Senolytic-Resistant Senescent Cells Have a Distinct SASP Profile and Functional Impact: The Path to Developing Senosensitizers

The senescent cell (SC) fate is linked to aging, multiple disorders and diseases, and physical dysfunction. Senolytics, agents that selectively eliminate 30-70% of SCs, act by transiently disabling the senescent cell anti-apoptotic pathways (SCAPs), which defend those SCs that are pro-apoptotic from their own senescence-associated secretory phenotype (SASP). Consistent with this, a JAK/STAT inhibitor, Ruxolitinib, which attenuates the pro-apoptotic SASP of senescent human preadipocytes, caused them to become “senolytic-resistant”. Administering senolytics to obese mice selectively decreased abundance of the subset of SCs that is pro-inflammatory. In cell cultures, the 30-70% of human senescent preadipocytes or human umbilical vein endothelial cells (HUVECs) that are senolytic-resistant (to Dasatinib or Quercetin, respectively) had increased p16INK4a, p21CIP1, senescence-associated ß-galactosidase (Saßgal), ?H2AX, and proliferative arrest similar to the total SC population (comprising senolytic-sensitive plus -resistant SCs). However, the SASP of senolytic-resistant SCs entailed less pro-inflammatory/apoptotic factor production, induced less inflammation in non-senescent cells, and was equivalent or richer in growth/ fibrotic factors. Senolytic-resistant SCs released less mitochondrial DNA (mtDNA) and more highly expressed the anti-inflammatory, immune evasion signal, glycoprotein non-melanoma-B (GPNMB). Transplanting senolytic-resistant SCs intraperitoneally into younger mice caused less physical dysfunction than transplanting the total SC population. Since Ruxolitinib attenuates SC release of pro-apoptotic SASP factors while pathogen-associated molecular pattern factors (PAMPs) can amplify release of these factors rapidly (acting as “senosensitizers”), senolytic-resistant and senolytic-sensitive SCs appear to be interconvertible. Overall design: Preadipocytes (also known as adipose-derived stromal cells or adipose mesenchymal “stem” cells [MSCs]) were isolated from abdominal subcutaneous fat biopsies obtained from subjects donating kidneys for transplantation. In human preadipocytes, senescence was induced by 20 Gy X-irradiation and experiments were performed 30 days after radiation. Cells were treated with DMSO or 800 nM dasatinib and incubated for 24 hrs. Next, the remaining cells were washed with PBS 4X and subsequently incubated 3 days and analyzed via RNA SEQ