Inhibition of gastric acid secretion in rat stomach by PACAP is mediated by secretin, somatostatin, and PGE(2)

Pituitary adenylate cyclase-activating polypeptide (PACAP), existing in two variants, PACAP-27 and PACAP-38, is found in the enteric nervous system and regulates function of the digestive system. However, the regulatory mechanism of PACAP on gastric acid secretion has not been well elucidated. We investigated the inhibitory action of PACAP-27 on acid secretion and its mechanism in isolated vascularly perfused rat stomach. PACAP-27 in four graded doses (5, 10, 20, and 50 microg/h) was vascularly infused to determine its effect on basal and pentagastrin (50 ng/h)-stimulated acid secretion. To study the inhibitory mechanism of PACAP-27 on acid secretion, a rabbit antisecretin serum, antisomatostatin serum, or indomethacin was administered. Concentrations of secretin, somatostatin, PGE(2), and histamine in portal venous effluent were measured by RIA. PACAP-27 dose-dependently inhibited both basal and pentagastrin-stimulated acid secretion. PACAP-27 at 10 microg/h significantly increased concentrations of secretin, somatostatin, and PGE(2) in basal or pentagastrin-stimulated state. The inhibitory effect of PACAP-27 on pentagastrin-stimulated acid secretion was reversed 33% by an antisecretin serum, 80.0% by an antisomatostatin serum, and 46.1% by indomethacin. The antisecretin serum partially reduced PACAP-27-induced local release of somatostatin and PGE(2). PACAP-27 at 10 microg/h elevated histamine level in portal venous effluent, which was further increased by antisomatostatin serum. However, antisomatostatin serum did not significantly increase acid secretion. It is concluded that PACAP-27 inhibits both basal and pentagastrin-stimulated gastric acid secretion. The effect of PACAP-27 is mediated by local release of secretin, somatostatin, and PGE(2) in isolated perfused rat stomach. The increase in somatostatin and PGE(2) levels in portal venous effluent is, in part, attributable to local action of the endogenous secretin.     

Proadrenomedullin NH(2)-terminal 20 peptide (PAMP) and adrenomedullin bind to teratocarcinoma cells

Proadrenomedullin NH(2-)terminal 20 peptide (PAMP) and adrenomedullin (ADM) bind to teratocarcinoma cells. The effects of PAMP and ADM on teratocarcinoma cells were investigated. (125)I-PAMP bound to PA1 cells with moderate affinity (K(d) = 110 nM) to a single class of sites (B(max) = 110 000/cell). Specific (125)I-PAMP binding was inhibited by PAMP (IC(50) of 100 nM) but not ADM, calcitonin gene-related peptide (CGRP), or amylin. Specific (125)I-ADM binding was inhibited with high affinity by ADM, CGRP, and CGRP(8-37) (IC(50) values of 10, 10, and 15 nM respectively) but not PAMP or amylin. ADM elevated cAMP (ED(50) value of 100 nM), whereas PAMP had no effect on basal cAMP but inhibited the increase in cAMP caused by 10 nM ADM. Also, the increase in cAMP caused by ADM was inhibited CGRP(8-37), suggesting that ADM is binding to CGRP receptors. ADM (100 nM) stimulated transiently c-fos mRNA, whereas PAMP (1000 nM) had little effect; however, PAMP inhibited the increase in c-fos mRNA caused by ADM. ADM stimulated [(3)H]thymidine uptake into PA1 cells, whereas PAMP inhibited the increase in thymidine uptake caused by ADM. These results indicate that ADM and PAMP are both biologically active in teratocarcinoma cells.     

Surgical Clipping versus Endovascular Intervention for the Treatment of Subarachnoid Hemorrhage Patients in New York State

Object

Randomized trials have demonstrated a survival benefit for endovascular treatment of ruptured cerebral aneurysms. We investigated the association of surgical clipping and endovascular coiling with outcomes in subarachnoid hemorrhage (SAH) patients in a real-world regional cohort.

Methods

We performed a cohort study involving patients with ruptured cerebral aneurysms, who underwent surgical clipping, or endovascular coiling from 2009–2013 and were registered in the Statewide Planning and Research Cooperative System (SPARCS) database. An instrumental variable analysis was used to investigate the association of treatment technique with outcomes.

Results

Of the 4,098 patients undergoing treatment, 2,585 (63.1%) underwent coiling, and 1,513 (36.9%) underwent clipping. Using an instrumental variable analysis, we did not identify a difference in inpatient mortality [marginal effect (ME), -0.56; 95% CI, -1.03 to 0.02], length of stay (LOS) (ME, 1.72; 95% CI, -3.39 to 6.84), or the rate of 30-day readmissions (ME, -0.30; 95% CI, -0.82 to 0.22) between the two treatment techniques for patients with SAH. Clipping was associated with a higher rate of discharge to rehabilitation (ME, 0.63; 95% CI, 0.24 to 1.01). In sensitivity analysis, mixed effect regression, and propensity score adjusted regression models demonstrated identical results.

Conclusions

Using a comprehensive all-payer cohort of patients in New York State presenting with aneurysmal SAH we did not identify an association of treatment method with mortality, LOS or 30-day readmission. Clipping was associated with a higher rate of discharge to rehabilitation.     

Effect of lipids on avian satellite cell proliferation, differentiation and heparan sulfate proteoglycan expression

The objective of this study was to determine the effects of fatty acids on the proliferation, differentiation, and expression of syndecan-4 and glypican-1 in avian myogenic satellite cells (SC). SC derived from the pectoralis major (PM) and biceps femoris (BF) muscles of the turkey and chicken were individually administered 8 different fatty acids in defined medium during proliferation. A parallel set of turkey SC was induced to differentiate. Highest levels of proliferation of turkey PM and BF SC occurred in cultures containing oleate. Linoleate and oleate were equipotent in supporting proliferation of chicken SC. Microscopic examination revealed that inclusion of docosahexaenoate or eicosapentaenoate was toxic towards both PM and BF SC from both species. Linolenate and arachidonate diminished levels of differentiation. Expression of glypican-1 varied between treatments to a greater extent with turkey BF than with PM SC. Expression in chicken PM and BF SC demonstrated a similar pattern in response to treatments. Turkey PM syndecan-4 expression varied between treatments, whereas expression in turkey BF SC was similar between treatments. Expression in chicken SC varied little between treatments. The results demonstrate species and muscle-specific differences in the parameters examined. It is proposed that changes in lipid raft receptor interactions may contribute to these observed differences.     

Camptothecin-somatostatin conjugates inhibit the growth of small cell lung cancer cells

The effects of camptothecin-somatostatin (CPT-SS) conjugates were investigated on small cell lung cancer (SCLC) cells. CPT was coupled to a SS agonist (SSA), c(Cys-Phe-DTrp-Lys-Thr-Cys)Thr-NH2 using the built in nucleophile assisted-releasing group (L1) N-methyl-aminoethyl-Gly-Dser-Nle-Dtyr-Dser or (L2) aminoethyl-Gly-Dser-Nle-Dtyr-Dser. The resulting CPT-L1-SSA and CPT-L2-SSA inhibited the specific binding of [125I-Tyr11]SS to NCI-H69 cell membranes with IC50 values of 0.2 and 2.1 nM, respectively. [125I]CPT-L1-SSA was internalized by SCLC cells at 37 degrees C but not at 4 degrees C. CPT-L1-SSA and CPT-L2-SSA inhibited in a dose-dependent manner the increase in adenylylcyclase activity caused by 25 microM forskolin. In vitro, 0.3 microM CPT-L1-SSA half-maximally inhibited the clonal growth of SCLC cells and 1 microM CPT-L1-SSA strongly inhibited 3H-thymidine incorporation into DNA and trypan-blue exclusion. These results suggest that CPT conjugated peptides such as CPT-L1-SSA may prove useful for exploring the efficacy of receptor-directed cytotoxicity to inhibit the proliferation of SCLC cells.     

Advanced glycation end products enhance expression of pro-apoptotic genes and stimulate fibroblast apoptosis through cytoplasmic and mitochondrial pathways

Both aging and diabetes are characterized by the formation of advanced glycation end products (AGEs). Both exhibit other similarities including deficits in wound healing that are associated with higher rates of fibroblast apoptosis. In order to investigate a potential mechanism for enhanced fibroblast apoptosis in diabetes and aged individuals, experiments were carried out to determine whether the predominant advanced glycation end product in skin, N-epsilon-(carboxymethyl) lysine (CML)-collagen, could induce fibroblast apoptosis. In vivo experiments established that CML-collagen but not unmodified collagen induced fibroblast apoptosis and that apoptosis was dependent upon caspase-3, -8, and -9 activity. In vitro experiments demonstrated that CML-collagen but not control collagen induced a time- and dose-dependent increase in fibroblast apoptosis. By use of blocking antibodies, apoptosis was shown to be mediated through receptor for AGE signaling. AGE-induced apoptosis was largely dependent on the effector caspase, caspase-3, which was activated through both cytoplasmic (caspase-8-dependent) and mitochondrial (caspase-9) pathways. CML-collagen had a global effect of enhancing mRNA levels of pro-apoptotic genes that included several classes of molecules including ligands, receptors, adaptor molecules, mitochondrial proteins, and others. However, the pattern of expression was not identical to the pattern of apoptotic genes induced by tumor necrosis factor alpha.     

Development of high affinity camptothecin-bombesin conjugates that have targeted cytotoxicity for bombesin receptor-containing tumor cells

Mammalian bombesin (BN) receptors are among those most frequently overexpressed by a number of common tumors including prostate, breast, lung, and colon cancers. The aim of this study was to develop a camptothecin-bombesin (CPT-BN) conjugate that interacts with all classes of BN receptors and possibly functions as a prodrug via a labile linker with site-specific cytotoxicity for cancer cells bearing these receptors. CPT was coupled to analogs of [D-Tyr6,beta-Ala11,Phe13,Nle14]BN-(6-14) (BA0) using carbamate linkers (L1 and L2) with built-in nucleophile-assisted releasing groups for intracellular cleavage of free cytotoxic agents. One conjugate, CPT-L2-BA3, bound to all three BN receptor classes with high affinity and functioned as a full agonist at each. 125I-CPT-L2-BA3 was rapidly internalized by cells expressing each BN receptor class and, using fluorescent imaging, was found to co-localize with BN receptors initially and later to be internalized in cytoplasmic compartments. HPLC analysis of internalized ligand showed that 40% was intact, 25% was metabolized by releasing free CPT, and 35% was metabolized to other breakdown products. CPT-L2-BA3 inhibited the growth of NCI-H1299 non-small cell lung cancer cells in 3-(4,5-dimethylthiazol-2-yl)-2.5-diphenyl-2H-tetrazolium bromide (MTT) and clonal growth assays. CPT-L2-BA3 was cytotoxic in an MTT assay for cells transfected with each class of BN receptor; however, it had significantly less effect in cells lacking BN receptors. These results indicate that CTP-L2-BA3 is a potent agonist that is cytotoxic for cells overexpressing any of the three BN receptor classes and functions as a prodrug for receptor-mediated cytoxicity. It therefore should be a useful prototype to explore the effectiveness of tumor-specific cytotoxicity delivery using a receptor-mediated mechanism.     

Deletion of ripA alleviates suppression of the inflammasome and MAPK by Francisella tularensis

Francisella tularensis is a facultative intracellular pathogen and potential biothreat agent. Evasion of the immune response contributes to the extraordinary virulence of this organism although the mechanism is unclear. Whereas wild-type strains induced low levels of cytokines, an F. tularensis ripA deletion mutant (LVSΔripA) provoked significant release of IL-1β, IL-18, and TNF-α by resting macrophages. IL-1β and IL-18 secretion was dependent on inflammasome components pyrin-caspase recruitment domain/apoptotic speck-containing protein with a caspase recruitment domain and caspase-1, and the TLR/IL-1R signaling molecule MyD88 was required for inflammatory cytokine synthesis. Complementation of LVSΔripA with a plasmid encoding ripA restored immune evasion. Similar findings were observed in a human monocytic line. The presence of ripA nearly eliminated activation of MAPKs including ERK1/2, JNK, and p38, and pharmacologic inhibitors of these three MAPKs reduced cytokine induction by LVSΔripA. Animals infected with LVSΔripA mounted a stronger IL-1β and TNF-α response than that of mice infected with wild-type live vaccine strain. This analysis revealed novel immune evasive mechanisms of F. tularensis.