IU Indianapolis ScholarWorks :: Browsing by Subject "Epithelial Cells"

Browsing by Subject "Epithelial Cells"

Now showing 1 - 10 of 13

Analyses of the pathways involved in early- and late-phase induction of IFN-beta during C. muridarum infection of oviduct epithelial cells
(PLoS, 2015-03-23) Hu, Sishun; Hosey, Kristen L.; Derbigny, Wilbert A.; Department of Microbiology and Immunology, IU School of Medicine
We previously reported that the IFN-β secreted by Chlamydia muridarum-infected murine oviduct epithelial cells (OE cells) was mostly dependent on the TLR3 signaling pathway. To further characterize the mechanisms of IFN-β synthesis during Chlamydia infection of OE cells in vitro, we utilized specific inhibitory drugs to clarify the roles of IRF3 and NF-κB on both early- and late-phase C. muridarum infections. Our results showed that the pathways involved in the early-phase of IFN-β production were distinct from that in the late-phase of IFN-β production. Disruption of IRF3 activation using an inhibitor of TBK-1 at early-phase Chlamydia infection had a significant impact on the overall synthesis of IFN-β; however, disruption of IRF3 activation at late times during infection had no effect. Interestingly, inhibition of NF-κB early during Chlamydia infection also had a negative effect on IFN-β production; however, its impact was not significant. Our data show that the transcription factor IRF7 was induced late during Chlamydia infection, which is indicative of a positive feedback mechanism of IFN-β synthesis late during infection. In contrast, IRF7 appears to play little or no role in the early synthesis of IFN-β during Chlamydia infection. Finally, we demonstrate that antibiotics that target chlamydial DNA replication are much more effective at reducing IFN-β synthesis during infection versus antibiotics that target chlamydial transcription. These results provide evidence that early- and late-phase IFN-β production have distinct signaling pathways in Chlamydia-infected OE cells, and suggest that Chlamydia DNA replication might provide a link to the currently unknown chlamydial PAMP for TLR3.
Eradication of Pseudomonas aeruginosa biofilms on cultured airway cells by a fosfomycin/tobramycin antibiotic combination
(Wiley Blackwell (Blackwell Publishing), 2013-02) Anderson, Gregory G.; Kenney, Thomas F.; Macleod, David L.; Henig, Noreen R.; O'Toole, George A.; Department of Biology, School of Science
Chronic biofilm formation by Pseudomonas aeruginosa in cystic fibrosis (CF) lungs is a major cause of morbidity and mortality for patients with CF. To gain insights into effectiveness of novel anti-infective therapies, the inhibitory effects of fosfomycin, tobramycin, and a 4:1 (wt/wt) fosfomycin/tobramycin combination (FTI) on Pseudomonas aeruginosa biofilms grown on cultured human CF-derived airway cells (CFBE41o-) were investigated. In preformed biofilms treated for 16 h with antibiotics, P. aeruginosa CFU per mL were reduced 4 log10 units by both FTI and tobramycin at 256 mg L(-1) , while fosfomycin alone had no effect. Importantly, the FTI treatment contained five times less tobramycin than the tobramycin-alone treatment. Inhibition of initial biofilm formation was achieved at 64 mg L(-1) FTI and 16 mg L(-1) tobramycin. Fosfomycin (1024 mg L(-1)) did not inhibit biofilm formation. Cytotoxicity was also determined by measuring lactate dehydrogenase (LDH). Intriguingly, sub-inhibitory concentrations of FTI (16 mg L(-1)) and tobramycin (4 mg L(-1)) and high concentrations of fosfomycin (1024 mg L(-1)) prevented bacterially mediated airway cell toxicity without a corresponding reduction in CFU. Overall, it was observed that FTI and tobramycin demonstrated comparable activity on biofilm formation and disruption. Decreased administration of tobramycin upon treatment with FTI might lead to a decrease in negative side effects of aminoglycosides.
Functional Effects of Nanoparticle Exposure on Calu-3 Airway Epithelial Cells
(2012) Banga, Amiraj; Witzmann, Frank A.; Petrache, Horia I.; Blazer-Yost, Bonnie
High concentrations of manufactured carbon nanoparticles (CNP) are known to cause oxidative stress, inflammatory responses and granuloma formation in respiratory epithelia. To examine the effects of lower, more physiologically relevant concentrations, the human airway epithelial cell line, Calu-3, was used to evaluate potential alterations in transepithelial permeability and cellular function of airway epithelia after exposure to environmentally realistic concentrations of carbon nanoparticles. Three common carbon nanoparticles, fullerenes, single- and multi-wall carbon nanotubes (SWCNT, MWCNT) were used in these experiments. Electrophysiological measurements were performed to assay transepithelial electrical resistance (TEER) and epinephrine-stimulated chloride (Cl(-)) ion secretion of epithelial cell monolayers that had been exposed to nanoparticles for three different times (1 h, 24 h and 48 h) and over a 7 log unit range of concentrations. Fullerenes did not have any effect on the TEER or stimulated ion transport. However, the carbon nanotubes (CNT) significantly decreased TEER and inhibited epinephrine-stimulated Cl(-) secretion. The changes were time dependent and at more chronic exposures caused functional effects which were evident at concentrations substantially lower than have been previously examined. The functional changes manifested in response to physiologically relevant exposures would inhibit mucociliary clearance mechanisms and compromise the barrier function of airway epithelia.
GT198 Expression Defines Mutant Tumor Stroma in Human Breast Cancer
(Elsevier, 2016-05) Yang, Zheqiong; Peng, Min; Cheng, Liang; Jones, Kimya; Maihle, Nita J.; Mivechi, Nahid F.; Ko, Lan; Department of Pathology and Laboratory Medicine, IU School of Medicine
Human breast cancer precursor cells remain to be elucidated. Using breast cancer gene product GT198 (PSMC3IP; alias TBPIP or Hop2) as a unique marker, we revealed the cellular identities of GT198 mutant cells in human breast tumor stroma. GT198 is a steroid hormone receptor coactivator and a crucial factor in DNA repair. Germline mutations in GT198 are present in breast and ovarian cancer families. Somatic mutations in GT198 are present in ovarian tumor stromal cells. Herein, we show that human breast tumor stromal cells carry GT198 somatic mutations and express cytoplasmic GT198 protein. GT198(+) stromal cells share vascular smooth muscle cell origin, including myoepithelial cells, adipocytes, capillary pericytes, and stromal fibroblasts. Frequent GT198 mutations are associated with GT198(+) tumor stroma but not with GT198(-) tumor cells. GT198(+) progenitor cells are mostly capillary pericytes. When tested in cultured cells, mutant GT198 induces vascular endothelial growth factor promoter, and potentially promotes angiogenesis and adipogenesis. Our results suggest that multiple lineages of breast tumor stromal cells are mutated in GT198. These findings imply the presence of mutant progenitors, whereas their descendants, carrying the same GT198 mutations, are collectively responsible for forming breast tumor microenvironment. GT198 expression is, therefore, a specific marker of mutant breast tumor stroma and has the potential to facilitate diagnosis and targeted treatment of human breast cancer.
The HMGB1-RAGE axis mediates traumatic brain injury-induced pulmonary dysfunction in lung transplantation
(American Association for the Advancement of Science, 2014-09-03) Weber, Daniel J.; Gracon, Adam S.A.; Ripsch, Matthew S.; Fisher, Amanda J.; Cheon, Bo M.; Pandya, Pankita H.; Vittal, Ragini; Capitano, Maegan L.; Kim, Youngsong; Allete, Yohance M.; Riley, Amanda A.; McCarthy, Brian P.; Territo, Paul R.; Hutchins, Gary D.; Broxmeyer, Hal E.; Sandusky, George E.; White, Fletcher A.; Wilkes, David S.; Medicine, School of Medicine
Traumatic brain injury (TBI) results in systemic inflammatory responses that affect the lung. This is especially critical in the setting of lung transplantation, where more than half of donor allografts are obtained postmortem from individuals with TBI. The mechanism by which TBI causes pulmonary dysfunction remains unclear but may involve the interaction of high-mobility group box-1 (HMGB1) protein with the receptor for advanced glycation end products (RAGE). To investigate the role of HMGB1 and RAGE in TBI-induced lung dysfunction, RAGE-sufficient (wild-type) or RAGE-deficient (RAGE(-/-)) C57BL/6 mice were subjected to TBI through controlled cortical impact and studied for cardiopulmonary injury. Compared to control animals, TBI induced systemic hypoxia, acute lung injury, pulmonary neutrophilia, and decreased compliance (a measure of the lungs' ability to expand), all of which were attenuated in RAGE(-/-) mice. Neutralizing systemic HMGB1 induced by TBI reversed hypoxia and improved lung compliance. Compared to wild-type donors, lungs from RAGE(-/-) TBI donors did not develop acute lung injury after transplantation. In a study of clinical transplantation, elevated systemic HMGB1 in donors correlated with impaired systemic oxygenation of the donor lung before transplantation and predicted impaired oxygenation after transplantation. These data suggest that the HMGB1-RAGE axis plays a role in the mechanism by which TBI induces lung dysfunction and that targeting this pathway before transplant may improve recipient outcomes after lung transplantation.
Modulation of basal and peptide hormone-stimulated Na transport by membrane cholesterol content in the A6 epithelial cell line
(2005) West, Aaron; Blazer-Yost, Bonnie
These studies examined the effect of altering plasma membrane cholesterol on basal Na+ flux as well as on the natriferic responses to the peptide hormones, insulin and anti-diuretic hormone (ADH) in the A6 model renal cell line. Membrane cholesterol concentrations were depleted or enriched using methyl-beta-cyclodextrin (MbetaCD) or a MbetaCD/cholesterol inclusion complex respectively. Effects of changes in the apical and basolateral plasma membranes were examined independently. Apical membrane cholesterol removal or supplementation had no effect on the basal Na+ transport rate. Short-term apical membrane cholesterol supplementation also had no effect on insulin-stimulated Na+ transport or on the initial phase of the ADH response. Interestingly, the additional apical membrane cholesterol had an inhibitory effect on the ADH response after 30 minutes. Apical membrane cholesterol depletion partially inhibited the responses to both insulin and ADH. Conversely, supplementation of basolateral cholesterol caused a significant increase in basal Na+ flux. Removal of cholesterol from the basolateral plasma membrane caused a decrease in basal Na+ flux with a time course analogous to channel turnover and completely inhibited peptide hormone responses. None of the changes in membrane cholesterol content decreased transcellular resistance. These results indicate an important role for membrane cholesterol content in the regulation of ENaC-mediated Na+ uptake.
Normal Breast-Derived Epithelial Cells with Luminal and Intrinsic Subtype-Enriched Gene Expression Document Interindividual Differences in Their Differentiation Cascade
(American Association for Cancer Research, 2018-09) Kumar, Brijesh; Prasad, Mayuri; Bhat-Nakshatri, Poornima; Anjanappa, Manjushree; Kalra, Maitri; Marino, Natascia; Storniolo, Anna Maria; Rao, Xi; Liu, Sheng; Wan, Jun; Liu, Yunlong; Nakshatri, Harikrishna; Surgery, School of Medicine
Cell-type origin is one of the factors that determine molecular features of tumors, but resources to validate this concept are scarce because of technical difficulties in propagating major cell types of adult organs. Previous attempts to generate such resources to study breast cancer have yielded predominantly basal-type cell lines. We have created a panel of immortalized cell lines from core breast biopsies of ancestry-mapped healthy women that form ductal structures similar to normal breast in 3D cultures and expressed markers of major cell types, including the luminal-differentiated cell-enriched ERα-FOXA1-GATA3 transcription factor network. We have also created cell lines from PROCR (CD201)+/EpCAM- cells that are likely the "normal" counterpart of the claudin-low subtype of breast cancers. RNA-seq and PAM50-intrinsic subtype clustering identified these cell lines as the "normal" counterparts of luminal A, basal, and normal-like subtypes and validated via immunostaining with basal-enriched KRT14 and luminal-enriched KRT19. We further characterized these cell lines by flow cytometry for distribution patterns of stem/basal, luminal-progenitor, mature/differentiated, multipotent PROCR+ cells, and organogenesis-enriched epithelial/mesenchymal hybrid cells using CD44/CD24, CD49f/EpCAM, CD271/EpCAM, CD201/EpCAM, and ALDEFLUOR assays and E-cadherin/vimentin double staining. These cell lines showed interindividual heterogeneity in stemness/differentiation capabilities and baseline activity of signaling molecules such as NF-κB, AKT2, pERK, and BRD4. These resources can be used to test the emerging concept that genetic variations in regulatory regions contribute to widespread differences in gene expression in "normal" conditions among the general population and can delineate the impact of cell-type origin on tumor progression.Significance: In addition to providing a valuable resource for the breast cancer research community to investigate cell-type origin of different subtypes of breast cancer, this study highlights interindividual differences in normal breast, emphasizing the need to use "normal" cells from multiple sources as controls to decipher the effects of cancer-specific genomic aberrations.
Oral epithelial expression of angiotensin converting enzyme-2: Implications for COVID-19 diagnosis and prognosis
(2020-06-23) Srinivasan, Mythily; Zunt, Susan L.; Goldblatt, Lawrence I.; Oral Pathology, Medicine and Radiology, School of Dentistry
The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) uses the angiotensin converting enzyme (ACE)-2 as the host receptor for target cell entry. The extent and distribution of ACE-2 has been associated with the clinical symptoms of coronavirus disease (COVID)-19. Here we show by immunofluorescence analysis that the ACE2 is abundantly expressed in oral mucosa, particularly in the surface epithelial cells suggesting that these cells could represent sites of entry for SARS-CoV-2. Further, together with the reports on ACE2 ectodomain shedding, we discuss the rationale for the hypothesis that the ACE-2 measurement in saliva could be a marker for COVID-19 infection during early phase following SARS-CoV-2 exposure.
Phosphoinositide lipid second messengers: new paradigms for transepithelial signal transduction
(Bonnie Blazer-Yost and Charity Nofziger. Phosphoinositide lipid second messengers: new paradigms for transepithelial signal transduction. Pflügers Archiv - European Journal of Physiology. (2005)., 2005-03) Blazer-Yost, Bonnie; Nofziger, Charity
Multiple forms of phosphatidylinositol are generated by differential phosphorylation of the inositol headgroup. These phosphoinositides, specifically PI(4,5)P2, have been implicated as modulators in a variety of transport processes. The data indicate that phosphoinositides can modulate transporters directly or via the activation of down-stream signaling components. The phosphoinositide pathway has been linked to changes in transporter kinetics, intracellular signaling, membrane targeting and membrane stability. Recent results obtained for several of the well-characterized transport systems suggest the need to reassess the role of PI(4,5)P2 and question whether lower abundance forms of the phosphoinositides, notably PI(3,4,5)P3 (PIP3) and PI(3,4)P2, are the pertinent transport regulators. In contrast to PI(4,5)P2, these latter forms represent a dynamic, regulated pool, the characteristics of which are more compatible with the nature of signaling intermediates. A recently described, novel transepithelial signaling pathway has been demonstrated for PIP3 in which a signal initiated on the basolateral membrane is transduced to the apical membrane entirely within the planar face of the inner leaflet of the plasma membrane. The new paradigms emerging from recent studies may be widely applicable to transporter regulation in other cell types and are particularly relevant for signaling in polarized cells.
Role of STAT1 in Chlamydia-Induced Type-1 Interferon Production in Oviduct Epithelial Cells
(Mary Ann Liebert, 2015-11) Hosey, Kristen Lynette; Hu, Sishun; Derbigny, Wilbert Alfred; Department of Microbiology & Immunology, IU School of Medicine
We previously reported that Chlamydia muridarum-infected murine oviduct epithelial cells (OE cells) secrete interferon β (IFN-β) in a mostly TLR3-dependent manner. However, C. muridarum-infected TLR3-deficient OE cells were still able to secrete detectable levels of IFN-β into the supernatants, suggesting that other signaling pathways contribute to Chlamydia-induced IFN-β synthesis in these cells. We investigated the role of STAT1 as a possible contributor in the Chlamydia-induced type-1 IFN production in wild-type (WT) and TLR3-deficient OE cells to ascertain its putative role at early- and late-times during Chlamydia infection. Our data show that C. muridarum infection significantly increased STAT1 gene expression and protein activation in WT OE cells; however, TLR3-deficient OE cells showed diminished STAT1 protein activation and gene expression. There was significantly less IFN-β detected in the supernatants of C. muridarum-infected OE cells derived from mice deficient in STAT1 when compared with WT OE cells, which suggest that STAT1 is required for the optimal synthesis of IFN-β during infection. Real-time quantitative polymerase chain reaction analyses of signaling components of the type-1 IFN signaling pathway demonstrated equal upregulation in the expression of STAT2 and IRF7 genes in the WT and TLR3-deficient OE cells, but no upregulation in these genes in the STAT1-deficient OE cells. Finally, experiments in which INFAR1 was blocked with neutralizing antibody revealed that IFNAR1-mediated signaling was critical to the Chlamydia-induced upregulation in IFN-α gene transcription, but had no role in the Chlamydia-induced upregulation in IFN-β gene transcription.

Browsing by Subject "Epithelial Cells"

Results Per Page

Sort Options