Short‐term Inflammation Increases Proliferative Capacity of Human Skeletal Muscle Progenitor Cells from Young and Old Female Donors

Rationale While short‐term inflammation is required for myogenic progenitor cell (MPC) expansion following damage, chronic or dysregulation of inflammation impairs regenerative capacity. In old (vs. young) skeletal muscle (SkM), the inflammatory response to damage is elevated, which could underlie impaired SkM regeneration observed in older adults. We previously observed that MPCs from older female donors had increased confluency at 240 h in growth medium when proinflammatory cytokine, TNFα, was administered at 96h and maintained for 48 hours (short‐term); this response was not observed when TNFα was administered at earlier or later time points. However, it was unclear whether increased confluence was due to increased proliferation, increased protein synthesis, decreased cell death or some combination of these three. Further, it was unclear whether this response was specific to older MPCs. Objectives The objective was to examine the effects of short‐term TNFα treatment on the proliferative capacity of young and old human female MPCs. Methods Primary human MPCs were harvested from SkM biopsies taken from the vastus lateralis of healthy young females (n=6) and healthy old females (n=6). MPCs were cultured in growth medium [(GM), Ham's F12, 20% FBS, 5ng/ml bFGF] for a total of 240 hours. From t=96 to 144 hours a subset of cells were exposed to 10ng/ml TNFα followed by GM (t=144 to 240 hour). % confluence, # of nuclei, and % cell death were tracked using the Celigo® Cell Imaging Cytometer from seeding (t=0) to t=240 hours. Cell cycle phase was determined using BrdU incorporation and Hoechst staining at t=96, 144, 192 hours. qPCR was used to measure the expression of select genes associated with inflammation and myogenesis. Results Net area under the confluence curve was significantly increased in TNFα treated cells of only young females at t=144, and of both old and young females at t=192 and t=240 (p<0.05). This was mostly attributed to increased nuclei count, which was increased at t=192 in both young and old MPCs However, old MPCs also had reduced cell death (p<0.05). Further supporting MPC proliferation and potentially commitment in young MPCs, the G2/M phase of the cell cycle was elevated with TNFα treatment at t=144 (p<0.05). Independent of age, TNFα increased the percentage of cells in G0/G1 and reduced the percentage in S phase of the cell cycle at t=240 (p<0.05). Additionally, TNFα treatment increased IL6 expression and reduced IL6 receptor (IL6r) and MyoD expression at t=192 hours (p<0.05) in both older and younger females. Older females had reduced IL6r compared to younger cells at t=192 (p<0.05). Conclusions Short‐term inflammation promotes progenitor cell expansion and potentially commitment to the myoblast lineage, independent of the age of the MPC donor. Intriguingly the responses to short‐term inflammation may be delayed in MPCs from the older female donors. Support or Funding Information Cornell University