Senescent cells expose and secrete an oxidized form of membrane-bound vimentin as revealed by a natural polyreactive antibody

Abstract

Studying the phenomenon of cellular senescence has been hindered by the lack of senescence-specific markers. As such, detection of proteins informally associated with senescence accompanies the use of senescence-associated β-galactosidase as a collection of semiselective markers to monitor the presence of senescent cells. To identify novel biomarkers of senescence, we immunized BALB/c mice with senescent mouse lung fibroblasts and screened for antibodies that recognized senescence-associated cell-surface antigens by FACS analysis and a newly developed cell-based ELISA. The majority of antibodies that we isolated, cloned, and sequenced belonged to the IgM isotype of the innate immune system. In-depth characterization of one of these monoclonal, polyreactive natural antibodies, the IgM clone 9H4, revealed its ability to recognize the intermediate filament vimentin. By using 9H4, we observed that senescent primary human fibroblasts express vimentin on their cell surface, and MS analysis revealed a posttranslational modification on cysteine 328 (C328) by the oxidative adduct malondialdehyde (MDA). Moreover, elevated levels of secreted MDA-modified vimentin were detected in the plasma of aged senescence-accelerated mouse prone 8 mice, which are known to have deregulated reactive oxygen species metabolism and accelerated aging. Based on these findings, we hypothesize that humoral innate immunity may recognize senescent cells by the presence of membrane-bound MDA-vimentin, presumably as part of a senescence eradication mechanism that may become impaired with age and result in senescent cell accumulation.aging | oxidative posttranslational modifications | biomarker | SAMP8 | malondialdehyde

Senescent cells: an emerging target for diseases of ageing

“…For example, oxidized vimentin 157 , DCR2 (REF. 158 ), density-enhanced phosphatase 1 (DEP1; also known as PTPRJ) 159 and β2 microglobulin (B2MG) 159 are overexpressed at the SNC cell surface and could be used for directing local accumulation of senolytics.…”

Plant defensin PvD1 modulates the membrane composition of breast tumour-derived exosomes

“…66 The increased zeta potential of MCF-7 cells does not likely arise from the interaction with PvD 1 only, but also from the secretion of senescence-associated-secretoryphenotype (SASP) factors, including vimentin. This protein is evidenced to accumulate on the surface of senescent cells 67 and is known to have preferential and strong affinity to negatively charged phospholipids. 68 Therefore, vimentin, when presented on the cell surface of senescent cells, associates with anionic POPS and phosphatidylinositol (PI) shielding their negative charge and thus contributing to the partial neutralization of the membrane charge density.…”

Isolation of senescent cells by iodixanol (OptiPrep) density gradient‐based separation

“…Recent high-throughput approaches to isolate senescent cells from tissues or body fluids, based on their increased cell size, appear more scalable, robust and specific than the ones based on markers that are specific to senescent cells. 6,7,[10][11][12][13] Our optimized iodixanol gradient-based centrifugation protocol to isolate DXR-induced senescent HCC cells is simple, quick (~1 hour) and robust. Our protocol is also unique because, unlike previous approaches, it permits cells to be re-plated on collagencoated flasks, despite their low or null proliferative capacity.…”

“…Attempts to isolate senescent cells using surface markers (DcR2, DPP4, oxidized vimentin) or fluorescent ubiquitination‐based cell‐cycle indicator technology have shown promise. However, simple methods to isolate and culture senescent cells—despite their low or null proliferative capacity—are lacking, particularly in the context of chemotherapy‐induced senescence for cancer treatment.…”