Research Project
10490214
PROJECT SUMMARY DNA damage response (DDR) and repair pathways are attenuated with aging. Accumulated damaged DNA, if left unrepaired, may disrupt tissue homeostasis, enabling an aging milieu favoring tissue degeneration, cancer, and metabolic dysfunction. Unrepaired DNA damage may also result in cell senescence, with senescence- associated secretory phenotype (SASP) promoting DNA damage and cell proliferation in neighboring cells, or in apoptosis. Non-pituitary growth hormone (npGH), synthesized locally in peripheral tissues, is recognized by widely expressed GH receptors (GHR), and acts through autocrine/paracrine mechanisms. Our preliminary results show that, in aged human tissues with high DNA damage, aged induced pluripotent stem cell (iPSC)- derived human colon 3D organoids, hypophysectomized rats lacking endocrine GH, and senescent cells, npGH is induced and npGH suppresses DDR, further reinforcing DNA damage. We propose a novel hypothesis supporting the role of local epithelial GH as an adverse determinant of age-associated DNA damage accumulation and as a marker of aging. We will assess npGH induction and DNA damage in aged normal human colon cells and aged colon organoids. We will explore mechanisms of npGH induction and assess DDR activity, senescence pathways, and DNA damage repair by blocking GHR signaling in vitro with pegvisomant, JAK2 inhibitor, and metformin, and in hypophysectomized rats devoid of pituitary GH. We will assess npGH actions in aged tissues derived from colon-specific and universal GHR knockout, and in GH overexpressing transgenic mice, and also assess colon epithelial copy number variation (CNV) and genes affected by CNV mutations in mice overexpressing the GH transgene. We will examine npGH expression in human colon tissue specimens derived from age-determined cohorts and assess spatial genomic profiles in conjunction with cells expressing npGH. We will study whether npGH is expressed in senescent cells, and whether it affects DNA damage and proliferation of adjacent cells. We will determine whether npGH, as a component of SASP, may serve as a specific marker of DNA-damage?induced senescence. We will test paracrine npGH effects on senescence in human Colon Intestine-Chip microfluidic devices co-cultured with human colon fibroblast npGH transfectants, and will confirm in vitro results in transgenic models overexpressing GH or in GHR knockout mice. We largely focus on validated human colon epithelial models, yet experiments in this proposal are designed to elucidate a universal mechanism whereby local npGH reinforces epithelial age-associated DNA damage leading to tissue degeneration.
