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Immune clearance of senescent cells contributes to their control in aging tissues; however, it remains unclear how immunity against senescent cells is regulated in humans. Here, we provide a protocol to measure the cytotoxicity of autologous human skin immune cells against senescent dermal fibroblasts. We describe the ex vivo co-culture experiments using normal and senescent fibroblasts together with autologous immune cells isolated from the human skin. This protocol is applicable to understanding the mechanism of immune surveillance and clearance of senescent cells in humans. For complete details on the use and execution of this protocol, please refer to Hasegawa et al. 1 • Isolation of human dermal fibroblasts and generation of senescent fibroblasts • Isolation of skin immune cells from the same donor as fibroblasts • Establishment of a co-culture system of autologous human immune cells with fibroblasts • Evaluation of immune cell-mediated cytotoxicity against senescent fibroblasts Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. Immune clearance of senescent cells contributes to their control in aging tissues; however, it remains unclear how immunity against senescent cells is regulated in humans. Here, we provide a protocol to measure the cytotoxicity of autologous human skin immune cells against senescent dermal fibroblasts. We describe the ex vivo co-culture experiments using normal and senescent fibroblasts together with autologous immune cells isolated from the human skin. This protocol is applicable to understanding the mechanism of immune surveillance and clearance of senescent cells in humans.
Hasegawa et al. (Tue,) studied this question.