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1.
Commun Biol ; 5(1): 47, 2022 01 12.
Artigo em Inglês | MEDLINE | ID: mdl-35022507

RESUMO

Lysosomal membrane permeabilization (LMP) and cathepsin release typifies lysosome-dependent cell death (LDCD). However, LMP occurs in most regulated cell death programs suggesting LDCD is not an independent cell death pathway, but is conscripted to facilitate the final cellular demise by other cell death routines. Previously, we demonstrated that Caenorhabditis elegans (C. elegans) null for a cysteine protease inhibitor, srp-6, undergo a specific LDCD pathway characterized by LMP and cathepsin-dependent cytoplasmic proteolysis. We designated this cell death routine, lysoptosis, to distinguish it from other pathways employing LMP. In this study, mouse and human epithelial cells lacking srp-6 homologues, mSerpinb3a and SERPINB3, respectively, demonstrated a lysoptosis phenotype distinct from other cell death pathways. Like in C. elegans, this pathway depended on LMP and released cathepsins, predominantly cathepsin L. These studies suggested that lysoptosis is an evolutionarily-conserved eukaryotic LDCD that predominates in the absence of neutralizing endogenous inhibitors.


Assuntos
Antígenos de Neoplasias/genética , Morte Celular , Células Epiteliais/fisiologia , Serpinas/genética , Animais , Antígenos de Neoplasias/metabolismo , Linhagem Celular Tumoral , Humanos , Camundongos , Serpinas/metabolismo
2.
Cell Rep Med ; 2(6): 100320, 2021 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-34195684

RESUMO

Necrotizing enterocolitis (NEC) is a deadly intestinal inflammatory disorder that primarily affects premature infants and lacks adequate therapeutics. Interleukin (IL)-22 plays a critical role in gut barrier maintenance, promoting epithelial regeneration, and controlling intestinal inflammation in adult animal models. However, the importance of IL-22 signaling in neonates during NEC remains unknown. We investigated the role of IL-22 in the neonatal intestine under homeostatic and inflammatory conditions by using a mouse model of NEC. Our data reveal that Il22 expression in neonatal murine intestine is negligible until weaning, and both human and murine neonates lack IL-22 production during NEC. Mice deficient in IL-22 or lacking the IL-22 receptor in the intestine display a similar susceptibility to NEC, consistent with the lack of endogenous IL-22 during development. Strikingly, treatment with recombinant IL-22 during NEC substantially reduces inflammation and enhances epithelial regeneration. These findings may provide a new therapeutic strategy to attenuate NEC.


Assuntos
Enterocolite Necrosante/imunologia , Interleucinas/genética , Mucosa Intestinal/imunologia , Proteínas Recombinantes/farmacologia , Regeneração/imunologia , Animais , Animais Recém-Nascidos , Quimiocina CXCL1/genética , Quimiocina CXCL1/imunologia , Quimiocina CXCL2/genética , Quimiocina CXCL2/imunologia , Modelos Animais de Doenças , Enterocolite Necrosante/tratamento farmacológico , Enterocolite Necrosante/microbiologia , Enterocolite Necrosante/patologia , Microbioma Gastrointestinal/imunologia , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Recém-Nascido , Doenças do Recém-Nascido/imunologia , Doenças do Recém-Nascido/microbiologia , Doenças do Recém-Nascido/patologia , Recém-Nascido Prematuro , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Interleucinas/imunologia , Mucosa Intestinal/metabolismo , Mucosa Intestinal/microbiologia , Camundongos , Camundongos Knockout , Isoformas de Proteínas/genética , Isoformas de Proteínas/imunologia , Receptores de Interleucina/genética , Receptores de Interleucina/imunologia , Regeneração/genética , Transdução de Sinais , Desmame , Interleucina 22
3.
Immunohorizons ; 5(4): 193-209, 2021 04 27.
Artigo em Inglês | MEDLINE | ID: mdl-33906960

RESUMO

Necrotizing enterocolitis (NEC) causes significant morbidity and mortality in premature infants; therefore, the identification of therapeutic and preventative strategies against NEC remains a high priority. The ligand-dependent transcription factor aryl hydrocarbon receptor (AhR) is well known to contribute to the regulation of intestinal microbial communities and amelioration of intestinal inflammation. However, the role of AhR signaling in NEC is unclear. Experimental NEC was induced in 4-d-old wild-type mice or mice lacking AhR expression in the intestinal epithelial cells or AhR expression in CD11c+ cells (AhRΔCD11c) by subjecting animals to twice daily hypoxic stress and gavage feeding with formula supplemented with LPS and enteric bacteria. During NEC, compared with wild-type mice treated with vehicle, littermates treated with an AhR proligand, indole-3-carbinol, had reduced expression of Il1b and Marco, a scavenger receptor that mediates dendritic cell activation and the recognition and clearance of bacterial pathogens by macrophages. Furthermore, indole-3-carbinol treatment led to the downregulation of genes involved in cytokine and chemokine, as revealed by pathway enrichment analysis. AhR expression in the intestinal epithelial cells and their cre-negative mouse littermates were similarly susceptible to experimental NEC, whereas AhRΔCD11c mice with NEC exhibited heightened inflammatory responses compared with their cre-negative mouse littermates. In seeking to determine the mechanisms involved in this increased inflammatory response, we identified the Tim-4- monocyte-dependent subset of macrophages as increased in AhRΔCD11c mice compared with their cre-negative littermates. Taken together, these findings demonstrate the potential for AhR ligands as a novel immunotherapeutic approach to the management of this devastating disease.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Enterocolite Necrosante/tratamento farmacológico , Indóis/farmacologia , Mucosa Intestinal/metabolismo , Receptores de Hidrocarboneto Arílico/metabolismo , Animais , Animais Recém-Nascidos , Modelos Animais de Doenças , Enterocolite Necrosante/metabolismo , Enterocolite Necrosante/microbiologia , Enterocolite Necrosante/patologia , Humanos , Indóis/uso terapêutico , Interleucina-1beta/metabolismo , Mucosa Intestinal/microbiologia , Mucosa Intestinal/patologia , Macrófagos/metabolismo , Macrófagos/patologia , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Transdução de Sinais/efeitos dos fármacos
4.
Sci Rep ; 10(1): 3842, 2020 03 02.
Artigo em Inglês | MEDLINE | ID: mdl-32123209

RESUMO

The development and physiologic role of small intestine (SI) vasculature is poorly studied. This is partly due to a lack of targetable, organ-specific markers for in vivo studies of two critical tissue components: endothelium and stroma. This challenge is exacerbated by limitations of traditional cell culture techniques, which fail to recapitulate mechanobiologic stimuli known to affect vessel development. Here, we construct and characterize a 3D in vitro microfluidic model that supports the growth of patient-derived intestinal subepithelial myofibroblasts (ISEMFs) and endothelial cells (ECs) into perfused capillary networks. We report how ISEMF and EC-derived vasculature responds to physiologic parameters such as oxygen tension, cell density, growth factors, and pharmacotherapy with an antineoplastic agent (Erlotinib). Finally, we demonstrate effects of ISEMF and EC co-culture on patient-derived human intestinal epithelial cells (HIECs), and incorporate perfused vasculature into a gut-on-a-chip (GOC) model that includes HIECs. Overall, we demonstrate that ISEMFs possess angiogenic properties as evidenced by their ability to reliably, reproducibly, and quantifiably facilitate development of perfused vasculature in a microfluidic system. We furthermore demonstrate the feasibility of including perfused vasculature, including ISEMFs, as critical components of a novel, patient-derived, GOC system with translational relevance as a platform for precision and personalized medicine research.


Assuntos
Capilares/crescimento & desenvolvimento , Técnicas de Cocultura/instrumentação , Intestino Delgado/citologia , Dispositivos Lab-On-A-Chip , Miofibroblastos/citologia , Humanos , Miofibroblastos/metabolismo , Oxigênio/metabolismo , Perfusão
5.
Pediatr Res ; 88(1): 66-76, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-31242501

RESUMO

BACKGROUND: Preterm infants are susceptible to unique pathology due to their immaturity. Mouse models are commonly used to study immature intestinal disease, including necrotizing enterocolitis (NEC). Current NEC models are performed at a variety of ages, but data directly comparing intestinal developmental stage equivalency between mice and humans are lacking. METHODS: Small intestines were harvested from C57BL/6 mice at 3-4 days intervals from birth to P28 (n = 8 at each age). Preterm human small intestine samples representing 17-23 weeks of completed gestation were obtained from the University of Pittsburgh Health Sciences Tissue Bank, and at term gestation during reanastamoses after resection for NEC (n = 4-7 at each age). Quantification of intestinal epithelial cell types and messenger RNA for marker genes were evaluated on both species. RESULTS: Overall, murine and human developmental trends over time are markedly similar. Murine intestine prior to P10 is most similar to human fetal intestine prior to viability. Murine intestine at P14 is most similar to human intestine at 22-23 weeks completed gestation, and P28 murine intestine is most similar to human term intestine. CONCLUSION: Use of C57BL/6J mice to model the human immature intestine is reasonable, but the age of mouse chosen is a critical factor in model development.


Assuntos
Epitélio/crescimento & desenvolvimento , Regulação da Expressão Gênica no Desenvolvimento , Intestinos/crescimento & desenvolvimento , Animais , Enterocolite Necrosante/metabolismo , Epitélio/patologia , Receptores ErbB/metabolismo , Perfilação da Expressão Gênica , Homeostase , Humanos , Enteropatias/metabolismo , Intestinos/patologia , Camundongos , Camundongos Endogâmicos C57BL
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