Mechanisms of arginine vasopressin-induced insulin secretion in RINm5F cells

The present study was to investigate the mechanism by which AVP increases insulin secretion in RINm5F cells. A specific PLC inhibitor, U-73122, and a PLA2 inhibitor, N-(p-amylcinnamoyl)anthranilic acid (ACA) were used. U-73122 (2-8 [mu]M) inhibited the AVP-induced increases in the intracellular concentration of inositol 1,3,4-trisphosphate and (Ca2+]i dose-dependently. U-73122 (8 [mu]M) abolished the AVP\u27s effect on IP3 and (Ca2+]i, but it only reduced the AVP-induced increase in insulin secretion by 35%. The discrepancy between the results of (Ca2+]i and insulin secretion may be due to the multiple signal transduction pathways. ACA (100 [mu]M) didn\u27t antagonize the AVP-induced increase in insulin release. These results suggested: (1) U-73122 blocks PLC activities but fails to block other signal transduction pathways that trigger insulin secretion in these cells, and (2) AVP increases insulin release from RINm5F cells through both the PLC mediated Ca2+-dependent and Ca2+-independent pathways;We investigated the effect of the antifungal antibiotic wortmannin, a PLD inhibitor, on AVP-induced increases in insulin secretion and (Ca2+]i. Wortmannin (0.1-1 [mu]M) inhibited AVP-induced increase of insulin secretion dose-dependently. The combination of wortmannin and U-73122, had an additive inhibition on the AVP-induced insulin secretion. However, wortmannin failed to change the AVP-induced increase of (Ca2+]i. These results suggested: (1) activation of PLD stimulated at least partially the AVP-induced insulin secretion, (2) PLD didn\u27t influence the AVP-elicited elevation of (Ca2+]i and, (3) PLC and PLD stimulate AVP-activated signal transduction in an independent manner;Ro 31-8220, a specific PKC inhibitor, dose-dependently potentiated the AVP-induced increase of insulin secretion and elevation of (Ca2+]i. The potentiation of AVP-induced elevation of (Ca2+]i by Ro 31-8220 was mediated by an increase in Ca2+ influx. Treatment with a DAG analog, OAG, inhibited AVP-induced insulin dose-dependently, but reduced slightly effect the AVP-induced elevation of (Ca2+]i. Activation of PKC by OAG had no effect on insulin secretion and (Ca2+]i. These results suggested: (1) PKC is a negative regulator of AVP\u27s action in beta-cells, (2) PKC inhibits the Ca2+ channels to regulate insulin secretion

Mechanisms underlying Actinobacillus pleuropneumoniae exotoxin ApxI induced expression of IL-1β, IL-8 and TNF-α in porcine alveolar macrophages

Actinobacillus pleuropneumoniae (A. pleuropneumoniae) causes fibrino-hemorrhagic necrotizing pleuropneumonia in pigs. Production of proinflammatory mediators in the lungs is an important feature of A. pleuropneumoniae infection. However, bacterial components other than lipopolysaccharide involved in this process remain unidentified. The goals of this study were to determine the role of A. pleuropneumoniae exotoxin ApxI in cytokine induction and to delineate the underlying mechanisms. Using real-time quantitative PCR analysis, we found native ApxI stimulated porcine alveolar macrophages (PAMs) to transcribe mRNAs of IL-1β, IL-8 and TNF-α in a concentration- and time-dependent manner. Heat-inactivation or pre-incubation of ApxI with a neutralizing antiserum attenuated ApxI bioactivity to induce cytokine gene expression. The secretion of IL-1β, IL-8 and TNF-α protein from PAMs stimulated with ApxI was also confirmed by quantitative ELISA. In delineating the underlying signaling pathways contributing to cytokine expression, we observed mitogen-activated protein kinases (MAPKs) p38 and cJun NH2-terminal kinase (JNK) were activated upon ApxI stimulation. Administration of an inhibitor specific to p38 or JNK resulted in varying degrees of attenuation on ApxI-induced cytokine expression, suggesting the differential regulatory roles of p38 and JNK in IL-1β, IL-8 and TNF-α production. Further, pre-incubation of PAMs with a CD18-blocking antibody prior to ApxI stimulation significantly reduced the activation of p38 and JNK, and subsequent expression of IL-1β, IL-8 or TNF-α gene, indicating a pivotal role of β2 integrins in the ApxI-mediated effect. Collectively, this study demonstrated ApxI induces gene expression of IL-1β, IL-8 and TNF-α in PAMs that involves β2 integrins and downstream MAPKs

Mutations in the Salmonella enterica serovar Choleraesuis cAMP-receptor protein gene lead to functional defects in the SPI-1 Type III secretion system

Salmonella enterica serovar Choleraesuis (Salmonella Choleraesuis) causes a lethal systemic infection (salmonellosis) in swine. Live attenuated Salmonella Choleraesuis vaccines are effective in preventing the disease, and isolates of Salmonella Choleraesuis with mutations in the cAMP-receptor protein (CRP) gene (Salmonella Choleraesuis ∆crp) are the most widely used, although the basis of the attenuation remains unclear. The objective of this study was to determine if the attenuated phenotype of Salmonella Choleraesuis ∆crp was due to alterations in susceptibility to gastrointestinal factors such as pH and bile salts, ability to colonize or invade the intestine, or cytotoxicity for macrophages. Compared with the parental strain, the survival rate of Salmonella Choleraesuis ∆crp at low pH or in the presence of bile salts was higher, while the ability of the mutant to invade intestinal epithelia was significantly decreased. In examining the role of CRP on the secretory function of the Salmonella pathogenicity island 1 (SPI-1) encoded type III secretion system (T3SS), it was shown that Salmonella Choleraesuis ∆crp was unable to secrete the SPI-1 T3SS effector proteins, SopB and SipB, which play a role in Salmonella intestinal invasiveness and macrophage cytotoxicity, respectively. In addition, caspase-1 dependent cytotoxicity for macrophages was significantly reduced in Salmonella Choleraesuis ∆crp. Collectively, this study demonstrates that the CRP affects the secretory function of SPI-1 T3SS and the resulting ability to invade the host intestinal epithelium, which is a critical element in the pathogenesis of Salmonella Choleraesuis

Mechanisms of arginine vasopressin-induced insulin secretion in RINm5F cells

The present study was to investigate the mechanism by which AVP increases insulin secretion in RINm5F cells. A specific PLC inhibitor, U-73122, and a PLA2 inhibitor, N-(p-amylcinnamoyl)anthranilic acid (ACA) were used. U-73122 (2-8 [mu]M) inhibited the AVP-induced increases in the intracellular concentration of inositol 1,3,4-trisphosphate and (Ca2+]i dose-dependently. U-73122 (8 [mu]M) abolished the AVP's effect on IP3 and (Ca2+]i, but it only reduced the AVP-induced increase in insulin secretion by 35%. The discrepancy between the results of (Ca2+]i and insulin secretion may be due to the multiple signal transduction pathways. ACA (100 [mu]M) didn't antagonize the AVP-induced increase in insulin release. These results suggested: (1) U-73122 blocks PLC activities but fails to block other signal transduction pathways that trigger insulin secretion in these cells, and (2) AVP increases insulin release from RINm5F cells through both the PLC mediated Ca2+-dependent and Ca2+-independent pathways;We investigated the effect of the antifungal antibiotic wortmannin, a PLD inhibitor, on AVP-induced increases in insulin secretion and (Ca2+]i. Wortmannin (0.1-1 [mu]M) inhibited AVP-induced increase of insulin secretion dose-dependently. The combination of wortmannin and U-73122, had an additive inhibition on the AVP-induced insulin secretion. However, wortmannin failed to change the AVP-induced increase of (Ca2+]i. These results suggested: (1) activation of PLD stimulated at least partially the AVP-induced insulin secretion, (2) PLD didn't influence the AVP-elicited elevation of (Ca2+]i and, (3) PLC and PLD stimulate AVP-activated signal transduction in an independent manner;Ro 31-8220, a specific PKC inhibitor, dose-dependently potentiated the AVP-induced increase of insulin secretion and elevation of (Ca2+]i. The potentiation of AVP-induced elevation of (Ca2+]i by Ro 31-8220 was mediated by an increase in Ca2+ influx. Treatment with a DAG analog, OAG, inhibited AVP-induced insulin dose-dependently, but reduced slightly effect the AVP-induced elevation of (Ca2+]i. Activation of PKC by OAG had no effect on insulin secretion and (Ca2+]i. These results suggested: (1) PKC is a negative regulator of AVP's action in beta-cells, (2) PKC inhibits the Ca2+ channels to regulate insulin secretion.</p

Determination of the quality of stripe-marked and cracked eggs during storage

Objective Stripe marks, which occasionally occur on the shell, do not cause breakage to the shell and shell membranes of eggs. This study investigated the quality of intact eggs (IEs), minor stripe-marked eggs (MEs), severe stripe-marked eggs (SEs), and cracked eggs (CEs) during 3-week storage at 25°C. Methods Shell eggs were collected the day after being laid and were washed. Among them, eggs without any visual cracks or stripe marks on the shells were evaluated as IEs by the plant employees using candling in a darkened egg storage room; the remaining eggs exhibited some eggshell defects. At day 3, the eggs were further categorized into IEs, MEs, SEs, CEs, and broken eggs (BEs) on the basis of the description given. Except BEs, which were discarded, the remaining eggs were stored at 25°C (approximate relative humidity 50%) and then analyzed. Results Stripe marks were observed primarily within the first 3 days after washing. At day 3, CEs had significantly (p<0.05) lower Haugh unit values, but all eggs had grades AA or A, according to the United States Department of Agriculture standard. As storage time increased, differences in egg quality between groups were more obvious. IEs had the highest eggshell breaking strength. During storage, the total plate counts and pathogens, namely Escherichia coli, Campylobacter spp., Staphylococcus aureus, and Salmonella spp., were not detectable in the internal content of IEs and SEs. Conclusion In conclusion, cracks degraded egg quality severely and minor stripe marks only slightly influenced the egg quality

Gastrointestinal Mast Cell Tumor in an African Dormouse (<i>Graphiurus</i> sp.)

Mast cell tumors (MCTs) are well-known neoplasms derived from either mucosal or connective tissue mast cells. While well studied in several domestic species, MCTs are rarely documented in rodents. A three-year-old, male African dormouse (Graphiurus sp.) presented with a history of vomiting and anorexia for 3 months. Sonography revealed thickened gastric mucosa and hyperperistalsis. The patient died after receiving symptomatic treatment for 2 months. At necropsy, locally extensive, pale, thickened mucosal foci obscuring the first half of the stomach lumen was noted. Histological examination revealed moderately polymorphic, round, oval to spindle cells with amphophilic cytoplasmic granules infiltrating the mucosa to tunica muscularis, with moderate numbers of eosinophils. The mucosa was severely ulcerated with the proliferation of granulation tissue. The granules in most tumor cells exhibited metachromasia with the toluidine blue stain. Neoplastic cells revealed positive membranous immunoreactivity to KIT. Herein, we report the first case report of MCT in dormouse but also the first gastrointestinal MCT in a rodent species

Genotyping, Plasmid Analysis, and Antimicrobial Susceptibility of Salmonella enterica Serotype Enteritidis Isolates from Humans and Chickens in Central Taiwan

Salmonella enterica serotype Enteritidis (SE) is the most frequent etiological agent of human salmonellosis. The molecular epidemiology and antimicrobial susceptibility of human and chicken isolates of SE were examined. Methods: A total of 27 human and 40 chicken isolates of SE were collected in 2005–2006. We examined these isolates by antimicrobial susceptibility testing, pulsed-field gel electrophoresis (PFGE), and plasmid analysis. Results: Most isolates were susceptible to the seven antibiotics tested, except chicken isolates in 2005, which showed 70% resistance to streptomycin and 75% to tetracycline. There were six plasmid profiles identified among these isolates. Almost all isolates (97%) harbored the 60-kb serotype-specific virulence plasmid. PFGE using Xba I digestion separated human isolates into eight subtypes (1a–1h) and chicken isolates into four subtypes (1a–1c and 1g). In 2005, 1a and 1c were predominant for human isolates and 1a for chicken isolates. However, in 2006, 1a and 1c remained predominant for human isolates and 1b and 1c for chicken isolates. Most 1b and 1c isolates belonged to plasmid type 2 or 4. Correlation between plasmid patterns and PFGE subtypes was obtained between a 36-kb plasmid and 1b and between another 3.6-kb plasmid and 1a. Conclusion: Plasmid profiling and PFGE were efficient for discriminating SE isolates from different sources. Our data support the notion that SE is transmitted from chickens to humans, presumably through the food chain, but it appears that chickens are not the sole reservoir for human infection with SE in Taiwan. SE remained susceptible to most antimicrobial agents