Jonathan D Kaunitz - Academia.edu (original) (raw)
Papers by Jonathan D Kaunitz
Frontiers in Medicine
BackgroundIodine and particularly its oxidated forms have long been recognized for its effective ... more BackgroundIodine and particularly its oxidated forms have long been recognized for its effective antiseptic properties. Limited in vitro and in vivo data suggest that iodine exposure may rapidly inactivate, reduce transmission, and reduce infectivity of SARS-CoV-2. We hypothesized that iodine exposure may be associated with decreased incident COVID-19 infection.MethodsA retrospective population-level cohort analysis was performed of the U.S. Veterans Health Administration between 1 March 2020 and 31 December 2020, before the widespread availability of vaccines against SARS-CoV-2. Multivariable logistic regression models estimated the adjusted odds ratios (OR) and 95% confidence intervals (CI) of the associations between iodinated contrast exposure and incident COVID-19 infection, adjusting for age, sex, race/ethnicity, place of residence, socioeconomic status, and insurance status.Results530,942 COVID-19 tests from 333,841 Veterans (mean ± SD age, 62.7 ± 15.2 years; 90.2% men; 61.9%...
Physiology News, 2010
Action points Grants The Society offers funding through the following grant schemes: Travel Grant... more Action points Grants The Society offers funding through the following grant schemes: Travel Grants, Non-Society Symposia Grants, Outreach Grants, International Teaching and Research Grants and the Vacation Studentship and Departmental Seminar Schemes. For full information, please visit: www.physoc.org/grants Membership applications Applications for membership to The Physiological Society are considered on a rolling basis, and a decision is normally made within 15 working days. For full information, please visit: www.physoc.org/membership Is your membership information correct? Please check and update your details at www.physoc.org, under 'My Physoc Profile'.
Journal of Clinical Investigation, 2021
Gastroenterology, 2018
Background: Previously we reported that several microRNAs (miRNAs), derived from peripheral blood... more Background: Previously we reported that several microRNAs (miRNAs), derived from peripheral blood mononuclear cells (PBMCs) in patients, were upregulated in colonic Crohn's disease (CD) relative to ulcerative colitis (UC). Several of these miRNAs are bioinformatically predicted to deregulate the autophagy pathway, a pathway known to be important in inflammatory bowel disease (IBD). The most overexpressed miRNA, miR-874-3p, is predicted to target the CD risk gene and key autophagy gene ATG16L1. This study aims to validate and evaluate miR-874-3p regulation of autophagy by targeting ATG16L1 in vitro. Methods: To validate that miR-874-3p targets ATG16L1 we conducted a Dual Luciferase Reporter (DLR) assay. HeLa cells were co-transfected with a 3'UTR reporter vector containing ATG16L1 and either a miR874-3p mimic miR874-3p inhibitor, a scrambled negative control miRNA, or no miRNA. To investigate the effect of miR-874-3p overexpression on autophagy, we performed immunofluorescence (IF) of microtubule-associated protein 1A/1B-light chain 3 (LC3) puncta in rapamycin treated (autophagy induced) and untreated HeLa cells. Transfected HeLa cells were treated with Rapamycin (100 ng/mL) for 4 hours or remained untreated. The number of puncta per cell was counted from 25 cells in triplicate from each condition. A 2-way ANOVA and Bonferroni post hoc test compared means within and between treatment groups. Results: One way ANOVA and Bonferroni post hoc test compared means among treatments in the DLR assay and showed miR-874-3p mimic significantly reduced ATG16L1 expression (p=0.0005). The miR-874-3p mimic significantly reduced the average number of LC3 puncta in the rapamycin group compared to inhibitor (p= 0.001) and negative control (p=0.006, Figure 1)). There was no significant difference between treatments in the untreated control group suggesting increased miR-874-3p expression inhibits autophagy. Quantitative PCR (qPCR) confirmed that mRNA transcripts of ATG16L1 were affected by miR874-3p overexpression. There was significantly fewer ATG16L1 transcripts with transfection of miR874-3p mimic compared to the inhibitor and negative control in untreated and rapamycin induced cells suggesting miR874-3p reduces transcription of ATG16L1 (Table 1). This was further confirmed in blood samples from 6 IBD patients (5 UC and 1 CD) and 1 healthy control, where higher levels of miR874-3p were associated with lower levels of ATG16L1 mRNA. Conclusions: We demonstrate the targeting of ATG16L1 by miR874-3p in vitro and the functional effect of this target interaction on autophagy. Confirmation of the inverse expression between miR874-3p and ATG16L1 was observed in the blood of a small cohort of IBD patients. Further experiments with an increased sample size of IBD patients are required to confirm the relationship between miR874-3p levels and disease subtype.
Nihon Shokakibyo Gakkai zasshi = The Japanese journal of gastro-enterology, 2018
The FASEB Journal, 2020
Although luminal short‐chain (SCFA) and long‐chain fatty acids (LCFA) stimulate mucosal protectiv... more Although luminal short‐chain (SCFA) and long‐chain fatty acids (LCFA) stimulate mucosal protective duodenal bicarbonate secretion (DBS), the role of medium‐chain fatty acids (MCFA) in DBS has not been studied. Thus, we examined the effects of MCFA on DBS in vivo and short‐circuit current (Isc) in the Ussing chamber.MCFA (C6‐C12, 0.1 – 30 mM) were applied to the mucosal or serosal bath of the Ussing chamber mounted with the muscle‐stripped mucosa‐submucosa preparations of rat proximal duodenum. A duodenal loop was perfused with sodium caprate (C10, 1 mM) in anesthetized rats with total CO2 output measured.Among MCFA, sodium caprate (C10) dose‐dependently (1 – 30 mM luminally and 0.1 – 3 mM serosally) increased Isc, which was sustained > 10 min after reaching a plateau, whereas other MCFA (C6, C8, and C12) had little effect. The luminal caprate (10 mM)‐induced Isc increase was delayed by luminal addition of ibuprofen (1 mM), a sodium‐dependent monocarboxylate transporter‐1 (SMCT‐1)...
The FASEB Journal, 2020
Lipopolysaccharides (LPS) entry from the intestinal lumen to the circulation is associated with s... more Lipopolysaccharides (LPS) entry from the intestinal lumen to the circulation is associated with systemic inflammation. We have found that LPS is transcellularly transported to portal vein during long‐chain fatty acid (LCFA) exposure via CD36‐ and lipid raft‐mediated pathways. We examined the effects of medium‐chain fatty acids (MCFA) on LPS transport in rat jejunum.FITC‐LPS was applied to the mucosal bath of Ussing chambered muscle‐stripped jejunal and distal colonic mucosa‐submucosa preparations, in which short‐circuit current (Isc) and tissue electrical resistance (TER) were measured. Serosal appearance of FITC‐LPS was measured with or without luminal application of sodium caprate (C10, 10 mM). FITC‐dextran 4kDa (FD4) m‐to‐s movement was also separately measured.Luminal application of caprate increased FITC‐LPS m‐to‐s transport without any changes in FD4 m‐to‐s movement and TER in the jejunum, whereas caprate remarkably increased FITC‐LPS transport with increased FD4 movement and ...
Gastroenterology, 2019
Methods: An acute model of pancreatitis was induced in 6-7 week-old B6129 (WT) and IL-4Rα-/mice b... more Methods: An acute model of pancreatitis was induced in 6-7 week-old B6129 (WT) and IL-4Rα-/mice by eight hourly injections of cerulein (50 mg/kg) every other day for 7 days. Pancreata were harvested 1, 4, 8, and 15 days after injury. Immunohistochemical analyses were performed employing antibodies specific to ductal cells (cytokeratin 19) and a marker of proliferation (Ki67). Proliferative index quantified as the proportion of ductal cells that stained for Ki67. A two-tailed student t-test was used to determine significance (p= 0.05). Results: Non-injured mice exhibited a relatively low proliferative index of 16.1% in the common channel, and 19.3% in the intrapancreatic biliary duct. in response to pancreatitis, ductal epithelial proliferation in the common channel increased to 23.8% by day 1, and 33.2% (p = 0.018) by day 4 post-injury. Proliferation remained elevated until day 15. Similarly, proliferation within the intrapancreatic biliary duct increased over time, although with delayed kinetics compared to the common channel. Proliferation remained unchanged at 18.5% on day 1 post-injury but increased to 30.4% by day 4 and 35.4% (p = 0.007) by day 15. To investigate whether IL-4Rα contributes to proliferation in the intrapancreatic biliary ductal system, we employed mice deficient in this receptor (IL-4Rα-/-). Similar to WT mice, non-injured IL-4Rα-/mice exhibited a proliferation index of 10.6% in the common channel and 11.8% in the intrapancreatic biliary duct. However, in contrast, a dramatic reduction in epithelial proliferation was observed upon injury. The proliferative index of the common channel decreased by day 1 post-injury (6.8%, p = 0.041), continued to decrease until day 4 (3.5%, p = 0.002), and maintained a significant reduction in proliferation through day 15. The proliferative index of the intrapancreatic biliary duct exhibited a stepwise decrease as well, decreasing to 4.3% by day 4 (p = 0.017) which was maintained through day 15 post-injury. Conclusion: Increased epithelial proliferation of the intrapancreatic ductal system of WT mice reveals the regenerative response of the epithelium after injury. Significantly decreased proliferation in mice lacking the IL-4Rα suggests an important role for cytokine signaling in maintaining the protective epithelial barrier of the intrapancreatic ductal system after pancreatitis.
Gastroenterology, 2019
or RAW 264.7 macrophages were infected or not with Hp PMSS1 for 24h ± aminooxy-acetic acid (AOAA)... more or RAW 264.7 macrophages were infected or not with Hp PMSS1 for 24h ± aminooxy-acetic acid (AOAA), a CTH inhibitor. Polyamine levels were measured by LC/MS. Metabolomics was performed by LC/MS/MS and analyzed using IMPaLA. Results: CTH expression was increased and localized to gastric CD68 + cells in Hp-infected WT mice compared to uninfected. There was no expression of CTH in Cth-/mice. In WT vs Cth-/mice, gastritis severity was reduced from 3.8±0.4 to 2.2±0.3 (p<0.05) with PMSS1, and from 3.0±0.9 to 0.5±0.2 (p<0.01) with SS1. The upregulation of genes encoding the M1 macrophage marker NO synthase 2 (NOS2), the prototype Th1 (IFN-γ) and Th17 (IL-17A) cytokines, and the Mreg/Treg cytokine IL-10 that occurred in infected WT mice was completely eliminated in Cth-/mice. Consistent with the attenuated immune response, Hp colonization was increased by a log order (p<0.05) in Cth-/mice with both strains. We found that CTH activity in Gmacs alters polyamine synthesis, as deletion of Cth resulted in a 25% decrease in Put and a 22% increase in Spd in infected mice. Similarly, in Bmacs or RAW 264.7 cells infected with Hp, AOAA reduced Put and increased Spd. This inhibitor also increased dcSAM in Hp-infected macrophages, indicating that blocking CTH re-establishes the metabolism of SAM through the Spd pathway. Additionally, CTH inhibition in infected macrophages modulated biochemical pathways involving polyamines, as well as methionine and cysteine, as determined by a metabolomics approach. Conclusions: Increased activity of the RTP in Gmacs regulates immunometabolism and contributes to Hp-induced inflammation. Thus, CTH represents a new therapeutic target to limit gastritis and potentially, the related risk for carcinogenesis.
Gastroenterology, 2020
Background and Aims: Although cAMP signaling is well known to regulate epithelial ion transports,... more Background and Aims: Although cAMP signaling is well known to regulate epithelial ion transports, the role of Ca 2+ signaling in cAMP-mediated ion secretion is poorly understood. We aimed to investigate the role Ca 2+ signaling in cAMP-mediated intestinal epithelial ion secretion and the cross-talk mechanisms between these two signaling. Methods: Functional, biochemical and immunofluoresence techniques were performed to examine anion secretion and protein expression and localization in mouse duodenal epithelium. Results: Activation of adenaly cyclase by foskolin (10 uM) to raise intracellular cAMP or direct application of dibuty-cAMP (300 uM), a well-known cell-permeable cAMP analogy, stimulated mouse duodenal I sc when added to serosal side but not mucosal side (p<0.001, n=7). Surprisingly, cAMP-mediated intestinal I sc was significantly reduced by external Ca 2+ omission at serosal side of the tissues (p<0.01, n=6), and was inhibited by several selective blockers of storeoperated Ca 2+ channels (SOC) and CRAC/Orai channels at serosal side but not at the mucosal side (p<0.01, n=6). The ER Ca 2+ chelation by TPEN could also significantly reduced cAMPmediated intestinal I sc (p<0.001, n=6). cAMP-mediated intestinal I sc was promoted by activator of ryanodine receptor on the ER (1 mM caffeine, p<0.01, n=6), but reduced by its blocker (300 uM dantrolene, p<0.01, n=6). Furthermore, cAMP-mediated intestinal I sc was reduced by serosal addition but not mucosal addition of H-89 (20 uM, p<0.001, n=6), a PKA inhibitor. Finally, STIM1/Orai1 proteins, the molecular components of SOC, were identified on serosal side of mouse duodenal epithelium. Conclusion: Forskolin/cAMP induces intestinal ion secretion in a polarized manner. It may trigger SOC via PKA phosphrylation of ryanodine receptor to deplete the ER Ca 2+ store and cause Ca 2+ entry via STIM1/ Orai1, which plays a critical role in cAMP-mediated intestinal ion secretion. Our findings provide a new insight to a cross-talk mechanism of cAMP and Ca 2+ signaling in epithelial ion secretion.
Gastroenterology, 2020
in intestinal epithelial cells and in mouse intestine via a transcriptional mechanism. We specula... more in intestinal epithelial cells and in mouse intestine via a transcriptional mechanism. We speculate that CP-induced decrease in PAT-1 function and expression may contribute to a higher occurrence of nephrolithiasis in immunocompromised patients. (Supported by NIDDK and Department of Veterans Affairs) 582 TEDUGLUTIDE INHIBITS SODIUM CAPRATE-INDUCED CLATHRIN-MEDIATED LPS TRANSPORT IN RAT JEJUNUM Yasutada Akiba, Jonathan D. Kaunitz Lipopolysaccharides (LPS) entry from the intestinal lumen to the circulation is associated with systemic inflammation. We have found that LPS is transcellularly transported to the portal vein (PV) during luminal perfusion with long-chain fatty acid (LCFA) via CD36-and lipid raft-mediated pathways, which are inhibited by and the stable glucagon-like peptide-2 (GLP-2) analog teduglutide (TDG). We examined the effects of medium-chain fatty acids (MCFA) on LPS transport in rat jejunum. FITC-LPS was applied to the mucosal bath of Ussing chambered muscle-stripped jejunal and distal colonic mucosa-submucosa preparations, in which short-circuit current (I sc) and tissue electrical resistance (TER) were measured. Serosal appearance of FITC-LPS was measured with or without luminal application of sodium caprate (C10, 10 mM). FITC-dextran 4kDa (FD4) m-to-s movement was also separately measured. The effects of serosal GLP-2 (100 nM) with a dipeptidyl peptidase 4 inhibitor NVP728 (10 µM) or TDG (10 µg/ml) on C10-mediated FITC-LPS transport were also examined. Luminal application of C10 increased FITC-LPS m-to-s transport without any changes in FD4 m-to-s movement and TER in the jejunum, whereas C10 increased FITC-LPS transport with increased FD4 movement and decreased TER in the distal colon, suggesting that C10 enhances transcellular LPS transport in the jejunum, but increases paracellular permeability in the distal colon. In the jejunum, C10-mediated FITC-LPS m-to-s transport was inhibited by pretreatment with luminal Pitstop2 (30 µM), a selective inhibitor of clathrinmediated endocytosis (CME), whereas the CD36 inhibitor sulfosuccinimidyl oleate (SSO, 0.1 mM), the lipid raft inhibitor methyl-b-cyclodextrin (MbCD, 1 mM), or the dynamin inhibitor Dynasore (50 µM) had no effect. In contrast, luminal OA (30 mM) with taurocholate (TCA, 0.1 mM) also increased FITC-LPS transport with SSO, MbCD and Dynasore-sensitive, but Pitstop2-insensitive manner. Serosal application of GLP-2 with NVP728 or TDG abolished C10-mediated FITC-LPS transport. Furthermore, serosal pretreatment of VIP (30 nM) or a nitric oxide donor (S)-Nitroso-N-acetylpenicillamine SNAP (0.1 mM) reduced C10-mediated FITC-LPS transport, suggesting that GLP-2-induced reduction of LPS transport is mediated via VIP and NO pathways. In vivo, jejunal luminal perfusion of C10 (10 mM) with FITC-LPS increased PV FITC-LPS appearance 15 min after perfusion, which was inhibited by coperfusion of Pitstop2. These results suggest that luminal MCFA and LCFA differentially induce transcellular LPS transport in the jejunum; MCFA via CME, but LCFA via the CD36/ lipid raft/caveolin pathway. GLP-2 and TDG inhibited MCFA-induced LPS transport possibly via VIP and NO pathways. Dietary fatty acids may exacerbate LPS-associated diseases, including the metabolic syndrome and multiple organ failure, that can be treated with TDG.
F1000Research, 2019
Vasoactive intestinal peptide (VIP), a gut peptide hormone originally reported as a vasodilator i... more Vasoactive intestinal peptide (VIP), a gut peptide hormone originally reported as a vasodilator in 1970, has multiple physiological and pathological effects on development, growth, and the control of neuronal, epithelial, and endocrine cell functions that in turn regulate ion secretion, nutrient absorption, gut motility, glycemic control, carcinogenesis, immune responses, and circadian rhythms. Genetic ablation of this peptide and its receptors in mice also provides new insights into the contribution of VIP towards physiological signaling and the pathogenesis of related diseases. Here, we discuss the impact of VIP on gastrointestinal function and diseases based on recent findings, also providing insight into its possible therapeutic application to diabetes, autoimmune diseases and cancer.
The Journal of Infectious Diseases, 2019
BackgroundCholera toxin (CT)–induced diarrhea is mediated by cyclic adenosine monophosphate (cAMP... more BackgroundCholera toxin (CT)–induced diarrhea is mediated by cyclic adenosine monophosphate (cAMP)–mediated active Cl– secretion via the cystic fibrosis transmembrane conductance regulator (CFTR). Although the constitutive activation of adenylyl cyclase (AC) in response to CT is due to adenosine diphosphate ribosylation of the small G protein α-subunit activating CFTR with consequent secretory diarrhea, the AC isoform(s) involved remain unknown.MethodsWe generated intestinal epithelial cell–specific adenylyl cyclase 6 (AC6) knockout mice to study its role in CT-induced diarrhea.ResultsAC6 messenger RNA levels were the highest of all 9 membrane-bound AC isoforms in mouse intestinal epithelial cells. Intestinal epithelial-specific AC6 knockout mice (AC6loxloxVillinCre) had undetectable AC6 levels in small intestinal and colonic epithelial cells. No significant differences in fluid and food intake, plasma electrolytes, intestinal/colon anatomy and morphology, or fecal water content wer...
British Journal of Pharmacology, 2015
The bioactive monoamine 5-HT, implicated in the pathogenesis of functional gastrointestinal disor... more The bioactive monoamine 5-HT, implicated in the pathogenesis of functional gastrointestinal disorders, is abundantly synthesized and stored in rat proximal colonic mucosa and released to the gut lumen and subepithelial space. Despite much data regarding its expression and function, the effects of luminal 5-HT on colonic anion secretion have not been fully investigated. EXPERIMENTAL APPROACH We measured short-circuit current (Isc) as an indicator of ion transport in mucosa-submucosa or mucosa-only preparations of rat proximal colon. Total CO2 output was measured in vitro and in vivo. Immunohistochemistry was performed to investigate the localization of 5-HT4, NOS1 and NOS2. KEY RESULTS Luminal 5-HT gradually increased the amplitude and sustained the elevation of Isc. Luminal 5-HT-evoked ΔIsc was acetazolamide sensitive and HCO3 − dependent, consistent with cytosolic carbonic anhydrase-dependent electrogenic HCO3 − secretion, while not affected by tetrodotoxin (TTX), atropine or indomethacin. Pretreatment with the selective 5-HT4 antagonist GR113808, but not antagonists for 5-HT3, 5-HT6 or 5-HT7, inhibited luminal 5-HT-evoked ΔIsc. Furthermore, luminal cisapride and tegaserod increased Isc to the same extent as did 5-HT in the presence of indomethacin and TTX. Removal of the submucosa or pretreatment with NOS inhibitors enhanced luminal 5-HT-evoked ΔIsc, suggesting that NO synthesized in the submucosa suppresses mucosal anion secretion. NOS1 and NOS2 were immunostained in the submucosal neurons and glial cells respectively. Luminal 5-HT-evoked HCO3 − secretion was confirmed in vivo, inhibited by co-perfusion of GR113808, but not by ondansetron. CONCLUSIONS AND IMPLICATIONS A novel apical 5-HT4-mediated HCO3 − secretory pathway and an NO-dependent inhibitory mechanism are present in the proximal colon. Luminal 5-HT-evoked HCO3 − secretion may be important for the maintenance of mucosal integrity by regulating luminal pH.
Gastroenterology, 2017
Background: Parkinson's disease (PD) is characterized by alpha-synucleinopathy at all levels of t... more Background: Parkinson's disease (PD) is characterized by alpha-synucleinopathy at all levels of the brain-gut axis including the enteric nervous system (ENS). Lesions in the ENS may be associated with gut inflammation, increased intestinal permeability and dysmotility contributing to the pathogenesis of PD and gastrointestinal manifestations. Aims: To evaluate fecal calprotectin and zonulin as biomarkers of gut inflammation and intestinal barrier dysfunction, respectively, in patients with PD. Methods: Ten consecutive patients with PD (mean age 65; 5 F, 5 M) and 10 healthy controls (mean age 53; 8 F, 2 M) participated in the study. The quantitative evaluation of calprotectin and zonulin in stool samples was performed by ready-to-use sandwich ELISA tests. Additionally, patients filled out a short questionnaire concerning gastrointestinal symptoms. The Kruskal-Wallis test was used for the comparison of differences between the groups. Results: A mean duration of PD in the studied group was 8 years (range 3-20 years). None of the patients was diagnosed with inflammatory bowel disease. Patients with PD reported constipation (40%), alternating bowel movement pattern (40%), and normal bowel habit (20%), in addition, abdominal pain (20%) and distension (50%). The increased levels of calprotectin and zonulin in stool samples were found in 60% and 40% of PD patients, respectively. Fecal calprotectin level was within the normal range in all the controls, while zonulin level was slightly elevated only in one out of 10 control subjects. The biomarker levels are presented as median and the lower quartile (25Q)-upper quartile (75Q). The fecal calprotectin level (µg/g) was significantly higher in PD patients compared to the controls 134.6 (89.1-145.5) vs 24.8 (18.0-27.9), p= 0.0025. The fecal zonulin level (ng/ml) was also higher in PD patients compared to the controls, but the p-value did not reach statistical significance: 105.5 (63.5-166.0) vs 54.5 (44.0-73.0), p=0.0588. Conclusions: Gastrointestinal symptoms are frequently reported by PD patients. The signs of gut inflammation and increased intestinal permeability are present in a remarkable number of PD patients. This is the first study using stool ELISA tests which confirmed the gut immune system activation and changes in the intestinal epithelial barrier integrity observed in PD patients.
Frontiers in Medicine
BackgroundIodine and particularly its oxidated forms have long been recognized for its effective ... more BackgroundIodine and particularly its oxidated forms have long been recognized for its effective antiseptic properties. Limited in vitro and in vivo data suggest that iodine exposure may rapidly inactivate, reduce transmission, and reduce infectivity of SARS-CoV-2. We hypothesized that iodine exposure may be associated with decreased incident COVID-19 infection.MethodsA retrospective population-level cohort analysis was performed of the U.S. Veterans Health Administration between 1 March 2020 and 31 December 2020, before the widespread availability of vaccines against SARS-CoV-2. Multivariable logistic regression models estimated the adjusted odds ratios (OR) and 95% confidence intervals (CI) of the associations between iodinated contrast exposure and incident COVID-19 infection, adjusting for age, sex, race/ethnicity, place of residence, socioeconomic status, and insurance status.Results530,942 COVID-19 tests from 333,841 Veterans (mean ± SD age, 62.7 ± 15.2 years; 90.2% men; 61.9%...
Physiology News, 2010
Action points Grants The Society offers funding through the following grant schemes: Travel Grant... more Action points Grants The Society offers funding through the following grant schemes: Travel Grants, Non-Society Symposia Grants, Outreach Grants, International Teaching and Research Grants and the Vacation Studentship and Departmental Seminar Schemes. For full information, please visit: www.physoc.org/grants Membership applications Applications for membership to The Physiological Society are considered on a rolling basis, and a decision is normally made within 15 working days. For full information, please visit: www.physoc.org/membership Is your membership information correct? Please check and update your details at www.physoc.org, under 'My Physoc Profile'.
Journal of Clinical Investigation, 2021
Gastroenterology, 2018
Background: Previously we reported that several microRNAs (miRNAs), derived from peripheral blood... more Background: Previously we reported that several microRNAs (miRNAs), derived from peripheral blood mononuclear cells (PBMCs) in patients, were upregulated in colonic Crohn's disease (CD) relative to ulcerative colitis (UC). Several of these miRNAs are bioinformatically predicted to deregulate the autophagy pathway, a pathway known to be important in inflammatory bowel disease (IBD). The most overexpressed miRNA, miR-874-3p, is predicted to target the CD risk gene and key autophagy gene ATG16L1. This study aims to validate and evaluate miR-874-3p regulation of autophagy by targeting ATG16L1 in vitro. Methods: To validate that miR-874-3p targets ATG16L1 we conducted a Dual Luciferase Reporter (DLR) assay. HeLa cells were co-transfected with a 3'UTR reporter vector containing ATG16L1 and either a miR874-3p mimic miR874-3p inhibitor, a scrambled negative control miRNA, or no miRNA. To investigate the effect of miR-874-3p overexpression on autophagy, we performed immunofluorescence (IF) of microtubule-associated protein 1A/1B-light chain 3 (LC3) puncta in rapamycin treated (autophagy induced) and untreated HeLa cells. Transfected HeLa cells were treated with Rapamycin (100 ng/mL) for 4 hours or remained untreated. The number of puncta per cell was counted from 25 cells in triplicate from each condition. A 2-way ANOVA and Bonferroni post hoc test compared means within and between treatment groups. Results: One way ANOVA and Bonferroni post hoc test compared means among treatments in the DLR assay and showed miR-874-3p mimic significantly reduced ATG16L1 expression (p=0.0005). The miR-874-3p mimic significantly reduced the average number of LC3 puncta in the rapamycin group compared to inhibitor (p= 0.001) and negative control (p=0.006, Figure 1)). There was no significant difference between treatments in the untreated control group suggesting increased miR-874-3p expression inhibits autophagy. Quantitative PCR (qPCR) confirmed that mRNA transcripts of ATG16L1 were affected by miR874-3p overexpression. There was significantly fewer ATG16L1 transcripts with transfection of miR874-3p mimic compared to the inhibitor and negative control in untreated and rapamycin induced cells suggesting miR874-3p reduces transcription of ATG16L1 (Table 1). This was further confirmed in blood samples from 6 IBD patients (5 UC and 1 CD) and 1 healthy control, where higher levels of miR874-3p were associated with lower levels of ATG16L1 mRNA. Conclusions: We demonstrate the targeting of ATG16L1 by miR874-3p in vitro and the functional effect of this target interaction on autophagy. Confirmation of the inverse expression between miR874-3p and ATG16L1 was observed in the blood of a small cohort of IBD patients. Further experiments with an increased sample size of IBD patients are required to confirm the relationship between miR874-3p levels and disease subtype.
Nihon Shokakibyo Gakkai zasshi = The Japanese journal of gastro-enterology, 2018
The FASEB Journal, 2020
Although luminal short‐chain (SCFA) and long‐chain fatty acids (LCFA) stimulate mucosal protectiv... more Although luminal short‐chain (SCFA) and long‐chain fatty acids (LCFA) stimulate mucosal protective duodenal bicarbonate secretion (DBS), the role of medium‐chain fatty acids (MCFA) in DBS has not been studied. Thus, we examined the effects of MCFA on DBS in vivo and short‐circuit current (Isc) in the Ussing chamber.MCFA (C6‐C12, 0.1 – 30 mM) were applied to the mucosal or serosal bath of the Ussing chamber mounted with the muscle‐stripped mucosa‐submucosa preparations of rat proximal duodenum. A duodenal loop was perfused with sodium caprate (C10, 1 mM) in anesthetized rats with total CO2 output measured.Among MCFA, sodium caprate (C10) dose‐dependently (1 – 30 mM luminally and 0.1 – 3 mM serosally) increased Isc, which was sustained > 10 min after reaching a plateau, whereas other MCFA (C6, C8, and C12) had little effect. The luminal caprate (10 mM)‐induced Isc increase was delayed by luminal addition of ibuprofen (1 mM), a sodium‐dependent monocarboxylate transporter‐1 (SMCT‐1)...
The FASEB Journal, 2020
Lipopolysaccharides (LPS) entry from the intestinal lumen to the circulation is associated with s... more Lipopolysaccharides (LPS) entry from the intestinal lumen to the circulation is associated with systemic inflammation. We have found that LPS is transcellularly transported to portal vein during long‐chain fatty acid (LCFA) exposure via CD36‐ and lipid raft‐mediated pathways. We examined the effects of medium‐chain fatty acids (MCFA) on LPS transport in rat jejunum.FITC‐LPS was applied to the mucosal bath of Ussing chambered muscle‐stripped jejunal and distal colonic mucosa‐submucosa preparations, in which short‐circuit current (Isc) and tissue electrical resistance (TER) were measured. Serosal appearance of FITC‐LPS was measured with or without luminal application of sodium caprate (C10, 10 mM). FITC‐dextran 4kDa (FD4) m‐to‐s movement was also separately measured.Luminal application of caprate increased FITC‐LPS m‐to‐s transport without any changes in FD4 m‐to‐s movement and TER in the jejunum, whereas caprate remarkably increased FITC‐LPS transport with increased FD4 movement and ...
Gastroenterology, 2019
Methods: An acute model of pancreatitis was induced in 6-7 week-old B6129 (WT) and IL-4Rα-/mice b... more Methods: An acute model of pancreatitis was induced in 6-7 week-old B6129 (WT) and IL-4Rα-/mice by eight hourly injections of cerulein (50 mg/kg) every other day for 7 days. Pancreata were harvested 1, 4, 8, and 15 days after injury. Immunohistochemical analyses were performed employing antibodies specific to ductal cells (cytokeratin 19) and a marker of proliferation (Ki67). Proliferative index quantified as the proportion of ductal cells that stained for Ki67. A two-tailed student t-test was used to determine significance (p= 0.05). Results: Non-injured mice exhibited a relatively low proliferative index of 16.1% in the common channel, and 19.3% in the intrapancreatic biliary duct. in response to pancreatitis, ductal epithelial proliferation in the common channel increased to 23.8% by day 1, and 33.2% (p = 0.018) by day 4 post-injury. Proliferation remained elevated until day 15. Similarly, proliferation within the intrapancreatic biliary duct increased over time, although with delayed kinetics compared to the common channel. Proliferation remained unchanged at 18.5% on day 1 post-injury but increased to 30.4% by day 4 and 35.4% (p = 0.007) by day 15. To investigate whether IL-4Rα contributes to proliferation in the intrapancreatic biliary ductal system, we employed mice deficient in this receptor (IL-4Rα-/-). Similar to WT mice, non-injured IL-4Rα-/mice exhibited a proliferation index of 10.6% in the common channel and 11.8% in the intrapancreatic biliary duct. However, in contrast, a dramatic reduction in epithelial proliferation was observed upon injury. The proliferative index of the common channel decreased by day 1 post-injury (6.8%, p = 0.041), continued to decrease until day 4 (3.5%, p = 0.002), and maintained a significant reduction in proliferation through day 15. The proliferative index of the intrapancreatic biliary duct exhibited a stepwise decrease as well, decreasing to 4.3% by day 4 (p = 0.017) which was maintained through day 15 post-injury. Conclusion: Increased epithelial proliferation of the intrapancreatic ductal system of WT mice reveals the regenerative response of the epithelium after injury. Significantly decreased proliferation in mice lacking the IL-4Rα suggests an important role for cytokine signaling in maintaining the protective epithelial barrier of the intrapancreatic ductal system after pancreatitis.
Gastroenterology, 2019
or RAW 264.7 macrophages were infected or not with Hp PMSS1 for 24h ± aminooxy-acetic acid (AOAA)... more or RAW 264.7 macrophages were infected or not with Hp PMSS1 for 24h ± aminooxy-acetic acid (AOAA), a CTH inhibitor. Polyamine levels were measured by LC/MS. Metabolomics was performed by LC/MS/MS and analyzed using IMPaLA. Results: CTH expression was increased and localized to gastric CD68 + cells in Hp-infected WT mice compared to uninfected. There was no expression of CTH in Cth-/mice. In WT vs Cth-/mice, gastritis severity was reduced from 3.8±0.4 to 2.2±0.3 (p<0.05) with PMSS1, and from 3.0±0.9 to 0.5±0.2 (p<0.01) with SS1. The upregulation of genes encoding the M1 macrophage marker NO synthase 2 (NOS2), the prototype Th1 (IFN-γ) and Th17 (IL-17A) cytokines, and the Mreg/Treg cytokine IL-10 that occurred in infected WT mice was completely eliminated in Cth-/mice. Consistent with the attenuated immune response, Hp colonization was increased by a log order (p<0.05) in Cth-/mice with both strains. We found that CTH activity in Gmacs alters polyamine synthesis, as deletion of Cth resulted in a 25% decrease in Put and a 22% increase in Spd in infected mice. Similarly, in Bmacs or RAW 264.7 cells infected with Hp, AOAA reduced Put and increased Spd. This inhibitor also increased dcSAM in Hp-infected macrophages, indicating that blocking CTH re-establishes the metabolism of SAM through the Spd pathway. Additionally, CTH inhibition in infected macrophages modulated biochemical pathways involving polyamines, as well as methionine and cysteine, as determined by a metabolomics approach. Conclusions: Increased activity of the RTP in Gmacs regulates immunometabolism and contributes to Hp-induced inflammation. Thus, CTH represents a new therapeutic target to limit gastritis and potentially, the related risk for carcinogenesis.
Gastroenterology, 2020
Background and Aims: Although cAMP signaling is well known to regulate epithelial ion transports,... more Background and Aims: Although cAMP signaling is well known to regulate epithelial ion transports, the role of Ca 2+ signaling in cAMP-mediated ion secretion is poorly understood. We aimed to investigate the role Ca 2+ signaling in cAMP-mediated intestinal epithelial ion secretion and the cross-talk mechanisms between these two signaling. Methods: Functional, biochemical and immunofluoresence techniques were performed to examine anion secretion and protein expression and localization in mouse duodenal epithelium. Results: Activation of adenaly cyclase by foskolin (10 uM) to raise intracellular cAMP or direct application of dibuty-cAMP (300 uM), a well-known cell-permeable cAMP analogy, stimulated mouse duodenal I sc when added to serosal side but not mucosal side (p<0.001, n=7). Surprisingly, cAMP-mediated intestinal I sc was significantly reduced by external Ca 2+ omission at serosal side of the tissues (p<0.01, n=6), and was inhibited by several selective blockers of storeoperated Ca 2+ channels (SOC) and CRAC/Orai channels at serosal side but not at the mucosal side (p<0.01, n=6). The ER Ca 2+ chelation by TPEN could also significantly reduced cAMPmediated intestinal I sc (p<0.001, n=6). cAMP-mediated intestinal I sc was promoted by activator of ryanodine receptor on the ER (1 mM caffeine, p<0.01, n=6), but reduced by its blocker (300 uM dantrolene, p<0.01, n=6). Furthermore, cAMP-mediated intestinal I sc was reduced by serosal addition but not mucosal addition of H-89 (20 uM, p<0.001, n=6), a PKA inhibitor. Finally, STIM1/Orai1 proteins, the molecular components of SOC, were identified on serosal side of mouse duodenal epithelium. Conclusion: Forskolin/cAMP induces intestinal ion secretion in a polarized manner. It may trigger SOC via PKA phosphrylation of ryanodine receptor to deplete the ER Ca 2+ store and cause Ca 2+ entry via STIM1/ Orai1, which plays a critical role in cAMP-mediated intestinal ion secretion. Our findings provide a new insight to a cross-talk mechanism of cAMP and Ca 2+ signaling in epithelial ion secretion.
Gastroenterology, 2020
in intestinal epithelial cells and in mouse intestine via a transcriptional mechanism. We specula... more in intestinal epithelial cells and in mouse intestine via a transcriptional mechanism. We speculate that CP-induced decrease in PAT-1 function and expression may contribute to a higher occurrence of nephrolithiasis in immunocompromised patients. (Supported by NIDDK and Department of Veterans Affairs) 582 TEDUGLUTIDE INHIBITS SODIUM CAPRATE-INDUCED CLATHRIN-MEDIATED LPS TRANSPORT IN RAT JEJUNUM Yasutada Akiba, Jonathan D. Kaunitz Lipopolysaccharides (LPS) entry from the intestinal lumen to the circulation is associated with systemic inflammation. We have found that LPS is transcellularly transported to the portal vein (PV) during luminal perfusion with long-chain fatty acid (LCFA) via CD36-and lipid raft-mediated pathways, which are inhibited by and the stable glucagon-like peptide-2 (GLP-2) analog teduglutide (TDG). We examined the effects of medium-chain fatty acids (MCFA) on LPS transport in rat jejunum. FITC-LPS was applied to the mucosal bath of Ussing chambered muscle-stripped jejunal and distal colonic mucosa-submucosa preparations, in which short-circuit current (I sc) and tissue electrical resistance (TER) were measured. Serosal appearance of FITC-LPS was measured with or without luminal application of sodium caprate (C10, 10 mM). FITC-dextran 4kDa (FD4) m-to-s movement was also separately measured. The effects of serosal GLP-2 (100 nM) with a dipeptidyl peptidase 4 inhibitor NVP728 (10 µM) or TDG (10 µg/ml) on C10-mediated FITC-LPS transport were also examined. Luminal application of C10 increased FITC-LPS m-to-s transport without any changes in FD4 m-to-s movement and TER in the jejunum, whereas C10 increased FITC-LPS transport with increased FD4 movement and decreased TER in the distal colon, suggesting that C10 enhances transcellular LPS transport in the jejunum, but increases paracellular permeability in the distal colon. In the jejunum, C10-mediated FITC-LPS m-to-s transport was inhibited by pretreatment with luminal Pitstop2 (30 µM), a selective inhibitor of clathrinmediated endocytosis (CME), whereas the CD36 inhibitor sulfosuccinimidyl oleate (SSO, 0.1 mM), the lipid raft inhibitor methyl-b-cyclodextrin (MbCD, 1 mM), or the dynamin inhibitor Dynasore (50 µM) had no effect. In contrast, luminal OA (30 mM) with taurocholate (TCA, 0.1 mM) also increased FITC-LPS transport with SSO, MbCD and Dynasore-sensitive, but Pitstop2-insensitive manner. Serosal application of GLP-2 with NVP728 or TDG abolished C10-mediated FITC-LPS transport. Furthermore, serosal pretreatment of VIP (30 nM) or a nitric oxide donor (S)-Nitroso-N-acetylpenicillamine SNAP (0.1 mM) reduced C10-mediated FITC-LPS transport, suggesting that GLP-2-induced reduction of LPS transport is mediated via VIP and NO pathways. In vivo, jejunal luminal perfusion of C10 (10 mM) with FITC-LPS increased PV FITC-LPS appearance 15 min after perfusion, which was inhibited by coperfusion of Pitstop2. These results suggest that luminal MCFA and LCFA differentially induce transcellular LPS transport in the jejunum; MCFA via CME, but LCFA via the CD36/ lipid raft/caveolin pathway. GLP-2 and TDG inhibited MCFA-induced LPS transport possibly via VIP and NO pathways. Dietary fatty acids may exacerbate LPS-associated diseases, including the metabolic syndrome and multiple organ failure, that can be treated with TDG.
F1000Research, 2019
Vasoactive intestinal peptide (VIP), a gut peptide hormone originally reported as a vasodilator i... more Vasoactive intestinal peptide (VIP), a gut peptide hormone originally reported as a vasodilator in 1970, has multiple physiological and pathological effects on development, growth, and the control of neuronal, epithelial, and endocrine cell functions that in turn regulate ion secretion, nutrient absorption, gut motility, glycemic control, carcinogenesis, immune responses, and circadian rhythms. Genetic ablation of this peptide and its receptors in mice also provides new insights into the contribution of VIP towards physiological signaling and the pathogenesis of related diseases. Here, we discuss the impact of VIP on gastrointestinal function and diseases based on recent findings, also providing insight into its possible therapeutic application to diabetes, autoimmune diseases and cancer.
The Journal of Infectious Diseases, 2019
BackgroundCholera toxin (CT)–induced diarrhea is mediated by cyclic adenosine monophosphate (cAMP... more BackgroundCholera toxin (CT)–induced diarrhea is mediated by cyclic adenosine monophosphate (cAMP)–mediated active Cl– secretion via the cystic fibrosis transmembrane conductance regulator (CFTR). Although the constitutive activation of adenylyl cyclase (AC) in response to CT is due to adenosine diphosphate ribosylation of the small G protein α-subunit activating CFTR with consequent secretory diarrhea, the AC isoform(s) involved remain unknown.MethodsWe generated intestinal epithelial cell–specific adenylyl cyclase 6 (AC6) knockout mice to study its role in CT-induced diarrhea.ResultsAC6 messenger RNA levels were the highest of all 9 membrane-bound AC isoforms in mouse intestinal epithelial cells. Intestinal epithelial-specific AC6 knockout mice (AC6loxloxVillinCre) had undetectable AC6 levels in small intestinal and colonic epithelial cells. No significant differences in fluid and food intake, plasma electrolytes, intestinal/colon anatomy and morphology, or fecal water content wer...
British Journal of Pharmacology, 2015
The bioactive monoamine 5-HT, implicated in the pathogenesis of functional gastrointestinal disor... more The bioactive monoamine 5-HT, implicated in the pathogenesis of functional gastrointestinal disorders, is abundantly synthesized and stored in rat proximal colonic mucosa and released to the gut lumen and subepithelial space. Despite much data regarding its expression and function, the effects of luminal 5-HT on colonic anion secretion have not been fully investigated. EXPERIMENTAL APPROACH We measured short-circuit current (Isc) as an indicator of ion transport in mucosa-submucosa or mucosa-only preparations of rat proximal colon. Total CO2 output was measured in vitro and in vivo. Immunohistochemistry was performed to investigate the localization of 5-HT4, NOS1 and NOS2. KEY RESULTS Luminal 5-HT gradually increased the amplitude and sustained the elevation of Isc. Luminal 5-HT-evoked ΔIsc was acetazolamide sensitive and HCO3 − dependent, consistent with cytosolic carbonic anhydrase-dependent electrogenic HCO3 − secretion, while not affected by tetrodotoxin (TTX), atropine or indomethacin. Pretreatment with the selective 5-HT4 antagonist GR113808, but not antagonists for 5-HT3, 5-HT6 or 5-HT7, inhibited luminal 5-HT-evoked ΔIsc. Furthermore, luminal cisapride and tegaserod increased Isc to the same extent as did 5-HT in the presence of indomethacin and TTX. Removal of the submucosa or pretreatment with NOS inhibitors enhanced luminal 5-HT-evoked ΔIsc, suggesting that NO synthesized in the submucosa suppresses mucosal anion secretion. NOS1 and NOS2 were immunostained in the submucosal neurons and glial cells respectively. Luminal 5-HT-evoked HCO3 − secretion was confirmed in vivo, inhibited by co-perfusion of GR113808, but not by ondansetron. CONCLUSIONS AND IMPLICATIONS A novel apical 5-HT4-mediated HCO3 − secretory pathway and an NO-dependent inhibitory mechanism are present in the proximal colon. Luminal 5-HT-evoked HCO3 − secretion may be important for the maintenance of mucosal integrity by regulating luminal pH.
Gastroenterology, 2017
Background: Parkinson's disease (PD) is characterized by alpha-synucleinopathy at all levels of t... more Background: Parkinson's disease (PD) is characterized by alpha-synucleinopathy at all levels of the brain-gut axis including the enteric nervous system (ENS). Lesions in the ENS may be associated with gut inflammation, increased intestinal permeability and dysmotility contributing to the pathogenesis of PD and gastrointestinal manifestations. Aims: To evaluate fecal calprotectin and zonulin as biomarkers of gut inflammation and intestinal barrier dysfunction, respectively, in patients with PD. Methods: Ten consecutive patients with PD (mean age 65; 5 F, 5 M) and 10 healthy controls (mean age 53; 8 F, 2 M) participated in the study. The quantitative evaluation of calprotectin and zonulin in stool samples was performed by ready-to-use sandwich ELISA tests. Additionally, patients filled out a short questionnaire concerning gastrointestinal symptoms. The Kruskal-Wallis test was used for the comparison of differences between the groups. Results: A mean duration of PD in the studied group was 8 years (range 3-20 years). None of the patients was diagnosed with inflammatory bowel disease. Patients with PD reported constipation (40%), alternating bowel movement pattern (40%), and normal bowel habit (20%), in addition, abdominal pain (20%) and distension (50%). The increased levels of calprotectin and zonulin in stool samples were found in 60% and 40% of PD patients, respectively. Fecal calprotectin level was within the normal range in all the controls, while zonulin level was slightly elevated only in one out of 10 control subjects. The biomarker levels are presented as median and the lower quartile (25Q)-upper quartile (75Q). The fecal calprotectin level (µg/g) was significantly higher in PD patients compared to the controls 134.6 (89.1-145.5) vs 24.8 (18.0-27.9), p= 0.0025. The fecal zonulin level (ng/ml) was also higher in PD patients compared to the controls, but the p-value did not reach statistical significance: 105.5 (63.5-166.0) vs 54.5 (44.0-73.0), p=0.0588. Conclusions: Gastrointestinal symptoms are frequently reported by PD patients. The signs of gut inflammation and increased intestinal permeability are present in a remarkable number of PD patients. This is the first study using stool ELISA tests which confirmed the gut immune system activation and changes in the intestinal epithelial barrier integrity observed in PD patients.