Change, Source, and Effect of GDF11 in Mouse Spleen

Abstract

Growth differentiation factor 11 (GDF11) is a secreted ligand of the transforming growth factor-β (TGF-β) superfamily of proteins and regulates gene expression through AKT-MTORC1 pathway. GDF11 has been proposed as a “rejuvenating factor” because it can restore regenerative functions in old mice. Specifically, previous works have demonstrated that restoring GDF11 levels in old mice not only reverses aged-related cardiac hypertrophy and age-related skeletal muscle dysfunction but also improves cerebral vasculature and neurogenesis. Furthermore, GDF11 also has notable impacts on the immune system since TGF-β molecules are important drivers of T cell differentiation and the role of GDF11 has been implicated in many immune-related diseases. However, there is no absolute consensus regarding the direction of change with age of GDF11 levels due to technical difficulties in specifically distinguishing GDF11 from other closely related TGF-beta ligands and the importance of GDF11 in regulating immune cell populations is unknown. Therefore, utilizing the GDF11-IRES-GFP mouse model generated by Goldstein et al. (2019) and fluorescent immunohistochemistry, I planned to unequivocally show whether GFP level, thus the GDF11 level, rise or decline with age in mouse spleen. Moreover, I planned to use flow cytometry and co-staining of GFP with the vasculature, lymphatic vessel, and hematopoietic cell subsets to interrogate the major cell types that are responsible for GDF11 production in spleen and reveal the localization pattern of GDF11 in spleen. Finally, I planned to study the GDF11-WE mice that endogenously overexpress a functional variant of GDF11 protein and investigate whether increased level of GDF11 can “rejuvenate” both GDF11-WE mice and wild type old mice when a parabiosis surgery created a shared circulation system between them. However, I encountered technical difficulties in immunohistochemical staining of spleen that prevented me from obtaining valid results from these experiments and I was not able to solve the issue despite strong endeavors. Nevertheless, I analyzed how heterozygous knockout of GDF11 alters proportion of splenic immune cell populations and discovered that none of the percentages of 10 splenic immune cell types analyzed as well as lymphatic endothelial cells and fibroblastic reticular cells among live cells of GDF11 heterozygous knockout mice was significantly different from those of wild type littermates. Altogether, my originally planned research on the GDF11-IRES-GFP mice can demonstrate the direction of change of GDF11 with ageing and localization pattern of GDF11 in spleen. Investigation of GDF11-WE mice could serve an initial evaluation of the potential therapeutic value of GDF11 in anti-aging. Furthermore, the difference in percentages of certain immune cells between GDF11 heterozygous knockout mice and wild type mice provided a new perspective on GDF11’s effect on the immune system and encouraged further research about the mechanisms of GDF11’s influence on such particular splenocytes.