Screening for Vitamin D Deficiency in Black Americans: Comparison of Total, Free, Bioavailable 25 Hydroxy Vitamin D Levels with Parathyroid Hormone Levels and Bone Mineral Density.

OBJECTIVES: There is debate surrounding the adequacy of total and free 25 hydroxy vitamin D [25(OH)D] levels in black Americans who have inherently high bone mineral density [BMD] and low serum concentration of vitamin D binding proteins [VDBP]. DESIGN: Retrospective analysis of serum samples and BMD analyses from the African American Health Study [AAHS] cohort. SETTING: The AAHS is a population-based longitudinal study initiated to examine issues of disability and frailty among urban-dwelling black Americans in the city of Saint Louis, Missouri. PARTICIPANTS: 122 men and 206 women, age 60.2 ± 4.3 years. INTERVENTION: Retrospective analysis. MEASUREMENTS: Total 25(OH)D, VDBP, PTH, and BMD of the lumbar spine and hip by dual energy x-ray photometry (DXA). Free and bioavailable vitamin D levels were calculated using serum concentrations and affinity constants for the VDBP (Gc1F and Gc1S) phenotypes. RESULTS: Serum total 25(OH)D levels were 14.6 ± 8.9 ng/mL (36 ± 22 nmol/L). Vitamin D insufficiency was estimated by compensatory elevations of PTH above the normal range (> 65 pg/mL). PTH levels were within the normal reference range in > 95% of the samples at total 25(OH)D levels ≥ 20 ng/mL (≥50 nmol/L). There was no difference in the correlation of the reciprocal relationship of vitamin D vs parathyroid hormone between the VDBP phenotypes. Receiver operating characteristic curve analyses indicated that serum total 25(OH)D discriminated sufficiency from insufficiency at least as well as the calculated levels of the free and bioavailable vitamin D. Very low levels of total 25(OH)D (≤ 8 ng/mL, ≤20 nmol/L) were associated with decreased BMD (p=0.02), but higher levels of 25(OH)D did not show statistical differences in BMD. CONCLUSION: Total 25(OH)D levels of ≤ 8ng/mL (≤20 nmol/L) are associated with clinically significant changes in BMD, whereas total 25(OH)D levels ≥ 20 ng/mL (≥50 nmol/L) suppressed PTH and were not associated with deficiencies in BMD. Lower levels of 25(OH)D may be acceptable for bone health in black than in white Americans.

Triglycerides cross the blood-brain barrier and induce central leptin and insulin receptor resistance.

OBJECTIVE: Resistance at the brain receptors for leptin and insulin has been associated with increased feeding, obesity and cognitive impairments. The causal agent for central resistance is unknown but could be derived from the blood. Here we postulate whether hypertriglyceridemia, the major dyslipidemia of the metabolic syndrome, could underlie central leptin and insulin resistance. DESIGN: We used radioactively labeled triglycerides to measure blood-brain barrier (BBB) penetration, western blots to measure receptor activation, and feeding and cognitive tests to assess behavioral endpoints. RESULTS: Human CSF was determined to contain triglycerides, a finding previously unclear. The radioactive triglyceride triolein readily crossed the BBB and centrally administered triolein and peripherally administered lipids induced in vivo leptin and/or insulin resistance at hypothalamic receptors. Central triolein blocked the satiety effect of centrally administered leptin. Decreasing serum triglycerides with gemfibrozil improved both learning and memory inversely proportionate to triglyceride levels. CONCLUSIONS: Triglycerides cross the blood-brain barrier rapidly, are found in human cerebrospinal fluid, and induce central leptin and insulin receptor resistance, decreasing satiety and cognition.

High Definition Oscillometry: a novel technique for non-invasive blood pressure monitoring in the cynomolgus monkey (Macaca fascicularis).

BACKGROUND: Current approaches for accurate blood pressure determination rely predominantly on invasive techniques. High Definition Oscillometry (HDO) was evaluated as a potential non-invasive approach for accurate blood pressure recordings in cynomolgus monkeys. METHODS: In conscious animals, systolic, diastolic, mean arterial blood pressure (MAP) and pulse/minute were determined 15 times within approx. 9 minutes per individual. This session was performed during 3 consecutive days. Anaesthesia induced hypotension was controlled simultaneously with HDO and telemetry as reference. RESULTS: Repeated measurements were highly reproducible. After procedural habituation, mean MAP was 96.2 +/- 13.7 mmHg in males and 86.9 +/- 4.3 mmHg in females. Mean intraindividual coefficients of variation ranged between 10.8% and 2.4% depending on the session and parameter. Values determined by HDO corresponded to those reported for invasive techniques. CONCLUSION: Our results demonstrate, using telemetry as reference, the accuracy of HDO-based non-invasive blood pressure measurements in macaques to detect drug-related cardiovascular changes.

Measurement of DNA damage after acute exposure to pulsed-wave 2450 MHz microwaves in rat brain cells by two alkaline comet assay methods.

PURPOSE: To investigate the effect of 2450 MHz pulsed-wave microwaves on the induction of DNA damage in brain cells of exposed rats and to discover whether proteinase K is needed to detect DNA damage in the brain cells of rats exposed to 2450 MHz microwaves. MATERIALS AND METHODS: Sprague-Dawley rats were exposed to 2450 MHz pulsed-wave microwaves and sacrificed 4 h after a 2-h exposure. Rats irradiated whole-body with 1 Gy (137)Cs were included as positive controls. DNA damage was assayed by two variants of the alkaline comet assay on separate aliquots of the same cell preparation. RESULTS: Significant DNA damage was observed in the rat brain cells of rats exposed to gamma-rays using both versions of the alkaline comet assay independent of the presence or absence of proteinase K. However, neither version of the assay could detect any difference in comet length and/or normalized comet moment between sham- and 2450 MHz pulsed-wave microwave-exposed rats, regardless of the inclusion or omission of proteinase K in the comet assay. CONCLUSIONS: No DNA damage in brain cells was detected following exposure of rats to 2450 MHz microwaves pulsed-wave at a specific absorption rate of 1.2 W kg(-1) regardless of whether or not proteinase K was included in the assay. Thus, the results support the conclusion that low-level 2450 MHz pulsed-wave microwave exposures do not induce DNA damage detectable by the alkaline comet assay.

Effect of LPS on the permeability of the blood-brain barrier to insulin.

Insulin has emerged as an important neuropeptide. Central actions of insulin appear to oppose those in the periphery. Insulin is transported across the blood-brain barrier (BBB) by a saturable transport system. The permeability of the BBB to insulin is altered by various events, but no studies exist that have examined the permeability of the BBB to insulin during infection or inflammation, states which can induce peripheral insulin resistance. We looked at the effects of lipopolysaccharide (LPS), a bacterial endotoxin and a powerful cytokine releaser, on the permeability of the BBB to human insulin in CD-1 mice. Intraperitoneal injections of LPS significantly increased the uptake by the brain of 131I-insulin and disrupted the BBB to 125I-albumin. After subtraction of the brain/serum ratio for 125I-albumin, brain/serum ratios for insulin were increased: 10.38 +/- 0.70 microl/g (LPS) vs. 3.62 +/- 0.27 microl/g (no LPS), P<0.0001, showing that LPS increased the uptake of insulin independent of BBB disruption. This increase in insulin uptake was due to enhanced saturable transport. Pretreatment with indomethacin 10 min before LPS injections enhanced BBB disruption, but not insulin transport. Pretreatment with the nitric oxide (NO) synthase inhibitor aminoguanidine had no effect on insulin or albumin uptake, but pretreatment with NG-nitro-L-arginine methyl ester (L-NAME) enhanced insulin transport, but not BBB disruption. We conclude that LPS increases the saturable transport of insulin across the BBB independent of disruption and prostaglandins with potentiation by NO inhibition. Such increased transport could potentiate the central effects of insulin and so contribute to the peripheral insulin resistance seen with infection and inflammation.

Laparoscopic v laparoscopy-assisted donor nephrectomy in the porcine model.

PURPOSE: Laparoscopic donor nephrectomy is an established procedure in the porcine model. We sought to compare intraoperative variables between live laparoscopic (LAP) and laparoscopy-assisted (LAP-A) donor nephrectomy. MATERIALS AND METHODS: Eight domestic pigs underwent either traditional laparoscopic donor nephrectomy (N = 4) or laparoscopy-assisted donor nephrectomy (N = 4) using the Pneumosleeve followed by conventional heterotopic autotransplantation. RESULTS: No significant differences were noted between the groups with regard to vessel length, ureteral length, or postoperative urine output. The operating room time was 108+/-12 minutes in the LAP group v 75.8+/-10.3 minutes in the LAP-A group (P = 0.0065). Although the difference was not statistically significant, warm ischemic time, tended to be lower in the LAP-A than the LAP group: 70+/-3.0 seconds v 135+/-57 seconds, respectively (P = 0.059). Graft survival was identical in the two groups. CONCLUSION: Laparoscopy-assisted (via Pneumosleeve) live donor nephrectomy shortens the operative time without affecting graft survival in the domestic swine model.

Effects of tetrodotoxin and OKY-046 in renal ischemia reperfusion.

Ischemia reperfusion injury (IRI) contributes significantly to posttransplant graft dysfunction. An emphasis, therefore, has been directed toward the identification of novel renoprotective agents. In this study, the renoprotective effect of tetrodotoxin (TTX) alone, or in combination with a thromboxane synthetase inhibitor (OKY-046), was investigated in a 60-min warm ischemia, 72-h reperfusion, IRI rodent model. Unilateral nephrectomized rats were treated with the test vehicle alone, 1, 2, or 4 microgram/kg of TTX or 2 mg/kg of OKY-046 intravenously, either 15 min pre- or postischemia, or 2 microgram/kg TTX administered simultaneously with OKY-046 (2 mg/kg), following the ischemic interval. Baseline, 24, and 72 h mean plasma creatinine (Cr) and urea nitrogen (BUN) were compared. Maximal renoprotection was demonstrated by significantly improved 72-h Cr and BUN levels with the 2 microgram/kg of TTX or with 2 mg/kg of OKY-046, each administered after ischemia (ischemic control Cr = 8. 01 +/- 1.07 mg/dl vs TTX = 3.84 +/- 0.80 mg/dl, P = 0.008; vs OKY-046 = 4.0 +/- 1.5, P + 0.008; ischemic control BUN = 241.3 mg/dl +/- 32.8 vs TTX = 85.7 mg/dl +/- 18.7, P < 0.008; vs OKY-046 = 52.6 +/- 22.5, P = 0.008). The combination therapy utilizing TTX with OKY-046 resulted in reduced animal survival, demonstrating no renoprotection as measured with the biochemical parameters. These results support the renoprotective effects of TTX in a severe, rodent IRI model. The exact mechanism of action, as well as the therapeutic potential of TTX in preservation/transplantation, warrants further study.

Evaluation of benzalkonium chloride chemoneurolytic proximal gastric vagotomy.

BACKGROUND: Transmucosal chemoneurolytic injection of benzalkonium chloride (BAC) has previously been shown to duplicate operative proximal gastric vagotomy (PGV) in controlling gastric acid secretion. In this study, BAC was evaluated as to efficacious dose, methods of delivery, and systemic toxicities. METHODS: Sham celiotomy, operative PGV controls, transmucosal injections through a gastrotomy, and transserosal injections of BAC (saline controls, 0. 625, 1.25, 2.5, 5.0, 10 mg BAC/kg body wt) were administered to Sprague-Dawley rats. After 3 months the rats underwent Congo red testing (CRT), horseradish peroxidase (HRP) neuronal staining, and necropsy. The color density change of the gastric mucosa from basic to acidic demonstrated by the CRT at the time of necropsy was used to calculate the residual anatomic acid-secreting area. Prior to necropsy, subserosal HRP injections into the anterior and posterior stomach walls assayed vagal neuronal viability via retrograde axonal flow. Results were compared by an ANOVA. RESULTS: The results demonstrated that 1.25-10 mg/kg transmucosal BAC replicated the results of operative PGV; 2.5 mg/kg was found to be the most effective dose. All injection groups including saline controls demonstrated similar diminished vagal retrograde axonal flow by HRP testing consistent with local BAC chemoneurolytic effects. No systemic toxic symptoms were observed after tail vein intravenous BAC 1.25, 2.5, and 5.0 mg/kg. CONCLUSIONS: These efficacy studies have demonstrated BAC's potential utility in the performance of endoscopic transmucosal chemoneurolytic PGV.

Effect of octreotide on stimulated insulin release from pancreatic tissue slices.

This study was designed to investigate a possible mechanism of action by which octreotide acetate causes insulin suppression in the denervated pancreas. Canine tissue slices were placed in a pH-adjusted medium with varying concentrations of glucose and octreotide acetate: Experiment 1, 30 min in basal medium with 0.6 mg/ml glucose; Experiment 2, addition of 6.0 mg/ml glucose; Experiment 3, addition of 4 microg octreotide acetate/70 ml (comparable to 100 microg/25 kg body weight); Experiment 4, addition of 16 microg octreotide acetate/70 ml; Experiment 5, incubation with 6.0 mg glucose/ml and 4 microg octreotide acetate/70 ml; Experiment 6, incubation with 6.0 mg glucose/ml and 16 microg octreotide acetate/70 ml; Experiment 7, preincubation with 4 microg octreotide acetate/70 ml, then with 6.0 mg glucose/ml; and Experiment 8, preincubation with 16 microg octreotide acetate/70 ml, then with 6.0 mg glucose/ml. Medium levels of insulin, glucagon, and amylase were collected at intervals during the incubation periods. There was an appropriate increase in the rate of insulin release to glucose stimulation in the high-glucose (6.0 mg/ml) group. There was no significant inhibition of basal or glucose-stimulated insulin release with either simultaneous or pretreatment of the canine pancreatic tissue slices with either concentration of octreotide acetate. These studies support an indirect mechanism by which octreotide acetate exerts its inhibitory effect on endocrine and exocrine function in the canine pancreas transplant model.

Renoprotective effects of the 21-aminosteroid U74389G in ischemia-reperfusion injury and cold storage preservation.

Free radical mediated lipid peroxidation (LPO) has been implicated in the pathogenesis of ischemic-reperfusion injury (IRI). To address the renoprotective effect(s) of LPO inhibition, the efficacy of the 21 aminosteroid U74389G was evaluated in three IRI models. In Model 1 51 unilateral nephrectomized rats that underwent 60 min of warm ischemia followed by a 72-hr reperfusion interval were treated with the test vehicle only, or 3, 6, or 12 mg/kg of U74389G intravenously, 5 min pre- or postischemia. In Model 2 Sprague-Dawley rats underwent sham operation (n=9), or 45 min of warm ischemia and 10 min of reperfusion with U74389G (6 mg/kg; n=10) or test vehicle only (n=10) administered intravenously over 10 min beginning 5 min prior to clamp release. After reperfusion, LPO was determined by assay of snap frozen tissue for thiobarbituric acid (TBA) concentrations (nmol/g tissue weight). In Model 3 domestic lean maid pigs (14-18 kg) underwent left nephrectomy with 30 min of warm ischemia, Collins C-4 flush, and 24 hr of cold storage preservation. Heterotopic autotransplantation and immediate contralateral nephrectomy was then performed in Group A-nonischemic controls (n=4), Group B-ischemic controls (n=5), and Group C-U74389G (6 mg/kg) administered preischemia and at autotransplantation (n=5). In Model 1 maximal renoprotection was demonstrated with the 6 mg/kg dose of U74389G administered after ischemia (ischemic control 72-hr serum creatinine (Cr) = 8.01+/-1.1 mg% vs. 3.32+/-0.96 mg%; ischemic control creatinine clearance = 0.069+/-0.03 ml/min vs. 0.206+/-0.04 ml/min; P<0.05). In Model 2 TBA levels were significantly lower in U74389G treated animals (88.5+/-10.0 vs. ischemic controls = 296.8+/-81.4; P=0.02). In Model 3 graft survivals were 100%, 0%, and 60% respectively. Peak Cr and BUN (mg%) were significantly greater in Group C vs. Group A, (Group A Cr = 8.59+/-0.63 vs. Group C = 12.8+/-1.01; Group A BUN = 64.1+/-2.73 vs. Group C = 104.9+/-12.21)--however, by day 10, thee were no significant differences in renal function: (Group A Cr = 2.15+/-0.3 vs. Group C = 2.10+/-0.06; Group A BUN = 27.0+/-6.0 vs. Group C = 31.1+/-6.4). These results support the beneficial effects of LPO inhibitors in models of ischemia-reperfusion, as well as preservation/transplantation, and suggest that this renoprotection correlates with decreased membrane lipid peroxidation.

Effect of somatostatin and octreotide acetate on OP-CCK-stimulated exocrine secretion in the denervated canine pancreas.

Somatostatin and its analogue, octreotide acetate (Sandostatin), have been demonstrated to suppress exocrine secretion in a denervated canine pancreatic autograft model. To help define this inhibitory mechanism, the effect of these agents on cholecystokinin (CCK)-stimulated acinar cell secretion was evaluated. In vitro assessment evaluated the effect of somatostatin on octapeptide (OP)-CCK-stimulated amylase release of pancreatic tissue slices. In vivo assessment employed animals with pancreatic autografts and pancreaticocystostomies, evaluating the effect of a bolus intravenous injection of 100 micrograms of octreotide acetate on the basal and OP-CCK-stimulated (125 ng/kg/h) secretion of urinary (autograft) amylase and bicarbonate. Incubation of tissue slices with 0.16, 0.24, or 0.32 microgram/ml somatostatin had no significant effect on in vitro OP-CCK-simulated amylase release. Intravenous octreotide acetate resulted in a significant decrease in the basal rate of amylase secretion but had no significant effect on OP-CCK-stimulated autograft amylase or bicarbonate release. These studies demonstrate that octreotide acetate has an in vivo inhibitory effect on basal amylase release of pancreatic autografts but cannot counteract maximal stimulation with exogenous OP-CCK. Also, somatostatin does not inhibit OP-CCK-stimulated acinar cell secretion of pancreatic tissue slices. These results indicate that the exocrine inhibition produced by somatostatin analogues in the grafted pancreas occurs via an indirect mechanism.

Effect of L-364,718 on pancreatic endocrine function following partial pancreatectomy.

This study was designed to determine the effect of a potent cholecystokinin antagonist, L-364,718, on canine pancreatic endocrine function following partial pancreatectomy. Plasma glucose, insulin, and glucagon were determined over a 2-hr interval following an intravenous bolus of 0.5 g/kg glucose in a 50% solution. The following groups were established: normal animals (group A, n = 5), normal animals pretreated with 20 nmole/kg L-364,178 (group B, n = 5), partially pancreatectomized animals (group C, n = 5), and partially pancreatectomized animals pretreated with 20 nmole/kg L-364,178 (group D, n = 5). In contrast to animals with an intact pancreas, pretreatment with L-364,718 following partial pancreatectomy resulted in a significant decrease in peak insulin (group C = 132.8 +/- 13.0 microU/ml vs Group D = 90.4 +/- 16.1 microU/ml, P < 0.05) and the basal-to-peak insulin difference (group C = 111.9 +/- 11.5 microU/ml vs group D = 77.5 +/- 16.6 microU/ml, P < 0.05). Despite this, the rate of glucose utilization (K value) was significantly increased in the partially pancreatectomized animals given the antagonist (group C = -1.22 +/- 0.22%/min vs group D = -2.79 +/- 0.427%/min) and there were no significant differences in basal or peak glucose when comparing the groups given L-364,718 with the groups given placebo (group A vs B and group C vs D). Thus, the CCK antagonist L-364,718 significantly decreases peak insulin in partially pancreatectomized animals but not in nonoperative control animals. There is a paradoxical increase in the rate of glucose utilization but no effect on glucose homeostasis. The effect of this antagonist in other models of reduced islet cell reserve (i.e., pancreas transplantation) remains to be determined.

Evaluation of the thromboxane A2 synthetase inhibitor OKY-046 in a warm ischemia-reperfusion rat model.

The pathophysiology of ischemia-reperfusion renal injury is mediated, in part, by the generation of the vasoconstricting prostanoid thromboxane A2 (TXA2). This study was undertaken to evaluate the renoprotective effects, as well as the optimal timing and dosage, of a selective thromboxane synthetase inhibitor, OKY-046, in a unilateral nephrectomized, 60 min ischemia, 72 hr reperfusion, rodent model. Forty-one rats were subjected to right nephrectomy only (group A), or right nephrectomy with 60 min of left renal ischemia and treatment with inactive vehicle only (group B), or 2 mg/kg or 4 mg/kg of OKY-046 administered intravenously before (groups C and D) or after (groups E and F) pedicle clamping. Outcome variables included animal survival; change in kidney weight; 0, 24, and 72 hr plasma creatinine (CR); urea nitrogen (BUN); thromboxane B2 (TXB2) and 6-keto prostaglandin F(1alpha) (6 kPGF(2alpha)) levels; creatinine clearance (CRCL); and histologic evidence of renal injury. Animal survival and postperfusion kidney weight were not significantly different among the groups. However, renal functional parameters were significantly improved with the 2 mg/kg dose of OKY-046 administered after renal ischemia. (group B 72 hr Cr= 8.01 +/- 1.1 mg% vs. group E=3.99 +/- 1.5 mg%, and group B 72 hr BUN=241.3 +/- 32.8 mg% vs. group E=52.6 +/- 22.5 mg%). The CRCL was also improved in group E vs. group B, although these results did not reach statistical significance (group B=0.069 ml/min vs. group E=0.194 ml/ min). The 24 hr TXB2 levels were significantly increased in group B (0 hr=754.1 +/- 219.4 pg/ml vs. 24 hr=2055.9 +/- 550.0 pg/ml), and pre- or posttreatment with OKY-046 abrogated this increase (group C 0 hr=517.1 +/- 80.9 pg/ml vs. 24 hr=384.7 +/- 251.5 pg/ml, and group E 0 hr=781.6 +/- 390.4 pg/ml vs. 24 hr=183.0 +/- 81.4 pg/ml). The 24 hr 6 kPGF(1alpha) levels decreased in all groups, whereas 72 hr 6 kPGF(1alpha) levels increased above baseline in groups A, C, and E, but not in group B. These data demonstrate the beneficial effects of thromboxane A2 synthesis inhibition in the setting of ischemia-reperfusion injury and suggest that this renoprotection correlates with late vasodilatory prostanoid synthesis.

Effect of octreotide on pancreatic endocrine function in partial pancreatectomy.

Octreotide acetate (Sandostatin), a long-acting somatostatin analogue, has been demonstrated to inhibit pancreatic exocrine secretion. The effect of octreotide acetate on pancreatic endocrine function in patients undergoing pancreas surgery or pancreas transplantation has not been as well described, nor have the clinical implications been studied as systematically. This study was designed to investigate the effects of octreotide acetate on glucose metabolism and endocrine function in a partial pancreatectomized canine model, simulating reduced islet cell reserve. Serum levels of glucose, insulin, and glucagon were determined at intervals over 2 hr following an intravenous glucose tolerance test (0.5 g/kg intravenous bolus of 50% glucose) in: normal animals (Group A, n = 5), normal animals pretreated with an intravenous bolus of 400 micrograms of octreotide acetate (Group B, n = 5), partial pancreatectomized animals (Group C, n = 5), and partial pancreatectomized animals pretreated with an intravenous bolus of 400 micrograms of octreotide acetate (Group D, n = 5). Peak glucose concentration was significantly increased in Group D when compared to Group C (Group C = 304.2 +/- 13.5 mg/dl vs Group D = 353.2 +/- 12.9 mg/dl, P < 0.05), indicating an impairment of glucose metabolism by octreotide. In addition, octreotide significantly decreased peak insulin release in the partial pancreatectomy groups (Group C = 129 +/- 12.9 micro U/ml vs Group D = 47.5 +/- 6.8 micro U/ml, P < 0.05). There were no significant differences in the rate of glucose utilization or glucagon concentrations among the groups. These results demonstrate that octreotide does result in insulin suppression, with a resultant increase in stimulated glucose concentrations, in a canine model of reduced islet cell mass. Further studies are required to determine the mechanism of action of octreotide on endocrine function in the setting of pancreas transplant.

Exocrine effects of the CCK antagonist L-364,718 in canine pancreatic autografts.

This study evaluated the effect of the cholecystokinin antagonist L-364,718 on exocrine secretion in canine pancreatic autografts with pancreaticocystostomies. Urinary (autograft) amylase (U/min) and bicarbonate (mmole/min) secretion, over a 6 hr interval, were determined in the basal state (Group A), after a bolus injection of 20 nmoles/kg of L-364,718 (Group B), during a continuous cholecystokinin octapeptide (OP-CCK) infusion at 125 ng/kg/hr either alone (Group C), with a bolus injection of 20 nmoles/kg (Group D), or 30 nmoles/kg (Group E), of L-364,718 1 hr before initiating OP-CCK, or 20 nmoles/kg of L-364,718 1 hr after initiating OP-CCK (Group F). L-364,718 had no effect on basal or OP-CCK-stimulated secretion of bicarbonate. Basal amylase secretion was decreased 1 hr after L-364,718 and remained significantly lower than controls throughout the study interval. When compared to Group C (280.3 +/- 48.6), OP-CCK-stimulated amylase secretion was significantly lower for the first hour after L-364,718 in both Group D (157 +/- 46.7) and Group E (31.9 +/- 11.6). In Group E, 2, 3, and 4 hr post-L-364,718 amylase releases were 60.2 +/- 19.7, 77.7 +/- 25.1, and 87.2 +/- 28.3 compared to 335.5 +/- 85.9, 291.0 +/- 21.8, and 289.9 +/- 45.7 in Group C indicating a sustained significant inhibition of stimulated autograft amylase secretion with the higher L-364,718 dosage. In Group F, no significant change in amylase secretion was demonstrated, indicating that L-364,718 must be administered prior to CCK stimulation to be effective. These studies demonstrate that L-364,718 has a dose dependent, inhibitory effect on basal, and OP-CCK-stimulated amylase secretion in a denervated autograft model. The therapeutic potential of L-364,718 and other CCK receptor antagonists in pancreatic transplantation warrants further study.

Evidence for a colonic PAF receptor.

We sought to determine the effect of exogenously administered platelet-activating factor (PAF) on eicosanoid release from the left colon of the rabbit. Using an isolated buffer-perfused rabbit left colon preparation, 1.0- or 5.0-micrograms doses of PAF were infused into the inferior mesenteric artery. Effluents from the inferior mesenteric vein and colonic lumen were collected and the concentrations of the eicosanoids, prostaglandin E, 6-ketoprostaglandin F1 alpha, thromboxane B2, and leukotriene B4 (LTB4), were measured by ELISA. During PAF infusion there was a significant increase of all prostanoids, but not LTB4 into the venous effluent and colonic luminal perfusate when compared to control experiments. Additional studies were performed by pretreating the colons with the PAF antagonists WEB-2170 or alprazolam prior to PAF infusion. Both venous and luminal effluent prostanoid release was effectively blocked by WEB-2170, but not by alprazolam. Colons pretreated with WEB-2170 prior to PAF had markedly diminished tissue injury when compared to colons treated with PAF alone. Inhibition of PAF-stimulated prostanoid release by WEB-2170 suggests that a PAF-sensitive receptor is present in rabbit colonic tissue which may induce eicosanoid-mediated tissue injury.

Platelet-activating factor mediates trinitrobenzene induced colitis.

Platelet-activating factor (PAF) is an endogenous phospholipid which may be an important mediator of shock and inflammation. Recent evidence suggests that PAF plays a role in the development of ischemic colitis and inflammatory bowel disease. Its effects are mediated by second messengers, including the arachidonic acid metabolites. Using an ex vivo isolated left colon rabbit perfusion model, our aims were to determine whether exogenously administered trinitrobenzene sulfonic acid (TNB), which produces experimental colitis, stimulates both PAF and eicosanoid release in the colon, and if so, whether this effect can be blocked by a PAF antagonist. Colonic inflammation was induced by the intracolonic administration of 0.25 ml of 50% ethanol containing 30 mg of TNB. Tissue and perfusate concentrations of the eicosanoids, [prostaglandin E (PGE2), 6-keto-prostaglandin F1 alpha (6-keto-PGF1 alpha) and thromboxane B2 (TXB2), leukotriene B4 (LTB4)] and the autocoid PAF were measured by ELISA. During TNB infusion there was a significant increase in tissue levels of PAF compared to control colons. Additional studies performed pretreating the colons with the PAF receptor antagonist WEB-2170 prior to TNB infusion blocked PAF release. TNB stimulated release of luminal eicosanoids except LTB4 and suppressed release of tissue prostanoids. Pretreatment with WEB-2170 prior to TNB inhibited luminal eicosanoids, and inhibited PGE2 and prostacyclin, but not TX tissue suppression. Inhibition of TNB-stimulated PAF release by WEB-2170 suggests that PAF may play a role in TNB-induced colitis and this phenomenon may mediate tissue injury.

Effect of platelet-activating factor and its antagonists on colonic dysmotility and tissue levels of colonic neuropeptides.

We investigated whether platelet-activating factor (PAF) alters colonic tissue levels of substance P and vasoactive intestinal peptide (VIP), two neuropeptides that regulate colonic motility. Left colons were harvested from NZ White Rabbits and underwent vascular perfusion via the inferior mesenteric artery. Strain gauge transducers were sewn onto the serosal surface of the colon to evaluate colonic motility. Colons were perfused with either buffered saline alone or with 5.0 x 10(-5) M PAF. PAF administration increased tissue VIP and substance P levels and decreased the force of colonic contractions. Pretreatment with WEB-2170 or alprazolam decreased concentrations of both tissue neuropeptides, and decreased the force of colonic contractions and minute motility index. These results suggest that both VIP and substance P are stimulated by PAF and may participate in colonic dysmotility during inflammatory states.

A laboratory model for evaluation of posttransplant pancreatic exocrine secretion.

Monitoring of urinary enzymuria has been utilized to detect allograft dysfunction after pancreas transplantation with pancreaticocystostomy. In addition, pharmacologic exocrine suppression has been advocated to minimize bicarbonate and protein wasting. Ensuring the validity of these approaches requires controlling both for immunologic alterations in transplant function and for the renal excretion of amylase, bicarbonate, and protein. Toward this end, adult mongrel dogs were divided into two groups. Group A animals underwent distal pancreatectomy alone, and group B animals underwent distal pancreatectomy with autotransplantation and pancreaticocystostomy. In each group, amylase, bicarbonate, and protein output were determined over a 5-hour period in the basal state, during a continuous infusion of octapeptide-cholecystokinin (OP-CCK) at 125 ng/kg/hour, and during a continuous infusion of OP-CCK (125 ng/kg/hour) plus a bolus injection of one clinical unit of secretion per kilogram. Bicarbonate output was not significantly different in the groups with and without autografts. Compared to nonautograft experiments, a statistically significant increase in amylase output was demonstrated in the autograft animals. An increase in protein output was also demonstrated in the autograft experiments, and this increase was statistically significant in the OP-CCK group and the OP-CCK and secretin group. In addition, compared to basal autograft secretion, OP-CCK and OP-CCK plus secretin stimulation resulted in a sustained and significant increase in urinary amylase and protein secretion, indicating preserved sensitivity of the denervated pancreas to exogenous hormones. These results indicate that the canine segmental pancreatic autograft model with pancreaticocystostomy is a suitable model to identify agents associated with exocrine inhibition after transplantation.

The effect of octreotide acetate on meal-stimulated exocrine secretion in canine pancreatic autografts.

Octreotide acetate (Sandostatin), a long-acting somatostatin analogue, has been demonstrated to have an inhibitory effect on exocrine secretion in the neurally intact pancreas. This study was designed to evaluate the effect of this agent on exocrine secretion in the denervated canine pancreas, utilizing animals with pancreatic autografts and functioning pancreaticocystostomies. The rates of secretion of urinary (autograft) amylase (units/min) and bicarbonate (mM/min), over a five-hr interval, were determined in the basal state (group A, n = 10), after a bolus injection of 400 micrograms of Sandostatin (group B, n = 5), after a standard meal (group C, n = 5), or a meal preceded by 400 micrograms of Sandostatin (group D, n = 5). Basal secretion of amylase was decreased for 4 hr following Sandostatin, although this decrease was not significant. Conversely, basal bicarbonate secretion was not inhibited by Sandostatin. When compared with group C (22.4 +/- 3.2), a significant inhibition of meal-stimulated amylase release was demonstrated in group D (5.4 +/- 0.21, P = 0.0006) during the first hour after Sandostatin was given. This inhibition remained significant at 2 hr (group C = 38.5 +/- 5.2 versus group D = 9.4 +/- 0.8; P = 0.0006) and 3 hr (group C = 38.6 +/- 6.3 versus group D = 17.5 +/- 0.9; P = 0.0108) after Sandostatin was given. In addition, meal-stimulated bicarbonate secretion was significantly inhibited for 2 hr following Sandostatin (group C = 0.19 +/- 0.03 versus group D = 0.07 +/- 0.02, P = 0.0096; and group C = 0.23 +/- 0.03 versus group D = 0.10 +/- 0.01, P = 0.0018, respectively). These studies demonstrate that Sandostatin has a profound inhibitory effect on meal-stimulated enzyme and bicarbonate release in a denervated canine autograft model. Although the site of action of this agent remains to be defined, Sandostatin may have therapeutic potential in clinical pancreas transplantation.