{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,1,15]],"date-time":"2026-01-15T19:46:59Z","timestamp":1768506419437,"version":"3.49.0"},"posted":{"date-parts":[[2023,8,6]]},"group-title":"Pathology","reference-count":50,"publisher":"openRxiv","content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2023,8,6]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:sec>\n                  <jats:title>Background<\/jats:title>\n                  <jats:p>\n                    Retinal pigment epithelial cells (RPE) play vital role in the pathogenesis of age-related macular degeneration (AMD). Our laboratory has shown that RPE cellular senescence contributed to the pathophysiology of experimental AMD, and SASP members are involved in this process. Recently, we presented confirmatory evidence to earlier GWAS studies that dysregulation of tumor necrosis factor receptor superfamily 10A (TNFRSF10A) dysregulation leads to AMD development and is linked to RPE dysfunction. This study aims to investigate the contribution of RPE senescence to AMD pathophysiology using\n                    <jats:italic>TNFRSF10A<\/jats:italic>\n                    silenced human RPE (hRPE) cells and Tnfrsf10 KO mice.\n                  <\/jats:p>\n                <\/jats:sec>\n                <jats:sec>\n                  <jats:title>Methods<\/jats:title>\n                  <jats:p>\n                    Sub-confluent primary hRPE cells and\n                    <jats:italic>TNFRSF10A<\/jats:italic>\n                    silenced hRPE were exposed to stress-induced premature senescence with H2O2 (500 \u03bcM, 48h), and senescence-associated markers (\u03b2gal, p16, and p21) were analyzed by RT-PCR and WB analysis. The effect of H2O2-induced senescence in non-silenced and silenced hRPE on OXPHOS and glycolysis was determined using Seahorse XF96 analyzer. Male C57BL\/6J Tnfrsf10 KO (\n                    <jats:italic>\n                      Tnfrsf10\n                      <jats:sup>-\/-<\/jats:sup>\n                    <\/jats:italic>\n                    ) mice were used to study the regulation of senescence by TNFRSF10A\n                    <jats:italic>in vivo<\/jats:italic>\n                    . Expression of p16 and p21 in control and KO mice of varying ages were determined by RT-PCR, WB, and immunostaining analysis.\n                  <\/jats:p>\n                <\/jats:sec>\n                <jats:sec>\n                  <jats:title>Results<\/jats:title>\n                  <jats:p>\n                    The senescence-associated p16 and p21 showed a significant (\n                    <jats:italic>p<\/jats:italic>\n                    &lt; 0.01) upregulation with H2O2 induction at the gene (1.8- and 3-fold) and protein (3.2- and 4-fold) levels in hRPE cells. The protein expression of p16 and p21 was further significantly increased by co-treatment with siRNA (\n                    <jats:italic>p<\/jats:italic>\n                    &lt; 0.05 vs. H2O2). Mitochondrial oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) (pmol\/min\/total DNA) increased with senescence induction by H2O2 for 48h in control RPE, and knockdown of\n                    <jats:italic>TNFRSF10A<\/jats:italic>\n                    caused a further increase in OCR and ECAR. In addition, co-treatment with PKC activator significantly improved all parameters. Similarly,\n                    <jats:italic>in vivo<\/jats:italic>\n                    studies showed upregulation of p16 and p21 by RT-PCR, WB, and immunostaining analysis in RPE\/choroid of Tnfrsf10 KO mice. When subjected to examination across distinct age groups, namely young (1-3 months), middle (6-9 months), and old (12-15 months) mice, a discernible age-related elevation in the expression of p16 and p21 was observed.\n                  <\/jats:p>\n                <\/jats:sec>\n                <jats:sec>\n                  <jats:title>Conclusions<\/jats:title>\n                  <jats:p>Our findings suggest that TNRSF10A is a regulator of regulates in RPE senescence. Further work on elucidating pathways of senescence will facilitate the development of new therapeutic targets for AMD.<\/jats:p>\n                <\/jats:sec>","DOI":"10.1101\/2023.08.04.552052","type":"posted-content","created":{"date-parts":[[2023,8,6]],"date-time":"2023-08-06T13:55:10Z","timestamp":1691330110000},"source":"Crossref","is-referenced-by-count":0,"title":["Characterization and contribution of RPE senescence to Age-related macular degeneration in\n                  <i>Tnfrsf10<\/i>\n                  knock out mice"],"prefix":"10.64898","author":[{"given":"Iori","family":"Wada","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Kenichiro","family":"Mori","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Parameswaran G","family":"Sreekumar","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Rui","family":"Ji","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Christine","family":"Spee","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Elise","family":"Hong","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Keijiro","family":"Ishikawa","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Koh-Hei","family":"Sonoda","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ram","family":"Kannan","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"54368","reference":[{"issue":"6","key":"2023080901551074000_2023.08.04.552052v1.1","doi-asserted-by":"crossref","first-page":"539","DOI":"10.1056\/NEJMcp2102061","article-title":"Age-Related Macular Degeneration","volume":"385","year":"2021","journal-title":"N Engl J Med"},{"key":"2023080901551074000_2023.08.04.552052v1.2","doi-asserted-by":"publisher","DOI":"10.1016\/S0140-6736(18)31550-2"},{"key":"2023080901551074000_2023.08.04.552052v1.3","doi-asserted-by":"publisher","DOI":"10.1038\/ng.3448"},{"key":"2023080901551074000_2023.08.04.552052v1.4","doi-asserted-by":"publisher","DOI":"10.1038\/ng.938"},{"issue":"4","key":"2023080901551074000_2023.08.04.552052v1.5","doi-asserted-by":"crossref","first-page":"361","DOI":"10.1016\/j.ophtha.2022.10.034","article-title":"Genome-Wide Association Study of Age-Related Macular Degeneration Reveals 2 New Loci Implying Shared Genetic Components with Central Serous Chorioretinopathy","volume":"130","year":"2023","journal-title":"Ophthalmology"},{"issue":"13","key":"2023080901551074000_2023.08.04.552052v1.6","doi-asserted-by":"crossref","first-page":"2194","DOI":"10.1093\/hmg\/ddac020","article-title":"TNFRSF10A downregulation induces retinal pigment epithelium degeneration during the pathogenesis of age-related macular degeneration and central serous chorioretinopathy","volume":"31","year":"2022","journal-title":"Hum Mol Genet"},{"issue":"6","key":"2023080901551074000_2023.08.04.552052v1.7","doi-asserted-by":"crossref","first-page":"2809","DOI":"10.1007\/s11357-021-00457-4","article-title":"Targeting senescent retinal pigment epithelial cells facilitates retinal regeneration in mouse models of age-related macular degeneration","volume":"43","year":"2021","journal-title":"Geroscience"},{"key":"2023080901551074000_2023.08.04.552052v1.8","doi-asserted-by":"publisher","DOI":"10.1016\/j.exer.2021.108918"},{"key":"2023080901551074000_2023.08.04.552052v1.9","doi-asserted-by":"publisher","DOI":"10.1016\/S0008-4182(04)80054-1"},{"key":"2023080901551074000_2023.08.04.552052v1.10","doi-asserted-by":"publisher","DOI":"10.1523\/JNEUROSCI.3580-11.2011"},{"key":"2023080901551074000_2023.08.04.552052v1.11","doi-asserted-by":"crossref","unstructured":"P.G. 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