3,5-dihydroxyphenylglycine is a highly selective agonist for phosphoinositide-linked metabotropic glutamate receptors in the rat hippocampus

Metabotropic glutamate receptors (mGluRs) are a heterogeneous family of G protein-coupled glutamate receptors that are linked to multiple second messenger systems in the CNS. In this study the selectivity of mGluR agonists for different mGluR second messenger effects was characterized in slices of the rat hippocampus. The mGluR agonists (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid and (2S,3S,4S)alpha-(carboxycyclopropyl)glycine produced multiple effects on second messengers that included enhanced phosphoinositide hydrolysis in both adult and neonatal rat hippocampus, inhibition of forskolin-stimulated cyclic AMP (cAMP) formation in adult tissue, and increases in basal cAMP formation in the neonatal hippocampus. In contrast, 3,5-dihydroxyphenylglycine was potent and effective in increasing phosphoinositide hydrolysis in both adult and neonatal hippocampus but unlike the other mGluR agonists did not inhibit forskolin-stimulated cAMP formation (in the adult) or substantially enhance basal cAMP formation (in the neonate). Thus, in the rat hippocampus mGluR agonist-mediated increases or decreases in cAMP formation are not secondary to mGluR-mediated changes in phosphoinositide hydrolysis. Furthermore, 3,5-dihydroxyphenylglycine can be used to activate subpopulations of mGluRs coupled to phosphoinositide hydrolysis with minimal effects on cAMP-mGluR second messenger systems.     

Potentiation by specific short-chain fatty acids of differentiation and apoptosis in human colonic carcinoma cell lines

The architecture of normal colonic mucosa suggest that terminally differentiated epithelial cells near the top of the crypt are extruded into the colonic lumen. Morphological studies have identified apoptotic cells among the differentiated phenotypes near the crypt-lumen interface, suggesting a link between pathways of differentiation, apoptosis, and cellular shedding. We studied these processes in HT29 and SW620 cells and found that compared to adherent cells, those cells which were shed during standard, uninduced culture conditions exhibited nonrandom DNA fragmentation characteristic of apoptosis. Moreover, these apoptotic cells, which accumulate in the media, exhibited a more differentiated phenotype. Because short-chain fatty acids (SCFAs) are natural effectors of colonic cell differentiation in vivo, we investigated the specificity of three 4-carbon atom SCFAs on potentiating differentiation and apoptosis, and thus accumulation of shed cells in the conditioned media, in these colonic carcinoma cell lines. Whereas the unbranched SCFA butyrate induced a more differentiated phenotype and enhanced apoptosis, two derivatives of butyrate, branched isobutyric acid and a nonmetabolizable fluorine-substituted analogue, heptafluorobutyric acid, were ineffective in inducing either differentiation or apoptosis. Thus, potentiated differentiation and apoptosis in colonic carcinoma cells were linked to SCFA structure and, most likely, utilization.     

Relationship between plasma phospholipid transfer protein activity and HDL subclasses among patients with low HDL and cardiovascular disease

NK lysin is a 9-kDa polypeptide that was originally isolated from porcine intestinal tissue based on its antibacterial activity. It is produced by cytolytic lymphocytes and is cytolytic against a number of different types of tumor cells. Here we report the binding of NK lysin to lipopolysaccharide (LPS) and its anti-LPS activity. NK lysin binds to matrix-coated LPS from Escherichia coli, Pseudomonas aeruginosa, and different strains of Salmonella enterica. Lipid A and polymyxin B inhibited the binding, demonstrating a preferential interaction of NK lysin with the lipid part of LPS. Chromium-labeled lymphoma cells were lysed by NK lysin, and LPS dose-dependently inhibited the cytolysis at equimolar amounts. In the same manner, NK lysin inhibited certain LPS-stimulated effects on mouse bone marrow cells as well as LPS binding to mouse granulocytes. These results suggest that NK lysin may be a another natural LPS-binding protein from lymphocytes that may participate in the endogenous defense response associated with elevated concentrations of LPS.     

A physical approach to reduce nonspecific adhesion in molecular recognition atomic force microscopy

Atomic force microscopy is one of the few techniques that allow analysis of biological recognition processes at the single-molecule level. A major limitation of this approach is the nonspecific interaction between the force sensor and substrate. We have modeled the nonspecific interaction by looking at the interaction potential between a conical Si3N4 tip with a spherical end face and a mica surface in solution, using DLVO (Derjaguin, Landau, Verwey, Overbeek) theory and numerical calculations. Insertion of the tip-sample potential in a simulation of an approach-retract cycle of the cantilever gives the well-known force-distance curve. Simulating a force-distance curve at low salt concentration predicts a discrete hopping of the tip, caused by thermal fluctuations. This hopping behavior was observed experimentally and gave rise to a novel approach to making measurements in adhesion mode that essentially works in the repulsive regime. The distance between tip and sample will still be small enough to allow spacer-involved specific interactions, and the percentage of nonspecific interactions of the bare tip with the mica is minimized. We have validated this physical model by imaging intercellular adhesion molecule 1 (ICAM-1) antigen with a tip functionalized with anti-ICAM-1 antibody. The measurement demonstrated that a significant decrease in the number of nonspecific interactions was realized, and the topographical image quality and the specific bonding capability of the tip were not affected.     

Multiple genetic aberrations including evidence of chromosome 11q13 rearrangement detected in pituitary adenomas by comparative genomic hybridization

OBJECT: This study was conducted to determine whether comparative genomic hybridization (CGH) is a more sensitive method for detecting genetic aberrations than other tests currently in use. METHODS: The authors used CGH to examine 40 primary and 13 recurrent adenomas obtained from 52 patients for loss and gain of genetic material. Copy number aberrations (CNAs) were detected in 25 (48%) of the 52 patients studied. The chromosomes affected were, in order of decreasing frequency, 11, 7, X, 1, 8, 13, 5, 14, 2, 6, 9, 10, 12, 3, 18, 21, 4, 16, 15, 19, 22, and Y. Endocrinologically active adenomas were more likely to contain (p = 0.009) and had a greater number (p = 0.003) of CNAs. Of 26 adenomas with CNAs, 18 showed multiple aberrations involving entire chromosomes or chromosome arms. The most frequent CNA involving a chromosome subregion, which was present in four (8%) of 53 adenomas, was the loss of all chromosome 11 material except for a preserved common segment containing 11q13. Immunoperoxidase staining did not detect cyclin D1 expression in those four cases, making cyclin D1 an unlikely target of this rearrangement. CONCLUSIONS: These findings indicate that genetic abnormalities are present in pituitary adenomas at a higher rate than previously reported, are associated with endocrinological activity, and often involve several chromosomes. Rearrangement at 11q13 may inactivate a tumor suppressor gene or activate an oncogene that is important in the initiation or progression of sporadic pituitary adenomas.     

Pharmacologic profile of survivors of acute myocardial infarction at United States academic hospitals

Optimal drug therapy for patients with acute myocardial infarction (AMI) is well described in the medical literature. However, data on the actual pharmacologic management of patients surviving AMI at academic hospitals is unavailable. The purpose of this study was to document treatment profiles in 500 patients surviving AMI at 12 academic hospitals in the United States. These profiles were compared with established guidelines and were evaluated for trends. Overall, thrombolytics (streptokinase > or = tissue-type plasminogen activator) were administered in 29% of the patients, with a greater proportion of patients receiving beta-blockers than calcium channel antagonists in the initial 72 hours (61% vs 40%; p < 0.005) and at discharge (51% vs 35%; p < 0.005). Further, women were less likely than men to receive thrombolytic therapy (odds ratio [OR] = 0.61; confidence interval [CI], 0.54 to 0.69) or beta-blocker therapy within the first 72 hours (OR = 0.61; CI, 0.55 to 0.67) or at hospital discharge (OR = 0.53; CI, 0.48 to 0.58). Overall, improvements could still be made in the number of patients who receive thrombolytic and acute and chronic beta-blocker therapies after AMI, particularly in women. Changes in treatment profiles may be a reflection of the publication of large clinical trials.     

Pharmacologic atrial natriuretic peptide reduces human leg capillary filtration

Atrial natriuretic peptide (ANP) is produced and secreted by atrial cells. We measured calf capillary filtration rate with prolonged venous-occlusion plethysmography of supine healthy male subjects during pharmacologic infusion of ANP (48 pmol/kg/min for 15 min; n = 6) and during placebo infusion (n = 7). Results during infusions were compared to prior control measurements. ANP infusion increased plasma [ANP] from 30 +/- 4 to 2,568 +/- 595 pmol/l. Systemic hemoconcentration occurred during ANP infusion: mean hematocrit and plasma colloid osmotic pressure increased 4.6 and 11.3%, respectively, relative to preinfusion baseline values (p < 0.05). Mean calf filtration, however, was significantly reduced from 0.15 to 0.08 ml/100 ml/min with ANP. Heart rate increased 20% with ANP infusion, whereas blood pressure was unchanged. Calf conductance (blood flow/arterial pressure) and venous compliance were unaffected by ANP infusion. Placebo infusion had no effect relative to prior baseline control measurements. Although ANP induced systemic capillary filtration, in the calf, filtration was reduced with ANP. Therefore, pharmacologic ANP infusion enhances capillary filtration from the systemic circulation, perhaps at upper body or splanchnic sites or both, while having the opposite effect in the leg.     

Differential consequences of lateral and central fluid percussion brain injury on receptor coupling in rat hippocampus

We have identified alterations in the responses of muscarinic and metabotropic receptors in rat hippocampus that persist for at least 15 days after central fluid percussion injury. This study compares the effect of lateral fluid percussion and central fluid percussion on these responses. Moderate injury was obtained by displacement and deformation of the brain within the closed cranial cavity using a fluid percussion device positioned either centrally or laterally. Carbachol and (+/-)-1-aminocyclopentane-trans-1,3-dicarboxylic acid (trans-ACPD)-stimulated polyphosphoinositide (PPI) hydrolysis was assayed in hippocampus from injured and sham-injured controls at 15 days following injury. At 15 days after central fluid percussion traumatic brain injury (TBI), the response to carbachol was enhanced by 30% and the response to trans-ACPD was enhanced by 75% compared to sham-injured animals. At 15 days after lateral fluid percussion TBI the response to trans-ACPD was enhanced by 40% both ipsilateral and contralateral to the side of injury. In contrast, the response to carbachol was enhanced by 29% contralateral to the side of injury but was diminished by 12% ipsilateral to the side of injury. Cresyl violet staining shows no hippocampal cell death after central fluid percussion injury or on the side contralateral to lateral fluid percussion injury but on the ipsilateral side cell death was identified in hippocampal area CA3. Thus, abnormal hippocampal cell signaling through the phosphoinositide pathway occurs in the absence of cell death and may contribute to cognitive impairment.