Project?Summary/Abstract? ? Hirschsprung?s? disease? (HSCR)? is? a? common? cause? of? neonatal? bowel? obstruction? that? results? from? failed? development? of? the? distal? enteric? nervous? system? (ENS).? The? pathophysiology,? approach? to? diagnosis? and? surgical?treatment?of?HSCR?has?been?well-?defined?over?the?last?60?years.?However,?Hirschsprung?s-?associated? enterocolitis? (HAEC),? the? most? serious? complication? of? HSCR,? remains? a? frequent? cause? of? pre-?? and? post-? operative? morbidity? and? mortality? in? HSCR? patients,? with? poorly? defined? pathophysiology? and? unchanged? treatment?guidelines?over?multiple?decades.?HAEC?is?the?presenting?symptom?of?HSCR?in?25%?of?infants?and? mortality?ranges?from?1-?10%,?with?the?majority?of?deaths?occurring?in?newborns?prior?to?definitive?operation.?Our? central?hypothesis?is?that?developmental?deficiency?of?the?ENS?in?HSCR?is?accompanied?by?defects?in?intestinal? homeostasis,?leading?to?increased?susceptibility?to?HAEC.?Advances?in?understanding?the?genetics?underlying? HSCR?have?allowed?development?of?animal?models?which?recapitulate?the?human?HAEC?phenotype.?Over?the? last? five? years,? studies? from? our? laboratory? utilizing? the? EdnrBNCC-?/-?? model? of? HSCR/HAEC? have? allowed? us? to? identify? defects? in? intestinal? homeostasis? that? contribute? to? HAEC.? We? have? identified? ENS? alterations? in? the? ?normal?,? ganglionated? colon? of? EdnrBNCC-?/-?? mice? which? include? decreased? neuronal? density? and? a? shift? in? neurotransmitter?phenotypes,?with?over-?representation?of?relaxation?neurotransmitters?and?under-?representation? of? contractility? neurotransmitters.? Additionally,? we? have? demonstrated? that? EdnrBNCC-?/-?? mice? develop? colonic? dysbiosis?prior?to?HAEC.?Most?recently,?preliminary?data?indicate?alterations?in?colonic?epithelial?tight?junctions? preceding?the?development?of?HAEC.?Furthermore,?EdnrBNCC-?/-??intestinal?segments?have?increased?susceptibility? to?invasion?by?pathogenic?E.coli,?with?decreased?production?of?innate?immune?defense?molecules.?Finally,?we? have? identified? impaired? cellular? immunity? (B-?cell? populations? and? decreased? secretory? IgA)? in? these? animals.? Taken?together,?intestinal?homeostasis?in?EdnrBNCC-?/-??mice?appears?to?governed?by?a?four-?way?interplay?between? the?enteric?nervous?system,?luminal?microbiota,?the?intestinal?epithelial?barrier,?and?the?mucosal?immune?system,? all?of?which?appear?to?be?perturbed?in?the?setting?of?altered?host?genetics.?However,?our?studies?to?date?have?not? allowed?us?to?distinguish?which?alterations?in?intestinal?homeostasis?are?causative?versus?associated?with?HAEC.? Recently,? intestinal? enteroids? have? emerged? as? a? tool? to? model? the? complexity? of? the? intestinal? epithelium? in? culture,?avoiding?confounding?influence?from?local?and?system?non-?epithelial?sources.?In?order?to?advance?our? understanding? of? the? complex? dynamics? that? contribute? to? HAEC,? we? propose? to? develop? an? experimental? paradigm? of? in? vivo,? ex? vivo? and? in? vitro? methods? that? will? allow? combinatorial? testing? of? the? components? of? intestinal?homeostasis.?This?proposal?will?evaluate?the?contributions?of?EdnrBNCC-?/-??microbiome?dysbiosis?and? epithelial?barrier?dysfunction?to?the?development?of?HAEC?and?will?determine?the?suitability?of?intestinal?enteroids? as?a?novel?system?to?study?the?pathogenesis?of?HAEC.?

R03DK114543

2017-08-15

2020-07-31