Browsing by Subject "Epithelial Cells"
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Item Open Access ACE-inhibition increases podocyte number in experimental glomerular disease independent of proliferation.(Journal of the renin-angiotensin-aldosterone system : JRAAS, 2015-06) Zhang, Jiong; Yanez, David; Floege, Anna; Lichtnekert, Julia; Krofft, Ronald D; Liu, Zhi-Hong; Pippin, Jeffrey W; Shankland, Stuart JObjective
The objective of this article is to test the effects of angiotensin-converting enzyme (ACE)-inhibition on glomerular epithelial cell number in an inducible experimental model of focal segmental glomerulosclerosis (FSGS).Background
Although ACE-inhibition has been shown to limit podocyte loss by enhancing survival, little is known about its effect on podocyte number following an abrupt decline in disease.Methods
Experimental FSGS was induced with cytotoxic antipodocyte antibody. Following induction, groups were randomized to receive the ACE-inhibitor enalapril, the smooth muscle relaxant hydralazine (blood pressure control) or drinking water. Blood pressure, kidney function and histology were measured seven and 14 days following disease induction.Results
Both glomerulosclerosis and urinary albumin-to-creatinine ratio were less in the ACE-inhibition arm at day 14. At day 7 of disease, mean podocyte numbers were 26% and 29% lower in the enalapril and hydralazine arms, respectively, compared to normal mice in which no antibody was injected. At day 14, the mean podocyte number was only 18% lower in the enalapril arm, but was 39% lower in the hydralazine arm compared to normal mice. Podocyte proliferation did not occur at any time in any group. Compared to water- or hydralazine-treated mice with FSGS, the enalapril arm had a higher mean number of glomerular parietal epithelial cells that co-expressed the podocyte proteins WT-1 and synaptopodin, as well as phospho-ERK.Conclusion
The results show following an abrupt decline in podocyte number, the initiation of ACE-inhibition but not hydralazine, was accompanied by higher podocyte number in the absence of proliferation. This was accompanied by a higher number of parietal epithelial cells that co-express podocyte proteins. Increasing podocyte number appears to be accompanied by reduced glomerulosclerosis.Item Open Access Age-related changes in the cellular composition and epithelial organization of the mouse trachea.(PloS one, 2014-01) Wansleeben, Carolien; Bowie, Emily; Hotten, Danielle F; Yu, Yen-Rei A; Hogan, Brigid LMWe report here senescent changes in the structure and organization of the mucociliary pseudostratified epithelium of the mouse trachea and main stem bronchi. We confirm previous reports of the gradual appearance of age-related, gland-like structures (ARGLS) in the submucosa, especially in the intercartilage regions and carina. Immunohistochemistry shows these structures contain ciliated and secretory cells and Krt5+ basal cells, but not the myoepithelial cells or ciliated ducts typical of normal submucosal glands. Data suggest they arise de novo by budding from the surface epithelium rather than by delayed growth of rudimentary or cryptic submucosal glands. In old mice the surface epithelium contains fewer cells per unit length than in young mice and the proportion of Krt5+, p63+ basal cells is reduced in both males and females. However, there appears to be no significant difference in the ability of basal stem cells isolated from individual young and old mice to form clonal tracheospheres in culture or in the ability of the epithelium to repair after damage by inhaled sulfur dioxide. Gene expression analysis by Affymetrix microarray and quantitative PCR, as well as immunohistochemistry and flow sorting studies, are consistent with low-grade chronic inflammation in the tracheas of old versus young mice and an increase in the number of immune cells. The significance of these changes for ARGL formation are not clear since several treatments that induce acute inflammation in young mice did not result in budding of the surface epithelium.Item Open Access Anti-phospholipid human monoclonal antibodies inhibit CCR5-tropic HIV-1 and induce beta-chemokines.(J Exp Med, 2010-04-12) Moody, MA; Liao, HX; Alam, SM; Scearce, RM; Plonk, MK; Kozink, DM; Drinker, MS; Zhang, R; Xia, SM; Sutherland, LL; Tomaras, GD; Giles, IP; Kappes, JC; Ochsenbauer Jambor, C; Edmonds, TG; Soares, M; Barbero, G; Forthal, DN; Landucci, G; Chang, C; King, SW; Kavlie, A; Denny, TN; Hwang, KK; Chen, PP; Thorpe, PE; Montefiori, DC; Haynes, BFTraditional antibody-mediated neutralization of HIV-1 infection is thought to result from the binding of antibodies to virions, thus preventing virus entry. However, antibodies that broadly neutralize HIV-1 are rare and are not induced by current vaccines. We report that four human anti-phospholipid monoclonal antibodies (mAbs) (PGN632, P1, IS4, and CL1) inhibit HIV-1 CCR5-tropic (R5) primary isolate infection of peripheral blood mononuclear cells (PBMCs) with 80% inhibitory concentrations of <0.02 to approximately 10 microg/ml. Anti-phospholipid mAbs inhibited PBMC HIV-1 infection in vitro by mechanisms involving binding to monocytes and triggering the release of MIP-1alpha and MIP-1beta. The release of these beta-chemokines explains both the specificity for R5 HIV-1 and the activity of these mAbs in PBMC cultures containing both primary lymphocytes and monocytes.Item Open Access C3a receptor antagonism as a novel therapeutic target for chronic rhinosinusitis.(Mucosal immunology, 2018-09) Mulligan, Jennifer K; Patel, Kunal; Williamson, Tucker; Reaves, Nicholas; Carroll, William; Stephenson, Sarah E; Gao, Peng; Drake, Richard R; Neely, Benjamin A; Tomlinson, Stephen; Schlosser, Rodney J; Atkinson, CarlChronic rhinosinusitis with nasal polyps (CRSwNP) is an inflammatory disease with an unknown etiology. Recent studies have implicated the complement system as a potential modulator of disease immunopathology. We performed proteomic pathway enrichment analysis of differentially increased proteins, and found an enrichment of complement cascade pathways in the nasal mucus of individuals with CRSwNP as compared to control subjects. Sinonasal mucus levels of complement 3 (C3) correlated with worse subjective disease severity, whereas no significant difference in systemic C3 levels could be determined in plasma samples. Given that human sinonasal epithelial cells were the predominate sinonasal source of C3 and complement anaphylatoxin 3a (C3a) staining, we focused on their role in in vitro studies. Baseline intracellular C3 levels were higher in CRSwNP cells, and following exposure to Aspergillus fumigatus (Af) extract, they released significantly more C3 and C3a. Inhibition of complement 3a receptor (C3aR) signaling led to a decrease in Af-induced C3 and C3a release, both in vitro and in vivo. Finally, we found in vivo that C3aR deficiency or inhibition significantly reduced inflammation and CRS development in a mouse model of Af-induced CRS. These findings demonstrate that local sinonasal complement activation correlates with subjective disease severity, and that local C3aR antagonism significantly ameliorates Af-induced CRS in a rodent model.Item Open Access Chlamydia trachomatis immune evasion via downregulation of MHC class I surface expression involves direct and indirect mechanisms.(Infectious diseases in obstetrics and gynecology, 2011-01) Ibana, Joyce A; Schust, Danny J; Sugimoto, Jun; Nagamatsu, Takeshi; Greene, Sheila J; Quayle, Alison JGenital C. trachomatis infections typically last for many months in women. This has been attributed to several strategies by which C. trachomatis evades immune detection, including well-described methods by which C. trachomatis decreases the cell surface expression of the antigen presenting molecules major histocompatibility complex (MHC) class I, MHC class II, and CD1d in infected genital epithelial cells. We have harnessed new methods that allow for separate evaluation of infected and uninfected cells within a mixed population of chlamydia-infected endocervical epithelial cells to demonstrate that MHC class I downregulation in the presence of C. trachomatis is mediated by direct and indirect (soluble) factors. Such indirect mechanisms may aid in priming surrounding cells for more rapid immune evasion upon pathogen entry and help promote unfettered spread of C. trachomatis genital infections.Item Open Access Chlamydia trachomatis Infection Leads to Defined Alterations to the Lipid Droplet Proteome in Epithelial Cells.(PLoS One, 2015) Saka, Hector Alex; Thompson, J Will; Chen, Yi-Shan; Dubois, Laura G; Haas, Joel T; Moseley, Arthur; Valdivia, Raphael HThe obligate intracellular bacterium Chlamydia trachomatis is a major human pathogen and a main cause of genital and ocular diseases. During its intracellular cycle, C. trachomatis replicates inside a membrane-bound vacuole termed an "inclusion". Acquisition of lipids (and other nutrients) from the host cell is a critical step in chlamydial replication. Lipid droplets (LD) are ubiquitous, ER-derived neutral lipid-rich storage organelles surrounded by a phospholipids monolayer and associated proteins. Previous studies have shown that LDs accumulate at the periphery of, and eventually translocate into, the chlamydial inclusion. These observations point out to Chlamydia-mediated manipulation of LDs in infected cells, which may impact the function and thereby the protein composition of these organelles. By means of a label-free quantitative mass spectrometry approach we found that the LD proteome is modified in the context of C. trachomatis infection. We determined that LDs isolated from C. trachomatis-infected cells were enriched in proteins related to lipid metabolism, biosynthesis and LD-specific functions. Interestingly, consistent with the observation that LDs intimately associate with the inclusion, a subset of inclusion membrane proteins co-purified with LD protein extracts. Finally, genetic ablation of LDs negatively affected generation of C. trachomatis infectious progeny, consistent with a role for LD biogenesis in optimal chlamydial growth.Item Open Access Chlamydia trachomatis Infection of Endocervical Epithelial Cells Enhances Early HIV Transmission Events.(PloS one, 2016-01) Buckner, Lyndsey R; Amedee, Angela M; Albritton, Hannah L; Kozlowski, Pamela A; Lacour, Nedra; McGowin, Chris L; Schust, Danny J; Quayle, Alison JChlamydia trachomatis causes a predominantly asymptomatic, but generally inflammatory, genital infection that is associated with an increased risk for HIV acquisition. Endocervical epithelial cells provide the major niche for this obligate intracellular bacterium in women, and the endocervix is also a tissue in which HIV transmission can occur. The mechanism by which CT infection enhances HIV susceptibility at this site, however, is not well understood. Utilizing the A2EN immortalized endocervical epithelial cell line grown on cell culture inserts, we evaluated the direct role that CT-infected epithelial cells play in facilitating HIV transmission events. We determined that CT infection significantly enhanced the apical-to-basolateral migration of cell-associated, but not cell-free, HIVBaL, a CCR5-tropic strain of virus, across the endocervical epithelial barrier. We also established that basolateral supernatants from CT-infected A2EN cells significantly enhanced HIV replication in peripheral mononuclear cells and a CCR5+ T cell line. These results suggest that CT infection of endocervical epithelial cells could facilitate both HIV crossing the mucosal barrier and subsequent infection or replication in underlying target cells. Our studies provide a mechanism by which this common STI could potentially promote the establishment of founder virus populations and the maintenance of local HIV reservoirs in the endocervix. Development of an HIV/STI co-infection model also provides a tool to further explore the role of other sexually transmitted infections in enhancing HIV acquisition.Item Open Access COVID-19 tissue atlases reveal SARS-CoV-2 pathology and cellular targets.(Nature, 2021-07) Delorey, Toni M; Ziegler, Carly GK; Heimberg, Graham; Normand, Rachelly; Yang, Yiming; Segerstolpe, Åsa; Abbondanza, Domenic; Fleming, Stephen J; Subramanian, Ayshwarya; Montoro, Daniel T; Jagadeesh, Karthik A; Dey, Kushal K; Sen, Pritha; Slyper, Michal; Pita-Juárez, Yered H; Phillips, Devan; Biermann, Jana; Bloom-Ackermann, Zohar; Barkas, Nikolaos; Ganna, Andrea; Gomez, James; Melms, Johannes C; Katsyv, Igor; Normandin, Erica; Naderi, Pourya; Popov, Yury V; Raju, Siddharth S; Niezen, Sebastian; Tsai, Linus T-Y; Siddle, Katherine J; Sud, Malika; Tran, Victoria M; Vellarikkal, Shamsudheen K; Wang, Yiping; Amir-Zilberstein, Liat; Atri, Deepak S; Beechem, Joseph; Brook, Olga R; Chen, Jonathan; Divakar, Prajan; Dorceus, Phylicia; Engreitz, Jesse M; Essene, Adam; Fitzgerald, Donna M; Fropf, Robin; Gazal, Steven; Gould, Joshua; Grzyb, John; Harvey, Tyler; Hecht, Jonathan; Hether, Tyler; Jané-Valbuena, Judit; Leney-Greene, Michael; Ma, Hui; McCabe, Cristin; McLoughlin, Daniel E; Miller, Eric M; Muus, Christoph; Niemi, Mari; Padera, Robert; Pan, Liuliu; Pant, Deepti; Pe'er, Carmel; Pfiffner-Borges, Jenna; Pinto, Christopher J; Plaisted, Jacob; Reeves, Jason; Ross, Marty; Rudy, Melissa; Rueckert, Erroll H; Siciliano, Michelle; Sturm, Alexander; Todres, Ellen; Waghray, Avinash; Warren, Sarah; Zhang, Shuting; Zollinger, Daniel R; Cosimi, Lisa; Gupta, Rajat M; Hacohen, Nir; Hibshoosh, Hanina; Hide, Winston; Price, Alkes L; Rajagopal, Jayaraj; Tata, Purushothama Rao; Riedel, Stefan; Szabo, Gyongyi; Tickle, Timothy L; Ellinor, Patrick T; Hung, Deborah; Sabeti, Pardis C; Novak, Richard; Rogers, Robert; Ingber, Donald E; Jiang, Z Gordon; Juric, Dejan; Babadi, Mehrtash; Farhi, Samouil L; Izar, Benjamin; Stone, James R; Vlachos, Ioannis S; Solomon, Isaac H; Ashenberg, Orr; Porter, Caroline BM; Li, Bo; Shalek, Alex K; Villani, Alexandra-Chloé; Rozenblatt-Rosen, Orit; Regev, AvivCOVID-19, which is caused by SARS-CoV-2, can result in acute respiratory distress syndrome and multiple organ failure1-4, but little is known about its pathophysiology. Here we generated single-cell atlases of 24 lung, 16 kidney, 16 liver and 19 heart autopsy tissue samples and spatial atlases of 14 lung samples from donors who died of COVID-19. Integrated computational analysis uncovered substantial remodelling in the lung epithelial, immune and stromal compartments, with evidence of multiple paths of failed tissue regeneration, including defective alveolar type 2 differentiation and expansion of fibroblasts and putative TP63+ intrapulmonary basal-like progenitor cells. Viral RNAs were enriched in mononuclear phagocytic and endothelial lung cells, which induced specific host programs. Spatial analysis in lung distinguished inflammatory host responses in lung regions with and without viral RNA. Analysis of the other tissue atlases showed transcriptional alterations in multiple cell types in heart tissue from donors with COVID-19, and mapped cell types and genes implicated with disease severity based on COVID-19 genome-wide association studies. Our foundational dataset elucidates the biological effect of severe SARS-CoV-2 infection across the body, a key step towards new treatments.Item Open Access Dectin 1 activation on macrophages by galectin 9 promotes pancreatic carcinoma and peritumoral immune tolerance.(Nature medicine, 2017-05) Daley, Donnele; Mani, Vishnu R; Mohan, Navyatha; Akkad, Neha; Ochi, Atsuo; Heindel, Daniel W; Lee, Ki Buom; Zambirinis, Constantinos P; Pandian, Gautam Sd Balasubramania; Savadkar, Shivraj; Torres-Hernandez, Alejandro; Nayak, Shruti; Wang, Ding; Hundeyin, Mautin; Diskin, Brian; Aykut, Berk; Werba, Gregor; Barilla, Rocky M; Rodriguez, Robert; Chang, Steven; Gardner, Lawrence; Mahal, Lara K; Ueberheide, Beatrix; Miller, GeorgeThe progression of pancreatic oncogenesis requires immune-suppressive inflammation in cooperation with oncogenic mutations. However, the drivers of intratumoral immune tolerance are uncertain. Dectin 1 is an innate immune receptor crucial for anti-fungal immunity, but its role in sterile inflammation and oncogenesis has not been well defined. Furthermore, non-pathogen-derived ligands for dectin 1 have not been characterized. We found that dectin 1 is highly expressed on macrophages in pancreatic ductal adenocarcinoma (PDA). Dectin 1 ligation accelerated the progression of PDA in mice, whereas deletion of Clec7a-the gene encoding dectin 1-or blockade of dectin 1 downstream signaling was protective. We found that dectin 1 can ligate the lectin galectin 9 in mouse and human PDA, which results in tolerogenic macrophage programming and adaptive immune suppression. Upon disruption of the dectin 1-galectin 9 axis, CD4+ and CD8+ T cells, which are dispensable for PDA progression in hosts with an intact signaling axis, become reprogrammed into indispensable mediators of anti-tumor immunity. These data suggest that targeting dectin 1 signaling is an attractive strategy for developing an immunotherapy for PDA.Item Open Access Discrimination of vanadium from zinc using gene profiling in human bronchial epithelial cells.(Environmental health perspectives, 2005-12) Li, Zhuowei; Stonehuerner, Jackie; Devlin, Robert B; Huang, Yuh-Chin TWe hypothesized that gene expression profiling may discriminate vanadium from zinc in human bronchial epithelial cells (HBECs). RNA from HBECs exposed to vehicle, V (50 microM), or Zn (50 microM) for 4 hr (n = 4 paired experiments) was hybridized to Affymetrix Hu133A chips. Using one-class t-test with p < 0.01, we identified 140 and 76 genes with treatment:control ratios > or = 2.0 or < or = 0.5 for V and Zn, respectively. We then categorized these genes into functional pathways and compared the number of genes in each pathway between V and Zn using Fisher's exact test. Three pathways regulating gene transcription, inflammatory response, and cell proliferation distinguished V from Zn. When genes in these three pathways were matched with the 163 genes flagged by the same statistical filtration for V:Zn ratios, 12 genes were identified. The hierarchical clustering analysis showed that these 12 genes discriminated V from Zn and consisted of two clusters. Cluster 1 genes (ZBTB1, PML, ZNF44, SIX1, BCL6, ZNF450) were down-regulated by V and involved in gene transcription, whereas cluster 2 genes (IL8, IL1A, PTGS2, DTR, TNFAIP3, CXCL3) were up-regulated and linked to inflammatory response and cell proliferation. Also, metallothionein 1 genes (MT1F, MT1G, MT1K) were up-regulated by Zn only. Thus, using microarray analysis, we identified a small set of genes that may be used as biomarkers for discriminating V from Zn. The novel genes and pathways identified by the microarray may help us understand the pathogenesis of health effects caused by environmental V and Zn exposure.Item Open Access Epigenetic basis of oncogenic-Kras-mediated epithelial-cellular proliferation and plasticity.(Developmental cell, 2022-02) Kadur Lakshminarasimha Murthy, Preetish; Xi, Rui; Arguijo, Diana; Everitt, Jeffrey I; Kocak, Dewran D; Kobayashi, Yoshihiko; Bozec, Aline; Vicent, Silvestre; Ding, Shengli; Crawford, Gregory E; Hsu, David; Tata, Purushothama Rao; Reddy, Timothy; Shen, XilingOncogenic Kras induces a hyper-proliferative state that permits cells to progress to neoplasms in diverse epithelial tissues. Depending on the cell of origin, this also involves lineage transformation. Although a multitude of downstream factors have been implicated in these processes, the precise chronology of molecular events controlling them remains elusive. Using mouse models, primary human tissues, and cell lines, we show that, in Kras-mutant alveolar type II cells (AEC2), FOSL1-based AP-1 factor guides the mSWI/SNF complex to increase chromatin accessibility at genomic loci controlling the expression of genes necessary for neoplastic transformation. We identified two orthogonal processes in Kras-mutant distal airway club cells. The first promoted their transdifferentiation into an AEC2-like state through NKX2.1, and the second controlled oncogenic transformation through the AP-1 complex. Our results suggest that neoplasms retain an epigenetic memory of their cell of origin through cell-type-specific transcription factors. Our analysis showed that a cross-tissue-conserved AP-1-dependent chromatin remodeling program regulates carcinogenesis.Item Open Access Epithelial cell plasticity: breaking boundaries and changing landscapes.(EMBO reports, 2021-07) Tata, Aleksandra; Chow, Ryan D; Tata, Purushothama RaoEpithelial tissues respond to a wide variety of environmental and genotoxic stresses. As an adaptive mechanism, cells can deviate from their natural paths to acquire new identities, both within and across lineages. Under extreme conditions, epithelial tissues can utilize "shape-shifting" mechanisms whereby they alter their form and function at a tissue-wide scale. Mounting evidence suggests that in order to acquire these alternate tissue identities, cells follow a core set of "tissue logic" principles based on developmental paradigms. Here, we review the terminology and the concepts that have been put forward to describe cell plasticity. We also provide insights into various cell intrinsic and extrinsic factors, including genetic mutations, inflammation, microbiota, and therapeutic agents that contribute to cell plasticity. Additionally, we discuss recent studies that have sought to decode the "syntax" of plasticity-i.e., the cellular and molecular principles through which cells acquire new identities in both homeostatic and malignant epithelial tissues-and how these processes can be manipulated for developing novel cancer therapeutics.Item Open Access Epithelial-mesenchymal transitions and hepatocarcinogenesis.(J Clin Invest, 2010-04) Jou, Janice; Diehl, Anna MaeEpithelial-mesenchymal transitions (EMTs) are believed to play a role in invasion and metastasis of many types of tumors. In this issue of the JCI, Chen et al. show that a gene that has been associated with aggressive biology in hepatocellular carcinomas initiates a molecular cascade that results in EMT.Item Open Access Evaluation of an epithelial plasticity biomarker panel in men with localized prostate cancer.(Prostate Cancer Prostatic Dis, 2016-03) Armstrong, AJ; Healy, P; Halabi, S; Vollmer, R; Lark, A; Kemeny, G; Ware, K; Freedland, SJBACKGROUND: Given the potential importance of epithelial plasticity (EP) to cancer metastasis, we sought to investigate biomarkers related to EP in men with localized prostate cancer (PC) for the association with time to PSA recurrence and other clinical outcomes after surgery. METHODS: Men with localized PC treated with radical prostatectomy at the Durham VA Medical Center and whose prostatectomy tissues were included in a tissue microarray (TMA) linked to long-term outcomes. We performed immunohistochemical studies using validated antibodies against E-cadherin and Ki-67 and mesenchymal biomarkers including N-cadherin, vimentin, SNAIL, ZEB1 and TWIST. Association studies were conducted for each biomarker with baseline clinical/pathologic characteristics an risk of PSA recurrence over time. RESULTS: Two hundred and five men contributed TMA tissue and had long-term follow-up (median 11 years). Forty-three percent had PSA recurrence; three died of PC. The majority had high E-cadherin expression (86%); 14% had low/absent E-cadherin expression. N-cadherin was rarely expressed (<4%) and we were unable to identify an E-to-N-cadherin switch as independently prognostic. No associations with clinical risk group, PSA recurrence or Gleason sum were noted for SNAIL, ZEB1, vimentin or TWIST, despite heterogeneous expression between patients. We observed an association of higher Ki-67 expression with Gleason sum (P=0.043), National Comprehensive Cancer Network risk (P=0.013) and PSA recurrence (hazard ratio 1.07, P=0.016). CONCLUSIONS: The expression of EP biomarkers in this cohort of men with a low risk of PC-specific mortality was not associated with aggressive features or PSA relapse after surgery.Item Open Access Ferroptotic stress promotes the accumulation of pro-inflammatory proximal tubular cells in maladaptive renal repair.(eLife, 2021-07-19) Ide, Shintaro; Kobayashi, Yoshihiko; Ide, Kana; Strausser, Sarah A; Abe, Koki; Herbek, Savannah; O'Brien, Lori L; Crowley, Steven D; Barisoni, Laura; Tata, Aleksandra; Tata, Purushothama Rao; Souma, TomokazuOverwhelming lipid peroxidation induces ferroptotic stress and ferroptosis, a non-apoptotic form of regulated cell death that has been implicated in maladaptive renal repair in mice and humans. Using single-cell transcriptomic and mouse genetic approaches, we show that proximal tubular (PT) cells develop a molecularly distinct, pro-inflammatory state following injury. While these inflammatory PT cells transiently appear after mild injury and return to their original state without inducing fibrosis, after severe injury they accumulate and contribute to persistent inflammation. This transient inflammatory PT state significantly downregulates glutathione metabolism genes, making the cells vulnerable to ferroptotic stress. Genetic induction of high ferroptotic stress in these cells after mild injury leads to the accumulation of the inflammatory PT cells, enhancing inflammation and fibrosis. Our study broadens the roles of ferroptotic stress from being a trigger of regulated cell death to include the promotion and accumulation of proinflammatory cells that underlie maladaptive repair.Item Open Access Histopathologic assessment of cultured human thymus.(PloS one, 2020-01) Hale, Laura P; Neff, Jadee; Cheatham, Lynn; Cardona, Diana; Markert, M Louise; Kurtzberg, JoanneThe maintenance and propagation of complex mixtures of cells in vitro in the form of native organs or engineered organoids has contributed to understanding mechanisms of cell and organ development and function which can be translated into therapeutic benefits. For example, allogeneic cultured postnatal human thymus tissue has been shown to support production of naïve recipient T cells when transplanted into patients with complete DiGeorge anomaly and other genetic defects that result in congenital lack of a thymus. Patients receiving such transplants typically exhibit reversal of their immunodeficiency and normalization of their peripheral blood T cell receptor V-beta repertoire, with long-term survival. This study was designed to assess the histopathologic changes that occur in postnatal human thymus slices when cultured according to protocols used for transplanted tissues. Results showed that as thymic organ cultures progressed from days 0 through 21, slices developed increasing amounts of necrosis, increasing condensation of thymic epithelium, and decreasing numbers of residual T cells. The architecture of the thymic epithelial network remained generally well-preserved throughout the 21 days of culture, with focal expression of cytokeratin 14, a putative biomarker of thymic epithelial cells with long-term organ-repopulating potential. All organ slices derived from the same donor thymus closely resembled one another, with minor differences in size, shape, and relative content of cortex versus medulla. Similarly, slices derived from different donors showed similar histopathologic characteristics when examined at the same culture time point. Taken together, these results demonstrate that diagnostic criteria based on structural features of the tissue identifiable via hematoxylin and eosin staining and cytokeratin immunohistochemistry can be used to evaluate the quality of slices transplanted into patients with congenital athymia.Item Open Access PDGFRα+ pericryptal stromal cells are the critical source of Wnts and RSPO3 for murine intestinal stem cells in vivo.(Proceedings of the National Academy of Sciences of the United States of America, 2018-04) Greicius, Gediminas; Kabiri, Zahra; Sigmundsson, Kristmundur; Liang, Chao; Bunte, Ralph; Singh, Manvendra K; Virshup, David MWnts and R-spondins (RSPOs) support intestinal homeostasis by regulating crypt cell proliferation and differentiation. Ex vivo, Wnts secreted by Paneth cells in organoids can regulate the proliferation and differentiation of Lgr5-expressing intestinal stem cells. However, in vivo, Paneth cell and indeed all epithelial Wnt production is completely dispensable, and the cellular source of Wnts and RSPOs that maintain the intestinal stem-cell niche is not known. Here we investigated both the source and the functional role of stromal Wnts and RSPO3 in regulation of intestinal homeostasis. RSPO3 is highly expressed in pericryptal myofibroblasts in the lamina propria and is several orders of magnitude more potent than RSPO1 in stimulating both Wnt/β-catenin signaling and organoid growth. Stromal Rspo3 ablation ex vivo resulted in markedly decreased organoid growth that was rescued by exogenous RSPO3 protein. Pdgf receptor alpha (PdgfRα) is known to be expressed in pericryptal myofibroblasts. We therefore evaluated if PdgfRα identified the key stromal niche cells. In vivo, Porcn excision in PdgfRα+ cells blocked intestinal crypt formation, demonstrating that Wnt production in the stroma is both necessary and sufficient to support the intestinal stem-cell niche. Mice with Rspo3 excision in the PdgfRα+ cells had decreased intestinal crypt Wnt/β-catenin signaling and Paneth cell differentiation and were hypersensitive when stressed with dextran sodium sulfate. The data support a model of the intestinal stem-cell niche regulated by both Wnts and RSPO3 supplied predominantly by stromal pericryptal myofibroblasts marked by PdgfRα.Item Open Access IL-27 Facilitates Skin Wound Healing through Induction of Epidermal Proliferation and Host Defense.(The Journal of investigative dermatology, 2017-05) Yang, Bin; Suwanpradid, Jutamas; Sanchez-Lagunes, Roberto; Choi, Hae Woong; Hoang, Peter; Wang, Donghai; Abraham, Soman N; MacLeod, Amanda SSkin wound repair requires a coordinated program of epithelial cell proliferation and differentiation as well as resistance to invading microbes. However, the factors that trigger epithelial cell proliferation in this inflammatory process are incompletely understood. In this study, we demonstrate that IL-27 is rapidly and transiently produced by CD301b+ cells in the skin after injury. The functional role of IL-27 and CD301b+ cells is demonstrated by the finding that CD301b-depleted mice exhibit delayed wound closure in vivo, which could be rescued by topical IL-27 treatment. Furthermore, genetic ablation of the IL-27 receptor (Il27Ra-/-) attenuates wound healing, suggesting an essential role for IL-27 signaling in skin regeneration in vivo. Mechanistically, IL-27 feeds back on keratinocytes to stimulate cell proliferation and re-epithelialization in the skin, whereas IL-27 leads to suppression of keratinocyte terminal differentiation. Finally, we identify that IL-27 potently increases expression of the antiviral oligoadenylate synthetase 2, but does not affect expression of antibacterial human beta defensin 2 or regenerating islet-derived protein 3-alpha. Together, our data suggest a previously unrecognized role for IL-27 in regulating epithelial cell proliferation and antiviral host defense during the normal wound healing response.Item Open Access Intercellular Protein Transfer from Thymocytes to Thymic Epithelial Cells.(PLoS One, 2016) Wang, Hong-Xia; Qiu, Yu-Rong; Zhong, Xiao-PingPromiscuous expression of tissue restricted antigens (TRAs) in medullary thymic epithelial cells (mTECs) is crucial for negative selection of self-reactive T cells to establish central tolerance. Intercellular transfer of self-peptide-MHC complexes from mTECs to thymic dendritic cells (DCs) allows DCs to acquire TRAs, which in turn contributes to negative selection and regulatory T cell generation. However, mTECs are unlikely to express all TRAs, such as immunoglobulins generated only in B cells after somatic recombination, hyper-mutation, or class-switches. We report here that both mTECs and cortical TECs can efficiently acquire not only cell surface but also intracellular proteins from thymocytes. This reveals a previously unappreciated intercellular sharing of molecules from thymocytes to TECs, which may broaden the TRA inventory in mTECs for establishing a full spectrum of central tolerance.Item Open Access Lung Regeneration: Cells, Models, and Mechanisms.(Cold Spring Harbor perspectives in biology, 2022-10) Konkimalla, Arvind; Tata, Aleksandra; Tata, Purushothama RaoLung epithelium, the lining that covers the inner surface of the respiratory tract, is directly exposed to the environment and thus susceptible to airborne toxins, irritants, and pathogen-induced damages. In adult mammalian lungs, epithelial cells are generally quiescent but can respond rapidly to repair of damaged tissues. Evidence from experimental injury models in rodents and human clinical samples has led to the identification of these regenerative cells, as well as pathological metaplastic states specifically associated with different forms of damages. Here, we provide a compendium of cells and cell states that exist during homeostasis in normal lungs and the lineage relationships between them. Additionally, we discuss various experimental injury models currently being used to probe the cellular sources-both resident and recruited-that contribute to repair, regeneration, and remodeling following acute and chronic injuries. Finally, we discuss certain maladaptive regeneration-associated cell states and their role in disease pathogenesis.