 1.27 (1.06, 1.53) at 6 mmol litre(-1) (108 mg dl(-1)), P=0.003}. Conversely, hyperglycaemic patients with diagnosed diabetes displayed a significantly lower 1 yr mortality than hyperglycaemic patients without diabetes [OR (95% CI) of 0.58 (0.44, 0.77) at 12 mmol litre(-1) (216 mg dl(-1)), P<0.001]. CONCLUSIONS: For elective non-cardiac surgery, preoperative hyperglycaemia should be given greater consideration in patients without diabetes than in those with diagnosed diabetes.

Cavity optomechanical magnetometer.

A cavity optomechanical magnetometer is demonstrated. The magnetic-field-induced expansion of a magnetostrictive material is resonantly transduced onto the physical structure of a highly compliant optical microresonator and read out optically with ultrahigh sensitivity. A peak magnetic field sensitivity of 400 nT Hz(-1/2) is achieved, with theoretical modeling predicting the possibility of sensitivities below 1 pT Hz(-1/2). This chip-based magnetometer combines high sensitivity and large dynamic range with small size and room temperature operation.

Modulation of insulin release by adenosine A1 receptor agonists and antagonists in INS-1 cells: the possible contribution of 86Rb+ efflux and 45Ca2+ uptake.

Due to the lack of specific agonists and antagonists the role of adenosine receptor subtypes with respect to their effect on the insulin secretory system is not well investigated. The A1 receptor may be linked to different 2nd messenger systems, i.e. cAMP, K+- and 45Ca2+ channel activity. Partial A1 receptor agonists are going to be developed in order to improve diabetes (increase in insulin sensitivity, lowering of FFA and triglycerides). In this study newly synthesized selective A1 receptor agonists and antagonists were investigated thereby integrating three parameters, insulin release (RIA), 45Ca2+ uptake and 86Rb+ efflux (surrogate for K+ efflux) of INS-1 cells, an insulin secretory cell line. The presence of A1-receptors was demonstrated by Western blotting. The receptor nonselective adenosine analogue NECA (5-N-ethylcarboxyamidoadenosine) at high concentration (10 microM) had no effect on insulin release and 45Ca2+ uptake which could be interpreted as the sum of effects mediated by mutual antagonistic adenosine receptor subtypes. However, an inhibitory effect mediated by A1 receptor agonism was detected at 10 nM NECA and could be confirmed by adding the A1 receptor antagonist PSB-36 (1-butyl-8-(3-noradamantyl)-3-(3-hydroxy-propyl)xanthine). NECA inhibited 86Rb+ efflux which, however, did not fit with the simultaneous inhibition of insulin secretion. The selective A1 receptor agonist CHA (N6-cyclohexyladenosine) inhibited insulin release; the simultaneously increased Ca2+ uptake (nifedipine dependent) and inhibition of 86Rb+ efflux did not fit the insulin release data. The CHA effect (even the maximum effect at 50 microM) can be increased by 10 microM NECA indicating that CHA and NECA have nonspecific and physiologically non-relevant effects on 86Rb+ efflux in addition to their A1-receptor interaction. Since PSB-36 did not influence the NECA-induced inhibition of 86Rb+ efflux, the NECA effect is not mediated by potassium channel-linked A1 receptors. The nonselective adenosine receptor antagonist caffeine increased insulin release which was reversed by CHA as expected when hypothesizing that both act via A1 receptors in this case. In conclusion, stimulation of A1 receptors by receptor selective and nonselective compounds reduced insulin release which is not coupled to opening of potassium channels (86Rb+ efflux experiments) or inhibition of calcium channels (45Ca2+ uptake experiments). It may be expected that of all pleiotropic 2nd messengers, the cAMP system (not tested here) is predominant for A1 receptor effects and the channel systems (K+ and Ca2+) are of minor importance and do not contribute to insulin release though being coupled to the receptor in other tissues.

End-capping of the modified melanocortin tetrapeptide (p-Cl)Phe-D-Phe-Arg-Trp-NH2 as a route to hMC4R agonists.

Of the 42 R'-X-(p-Cl)Phe-D-Phe-Arg-Trp-NH(2) (X=CO, SO(2), PO, PS) tested at the human (h)MC1, hMC3, and hMC4 receptors (R), the most potent MC4R agonists (EC(50) of 8-20 nM) were obtained by end-capping with R'=CH(2)CHCH(2) (9), NCCH(2) (16), NH(2)COCH(2) (17), HCONHCH(2) (18), CH(3)NH (19), CH(2)CHCH(2)NH (21), 2-Th (23), PhCH(2) (30) and X=CO. These compounds possess 35-60-fold hMC4 versus hMC1Rs selectivity with urea LK-71 (19) being the most potent at hMC4R and MC4/1R selective (EC(50)=8.5 nM, MC4/1R=100). LK-75 (16) combines high potency at hMC4R and MC4/3R selectivity (EC(50)=10.5 nM, MC4/3R=290). SAR is discussed.

Analogs of sub-nanomolar hMC1R agonist LK-184 [Ph(CH2)3CO-His-D-Phe-Arg-Trp-NH2]. An additional binding site within the human melanocortin receptor 1?

Twenty nine analogs of a superpotent MC1R agonist LK-184 (1) were tested at human melanocortin receptors (hMC1, hMC3, and hMC4Rs). All derivatives with the spacer between the N-terminus and the aromatic ring longer or shorter than C(3) were much less potent at hMC1R than 1. Only LK-312 PhCO(CH(2))(3)CO-His-d-Phe-Arg-Trp-NH(2) (3), partially mimicking the pi-system of 1, had an EC(50) of 0.05 nM at hMC1R, which confirms the localization of the pi-binding zone of the receptor. Truncation of 1 to Ph(CH(2))(3)CO-His-d-Phe-Arg-NH(2) gave a full MC1 agonist, LK-394 (30), with an EC(50) of 5 nM and a weak partial agonism at MC3/4Rs. This suggests the existence of an additional binding site within hMC1R next to that for the core sequence His-d-Phe-Arg-Trp-NH(2).

Sub-nanomolar hMC1R agonists by end-capping of the melanocortin tetrapeptide His-D-Phe-Arg-Trp-NH(2).

Twenty three derivatives of the core fragment His(6)-D-Phe(7)-Arg(8)-Trp(9)-NH(2) end-capped with carboxylic and sulfonic acids were synthesized and evaluated at human melanocortin receptors (hMC1, hMC3, and hMC4Rs). The SAR within this series allowed us to map the hMCRs near the His(6) binding site and design a superpotent MC1R agonist, LK-184, Ph(CH(2))(3)CO-His-D-Phe-Arg-Trp-NH(2) (19) with EC(50) 0.01 nM (5 nM at MC3 and MC4Rs).

Detection of Lawsonia intracellularis in swine using polymerase chain reaction methodology.

The polymerase chain reaction (PCR) was evaluated for its usefulness as a diagnostic tool to detect Lawsonia (ileal symbiont) intracellularis. Porcine ilea were collected from swine cases submitted to the Iowa State University Veterinary Diagnostic Laboratory between December 1, 1994, and June 30, 1995. Sampling was random, with no regard to health status. There were 621 ileum scrapings evaluated using the PCR technique. Thirty-five of the samples were positive, either by PCR or conventional diagnostic methods such as histology and Warthin-Starry silver stain. These 35 samples were further evaluated by Warthin-Starry silver stain and indirect immunofluorescent antibody test (IFAT) to confirm the presence of L. intracellularis in the tissue sections. Of the 26 samples positive by PCR, 22 were positive by IFAT. Sixteen of the 22 were also positive when stained with Warthin-Starry and evaluated microscopically for typical bacteria. Nine of the original samples were negative by all 3 techniques. PCR appears more sensitive and specific for L. intracellularis detection than Warthin-Starry stain and IFAT. This study provides evidence that PCR may be useful as a reference standard for the detection of L. intracellularis. PCR may be an appropriate monitoring tool for swine herds because it is a rapid procedure that could be applied to batch testing. Although the test is currently too laborious and expensive for routine diagnostic use, there may be situations in which it is justified because of the advantages of greater sensitivity and specificity.

The local minima method (LMM) of pharmacophore determination: a protocol for predicting the bioactive conformation of small, conformationally flexible molecules.

Software has been developed for potential energy surface analysis and the local minima method of pharmacophore determination. LMM is rigorous and systematic and employs multiple conformations which are the local minima from the potential energy surface of each compound in the data set. It produces a series of possible pharmacophores from a postulated set of pharmacophore elements. The best pharmacophore is then determined by performing a comparative molecular field analysis (CoMFA) on each one. The pharmacophore which produces the most self-consistent model is deemed the best. Local minima on the gas-phase potential energy surface are shown to be a reasonably close approximation to protein bound conformations, and these conformations can be found through systematic conformational searches followed by minimization of the local minima. LMM was used to develop a 3D-QSAR model for dopamine beta-hydroxylase (DBH) inhibitors which was highly predictive (predictive R2 = 0.71 and standard error of predictions = 0.41). The model predicted that the phenyl and thienyl series of inhibitors were acting as bioisosteres. Examination of compounds overlayed in the model indicated a possible hydrogen bond acceptor in the DBH active site. Three tyrosine residues previously labeled by mechanism based inhibitors may be acting as the acceptor and therefore represent excellent candidates for site-directed mutagenesis studies.

Evaluation of antemortem polymerase chain reaction and serologic methods for detection of Lawsonia intracellularis-exposed pigs.

OBJECTIVE: To evaluate polymerase chain reaction (PCR) for detection of Lawsonia intracellularis DNA in feces and an indirect fluorescent antibody test (IFAT) for detecting serum IgG antibodies in pigs exposed to L intracellularis. ANIMALS: 15 seven-week-old pigs and 42 three-week-old pigs. PROCEDURE: During 3 experiments, 23 pigs were inoculated with a pure culture of L intracellularis, 31 pigs served as noninoculated controls, and 3 pigs were used as sentinels. Fecal shedding of L intracellularis was monitored by use of PCR analysis at 7-day intervals. At euthanasia, the ileum was obtained for PCR and histologic analyses. Serum was obtained at 7-day intervals for use in the IFAT. RESULTS: Polymerase chain reaction analysis detected L intracellularis DNA in the feces of 39% of the inoculated pigs; by postinoculation days 21 to 28, 90% of inoculated pigs developed IgG antibodies detected by IFAT. Neither L intracellularis DNA nor IgG antibodies were detected in any of the noninoculated control pigs at euthanasia. Sera from pigs inoculated with enteric pathogens other than L intracellularis did not contain detectable antibodies that reacted with L intracellularis by use of the IFAT. CONCLUSION: The IFAT for L intracellularis IgG antibody detection appeared to be a more sensitive antemortem test for detecting pigs experimentally infected with L intracellularis than was a PCR method for direct detection of the organism in the feces. CLINICAL RELEVANCE: Not all animals that are infected with L intracellularis shed the organism in feces at detectable amounts.

39.5% conversion of low-power Q-switched Nd:YAG laser radiation to 266 nm by use of a resonant ring cavity.

A simple ring cavity consisting of mirrors reflecting at the second harmonic is used to generate the fourth harmonic of low-power Q-switched Nd:YAG laser pulses. A large beam waist of 1 mm minimized the effects of double refraction and thermal loading of the nonlinear crystals. By simulating the low-power pulses with a small portion of the output from a 30-Hz flash-lamp-pumped Q-switched Nd:YAG laser, we obtained IR-UV energy conversion as great as 39.5% to yield 84-mW average power at 266 nm from 213 mW of single-mode 1064-nm radiation.

Pancreastatin inhibits insulin secretion in RINm5F cells through obstruction of G-protein mediated, calcium-directed exocytosis.

To elucidate the regulatory pathway through which pancreastatin inhibits insulin secretion, RINm5F insulinoma cells were challenged with physiological and pharmacological probes known to stimulate insulin release through different mechanisms. Utilizing the electrophysiological technique of capacitance measurements as a correlate to exocytosis, pancreastatin was found to significantly diminish maximum capacitance changes evoked by glyceraldehyde, an effect which was attenuated in pertussis toxin-treated cells. In static incubations of this cell line, pancreastatin significantly inhibited insulin secretion stimulated by glyceraldehyde, carbachol and A23187, secretagogues known to directly elevate beta-cell cytosolic Ca2+. This peptide also inhibited insulin secretion stimulated by phorbol myristate acetate (PMA), but only at incubation times < or = 15 min. It was without effect on insulin secretion stimulated by mastoparan and longer incubations (30 min) with PMA, where the secretory mechanisms are not necessarily Ca(2+)-dependent. Additionally, pancreastatin had no effect on carbachol-generated inositol phosphate accumulation but inhibited simultaneously stimulated insulin secretion. All inhibitory effects of pancreastatin were pertussis toxin sensitive. These results suggest that pancreastatin inhibits insulin secretion in RINm5F cells through a G-protein regulated mechanism at a control point involved in the Ca(2+)-directed exocytotic machinery, a feature shared by other physiologic inhibitors of insulin secretion.

Mechanistic studies of sterol carrier protein-2 effects on L-cell fibroblast plasma membrane sterol domains.

The factors which regulate intermembrane sterol domains and exchange in biomembranes are not well understood. A new fluorescent sterol exchange assay allowed correlation of changes in polarization to sterol transfer. Analysis of spontaneous sterol exchange between L-cell plasma membranes indicated two exchangeable and one very slowly or nonexchangeable sterol domain. The exchangeable domains exhibited half-times of 23 and 140 min with fractional contributions of 5 and 30%, respectively. Sterol carrier protein-2 (SCP-2) enhanced sterol exchange between L-cell plasma membranes and altered sterol domain size in a concentration dependent manner. Previous model membrane studies indicate that SCP-2 alters sterol domains and exchange through interaction with anionic phospholipids. In contrast to these observations, the ionic shielding agents KCl, low pH, or neomycin were either totally or partially ineffective inhibitors of SCP-2 action in L-cell plasma membrane exchanges. Thus the mechanism of SCP-2 in sterol transfer appears to be less charge dependent in L-cell plasma membranes than in model membranes. The cholesterol lowering drug probucol was also capable of altering the sterol exchange kinetics.

Evidence for cholecystokinin receptor subtype in endocrine pancreas.

Cholecystokinin (CCK) is a gut hormone that regulates pancreatic endocrine functions via CCKA receptors. CCK4 (Trp-Met-Asp-Phe-NH2) has an insulinotropic effect, but is 1000-fold less potent than CCK8. The in vitro potencies and selectivity of newly synthesized CCK4 analogs were investigated. Exchanging various a amino acids, for example Met by Nle and modifying Phe and/or Trp, led to compounds that were much more effective than CCK4 itself and show insulinotropic effects comparable with those of CCK8. Compounds that possess electron withdrawing groups on the C-terminal phenylalanine were especially effective; compounds with electron-donating groups had no effect. In contrast to CCK8 the synthetic CCK4 compounds were selective for the endocrine pancreas: they had no agonistic or antagonistic effect on the contraction of the guinea pig ileum, amylase release from isolated acini, and no major effect on the feeding behavior of mice being supplied with either compound by an implantable AlzetR pump for 8 days. The data indicate that some of the synthetic tetrapeptides exhibit a high affinity for the CCK receptor of the endocrine pancreas and that they are highly selective for this (peripheral) CCKA receptor subtype. The beta-cell CCKA receptors are different from those in exocrine pancreas, smooth muscle, and those for regulating appetite; these peripheral receptor subtypes can be discriminated for the first time.

Membrane cholesterol dynamics: cholesterol domains and kinetic pools.

Nonreceptor mediated cholesterol uptake and reverse cholesterol transport in cells occur through cellular membranes. Thus, elucidation of cholesterol dynamics in membranes is essential to understanding cellular cholesterol accumulation and loss. To this end, it has become increasingly evident that cholesterol is not randomly distributed in either model or biologic membranes. Instead, membrane cholesterol appears to be organized into structural and kinetic domains or pools. Cholesterol-rich and poor domains can even be observed histochemically and physically isolated from epithelial cell surface membranes. The physiologic importance of these domains is 2-fold: (i) Select membrane proteins (receptors, transporters, etc.) are localized in either cholesterol-rich or cholesterol-poor domains. Consequently, the structure and properties of the domains rather than of the bulk lipid may selectively affect the function of proteins residing therein. (ii) Kinetic evidence suggests that cholesterol transport through and between membranes may occur through specific domains or pools. Regulation of the size and properties of such domains may be controlling factors of cholesterol transport or accumulation in cells. Recent technologic advances in the use of fluorescent sterols have allowed examination of cholesterol domain structure in model and biologic membranes. These techniques have been applied to examine the role of high-density lipoprotein, cholesterol lowering drugs, and intracellular lipid transfer proteins in membrane sterol domain structure and sterol movement between membranes.

Virus envelope-based peptide vaccines against virus-induced mammary tumors.

Previous studies by us and others established that mammary tumors induced by murine mammary tumor virus (MuMTV) could be prevented to various extents by prior vaccination with MuMTV-containing or subviral component immunogens. In this report, four predicted surface-accessible peptide regions (EP-1 to EP-4) of the major viral envelope glycoprotein (gp52) of C3H-MuMTV were tested as carrier-conjugated vaccines for the protection of Balb/c mice against a live virus challenge. With tumor incidence as an endpoint, vaccination with one of these synthetic peptides (EP-3) resulted in a significant reduction in the frequency of early onset tumors and 67% of the test animals remained tumor-free for the entire observation period (16 months). In contrast, only marginal protection was obtained by immunization with the intact glycoprotein (gp52). Immunologic interference may explain the lower protective efficacy of gp52, as compared to EP-3.

Stimulation of insulin secretion from pancreatic islets by the cholecystokinin-tetrapeptide analogs Trp-Pro-Asp-Phe-NH2 and Trp-Pro-Asp-Phe(4'-NO2)-NH2.

The cholecystokinin-tetrapeptide (CCK4) analogs Trp-Pro-Asp-Phe-NH2 (3) and Trp-Pro-Asp-Phe-(4'-NO2)-NH2 (4) were found to be nearly equipotent to cholecystokinin-octapeptide (CCK8) in potentiating glucose-induced insulin secretion from islets of Langerhans isolated from rat pancreas. This stimulatory action was found to be dose-dependent and, in the case of 4, to exhibit a biphasic dose-response curve; i.e., at concentrations greater than 1.0 nM, the stimulating effect of 4 is reversed. These results suggest that conformational restriction of CCK4 and/or modification of the phenylalanine residue could produce more potent analogs capable of stimulating insulin release. Such compounds could have potential therapeutic utility in the treatment of non-insulin-dependent diabetes mellitus (NIDDM).

Synthesis and resolution of novel 3'-substituted phenylalanine amides.

A convenient approach to the synthesis of ring-substituted phenylalanine amides is described. Phase transfer alkylation of N-(diphenylmethylene)amino acetonitrile or N-(diphenylmethylene)glycine ethyl ester provides alpha-substituted imines 2 and 6. After acid hydrolysis and esterification, resolution with alpha-chymotrypsin provided 3'-substituted phenylalanine analogs 9a-d which could easily be converted to N alpha-t-butyloxycarbonyl-phenylalanine amides for use in the synthesis of peptide analogs. The approach described here is amenable to the synthesis of 3-substituted phenylalanines with a wide variety of substituents for determination of structure-activity relationships of peptides.