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Open Access Publications from the University of California

Scholarly Works (22 results)

  • Article
  • Peer Reviewed
UC San Diego Previously Published Works (2020)
The tight junction protein claudin-2 is upregulated in inflammatory bowel disease, and yet its deficit worsens infectious and chemical colitis. In this issue of the JCI, Raju and Shashikanth et al. examined the contribution of claudin-2 to immune-mediated colitis. The authors used transgenic mouse models to show that claudin-2 deficiency attenuated colitis progression as well as a leak barrier defect, albeit at the risk of intestinal obstruction. Further, inhibition of claudin-2 by targeting casein kinase 2 (CK2) also ameliorated colitis. The findings reveal unsuspected links between the pore and leak pathways of intestinal permeability and immune responses leading to colitis. They additionally suggest potential targets for therapeutic intervention in inflammatory bowel disease.
    Cover page: Claudin-2 pore causes leak that breaches the dam in intestinal inflammation.
    • Thesis
    • Peer Reviewed
    UC San Diego Electronic Theses and Dissertations (2020)

    Non-typhoidal Salmonella is one of the most burdensome foodborne diarrheal pathogens. Previous work has shown a decrease in expression of the chloride/bicarbonate exchanger SLC26A3 (Down-Regulated in Adenoma; DRA) following Salmonella infection and it is thought that diarrheal symptoms could stem from the downregulation of DRA. However, despite its prevalence, the mechanism by which Salmonella elicits diarrhea is not entirely known. To elucidate this, we have developed an enteroid model to recapitulate both the absorptive and secretory lineages of the intestinal epithelium. Enteroid-derived monolayers (EDMs) were infected with Salmonella where relevant protein expression was studied using qRT-PCR and western blot. Infection reduced expression of DRA and Hes1 while upregulating ATOH1 and Muc2. Hes1 is a Notch pathway downstream signaling molecule and thus a precursor to the absorptive epithelial lineage that expresses DRA. ATOH1, in contrast, is a Wnt pathway downstream signaling molecule and a precursor to secretory lineages, including goblet cells that express Muc2. The involvement of Notch was further investigated by inhibiting Notch signaling using a γ-secretase inhibitor, which reproduced the downregulation in Hes1 and DRA and upregulation in ATOH1 and Muc2 seen with infection. Secretory transporters NKCC1 and CLCA1 were also upregulated following infection, which might contribute to the imbalance between absorption and secretion. Our findings suggest that the pathogenesis of diarrheal disease in the setting of Salmonella infection may reflect Notch inhibition and an accompanying shift in differentiation from absorptive to secretory cell types and transporters, a decreased capacity for absorption, and thus the accumulation of diarrheal fluid.

      Cover page: Salmonella biases epithelial differentiation through Wnt and Notch signaling, which may contribute to diarrheal pathogenesis
      • Article
      • Peer Reviewed
      UC San Diego Previously Published Works (2018)
      Every year, enteric infections and associated diarrhea kill millions of people. The situation is compounded by increases in the number of enteric pathogens that are acquiring resistance to antibiotics, as well as (hitherto) a relative paucity of information on host molecular targets that may contribute to diarrhea. Many forms of diarrheal disease depend on the dysregulation of intestinal ion transporters, and an associated imbalance between secretory and absorptive functions of the intestinal epithelium. A number of major transporters have been implicated in the pathogenesis of diarrheal diseases and thus an understanding of their expression, localization, and regulation after infection with various bacteria, viruses, and protozoa likely will prove critical in designing new therapies. This article surveys our understanding of transporters that are modulated by specific pathogens and the mechanism(s) involved, thereby illuminating targets that might be exploited for new therapeutic approaches.
        Cover page: The Role of Ion Transporters in the Pathophysiology of Infectious Diarrhea
        • Thesis
        • Peer Reviewed
        UC San Diego Electronic Theses and Dissertations (2017)

        Non-typhoidal Salmonella is the most burdensome foodborne illness worldwide, yet despite its prevalence, the mechanism through which Salmonella causes diarrhea is not entirely known. Intestinal ion transporters play important roles in water absorption in the intestine, and can be a therapeutic target in Salmonella induced diarrhea, but there is not a complete understanding of which ion transporters Salmonella regulates. We have previously shown that infection with Salmonella causes decreased expression of the chloride/bicarbonate exchanger SLC26A3 (DRA; Down-Regulated in Adenoma) in the colonic tissues of mice. To better understand the mechanism

        of Salmonella infection in intestinal epithelial cells, we developed an enteroid model that mimics the crypt-villus axis of the intestines, and contains the four different cell subtypes found in the intestines (enterocytes, enteroendocrine, goblet, and Paneth cells). These enteroids were converted to enteroid-derived monolayers (EDM) to allow access to the apical membrane. With this model, we looked at changes in the Wnt/NOTCH signaling pathways responsible for controlling cell fate commitment in the intestines in addition to changes in DRA in both colonic and small intestinal EDM. We hypothesized that EDM would show similar decreases in DRA to those seen in vivo, in addition to alterations in Wnt/NOTCH signaling that may be linked to DRA down-regulation. We found that expression of both DRA and NICD (of the NOTCH signaling pathway) were decreased 6 h post infection. Our work shows the possibility of using EDM as a model to uncover mechanisms that can be targeted for therapeutic developments against diarrhea caused by Salmonella infection.

          Cover page: Mechanisms Underlying Diarrheal Disease in Enteric Salmonella Infections
          • Article
          • Peer Reviewed
          UC Riverside Previously Published Works (2016)
          Reactive oxygen species (ROS) such as hydrogen peroxide (H2 O2 ) contribute to epithelial damage and ion transport dysfunction (key events in inflammatory diarrhoea) in inflammatory bowel disease (IBD). The aim of this study was to identify if H2 O2 mediates suppression of colonic ion transport function in the murine dextran sulfate sodium (DSS) colitis model by using the H2 O2 degrading enzyme, catalase. Colitis was induced by administering DSS (4%) in drinking water for 5 days followed by 3 days on normal H2 O. Mice were administered either pegylated catalase or saline at day -1, 0 and +1 of DSS treatment. Ion transport responses to the Ca2+ -dependent agonist, carbachol (CCh), or the cAMP-dependent agonist, forskolin, were measured across distal colonic mucosa mounted in Ussing chambers. Parameters of DSS-induced inflammation (loss in body weight, decreased colon length, altered stool consistency), were only partially alleviated by catalase while histology was only minimally improved. However, catalase significantly reversed the DSS-induced reduction in baseline ion transport as well as colonic Isc responses to CCh. However, ion transport responses to forskolin were not significantly restored. Catalase also reduced activation of ERK MAP kinase in the setting of colitis, and increased expression of the Na+ -K+ -2Cl- cotransporter, NKCC1, consistent with restoration of ion transport function. Ex vivo treatment of inflamed colonic mucosae with catalase also partially restored ion transport function. Therefore, catalase partially prevents, and rescues, the loss of ion transport properties in DSS colitis even in the setting of unresolved tissue inflammation. These findings indicate a prominent role for ROS in ion transport dysfunction in colitis and may suggest novel strategies for the treatment of inflammatory diarrhoea.
            Cover page: Hydrogen peroxide scavenger, catalase, alleviates ion transport dysfunction in murine colitis
            • Article
            • Peer Reviewed
            UC San Diego Previously Published Works (2017)
            Chronic watery diarrhea poses a diagnostic and therapeutic challenge and is often a disabling condition for patients. Although acute diarrhea is likely to be caused by infection, the causes of chronic diarrhea (>4 weeks in duration) are more elusive. We review the pathophysiology, diagnosis, and treatment of chronic diarrhea. Drawing on recent insights into the molecular mechanisms of intestinal epithelial transport and barrier function, we discuss how diarrhea can result from a decrease in luminal solute absorption, an increase in secretion, or both, as well as derangements in barrier properties. We also describe the various extraepithelial factors that activate diarrheal mechanisms. Finally, clinical evaluation and tests used in the assessment of patients presenting with chronic diarrhea are reviewed, and an algorithm guiding therapeutic decisions and pharmacotherapy is presented.
              Cover page: Pathophysiology, Evaluation, and Management of Chronic Watery Diarrhea.
              • Article
              • Peer Reviewed
              UC San Diego Previously Published Works (2020)
              Epidermal growth factor receptor tyrosine kinase inhibitors (EGFr TKIs) are first-line therapies for various cancers, and cause dose-limiting severe diarrhea in many patients. We hypothesized that diarrhea caused by EGFr TKIs might reflect actions on epithelial transport, barrier function, or both, which we tested using cell cultures including murine and human enteroid-derived monolayers (EDMs), analyzed using electrophysiological and other relevant methods. EGFr TKIs (such as afatinib, erlotinib, and osimertinib) reversed the acute inhibitory effect of EGF on chloride secretion induced by carbachol (CCh) across T84 human colonic epithelial cells, which correlated with the diarrhea-inducing effect of each agent clinically. EGFr TKIs also reduced transepithelial electrical resistance (TEER), whereas co-treatment with CCh delayed the decrease in TEER compared with that of cells co-treated with EGF. Furthermore, afatinib and erlotinib prevented EGF- or CCh-induced EGFr phosphorylation. EGFr TKIs also suppressed phosphorylation of extracellular signal-regulated kinase (Erk)1/2 in response to EGF, whereas they had weaker effects on CCh-induced Erk1/2 phosphorylation. In human EDMs, EGF potentiated ion transport induced by CCh, whereas afatinib reversed this effect. The ability of EGFr TKIs to reverse the effects of EGF on calcium-dependent chloride secretion could contribute to the diarrheal side effects of these agents, and their disruption of epithelial barrier dysfunction is likely also pathophysiologically significant. CCh-activated Erk1/2 phosphorylation was relatively insensitive to EGFr TKIs and delayed the deleterious effects of EGFr TKIs on barrier function. These findings confirm and extend those of other authors, and may be relevant to designing strategies to overcome the diarrheal side effects of EGFr TKIs.
                Cover page: Potentiation of calcium‐activated chloride secretion and barrier dysfunction may underlie EGF receptor tyrosine kinase inhibitor‐induced diarrhea
                • Article
                • Peer Reviewed
                UC San Diego Previously Published Works (2023)
                Diarrhea is an important cause of hospitalizations in Mozambique. However, little attention has been paid to the impact HIV infection on the prevalence or clinical manifestations of enteric bacterial infections. This study aimed to determine the prevalence of Salmonella spp., Shigella spp. and Campylobacter spp. in HIV-infected and HIV-uninfected patients with diarrhea, identify risk factors for infection, and explore the association between HIV status, viral load, and bacterial prevalence. We conducted a case-control study at the Centro de Saúde de Mavalane and Centro de Saúde 1° de Maio in Maputo, Mozambique, from November 2021 to May 2022. We recruited 300 patients, including 150 HIV-infected (cases) and 150 HIV-uninfected patients (controls), aged between 0-88 years, presenting with diarrhea. Stool samples were collected for bacterial isolation through culture, and for each HIV-infected patient, 4 ml of venous blood were obtained for viral load detection through PCR. A total of 129 patients (43.0%) had at least one bacterial infection. The prevalence of Salmonella spp., Shigella spp. and Campylobacter spp. was 33.0% (n = 99), 15.0% (n = 45) and 4.3% (n = 13), respectively. The prevalence of any bacterial infection did not differ significantly between HIV-infected (45.3%, n = 68) and HIV-uninfected patients (40.7%, = 61) (p = 0.414). Overall, having 2-3 symptoms of enteric disease (p = 0.008) and a basic education (p = 0.030) were factors associated with bacterial infection. Of the 148 patients for whom HIV-1 RNA levels were available, 115 had copy numbers ≤ 75. Another 13 had levels between 76 and 1,000 and the remaining 20 had an average of 327,218.45 copies/ml. Bivariate logistic regression found that Shigella spp. were associated with HIV (p = 0.038), although no association was found in the multivariate analysis. Enteric infections are common in both HIV-infected and -uninfected patients. Low schooling influences the occurrence of enteric infections, which highlights the need to raise awareness about their prevention.
                  Cover page: Microbiological assessment reveals that Salmonella, Shigella and Campylobacter infections are widespread in HIV infected and uninfected patients with diarrhea in Mozambique.
                  • Article
                  • Peer Reviewed
                  UC San Diego Previously Published Works (2015)

                  Background

                  Gastroenteritis is the most common manifestation of nontyphoidal Salmonella enterica infections, but little is known about the pathogenesis of diarrhea in this infection

                  Methods

                  To determine whether polymorphonuclear neutrophils (PMNs) are required for diarrhea for Salmonella colitis, we infected kanamycin-pretreated interleukin 8R (IL-8R) mutant mice and controls, both with nonmutant Slc11a1 (Nramp1, ItyR). We compared the 2 mouse strains for increases in fecal water content (diarrhea) 3 days after infection, changes in expression of ion transporters in colonic epithelial cells, proliferation of epithelial cells, and severity of infection as measured by colony-forming units (CFUs).

                  Results

                  The IL-8R knockout mice had fewer PMNs in the colon but the other variables we measured were unaffected except for an increase in CFUs in the colon. The pathologic changes in the cecum were similar in both groups except for the lack of PMNs in the IL-8R knockout mice. There was minimal damage to the colon more distally.

                  Conclusions

                  In the early stage of Salmonella colitis, PMNs are not required for diarrhea or for the decrease in expression of colonic epithelial cell apical ion transporters. They contribute to defense against infection in the cecum but not extracolonically at this stage of Salmonella colitis.
                    Cover page: Salmonella-induced Diarrhea Occurs in the Absence of IL-8 Receptor (CXCR2)-Dependent Neutrophilic Inflammation
                    • Article
                    • Peer Reviewed
                    UC San Diego Previously Published Works (2023)
                    BACKGROUND: Antibacterial resistance is a growing concern worldwide, including in Mozambique. Diarrhea is an important cause of mortality in Mozambique, yet few local studies have reported on the resistance of bacterial pathogens in this context. Therefore, this study aims to characterize antibiotic susceptibility patterns of Salmonella, Shigella and Campylobacter spp. among patients with diarrhea, including those who are HIV-infected and-uninfected. METHODS: We conducted antibiotic susceptibility testing on 157 stool isolates recovered from 129 patients aged between 0 and 80 years with diarrhea, including HIV infected (n = 68) and-uninfected individuals (n = 61), assisted at two health centers in Maputo city. The isolates comprised of 99 Salmonella, 45 Shigella and 13 Campylobacter strains. The Kirby-Bauer disk diffusion method was used on Mueller-Hinton II agar for Salmonella and Shigella spp., while Mueller-Hinton II agar with 5% defibrinated sheep blood was used for Campylobacter spp. We tested six antibiotics listed on the national essential medicines list, including ciprofloxacin, erythromycin, azithromycin, trimethoprim-sulfamethoxazole, gentamicin, and tetracycline. RESULTS: All isolates were resistant to at least one antibiotic. A high percentage of Salmonella spp. isolates were found to be resistant to trimethoprim-sulfamethoxazole (89.9%, n = 89), erythromycin (88.9%, n = 88) and tetracycline (76.8%, n = 76). In addition, 86.6% (n = 39) and 68.9% (n = 31) of Shigella isolates were resistant to trimethoprim-sulfamethoxazole and tetracycline, respectively. The majority of Campylobacter isolates (92.3%, n = 12) were resistant to erythromycin, azithromycin and tetracycline. Multidrug resistance (MDR) was observed in 79.8% of Salmonella spp., 76.9% of Campylobacter spp., and 57.8% of Shigella spp. Drug susceptibility profiles for Salmonella spp. and Campylobacter were similar in both HIV-1 infected and uninfected patients. However, Shigella spp. isolates obtained from patients without HIV infection were significantly more likely to be resistant to erythromycin, azithromycin or to exhibit multidrug resistance than those obtained from patients with HIV-1 infection (p < 0.05). All Shigella spp. and Campylobacter spp. isolates were susceptible to gentamicin. CONCLUSION: Our study highlights concerning rates of antibiotic resistance and MDR among diarrheal bacterial pathogens in Mozambique. Further research is needed to understand the impact of HIV, ART therapy and immunosuppression on antibiotic resistance. Urgent interventions are essential to prevent the spread of resistant strains.
                      Cover page: Susceptibility antibiotic screening reveals high rates of multidrug resistance of Salmonella, Shigella and Campylobacter in HIV infected and uninfected patients from Mozambique.
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