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Senescence Rewires Microenvironment Sensing Kom Yooj Yim Antitumor Immunity

Dec 15, 2023

TSAB NTAWV

Cellular senescence cuam tshuam nrog kev ruaj khov ntawm lub voj voog kaw ua ke nrog cov txheej txheem zais cia uas, qee zaus, txhawb kev tiv thaiv kab mob ntawm cov hlwb senescent. Siv tus qauv tiv thaiv kab mob qog noj ntshav uas muaj peev xwm ua rau mob qog noj ntshav ua rau CD8 T cell-mediated qog rejection, peb qhia tias senescence kuj hloov kho lub cell-nto proteome los hloov cov qog hlwb paub txog tej yam hauv ib puag ncig, raws li piv txwv los ntawm hom II interferon (IFN ). Piv nrog rau cov cell proliferating, senescent hlwb upregulate IFN receptor, ua hypersensitized rau microenvironmental IFN, thiab ntau robustly induce antigen-presenting machinery-ffects kuj recapitulated nyob rau hauv tib neeg qog hlwb nyob rau hauv txoj kev kho-induced senescence. Kev cuam tshuam ntawm IFN sensing nyob rau hauv senescent hlwb blunts lawv lub cev tiv thaiv kab mob kev tshem tawm yam tsis muaj kev cuam tshuam lub senescence xeev los yog nws cov yam ntxwv secretory kev pab cuam. Peb cov txiaj ntsig tau pom tias cov hlwb senescent muaj peev xwm txhim kho ob qho tib si xa thiab tau txais cov cim qhia ib puag ncig thiab qhia tias txhua tus txheej txheem yuav tsum tau ua rau lawv txoj kev tiv thaiv kab mob zoo.

Desert ginseng-Improve immunity (15)

cistanche cog-nce kev tiv thaiv kab mob

Nyem qhov no mus saib Cistanche Enhance Immunity khoom

【Nug ntxiv】 Email: cindy.xue@wecistanche.com / Whats App: 0086 18599088692 / Wechat: 18599088692

CEEB TOOM:

Peb txoj haujlwm nthuav tawm kev sib cuam tshuam ntawm kev hloov kho cov ntaub so ntswg thiab cov ntaub so ntswg-sensing cov kev pab cuam uas tuaj yeem koom nrog los ntawm senescence hauv cov qog noj ntshav siab kom ua rau cov qog hlwb pom ntau dua rau lub cev tiv thaiv kab mob. Qhov tshiab facet ntawm senescence tsim reciprocal heterotypic signaling kev sib cuam tshuam uas tuaj yeem raug ntxias kho mob los txhim kho kev tiv thaiv kab mob.

Taw qhia

Cellular senescence yog ib qho kev pabcuam kev ntxhov siab uas muaj lub cev ruaj khov ntawm lub voj voog kaw thiab cov txheej txheem zais cia muaj peev xwm hloov kho cov ntaub so ntswg ib puag ncig (1). Nyob rau hauv cov ntaub so ntswg ib txwm, senescence pab rau cov ntaub so ntswg homeostasis thaum lub qhov txhab kho; Txawm li cas los xij, nyob rau hauv cov laus los yog cov ntaub so ntswg puas, lub aberrant tsub zuj zuj ntawm senescent hlwb yuav ua rau mob o thiab txo cov ntaub so ntswg regenerative muaj peev xwm (2-4). Hauv kev mob qog noj ntshav, kev laus tau pom tias yuav kho tau ob qho tib si muaj txiaj ntsig thiab cuam tshuam rau cov ntaub so ntswg biology. Ntawm ib sab tes, senescence muab kev cuam tshuam rau oncogene-initiated tumorigenesis thiab pab txhawb rau kev ua haujlwm antitumor ntawm qee qhov kev kho mob qog noj ntshav (5, 6). Ntawm qhov tod tes, qhov kev pheej hmoo ntawm cov qog qog hlwb tom qab kho tuaj yeem tsim cov ntaub so ntswg ib puag ncig uas txhawb nqa rov qab thiab metastasis (7, 8). Cov molecular underpinnings ntawm cov opposing biological outputs tseem tsis to taub. Ib lub ntsej muag ntawm qhov kev pab cuam senescence uas yuav ua rau muaj kev sib txawv ntawm biology yog cov senescence-associated secretory phenotype (SASP; ref. 9). SASP yog qhib los ntawm lub ntiaj teb no chromatin remodeling txheej txheem uas evolved thiab tswj los ntawm tseem ceeb epigenetic regulators xws li BRD4 thiab proinflammatory transcription yam xws li NF-κB thiab C/EBP- (10–12). Qhov no, nyob rau hauv lem, ua rau induction ntawm cov noob uas encode cov ntaub so ntswg remodeling proteins xws li matrix metalloproteinases, loj hlob yam, thiab fibrinolytic yam uas paub ua si lub luag hauj lwm tseem ceeb nyob rau hauv lub qhov txhab kho (3, 13, 14). Lwm yam SASP cov khoom suav nrog cov tshuaj chemokines thiab cytokines uas tuaj yeem hloov pauv qhov muaj pes tsawg leeg thiab lub xeev ntawm lub cev tiv thaiv kab mob hauv cov ntaub so ntswg, ua rau lub cev tiv thaiv kab mob thiab kev tshem tawm ntawm cov hlwb senescent lawv tus kheej (15, 16). Txawm li cas los xij, qhov tsis txaus ntseeg ntawm cov hlwb senescent nyob rau hauv ntau yam kab mob pathologic txhais tau hais tias kev tiv thaiv kab mob hauv lub cev tsis yog qhov tshwm sim thoob ntiaj teb ntawm senescence lossis SASP thiab ua rau muaj peev xwm ntxiv cov txheej txheem tswj hwm cov txiaj ntsig tsis zoo thiab cuam tshuam ntawm senescence hauv cov ntaub so ntswg tiv thaiv kab mob. (17–19).

Muaj tseeb tiag, kev soj ntsuam kev tiv thaiv kab mob tsis muaj zog tuaj yeem muaj peev xwm tiv thaiv qog noj ntshav, txawm hais tias cov txheej txheem cuam tshuam meej sib txawv nrog cov ntaub so ntswg thiab hom cell (10, 15, 16, 20). Hauv cov qauv nas ntawm hepatocellular carcinoma (HCC), daim siab qog hlwb ua rau lub cev raug tshem tawm los ntawm kev tiv thaiv kab mob los ntawm cov tsiaj qus (WT) p53 (15). Hauv kev pom zoo, TP53 feem ntau hloov pauv hauv tib neeg HCC, tshwj xeeb tshaj yog nyob rau hauv cov qog nqaij hlav "proliferation class" uas qhia txog qhov tsis zoo (21, 22). Txawm hais tias kev siv tshuaj tiv thaiv kab mob thiab TP53-cov tshuaj tiv thaiv tau tshwm sim los ua cov tswv yim zoo los txhim kho cov txiaj ntsig ntawm tus kab mob, lub hauv paus molecular rau cov lus teb thiab kev tiv thaiv tseem tsis paub (23–25). Yog li ntawd, kev nkag siab txog cov txheej txheem uas ua rau cov qog nqaij hlav hauv lub siab tuaj yeem pom ntawm lub cev tiv thaiv kab mob tuaj yeem pab txhawb cov tswv yim los txhawb kev tiv thaiv kab mob hauv TP53-hloov HCC uas yuav txuas mus rau lwm hom qog.

Ntawm no, peb tau teeb tsa los tsim cov hauv paus ntsiab lus uas hloov kho lub cev tiv thaiv kab mob thiab tshem tawm cov hlwb senescent los txheeb xyuas cov txheej txheem ua kom muaj zog uas yuav raug siv los txhim kho kev tiv thaiv kab mob qog noj ntshav. Txog rau qhov kawg no, peb tau tsim ib qho tshiab "senescence-inducible" qauv nyob rau hauv uas lub siab mob qog noj ntshav tuaj yeem xaiv tau hloov mus rau lub xeev senescent los ntawm kev hloov kho caj ces ntawm endogenous p53. Peb xav tias qhov no yuav ua raws li cov txiaj ntsig ntawm kev kho mob uas ua rau muaj qhov tsis zoo (26, 27) thaum zam qhov tsis txaus ntseeg ntawm cov kev kho mob senescence-inducing kho ntawm lub cev tiv thaiv kab mob lossis lwm yam khoom ntawm cov ntaub so ntswg ib puag ncig. Siv cov qauv no thiab txuas ntxiv mus rau lwm lub tshuab, peb nthuav tawm tias, ntxiv rau SASP, kev ua kom muaj zog ua rau muaj kev hloov pauv loj ntawm cov cell-surface proteome thiab cov phiaj xwm teeb pom kev zoo raws li kev kwv yees los hloov pauv txoj kev hlwb thiab teb rau ib puag ncig. Cov teeb liab, piv txwv ntawm no los ntawm kev kub siab rau microenvironmental hom II IFN (IFN ). Cov txheej txheem no ua rau muaj kev txhim kho muaj zog dua ntawm cov txheej txheem antigen thiab nthuav tawm cov tshuab hauv cov qog hlwb uas ua rau lawv muaj kev tiv thaiv kab mob hauv vivo. Yog li, peb cov txiaj ntsig tau nthuav tawm cov txheej txheem rov ua dua cov ntaub so ntswg hauv cov hlwb uas ua haujlwm hauv kev hais kwv txhiaj nrog SASP los txhawb lawv lub peev xwm tiv thaiv kab mob, yog li pab txhawb kev tiv thaiv qog noj ntshav.

Desert ginseng-Improve immunity (2)

cistanche tubulosa- txhim kho lub cev tiv thaiv kab mob

TSEEM CEEB

Ib p53-Restorable Immunocompetent Tumor Model to Study Senescence Surveillance

Txhawm rau kawm yuav ua li cas senescence reprograms cellular thiab cov ntaub so ntswg hauv lub xeev, peb siv cov txheej txheem hydrodynamic tail-vein txhaj (HTVI) (28) los tsim cov qauv qog nqaij hlav-inducible daim siab uas tswj los ntawm cov qog nqaij hlav, kho tau p53 luv luv hairpin RNA (shRNA). Tshwj xeeb, cov neeg laus daim siab hepatocytes ntawm immunocompetent Bl/6 nas tau kis hauv vivo nrog kev pw tsaug zog zoo nkauj SB13 transposase vector thiab ob lub transposon tsim (encoding NrasG12D-IRES-rtTA thiab TRE-tRFP-shp53, lossis "NSP") uas koom ua ke hauv genome . Nyob rau hauv no Tet-On system, endogenous p53 yog suppressed nyob rau hauv lub xub ntiag ntawm doxycycline (Dox) los ntawm kev ua kom inducible shRNA txuas rau RFP (Fig. 1A), ua rau cov noob caj noob ces tswj senescence nyob rau hauv tsim cov qog. Raws li kev sib koom ua ke ntawm kev hloov pauv uas tsis ua haujlwm TP53 thiab ua kom cov xovtooj ntawm tes nthuav qhia txoj hauv kev (piv txwv li, PI3K / AKT thiab RAS / MAPK cascades) hauv tib neeg lub siab qog, kev koom tes ntawm oncogenic RAS thiab kev tawm tsam ntawm p53 coj mus rau hepatocyte transformation, nrog rau feem ntau. nas tsim cov qog uas tsis zoo sib txawv ntawm 5 mus rau 8 lub lis piam tom qab HTVI. Transcriptional profiling qhia tau hais tias cov qog murine zoo ib yam li cov chav kawm "proliferation" ntawm tib neeg HCC (Supplementary Fig. S1A–S1F), uas yog cov chav kawm ntawm tib neeg HCC harboring TP53 kev hloov pauv (21, 22, 29).

Raws li kev ua haujlwm dhau los (15), peb cia siab tias p53 reactivation nyob rau hauv cov txheej txheem saum toj no yuav ua rau muaj qhov tsis zoo thiab koom nrog kev tiv thaiv kab mob. Yog li ntawd, Dox tshem tawm ua rau mob qog noj ntshav ntau dhau ob peb lub lis piam, ua rau cov tsiaj muaj sia nyob ntev (Daim duab 1B thiab C). Kev soj ntsuam ntawm cov qog ntawm 14 hnub tom qab Dox tshem tawm tau qhia txog qhov kev cia siab ntawm p53 shRNA (raws li pom los ntawm tus kws tshaj xov xwm RFP txuas) thiab tsub zuj zuj ntawm senescence-associated -galactosidase (SA- -gal) tsis muaj qhov cuam tshuam tseem ceeb ntawm RAS-effector p-ERK (Fig. 1D). Ib yam li ntawd, kev nce hauv SA- -gal kev ua haujlwm thiab SASP-txuas nrog cov ntaub ntawv hloov pauv, ua ke nrog kev raug ntes ntawm cov qog nqaij hlav, tau pom nyob rau hauv cov qog nqaij hlav 6 mus rau 8 hnub tom qab p53 kho dua tshiab (Ntxiv Fig. S2A– S2H). Nco ntsoov, engraftment ntawm cov kab lis kev cai no ( khaws cia rau Dox kom tswj p53 silencing) rau hauv Dox-fed immunocompetent nas ua synchronous thiab focal secondary qog uas regressed nrog zoo sib xws kinetics raws li cov thawj qog thaum Dox tshem tawm (Fig. 1B; Supplementary Fig. S3A– S3E). Tswj kev sim siv lub Tet-Off system los yog sib koom ua ke p53 shRNA tau txiav txim siab tias Dox nws tus kheej muaj kev cuam tshuam rau qog tus cwj pwm hauv peb cov qauv (Cov duab ntxiv. S3F thiab S3G). Yog li, qhov system no tso cai rau kev ua kom muaj txiaj ntsig zoo ntawm cov qog nqaij hlav hauv cov qog nqaij hlav tsis muaj kev kho mob uas tuaj yeem hloov kho lub cev tiv thaiv kab mob. Muab nws qhov yooj yim ntxiv, peb siv cov qauv hloov pauv orthotopic (tom qab no hu ua "NSP") rau ntau qhov kev tshawb fawb txog kev siv tshuab tau piav qhia hauv qab no. Raws li kev cia siab, cov qog qog regressions tau sau tseg saum toj no yog kev tiv thaiv kab mob. Li no, NSP cov qog uas tshwm sim tom qab hloov pauv mus rau hauv cov tshuaj tiv thaiv kab mob ntshav qab zib thiab Rag2−/−Il2rg-/- (R2G2) nas tau txais cov lus teb cytostatic tseem ceeb tab sis ua tsis tiav, nrog cov tsiaj R2G2 qhia txog qhov tsis xws luag (Daim duab 1E–G; Ntxiv. Fig. S3H and S3I). Raws li cov nas liab qab muaj qhov tsis zoo hauv kev tiv thaiv kev tiv thaiv kab mob thiab R2G2 kuj raug cuam tshuam rau cov yam ntxwv ntawm kev tiv thaiv kab mob hauv lub cev, cov txiaj ntsig no qhia tau hais tias lub cev tiv thaiv kab mob yog qhov tseem ceeb rau kev ua haujlwm ntawm qog nqaij hlav hauv tus qauv thiab tsim kom muaj kev tswj xyuas cov ntsiab lus zoo los tshawb txog lub hauv paus rau kev siv tshuab. cov teebmeem no.

Cistanche deserticola-improve immunity (6)

cistanche cog-nce kev tiv thaiv kab mob

Senescence ua rau hloov pauv los ntawm cov qog tiv thaiv kab mob rau kev tiv thaiv kab mob

Txhawm rau ua tus yam ntxwv ntawm cov qog-tshwj xeeb paracrine cuam tshuam ntawm senescence, peb tom ntej no qhia txog kev tiv thaiv kab mob microenvironments ntawm cov qog harboring p53-suppressed (hu ua "proliferating") thiab p53- rov qab los (hu ua "senescent" ) qog hlwb tom qab 1 lub lis piam ntawm Dox tshem tawm, ib lub sij hawm thaum senescence yog tsim, tab sis cov qog tseem tsis tau regressed (Ntxiv Fig. S2, S3, thiab S4A). Cov kab mob harboring senescent qog hlwb pom ib tug ∼1.8-fold nce nyob rau hauv tag nrho CD45+ lub cev tiv thaiv kab mob piv nrog proliferating tswj (Fig. 2A; refs. 15, 16). Immunophenotypic thiab histologic tsom xam (thaum hnub 9 tom qab Dox tshem tawm) tau qhia tias qhov no cuam tshuam qhov nce ntawm feem pua ​​​​ntawm lymphocytes (B hlwb, CD4 T hlwb, thiab CD8 T hlwb) thiab txo qis hauv feem pua ​​​​ntawm Gr1+ myeloid-derived suppressor cells/neutrophils (CD11b+Gr1+Ly6Clo; Fig. 2B; Supplementary Fig. S4B). Txawm hais tias feem pua ​​​​ntawm cov macrophages raws li feem pua ​​​​ntawm tag nrho cov CD45 cov pej xeem tseem tsis tau hloov pauv, cov lej tseeb tau nce ntxiv (Fig. 2A thiab B; Ntxiv Fig. S4C–S4E). Nyob rau hauv cov neeg T-cell, accumulating CD8 T hlwb pom cov cim ntawm antigen kev (CD44+, CD69+) thiab harbored ib tug ntau ntawm cov pejxeem ntawm effector hlwb (CD44+CD62L−; Fig. . 2C; ref. 30). Qhov kev hloov kho tag nrho ntawm lub cev tiv thaiv kab mob no ua rau muaj qhov nce ntxiv hauv CD3: neutrophil piv rau cov qog nqaij hlav senescent hlwb (Sab Ntxiv Fig. S4F), cov teebmeem zoo ib yam nrog kev nce ntxiv hauv CD3: neutrophil piv uas tau cuam tshuam nrog kev tiv thaiv kab mob hauv tib neeg. kab mob siab (31) . Kev hloov kho tuaj yeem pom tau siv 3D duab tom qab tshem tawm cov ntaub so ntswg (Daim duab 2D; Ntxiv Fig. S4G thiab S4H; Ntxiv Video S1; ref 32).

Figure 1. A p53-restorable tumor model to study senescence immune surveillance. A, Generation of the p53-restorable, NRAS-driven mouse liver cancer model using the sleeping beauty transposon system delivered through HTVI. (Created with BioRender.com.) B, Representative ultrasonogram of HTVI and orthotopic injection liver cancer models at the indicated time after p53 restoration. C, Survival analysis of mice in the HTVI model. D, Representative hematoxylin and eosin (H&E), immunofluorescence (IF), and senescence-associated β-galactosidase (SA-β-gal) staining of p53-suppressed (p53 off) and -restored (p53 on for 14 days) tumor sections generated from the HTVI model. Scale bars, 50 μm. E–G, Orthotopic injection of GFP-luciferase vector-transduced NSP tumor cells into the livers of immunocompetent and immunodeficient mouse strains. E, Tumor size change measured by ultrasound upon p53 restoration. R2G2, Rag2-Il2rg double-knockout mouse. Data are presented as mean ± SEM. n ≥ 9 for each strain. F, Representative macroscopic pictures at 21 days of p53 on or endpoint p53 off the tumor. G, Representative IHC staining of GFP-labeled tumor cells at day 21 upon p53 restoration. Scale bars, 100 μm. **, P < 0.01; ***, P < 0.001.


Daim duab 1. Ib p53-cov qauv qog kho tau rov qab los kawm txog kev tiv thaiv kab mob tsis zoo. A, Generation of the p53-restorable, NRAS-tsav nas daim siab mob qog noj ntshav qauv siv lub cev pw tsaug zog zoo nkauj transposon xa los ntawm HTVI. (Tsim nrog BioRender.com.) B, Tus Neeg Sawv Cev Ultrasonogram ntawm HTVI thiab orthotopic txhaj rau daim siab mob qog noj ntshav ntawm lub sijhawm qhia tom qab p53 kho dua tshiab. C, Kev soj ntsuam ciaj sia ntawm cov nas hauv HTVI qauv. D, Tus Neeg Sawv Cev Hematoxylin thiab eosin (H&E), immunofluorescence (IF), thiab senescence-associated -galactosidase (SA- -gal) staining ntawm p53-suppressed (p53 tawm) thiab -restored (p53 rau 14 hnub) cov qog qog tsim los ntawm tus qauv HTVI. Scale bars, 5{{30}} μm. E-G, Orthotopic txhaj ntawm GFP-luciferase vector-transduced NSP qog hlwb mus rau hauv lub siab ntawm immunocompetent thiab immunodeficient nas hom. E, Cov qog loj hloov pauv ntsuas los ntawm ultrasound thaum kho p53. R2G2, Rag2-Il2rg ob-knockout nas. Cov ntaub ntawv raug nthuav tawm raws li txhais tau tias ± SEM. n Ntau dua lossis sib npaug rau 9 rau txhua hom. F, Cov duab sawv cev macroscopic ntawm 21 hnub ntawm p53 ntawm lossis qhov kawg p53 tawm ntawm cov qog. G, Tus Neeg Sawv Cev IHC staining ntawm GFP-labeled qog hlwb ntawm hnub 21 thaum p53 kho dua tshiab. Scale bar, 100 μm. **, P <0.01; ***, P <0.001.

Txhawm rau txhawm rau txheeb xyuas cov kab mob tiv thaiv kab mob tshwj xeeb uas muaj lub luag haujlwm rau kev tiv thaiv kev tiv thaiv kab mob ntawm cov qog hlwb, peb tau tsim cov kab ke sib luag ntawm cov nas harboring orthotopic NSP qog thiab tshuaj xyuas qhov cuam tshuam ntawm depleting ntau lub cev tiv thaiv kab mob ntawm cov qog regressions tom qab Dox tshem tawm. Thaum thaiv cov tshuaj tiv thaiv kab mob uas tsom rau neutrophils / monocytes (Gr1), natural killer (NK) hlwb (NK1.1), thiab CD4 T cells (GK1.5) tsis muaj txiaj ntsig, depletion ntawm CD8 T cells (2.43) thiab macrophages (siv liposomal clodronate). , uas xaiv lub hom phiaj macrophages (CD11b + F4 / 80+) tab sis tsis yog cov kab mob classical dendritic (CD11b-CD11c + MHC-II + CD103+; refs. 33, 34) ua rau mob qog nqaij hlav regression (Fig. 2E; Supplementary Fig. S4I). Txhawm rau ua tus yam ntxwv li cas p53-tsav qog senescence ua rau muaj kev tiv thaiv kab mob tiv thaiv kab mob, peb tau ua ib leeg-cell RNA-seq (scRNA-seq) kev tshuaj xyuas ntawm cov CD45 uas raug cais tawm tshiab los ntawm kev loj hlob thiab senescent NSP qog nqaij hlav ntxov tom qab Dox tshem tawm (8 hnub; Ntxiv Fig. S5A thiab S5B) thiab siv qhov sib txawv abundance testing algorithm Milo (35) los ntes cell xeev hloov nyob rau hauv lub cev tiv thaiv kab mob hom kev kho cov txheej txheem no (Supplementary Fig. S5C–S5F). kab nrog lawv txoj kev koom tes rau cov qog regression, CD8 T-cell thiab macrophage subpopulations pom tau tias muaj kev hloov pauv hauv qhov ntau thiab lub xeev. Hais txog T hlwb, proliferating (p53-suppressed) cov qog nqaij hlav tau loj hlob hauv CD8 T lub xeev uas muaj kev qhia siab ntawm ob qho kev ua haujlwm tsis zoo (Tox, Tigit, Lag3, Ctla4, Pdcd1 / PD1, thiab Cd160) thiab cov cim ua kom muaj zog (Prf1 ; Fig. 2F thiab G; Ntxiv Fig. S5G; Ntxiv Table S1). Cov CD8 T hlwb no kuj tau pom cov qib siab ntawm Tnfrsf9, ib qho cim paub tias delineate T-cell subsets uas muaj lub peev xwm los ua kom rov muaj zog hauv tib neeg HCC thiab lwm yam mob qog noj ntshav (36, 37). Hauv qhov tsis sib xws, hauv cov kab mob senescent (p53-reactivated) cov kab mob, CD8 T cov pej xeem tau tshwm sim heev, uas qhia tau hais tias cov cim tsis ua haujlwm qis thiab kev qhia siab ntawm cov tshuaj cytokines (piv txwv li, Ifng, Tnf; Supplementary Table S1). Raws li, qhov kev hloov pauv ntawm cov qog nqaij hlav ntau pom pom kev tiv thaiv kab mob thiab cov kab mob cytotoxic nyob rau hauv cov qog nqaij hlav senescent regression (Supplementary Fig. S5H; ref. 38).

Figure 2. Senescence triggers an immune evasion-to-immune recognition tumor switch. A, Representative images of CD45 and GFP staining marking immune cells and tumor cells, respectively, in p53-suppressed and p53-restored tumors (7 days after p53 restoration). Right, the quantification of the area of CD45+ staining was calculated from 3 random fields per mouse. Each dot represents a mouse. B, Flow cytometry analysis of the global immune landscape in an orthotopic NSP liver tumor model. Immunophenotyping of senescent tumors is performed 9 days after Dox withdrawal, a time point when the senescent state is fully established, yet preceding the massive tumor regression. G-MDSC, granulocytic myeloid-derived suppressor cells; M-MDSC, monocytic myeloid-derived suppressor cells. Data are pooled from 2 independent experiments, with n = 7 in the proliferating group and n = 9 in the senescent group. Note that, as the absolute number of CD45+ cells increases in senescent NSP tumor lesions (A), so do the total numbers of the indicated cell types. C, Flow cytometry analysis of CD8 T cells. Data are pooled from 2 independent experiments, with n = 11 in the proliferating and n = 10 in the senescent groups. Experiments were performed 9 days after Dox withdrawal. D, Representative tissue clearing images of the orthotopic NSP liver tumors. T cells, neutrophils, and vasculature are labeled by CD3, MPO, and CD31 staining, respectively. Samples were collected 9 days after Dox withdrawal. E, Tumor size change measured by ultrasound upon p53 restoration in mice after depleting specific immune cell types using antibodies or drugs. F, Left, uniform manifold approximation and projection (UMAP) plot of CD8 T cells isolated from p53-suppressed proliferating (PRO) and p53-reactivated senescent (SEN) tumors. Right, gene set enrichment analysis of T-cell exhaustion marker genes in CD8+ T cells from proliferating (p53-suppressed) versus senescent (p53-reactivated) tumors. NES, normalized enrichment score; Pval, P value. G, UMAP plot of the expression of selected genes (Cd8a, Cd44, Tnfrsf9, Cd69, Tox, and Fasl) between CD8 T cells isolated from senescent (p53-reactivated) and proliferating (p53-suppressed) tumors. H, Representative immunofluorescence images of CD8 T cells and F4/80-positive macrophage staining in the orthotopic NSP liver tumor. Tumor samples were collected 9 days after Dox withdrawal. Data are presented as mean ± SEM. All scale bars, 100 μm. A two-tailed Student t-test was used. *, P < 0.05; **, P < 0.01.


Daim duab 2. Senescence ua rau lub cev tiv thaiv kab mob evasion-rau-immune recognition qog hloov. A, Cov duab sawv cev ntawm CD45 thiab GFP staining cim lub cev tiv thaiv kab mob thiab cov qog hlwb, raws li, hauv p53-suppressed thiab p53-cov qog rov qab (7 hnub tom qab p53 kho). Txoj cai, qhov kom muaj nuj nqis ntawm thaj tsam ntawm CD45+ staining yog xam los ntawm 3 random teb rau ib tus nas. Txhua lub dot sawv cev rau tus nas. B, Flow cytometry tsom xam ntawm lub ntiaj teb kev tiv thaiv toj roob hauv pes hauv orthotopic NSP daim siab qog qauv. Immunophenotyping ntawm senescent qog yog ua tiav 9 hnub tom qab Dox tshem tawm, lub sijhawm taw qhia thaum lub xeev senescent tau tsim tag nrho, tab sis ua ntej lub qog loj heev. G-MDSC, granulocytic myeloid-derived suppressor hlwb; M-MDSC, monocytic myeloid-derived suppressor hlwb. Cov ntaub ntawv yog sib sau ua ke los ntawm 2 qhov kev sim ywj pheej, nrog n=7 hauv pawg proliferating thiab n=9 nyob rau hauv pawg senescent. Nco ntsoov tias, raws li tus naj npawb ntawm CD45+ hlwb nce hauv senescent NSP qog nqaij hlav (A), yog li ua tag nrho cov naj npawb ntawm cov hom cell qhia. C, Flow cytometry tsom xam ntawm CD8 T hlwb. Cov ntaub ntawv yog sib sau ua ke los ntawm 2 qhov kev sim ywj pheej, nrog n=11 hauv kev loj hlob thiab n=10 hauv pawg senescent. Kev sim tau ua 9 hnub tom qab Dox tshem tawm. D, Tus neeg sawv cev cov ntaub so ntswg tshem tawm cov duab ntawm orthotopic NSP daim siab qog. T hlwb, neutrophils, thiab vasculature yog sau los ntawm CD3, MPO, thiab CD31 staining, feem. Cov qauv raug sau 9 hnub tom qab Dox tshem tawm. E, Cov qog loj hloov pauv tau ntsuas los ntawm ultrasound thaum p53 kho dua tshiab hauv cov nas tom qab tshem tawm cov kab mob tiv thaiv kab mob tshwj xeeb uas siv cov tshuaj tiv thaiv lossis tshuaj. F, Sab laug, uniform manifold approximation and projection (UMAP) plot of CD8 T cells cais tawm ntawm p53-suppressed proliferating (PRO) and p{{30}}reactivated senescent (SEN) hlav. Txoj cai, gene teem enrichment tsom xam ntawm T-cell exhaustion marker noob nyob rau hauv CD8+ T hlwb los ntawm proliferating (p53-suppressed) piv rau senescent (p53-reactivated) qog. NES, normalized enrichment qhab nia; Pval, P tus nqi. G, UMAP zaj duab xis ntawm kev qhia ntawm cov noob xaiv (Cd8a, Cd44, Tnfrsf9, Cd69, Tox, thiab Fasl) nruab nrab ntawm CD8 T hlwb cais los ntawm senescent (p{{40}}reactivated) thiab proliferating (p{ {41}}suppressed) qog. H, Tus sawv cev immunofluorescence dluab ntawm CD8 T hlwb thiab F4/80- zoo macrophage staining nyob rau hauv orthotopic NSP daim siab qog. Cov qog qog tau sau 9 hnub tom qab Dox tshem tawm. Cov ntaub ntawv raug nthuav tawm raws li txhais tau tias ± SEM. Tag nrho cov nplai bar, 100 μm. Ib tug ob-tailed Student t-test tau siv. *, P <0.05; **, P <0.01.

Cov kev hloov pauv rau hauv macrophage compartment muab pov thawj ntxiv tias qog cell senescence ua rau lub cev tiv thaiv kab mob hloov mus rau kev soj ntsuam. Li no, scRNA-seq, immunophenotyping, thiab histology qhia tias cov qog txuam nrog macrophage phenotypes hloov ntawm F4/80lo; CD11chi xeev (pawg 8), suav nrog kev tiv thaiv kev tiv thaiv kab mob PD-L1+ cov pej xeem (tus yam ntxwv ntawm tib neeg HCC qog nqaij hlav nrog qhov tsis zoo; refs. 39, 40) rau F4/80hi; CD11c- xeev (cluster 0), txhais los ntawm kev nthuav qhia siab ntawm kev nthuav qhia cov noob caj noob ces kos npe (Cov duab ntxiv. S5E–S5J thiab S6A thiab S6B; Cov Lus Ntxiv S1). Nco ntsoov, cov senescence-sociated F4/80hi; CD11c- macrophages tshwj xeeb tshaj yog rau kev kho mob liposomal clodronate (Ntxiv Fig. S6C–S6E), uas kuj ua rau muaj qhov txo qis hauv feem ntawm CD8 nquag tab sis tsis yog CD4 T hlwb (Sab Ntxiv Fig. S6F thiab S6G), qhia txog CD8. T-nyob ntawm lub cev tiv thaiv kab mob cuam tshuam nrog kev koom tes nrog macrophages. Raws li, histologic tsom xam tau lees paub tias accumulating CD8 T hlwb thiab F4/80+ macrophages tau nquag coenriched tom qab senescence induction nyob rau hauv cov qog (Fig. 2H; Supplementary Fig. S4D). Ua ke, cov tshuaj lom neeg thiab cov tshuaj molecular no txhawb nqa tus qauv uas cov qog cell senescence ua rau muaj kev hloov pauv sai ntawm kev tiv thaiv kab mob tiv thaiv kab mob rau kev tiv thaiv kab mob los ntawm kev hloov pauv hauv macrophages thiab CD8 T-cell xeev, ua rau muaj kev tiv thaiv kab mob tiv thaiv kab mob thiab, thaum kawg, cov qog rejection.

Senescence Remodels Tissue-Sensing Programs thiab Cell-Surfaceome Landscape

Peb tom ntej no tau tsim tawm los siv cov qauv saum toj no kom nkag siab txog cov txheej txheem molecular lub luag haujlwm rau kev ua cov qog qog hlwb pom rau lub cev tiv thaiv kab mob. Senes cence induction cuam tshuam nrog chromatin remodeling program uas ntsiag to proliferative genes thiab activates ntau genes encoding SASP yam, nrog rau cov kev pab cuam tom kawg yog nyob ntawm ntau yam nyob rau hauv lub enhancer nyeem BRD4 (10). Yog li ntawd, peb tau ua cov kev sim transcriptional profiling ntawm NSP hlwb nyob rau hauv proliferating (p53-suppressed) piv rau senescent (p53- rov qab) tej yam kev mob nyob rau hauv lub tsis muaj thiab muaj JQ1, ib tug tshuaj uas inhibits BRD4 muaj nuj nqi (Sab ntxiv Table S2 ). Raws li qhov kev cia siab, p53 kev kho dua tshiab tau txo qis qhov kev nthuav qhia ntawm cov noob caj noob ces thiab ua rau muaj kev nthuav qhia ntawm SASP yam tseem ceeb (Fig. 3A; Ntxiv Fig. S7A; ref. 7), suav nrog ntau lub cytokines paub los txhawb T hlwb (Cxcl16, Il18). ) lossis macrophage activation thiab recruitment (Csf2, encoding protein GM-CSF) lossis yav dhau los txuas rau senescence (Igfbp7, Igfbp3, Pdgfa). Raws li kev cia siab los ntawm kev ua haujlwm dhau los (10), ntau qhov kev hloov kho SASP encoding transcripts (∼65%) yog BRD4-dependent (Supplementary Fig. S7B). Ib yam li ntawd, ntau yam kev loj hlob thiab kev tiv thaiv kab mob tau raug tso tawm los ntawm cov hlwb senescent, raws li kev soj ntsuam los ntawm multiplexed cytokine assays, suav nrog T-cell thiab macrophage attractants CCL5, CXCL9, thiab GMCSF, nrog rau vasculature remodeling factor VEGF (Ntxiv daim duab. . S7C). Yog li ntawd, senescence nyob rau hauv p53-rov qab NSP qog hlwb yog txuam nrog ib tug SASP robust, raws li cov cim remodeling ntawm lub cev tiv thaiv kab mob ecosystem yam ntxwv saum toj no.

Strikingly, kev soj ntsuam ntawm subcellular localization rau txawv cov noob caj noob ces (DEG) qhia tau hais tias senescent qog hlwb tsis tsuas yog nce lawv cov kev qhia ntawm secreted ("extracellular," EC) SASP yam, tab sis kuj tso saib cov kev hloov loj nyob rau hauv cov theem qhia ntawm transcripts encoding nto proteins. ("plasma membrane," PM; Fig. 3B). Tseeb, 25% ntawm tag nrho cov upregulated DEGs encoded PM proteins, ib tug tseem ceeb enrichment uas deviated los ntawm random faib (15%; Fig. 3B). Dynamic PM-DEGs tau txuas nrog cov protein tyrosine kinase signaling transduction (Nrp1, Egfr), cytokine receptor kev ua haujlwm (Ifngr1), extracellular matrix receptors (Itgb3, Cd44), thiab ion transporters (Slc12a1, Slc24a3) thiab ntes tau cov neeg paub txog senescated. Cd44, Vcam1, thiab Itgb3), tawm tswv yim cov hlwb muaj peev xwm ua kom muaj peev xwm cuam tshuam nrog thiab paub txog lawv ib puag ncig (Fig. 3C; Ntxiv Fig. S7D; refs. 41–43). Interestingly, lub senescence-koom nrog nce nyob rau hauv kev qhia ntawm ntau ntawm PM cov proteins no yog blunted los ntawm JQ1, qhia tias lawv induction tej zaum yuav yog ib feem ntawm qhov dav chromatin remodeling program ua ke rau SASP (Fig. 3D; ref. 10). Nco ntsoov, qhov kev hloov pauv loj hauv kev hloov pauv ntawm cov noob encoding PM proteins kuj tshwm sim hauv p53- tsis txaus NSP qog hlwb kho nrog cov tshuaj sib xyaw ua ke trametinib thiab palbociclib (Ntxiv daim duab S7E, sab saum toj vaj huam sib luag; Supplementary Table S2; ref. 20) thiab nyob rau hauv ib tug series ntawm 13 noob caj noob ces TP53 WT thiab TP53- mutant tib neeg kab mob qog noj ntshav muab tau los ntawm daim siab, mis, ntsws, thiab nyuv qog nqaij hlav induced rau senesce los ntawm ntau yam triggers (Fig. 3E; Supplementary Fig. S7F; ib. 44). Qhov no yog tshwj xeeb tshaj yog muaj zog rau upregulated (tab sis tsis downregulated) PM-DEGs, reminiscent ntawm cov teebmeem tshwm sim rau EC SASP yam (Fig. 3E; Supplementary Fig. S7E, hauv qab vaj huam sib luag). Yog li ntawd, qhov kev hloov pauv ntawm cov cell-surface proteins uas peb tau pom nyob rau hauv peb cov qauv txuas mus dhau p53-induced senescence thiab tej zaum yuav yog ib qho cim ntawm lub xeev senescent.

Figure 3. Senescence remodels tissue-sensing programs and cell-surfaceome landscape. A, Gene set enrichment analysis (Reactome) of RNA-seq data from proliferating (PRO, p53 off) versus senescent (SEN, p53 on for 8 days) NSP liver tumor cells in vitro. NES, normalized enrichment score. B, Subcellular localization of DEGs (P < 0.05; fold change > 2) in all detected genes [transcripts per kilobase million (TPM) > 1] from RNA-seq. C, Gene ontology (GO) analysis of DEGs encoding PM proteins upregulated in senescent cells. TM, transmembrane. D, Transcriptomic analysis of all DEGs (proliferating vs. senescent) in the presence or absence of JQ1 treatment. The C1 cluster (in red) contains the senescence-specific genes sensitive to JQ1, and the C4 cluster (in blue) contains the proliferation-specific genes sensitive to JQ1. E, Meta-analysis of RNA-seq dataset from SENESCopedia by performing subcellular localization of DEGs (same as Fig. 2D) and Fisher exact test to examine the relative enrichment of upregulated and downregulated EC/PM-DEGs deviated from the random distribution. See also Supplementary Fig. S7E and S7F. F, Mass spectrometry (MS) analysis of PM-enriched proteome in proliferating and senescent cells. The protein level is normalized to the mean expression of the protein of all samples. Controls are the samples without biotin labeling serving as background. Red and blue boxes represent proteins enriched in senescent and proliferating cells, respectively. n = 6 for both the senescent and proliferating experimental groups, and n = 3 and 4, respectively, for their control. G, Distribution of upregulated and downregulated GeneCards annotated PM proteins profiled by MS. NC, no change. H, Volcano plot of GeneCards-annotated PM proteins profiled by MS.


Daim duab 3. Senescence remodels cov ntaub so ntswg-sensing cov kev pab cuam thiab cell-surfaceome toj roob hauv pes. A, Gene teem enrichment tsom xam (Reactome) ntawm RNA-seq cov ntaub ntawv los ntawm proliferating (PRO, p53 tawm) piv rau senescent (SEN, p53 rau 8 hnub) NSP daim siab qog hlwb hauv vitro. NES, normalized enrichment score. B, Subcellular localization ntawm DEGs (P < 0}.05; hloov pauv> 2) hauv txhua cov noob caj noob ces [transcripts per kilobase lab (TPM) > 1] los ntawm RNA-seq. C, Gene ontology (GO) tsom xam ntawm DEGs encoding PM proteins upregulated nyob rau hauv senescent hlwb. TM, transmembrane ua. D, Kev tshuaj ntsuam xyuas ntawm txhua DEGs (proliferating vs. senescent) nyob rau hauv lub xub ntiag los yog tsis muaj JQ1 kev kho mob. C1 pawg (hauv liab) muaj cov noob caj noob ces tshwj xeeb rau JQ1, thiab C4 pawg (hauv xiav) muaj cov noob caj noob ces tshwj xeeb rau JQ1. E, Meta-analysis ntawm RNA-seq dataset los ntawm SENESCopedia los ntawm kev ua haujlwm subcellular localization ntawm DEGs (tib yam li Fig. 2D) thiab Fisher pes tsawg xeem los tshuaj xyuas cov txheeb ze enrichment ntawm upregulated thiab downregulated EC/PM-DEGs deviated los ntawm random faib. Saib ntxiv Fig. S7E thiab S7F. F, Mass spectrometry (MS) tsom xam ntawm PM-enriched proteome nyob rau hauv proliferating thiab senescent hlwb. Cov qib protein yog normalized rau lub ntsiab lus ntawm cov protein ntawm tag nrho cov qauv. Kev tswj hwm yog cov qauv uas tsis muaj biotin labeling ua haujlwm tom qab. Cov thawv liab thiab xiav sawv cev rau cov protein ntau ntxiv hauv cov hlwb senescent thiab proliferating, feem. n=6 rau ob pawg senescent thiab proliferating sim, thiab n=3 thiab 4, raws li, rau lawv tswj. G, Kev faib tawm ntawm upregulated thiab downregulated GeneCards annotated PM proteins profiled los ntawm MS. NC, no change. H, Volcano zajlus ntawm GeneCards-annotated PM proteins profiled los ntawm MS.

Txhawm rau txheeb xyuas lub ntiaj teb kev hloov kho PM yam nyob rau hauv senescence ntawm qib protein, peb tau ua cov txheej txheem saum npoo ntawm isogenic proliferating thiab senescent NSP qog hlwb, siv cov txheej txheem biotin-labeling enrichment, nyob rau hauv uas cell-dog proteins tau sau nrog membrane-impermeable biotin, purified, thiab raug rau huab hwm coj spectrometry (Fig. 3F; Supplementary Fig. S7G; ref. 45). Kev sib raug zoo ntawm cov khoom siv roj ntsha nyob hauv txhua qhov mob tau pom (Cov Lus Qhia Ntxiv. S7H), nrog rau cov proteins uas tau kuaj pom tau ntxiv rau cov ntsiab lus PM proteins los ntawm 60% tom qab induction ntawm p53-induced senescence. Ntawm 887 cov proteins uas tau kuaj pom dua, ntau dua 50% tau qhia txawv. Feem ntau qhov sib txawv ntawm cov proteins sib raug zoo nrog cov kev taw qhia tau pom nyob rau hauv peb cov ntaub ntawv transcriptional profiling, txawm hais tias qee qhov tau hais txawv txawv yam tsis muaj kev hloov pauv hauv cov ntawv sau tseg (Sab Ntxiv Fig. S7I).

Desert ginseng-Improve immunity (9)

cistanche cov txiaj ntsig rau txiv neej-ua kom muaj zog tiv thaiv kab mob

Annotated cell-surface proteins kuaj pom los ntawm huab hwm coj spectrometry raws li senescence induction suav nrog ob peb yav tas los txuas rau senescence (xws li, CD44 thiab VCAM1), ntau yam kev loj hlob thiab cytokine receptors (piv txwv li, EGFR, ICAM1, thiab IFNGR1), thiab lwm yam tsawg tus yam ntxwv (Daim duab . 3F–H; Supplementary Fig. S7J and S7K). Ntawm qhov kev ceeb toom, cov txheej txheem ntawm cov cell nto-enriched proteins pom nyob rau hauv peb cov qauv qhia tsis sib tshooj nrog cov neeg uas pom nyob rau hauv tib neeg fibroblasts undergoing oncogene-induced senescence (46), qhia heterogeneity ntawm cell hom los yog senescence triggers. Txawm li cas los xij, cov txiaj ntsig no qhia tau hais tias ntxiv rau kev rov ua dua tshiab hauv lawv txoj haujlwm zais cia, senescent hlwb tau hloov pauv ntau hauv cov ntsiab lus thiab kev nplua nuj ntawm cov cell-nto proteins thiab qhia tias cov hlwb senescent tau txais qhov tshwj xeeb microenvironment-sensing zoo uas yuav cuam tshuam lawv lub xeev thiab txoj hmoo hauv vivo. .

Senescent Cells yog Primed rau Sense IFNg thiab Amplify IFNg Signaling

Txhawm rau txheeb xyuas txoj hauv kev uas tuaj yeem cuam tshuam li cas cov hlwb senescent paub txog lawv qhov chaw nyob, peb tau mined cov ntaub ntawv hloov pauv thiab cov ntaub ntawv proteomic rau cov kev hloov pauv uas cuam tshuam nrog kev tiv thaiv kab mob antitumor. Interestingly, gene ontology (GO) tsom xam tau qhia tias hom II interferon gamma (IFN) cov lus teb (47) yog nyob rau saum 5 annotated pathways enriched thaum lub sij hawm senescence thiab nyob ntawm lub xeev ntawm cell-specific enhancer cov kev pab cuam (ie, JQ1-sensitive ; ie, "C1" of Fig. 3D; Supplementary Fig. S8A). Ntawm cov ntaub ntawv hloov pauv, peb tau sau tseg ntau qhov kev tswj hwm zoo ntawm IFN signaling, suav nrog IFN receptor subunit IFNGR1 (ib qho ntawm cov protein ntau tshaj plaws los ntawm peb cov ntaub ntawv proteomic) thiab ntau yam interferon effectors (Irf1, Irf7, thiab Irf9; refs. 47, 48). ; Fig. 4A–C; Supplementary Fig. S8B and S8C). Dhau li ntawm cov Brd 4-cov noob muaj txiaj ntsig-upregulated, cov ntaub ntawv sau tseg tsis zoo ntawm IFN signaling (Ptpn2, Socs1, thiab Socs3) tau txo qis (Fig. 4C; refs. 49, 50). Cov kev hloov zoo sib xws tau raug sau tseg hauv NSP qog hlwb kho nrog sib txawv senescence inducers (Fig. 4C; Ntxiv Fig. S8D-S8G) thiab, ntau qhov dav, nyob rau hauv ib lub vaj huam sib luag ntawm 13 tib neeg lub mis-, ntsws-, siab-, thiab mob qog noj ntshav. cov kab ntawm tes ua rau muaj kev cuam tshuam (Fig. 4D; ref. 44). Yog li ntawd, kev hloov pauv hauv kev qhia ntawm hom II IFN cov cim qhia cov khoom siv yog ib qho tseem ceeb ntawm cov hlwb senescent, ywj siab ntawm hom cell, cell genotype, hom, thiab qhov xwm txheej ntawm senescence inducer. Qhov kev nce ntxiv hauv IFN signaling effectors thiab txo qis hauv cov tswj tsis zoo coj peb mus rau kev xav tias cov hlwb senescent ua primed kom nkag siab IFN hauv lawv ib puag ncig. Txhawm rau kuaj qhov kev xav no ncaj qha, peb tau kho cov kab mob loj hlob thiab senescent NSP nrog recombinant IFN thiab ua cov tshuaj tiv thaiv kab mob ntawm JAK-STAT teeb liab ua kom muaj zog. Txawm hais tias IFN tau nce qib theem pib ntawm STAT1 hauv ob lub xeev, cov hlwb senescent tau sau ntau phosphorylated STAT1, tsis hais txog qhov tshwm sim ntawm senescence (Fig. 4E; Supplementary Fig. S8H). Tsis tas li ntawd, peb kuj pom muaj qib ntawm phosphorylated JAK1 nyob rau hauv p53- rov qab senescent hlwb, ntxiv dag zog rau peb txoj kev tshawb nrhiav ntawm JAK-STAT teeb liab txoj hauv kev hauv senescent hlwb sensing IFN (Sab ntxiv Fig. S8I). Raws li kev kwv yees los ntawm kev txheeb xyuas cov ntaub ntawv sau tseg, kev laus kuj ua rau txo qis hauv PTPN2 protein (51), txawm tias muaj exogenous IFN (Fig. 4E). Yog li, senescent hlwb ua haujlwm tau zoo dua IFN signaling nyob rau hauv cov lus teb rau kev txwv concentration ntawm IFN nyob rau hauv ib puag ncig.

Senescence thiab EC IFNg Cooperatively Upregulate Antigen Processing thiab Presentation Machinery

Txhawm rau kom nkag siab zoo dua qhov kev pab cuam ntawm IFN sensing rau qhov kev pab cuam senescence, peb txuas ntxiv piv cov phenotypic thiab transcriptional xeev ntawm proliferating thiab p53-rov qab senescent NSP qog hlwb kho nrog recombinant IFN ntawm qis (50 pg / mL) los yog ntau dua (1 ng / mL) koob tshuaj. Txawm hais tias qhov sib ntxiv ntawm exogenous IFN mus rau proliferating los yog senescent qog hlwb muaj ib tug negligible cuam tshuam rau lub viability, proliferation, los yog SASP noob qhia ntawm ob hom cell ntawm koob tshuaj ntsuam xyuas (Fig. 5A; Supplementary Fig. S9A–S9D), cim kev hloov nyob rau hauv IFN txoj kev gene qhia txuas nrog lub xeev senescent tau pom. Tshwj xeeb, kev saib xyuas pawg ntawm Hallmark "IFN cov lus teb kos npe" thoob plaws cov hlwb loj hlob thiab cov senescent tau nthuav tawm peb DEG modules: (i) cov noob uas tau txo qis thaum lub sij hawm senescence tsis hais txog IFN (xws li cov kev tswj tsis zoo li hais saum toj no); (ii) cov noob uas tau hloov kho thaum lub sij hawm senescence tsis hais txog IFN ; thiab, nthuav, (iii) ib qho tseem ceeb ntawm DEGs uas yog kev koom tes induced los ntawm kev sib xyaw ntawm senescence thiab IFN (Fig. 5B). Yog li ntawd, senescence ua rau kom muaj nuj nqis thiab zoo hloov pauv hauv cov lus teb rau IFN. Ib qho txiaj ntsig zoo tsim los ntawm IFN qhia txog kev tswj cov hlwb 'ua rau yoog raws kev tiv thaiv kab mob yog qhov muaj peev xwm ntxiv rau kev nthuav qhia antigen mediated los ntawm MHC class I molecules (MHC-I; refs. 47, 52). Qhov tseeb, ntau cov noob tau tsim kho hauv cov hlwb senescent (chav kawm ii noob) lossis superinduced nyob rau hauv lub xub ntiag ntawm exogenous IFN (chav kawm iii noob) suav nrog cov khoom ntawm cov khoom siv tshuaj tiv thaiv antigen. Ntawm cov genes induced thaum lub sij hawm senescence (chav kawm ii noob) yog Tap1, transporters txuam nrog antigen ua, thiab Psme1, ib tug proteasome yam cuam tshuam nrog antigen ua (53). Cov hypersensitive rau exogenous IFN (chav kawm iii noob) suav nrog Nlrc5, transcriptional coactivator ntawm MHC-I noob (54); MHC-I sib dhos ua ke Tapbp; thiab MHC-I subunit B2m. Ob chav kawm iii noob caj noob ces yog cov khoom ntawm immunoproteasome (Psmb8 thiab Psmb9) uas nws cov yeeb yam tuaj yeem hloov pauv cov repertoire ntawm cov peptides nthuav tawm thaum overexpressed thiab cuam tshuam nrog kev txhim kho qog cov lus teb rau lub cev tiv thaiv kab mob (55). Qhov kev nthuav tawm no ntawm IFN nyob rau hauv cov hlwb senescent tau lees paub los ntawm RT-qPCR thiab tau khaws cia nyob rau theem siab dua ntawm exogenous IFN (Fig. 5C; Supplementary Table S3). Raws li cov txheej txheem multifactorial tau piav qhia saum toj no, cov txiaj ntsig no tsis tau pom nyob rau hauv cov qog nqaij hlav proliferating, txawm tias cov overexpressing IFNGR1 cDNA thiab/los yog kho nrog IFN (Ntxiv Fig. S10A-S10D).

Figure 4. Senescent cells are primed to sense and amplify IFNγ signaling. A and B, IFNGR1 level on proliferating and senescent cells profiled by mass spectrometry (A) and validated by flow cytometry (B). AU is an arbitrary unit. Data are presented as mean ± SEM. n = 6 for both the proliferating and senescent groups. C, Transcriptomic analysis of selected genes regulating IFNγ signaling from RNA-seq data of 3 independent p53-restorable cell lines (NSP, NSM2, and NSP5) restoring p53 along with NSP cells treated with two other senescence triggers. SEN/PRO, senescent/proliferating; T + P, trametinib plus palbociclib. D, mRNA expression of selected genes involved in IFNγ signaling in human cell lines triggered to senesce. Treatment: Ali, alisertib; Eto, etoposide; the number indicates the length of treatment (days). Data are obtained from the public dataset SENESCopedia (44). E, Top, immunoblot analysis of NSP cells under different senescent triggers in the presence or absence of IFNγ (1 ng/mL). Bottom, quantification of the intensity of the signal from immunoblot. p-STAT1, phospho-STAT1 (Tyr701).


Daim duab 4. Senescent hlwb raug tsim los rau kev nkag siab thiab ua kom IFN signaling. A thiab B, IFNGR1 theem ntawm kev loj hlob thiab senescent hlwb profiled los ntawm huab hwm coj spectrometry (A) thiab validated los ntawm flow cytometry (B). AU yog ib lub koom haum arbitrary. Cov ntaub ntawv raug nthuav tawm raws li txhais tau tias ± SEM. n=6 rau ob pawg proliferating thiab senescent. C, Transcriptomic tsom xam ntawm cov noob caj noob ces tswj IFN signaling los ntawm RNA-seq cov ntaub ntawv ntawm 3 ywj siab p53-restorable cell kab (NSP, NSM2, thiab NSP5) restoring p53 nrog rau NSP hlwb kho nrog ob lwm yam senescence triggers. SEN / PRO, senescent / proliferating; T + P, trametinib plus palbociclib. D, mRNA qhia ntawm cov noob xaiv koom nrog hauv IFN signaling hauv tib neeg cov kab ntawm tes ua rau senesce. Kev kho mob: Ali, alisertib; Eto, etoposide; tus lej qhia qhov ntev ntawm kev kho mob (hnub). Cov ntaub ntawv tau txais los ntawm cov ntaub ntawv pej xeem SENESCOpedia (44). E, Sab saum toj, immunoblot tsom xam ntawm NSP hlwb nyob rau hauv txawv senescent tshwm sim nyob rau hauv lub xub ntiag los yog tsis muaj IFN (1 ng / mL). Hauv qab, kom muaj nuj nqis ntawm kev siv lub teeb liab los ntawm immunoblot. p-STAT1, phospho-STAT1 (Tyr701).

Tsis tas li ntawd ua raws li cov kev hloov pauv ntawm cov noob tau piav qhia saum toj no, cov qog nqaij hlav senescent ntau zog tau tswj hwm MHC-I hauv cov lus teb rau qib qis ntawm exogenous IFN piv nrog cov neeg sib tw proliferating. Yog li ntawd, thaum lub cell-surface theem ntawm MHC-I ntawm ob qho tib si proliferating thiab senescent hlwb yog tsawg nyob rau hauv lub hauv paus thiab induced los ntawm exogenous IFN, senescent hlwb pom ib tug tseem ceeb nce nyob rau hauv MHC-I protein qhia (Fig. 5D). Cov kev sib koom ua ke zoo sib xws tau raug soj ntsuam rau cov xov tooj ntawm tes-nto HLA qhia (zoo ib yam li MHC-I hauv cov nas) hauv tib neeg cov qog nqaij hlav qog noj ntshav los ntawm lub siab thiab lwm yam mob qog noj ntshav uas ua rau muaj qhov tsis zoo nrog Butlin, uas koom nrog ap53-dependent senescence program (56) , los yog trametinib/palbociclib, uas nyiam lub hom phiaj qog hlwb nrog txoj kev ua haujlwm MAPK (Ntxiv Fig. S11A–S11D; ref. 20). Ntawm qhov kev ceeb toom, cov teebmeem kev sib txuas ntawm cov tshuaj kho mob thiab IFN ntawm HLA qhia yuav tsum muaj qhov tshwm sim ntawm qhov tsis txaus ntseeg thiab tsis tshwm sim hauv daim siab qog hlwb uas ua tsis tau zoo vim muaj kev hloov pauv ntawm tus kheej lossis kev hloov pauv p53 (tsis teb rau cov lus piav qhia) lossis txoj hauv kev tsis yog MAPK (tsis teb. rau trametinib/palbociclib). Tsis tas li ntawd, txawm tias hom I thiab II IFN cov lus teb muaj xws li cov khoom sib tshooj, kev kho mob IFN tsis tuaj yeem hloov pauv rau IFN hauv kev tsim cov MHC-I induction muaj zog hauv cov hlwb los yog qhov sib txawv ntawm induction ntawm proliferating thiab senescent hlwb nyob rau hauv peb p53 cov qauv kho dua tshiab ( Ntxiv daim duab S11E thiab S11F). Cov ntaub ntawv no qhia tau hais tias murine thiab tib neeg lub hlwb ua rau senesce tau txais kev muaj peev xwm ntau ntxiv rau kev ua cov tshuaj tiv thaiv thiab kev nthuav qhia nyob rau hauv muaj kev txwv ntau ntawm IFN .

Senescent Tumor Cells Hyperactivate IFNg Signaling Pathway Hauv Vivo

Txhawm rau txiav txim siab qhov tshwm sim hauv vivo ntawm kev rov xa rov qab ntawm IFN cov cim qhia pom nyob rau hauv cov hlwb senescent, peb txuas ntxiv hloov kho IFN sensing (IGS) cov neeg sau xov xwm kom ncaj qha pom qhov intracellular IFN signaling activation hauv real-time (57). Cov neeg sau xov xwm no muaj ntau qhov kev pom zoo IFN -activated sequences, uas muaj qhov tshwj xeeb rau hom II IFN tshaj lwm cov cim (57), ua raws li cDNA ib ntus encoding ZsGreen1 fluorescent protein thiab txuas mus rau ib qho kev qhia RFP transgene kom pom cov hlwb transduced ( Fig. 6A). NSP qog hlwb uas qhia txog qhov tsim no yog RFP zoo thiab pom tias muaj koob tshuaj nce ntxiv hauv ZsGreen1 teeb liab thaum kho nrog IFN hauv vitro uas tau nce ntxiv tom qab p53 induction lossis tom qab kev kho mob nrog cov tshuaj senescence-inducing (Fig. 6B; Supplementary Fig. S12A thiab S12B) .

Peb tom ntej no siv cov kab ke no los saib xyuas kev ua haujlwm ntawm kev ua haujlwm tom qab senescence induction hauv cov qog. Tshaj tawm cov qog hlwb (ntawm Dox) uas qhia txog kev tsim RFP tau txhaj rau hauv daim siab ntawm Dox-fed syngeneic cov neeg tau txais, thiab, thaum pom cov qog, Dox raug tshem tawm kom ntxias p53 qhia thiab ua rau muaj qhov tsis zoo li saum toj no (saib daim duab 1 thiab 2). Cov qog nqaij hlav rov qab raug cais tawm 9 hnub tom qab Dox tshem tawm rau 3D duab ntawm cov neeg sau xov xwm kev ua ub no thiab kev ntsuam xyuas tib lub sijhawm ntawm IFN signaling nyob rau hauv kev sib piv nrog proliferating tswj (los ntawm nas tswj ntawm Dox). Raws li pom nyob rau hauv daim duab 6C, proliferating qog hlwb pom me ntsis, yog hais tias muaj, reporter qhia, whereas qog hlwb ua rau senesce nyob rau hauv vivo tso tawm ib tug tseem ceeb ZsGreen1 teeb liab (Fig. 6C thiab D; Ntxiv Video S2). Cov nyhuv no coincided nrog ib qho kev nce qib ntawm IFN protein (tab sis tsis yog hom I IFN) nyob rau hauv cov qog nqaij hlav extracts (Fig. 6E; Supplementary Fig. S12C).

Txhawm rau kuaj seb qhov hloov pauv ntawm lub cev tiv thaiv kab mob hauv cov qog nqaij hlav puas hlwb (Fig. 2; Supplementary Fig. S5–S6) tau pab txhawb rau lub teeb liab txhim kho ntawm IGS tus neeg sau xov xwm, peb tau ua hauv vitro coculture kev soj ntsuam uas tso cai rau kev nthuav tawm ntawm senescent lossis proliferating qog hlwb rau ib qho sib npaug ntawm cov kab mob CD8 T hlwb, uas peb txheeb xyuas los ntawm scRNA-seq cov ntaub ntawv raws li qhov tseem ceeb ntawm cov cellular ntawm IFN hauv vivo (Fig. 6F–H). Senescent hlwb tseem pom qhov nce ntxiv ntawm ZsGreen1 teeb liab raws li piv nrog cov tswj kev loj hlob (Fig. 6I). Raws li qhov tsis yog-cell-autonomous signaling activation, IFN tsis pom nyob rau hauv cov xov xwm txias los ntawm NSP qog hlwb nyob rau hauv proliferative los yog senescent tej yam kev mob (Supplementary Fig. S12D), tsis tau IFN tau nkag tau yooj yim thaum coculture nrog CD8 T hlwb, ib qho kev cuam tshuam uas yog kev txhim kho ntxiv los ntawm kev sib ntxiv ntawm macrophages thiab cuam tshuam nrog nce MHC chav kawm I ntawm cov hlwb senescent nrog rau kev ua kom muaj zog ntawm CD8 T hlwb (Supplementary Fig. S12E–S12J). Ua ke, cov ntaub ntawv no txhawb nqa tus qauv uas yog kev sib cuam tshuam ntawm heterotypic ntawm cov qog hlwb senescent thiab lub cev tiv thaiv kab mob rhiab cov qog mus rau exogenous IFN, ua rau txhim kho antigen kev nthuav qhia thiab kev tiv thaiv kev tiv thaiv zoo.

Figure 5. Senescence and EC IFNγ cooperate to upregulate antigen processing and presentation machinery. A and B, mRNA expression of genes in proliferating and senescent NSP cells in vitro in the presence or absence of IFNγ (50 pg/mL) treatment. mRNA level is normalized to the mean expression of the gene in all samples. A, DEG-encoding SASP factors in our model. B, IFNγ response genes from the Hallmark signature database. C, RT-qPCR of selected antigen presentation pathway genes in proliferating and senescent cells treated with low (50 pg/mL) or high (1 ng/mL) concentration of IFNγ. Samples are from 2 biological replicates. D, MHC-I level of proliferating and senescent cells treated with IFNγ for 24 hours measured by flow cytometry. MFI, median fluorescence intensity. Data are presented as mean ± SEM.

Daim duab 5. Senescence thiab EC IFN koom tes los txhim kho antigen ua thiab kev nthuav qhia machinery. A thiab B, mRNA qhia txog cov noob nyob rau hauv proliferating thiab senescent NSP hlwb nyob rau hauv vitro nyob rau hauv lub xub ntiag los yog tsis muaj IFN (50 pg / mL) kev kho mob. mRNA qib yog normalized rau qhov txhais tau hais tias cov noob hauv txhua qhov qauv. A, DEG-encoding SASP yam hauv peb cov qauv. B, IFN teb cov noob los ntawm Hallmark kos npe database. C, RT-qPCR ntawm kev xaiv antigen nthuav qhia txoj hauv kev noob hauv kev loj hlob thiab senescent hlwb kho nrog qis (50 pg / mL) lossis siab (1 ng / mL) concentration ntawm IFN . Cov qauv yog los ntawm 2 bioreplicates. D, MHC-I theem ntawm proliferating thiab senescent hlwb kho nrog IFN rau 24 teev ntsuas los ntawm flow cytometry. MFI, qhov nruab nrab fluorescence siv. Cov ntaub ntawv raug nthuav tawm raws li txhais tau tias ± SEM.

Figure 6. Senescence enhances IFNγ-mediated heterotypic signaling from activated immune cells to tumor cells. A Graphic illustration of the IGS reporter. (Created with BioRender.com.) B, Left, representative flow cytometry plots measuring ZsGreen1 signals in proliferating and senescent NSP cells treated with 1 ng/mL IFNγ. Right, quantification of the percentage of ZsGreen1-positive cells upon IFNγ treatment. MFI, median fluorescence intensity. C and D, Representative 3D imaging of tissue-cleared tumors from the orthotopically injected liver NSP cell line expressing IGS reporter (C). Quantification of 3 randomly selected fields from the liver tumor of each mouse (D). n = 5 and n = 3 for the proliferating and senescent groups (9 days after p53 restoration), respectively. Scale bars, 100 μm. E, Top, cytometric bead array assay for the IFNγ level from in vivo tumor tissue lysate samples (7 days after p53 restoration). Bottom, transcripts of indicated genes from RNA-seq of in vivo bulk samples of tumors generated by HTVI (PRO, p53 off; SEN, p53 restoration for 12 days). TPM, transcripts per kilobase million. Noted Ifna/b cluster contains 14 Ifna subtypes and 1 Ifnb gene. F and G, Expression of Ifng in tumor-infiltrating immune cells profiled by scRNA-seq in the NSP transplantable model (as in Fig. 2, a sample collected at day 8 after p53 restoration). H, Uniform manifold approximation and projection plot of the expression of Havcr2 (encoding TIM3) and Ifng in CD8 T cells harvested from proliferating (P) and senescent (S) tumor lesion. The top panel is replicated from Fig. 2F (left) to indicate cells corresponding to each condition. I, Quantification of ZsGreen1 intensity of NSP tumor cells in the OT-I T-cell and SIINFEKL-expressing tumor cell coculture experiment (effector-to-target ratio, 5:1) after 20 hours of coculture. Signal measured by flow cytometry. T + P, trametinib plus palbociclib. See experimental details in Supplementary Fig. S12E. Data are presented as mean ± SEM. Two-tailed Student t-test was used. *, P < 0.05; **, P < 0.01; ***, P < 0.001.


Daim duab 6. Senescence txhim kho IFN -mediated heterotypic signaling los ntawm lub cev tiv thaiv kab mob mus rau qog hlwb. Daim duab kos duab ntawm IGS reporter. (Tsim nrog BioRender.com.) B, Sab laug, tus neeg sawv cev ntws cytometry plots ntsuas ZsGreen1 cov cim hauv kev loj hlob thiab senescent NSP hlwb kho nrog 1 ng / mL IFN . Txoj cai, kom muaj nuj nqis ntawm feem pua ​​​​ntawm ZsGreen1- cov hlwb zoo thaum kho IFN. MFI, qhov nruab nrab fluorescence siv. C thiab D, Tus Neeg Sawv Cev 3D daim duab ntawm cov nqaij mos-cleared qog los ntawm orthotopically txhaj daim siab NSP cell kab qhia IGS reporter (C). Quantification ntawm 3 randomly xaiv teb los ntawm daim siab qog ntawm txhua tus nas (D). n=5 thiab n=3 rau pawg proliferating thiab senescent (9 hnub tom qab p53 restoration), feem. Scale bars, 100 μm. E, Sab saum toj, cytometric hlaws array assay rau qib IFN los ntawm hauv vivo qog nqaij hlav lysate kuaj (7 hnub tom qab p53 kho). Hauv qab, cov ntaub ntawv sau tseg ntawm cov noob qhia los ntawm RNA-seq ntawm hauv vivo cov qauv ntawm cov qog tsim los ntawm HTVI (PRO, p53 tawm; SEN, p53 kho dua tshiab rau 12 hnub). TPM, transcripts ib kilobase lab. Sau tseg Ifna/b pawg muaj 14 Ifna subtypes thiab 1 Ifnb gene. F thiab G, Kev nthuav qhia ntawm Ifng nyob rau hauv cov qog-infiltrating lub cev tiv thaiv kab mob profiled los ntawm scRNA-seq nyob rau hauv NSP transplantable qauv (raws li nyob rau hauv daim duab 2, ib tug qauv sau nyob rau hauv hnub 8 tom qab p53 kho). H, Uniform manifold approximation and projection plot of the expression of Havcr2 (encoding TIM3) and Ifng in CD8 T cells harvested from proliferating (P) and senescent (S) qog lesion. Lub vaj huam sib luag sab saum toj yog rov ua dua los ntawm daim duab 2F (sab laug) los qhia cov cell sib raug rau txhua yam mob. I, Quantification ntawm ZsGreen1 siv ntawm NSP qog hlwb nyob rau hauv OT-I T-cell thiab SIINFEKL-expressing qog cell coculture sim (effector-to-target ratio, 5:1) tom qab 20 teev ntawm coculture. Teeb liab ntsuas los ntawm flow cytometry. T + P, trametinib plus palbociclib. Saib cov ntsiab lus kev sim nyob rau hauv Ntxiv Fig. S12E. Cov ntaub ntawv raug nthuav tawm raws li txhais tau tias ± SEM. Ob-tailed Student t-test tau siv. *, P <0.05; **, P <0.01; ***, P <0.001.

IFNg Kev Tshaj Tawm hauv Senescent Tumor Cells yog qhov tsim nyog rau kev soj ntsuam kev tiv thaiv kab mob

Peb cov txiaj ntsig tau hais tias kev tiv thaiv kab mob sib kis ntawm senescent NSP qog hlwb cuam tshuam nrog kev sib xyaw ua ke ntawm SASP, paub los txhawb kev tiv thaiv kab mob hlwb (10, 20, 58), nrog rau yav dhau los tsis txaus siab lub peev xwm ntawm senescent hlwb rau kev paub zoo dua thiab cov lus teb rau EC cov cim, raws li qhia ntawm no nrog IFN . Txhawm rau kuaj qhov kev pab cuam ntawm senescence-associated IFN sensing program rau kev tiv thaiv kev tiv thaiv kab mob ntawm cov qog hlwb, peb tau tshuaj xyuas seb puas muaj kev cuam tshuam ntawm IFNGR hauv cov qog hlwb, lossis IFN depletion hauv tus tswv tsev, cuam tshuam rau kev tshem tawm ntawm NSP qog hlwb thaum senescence induction. . Tseeb, qog regression (tab sis tsis senescence ib se; Ntxiv Fig. S13A–S13D) yog impaired raws li knockout (KO) ntawm IFNGR1 (Fig. 7A thiab B; Supplementary Fig. S14A–S14C), ib tug nyhuv uas yog txawm ntau pronounced rau IFNGR-cov qog nqaij hlav tsis zoo tau nkag mus rau hauv Ifng-/- nas (Daim duab 7C thiab D) thiab cuam tshuam nrog qhov xav tias poob ntawm MHC-I hauv cov qog hlwb (Cov duab ntxiv S14D thiab S14E).

Raws li kev paub txog kev koom tes ntawm IFN signaling thiab qog cell MHC-I rau CD8+ kev koom tes (59), cov qog uas tsis muaj IFNGR1 los yog tsim hauv Ifng-/- cov neeg tau txais muaj tsawg dua CD8 T hlwb dua li lawv cov WT counterparts hauv ob qho tib si. proliferating thiab senescent xeev (Sab ntxiv Fig. S14F) thaum tseem inducing robust tiv thaiv kab mob infiltrate nrog rau ntau macrophages (Fig. 7E thiab F; Supplementary Fig. S14G). Txawm li cas los xij, qhov tsis zoo ntawm kev soj ntsuam phenotype tsis yog qhov tshwm sim ntawm qhov txo qis hauv CD8 T hlwb. Coculture assays muab cov kev sib raug zoo ntawm IFNGR1 KO thiab WT qog hlwb rau CD8 T hlwb thiab macrophages tseem pom IFNGR1- nyob ntawm kev tua ntawm cov qog nqaij hlav hlwb (Ntxiv Fig. S15A–S15E)-ib qho kev cia siab uas yuav tsum muaj ob qho tib si T. hlwb thiab macrophages thiab uas tsis tau pom nyob rau hauv proliferating qog hlwb nyob rau hauv tib yam mob. Muab ua ke, cov ntaub ntawv no qhia tau hais tias kev txhim kho lub peev xwm ntawm senescent hlwb kom nkag siab txog microenvironmental IFN ua hauv kev hais kwv txhiaj nrog SASP-stimulated lub cev tiv thaiv kab mob los pab txhawb kev sib koom ua ke heterotypic kev sib cuam tshuam ntawm cov qog hlwb, macrophages, thiab activated T hlwb uas txhim kho antigen kev nthuav qhia thiab kev tiv thaiv kab mob. , ua rau muaj zog qog regressions.

Kev sib tham

Pab tau los ntawm tus qauv murine qog nyob rau hauv uas mob qog noj ntshav tiv thaiv kab mob tiv thaiv kev soj ntsuam senescence yog nyob rau hauv kev tswj cov caj ces nruj, peb qhia seb cov hlwb senescent hloov pauv lawv lub peev xwm li cas rau ob qho tib si xa thiab tau txais ib puag ncig cov cim (Ntxiv Fig. S16). Raws li cov kev paub txog kev laus laus laus, p53-tsav kev ua kom tsis muaj zog ua rau lub cev tsis muaj zog ntawm cov noob caj noob ces thiab ua rau SASP. Txawm li cas los xij, peb kuj tau pom muaj txiaj ntsig zoo ntawm cov noob qhia rau PM cov proteins, suav nrog ntau qhov kev loj hlob zoo receptors thiab cytokine receptors uas tau kwv yees kom hloov pauv li cas cov hlwb senescent teb rau ib puag ncig cov cim. Qhov tseem ceeb, txawm hais tias peb siv tus qauv mob qog noj ntshav raws li peb qhov kev sim thawj zaug, qhov zoo sib xws hauv kev qhia ntawm cov xov tooj ntawm tes-nto sensors thiab cov kev pab cuam gene sensitizing rau ib puag ncig cov cim tau pom nyob rau hauv ntau yam murine thiab tib neeg qog hlwb kho nrog senescence-inducing. cov neeg sawv cev, txhais tau hais tias qhov kev hloov pauv hloov pauv tau yog ib qho kev qhia dav dav ntawm lub xeev senescent.

Desert ginseng-Improve immunity (12)

cistanche cov txiaj ntsig rau txiv neej-ua kom muaj zog tiv thaiv kab mob

Ib txoj hauv kev tseem ceeb ntawm kev hnov ​​​​qab hloov pauv hauv cov hlwb senescent suav nrog hom II IFN signaling. Hauv peb cov qauv mob qog noj ntshav thiab thoob plaws txhua lub xeev senescent peb tau tshuaj xyuas, senescence yog nrog los ntawm cell-intrinsic transcriptional thiab protein qhia kev hloov pauv tau kwv yees los txhim kho kev taw qhia los ntawm exogenous IFN. Tseeb tiag, senescent hlwb muaj zog ntau dua activated IFN effectors teb rau IFN hauv vitro thiab hauv vivo, thiab ob qho tib si IFN effector txoj hauv kev thiab IFN nyob rau hauv ib puag ncig yuav tsum tau ua kom zoo CD8 T cell-mediated clearance ntawm senescent qog hlwb. Txawm hais tias txoj kev tsom xam ntawm senescent cell transcriptomes invariably txheeb xyuas hom II IFN signaling raws li ib tug enriched feature, overlaps ntawm hom I thiab II signaling Cheebtsam thiab qhov tseeb hais tias IFN yog feem ntau tsis pom raws li ib tug SASP yam tseem tshuav cov lus nug txog yam II IFN signaling nyob rau hauv senescence. loj heev unexplored. Peb cov kev tshawb fawb pom tau hais tias qhov kev txhawb nqa zoo li no hauv IFN kos npe kos npe ntawm cov hlwb senescent qhia txog qhov muaj peev xwm txhim kho rau IFN sensing uas nws cov zis yog qhov tseem ceeb tshaj plaws hauv vivo.

Figure 7. IFNγ signaling in senescent tumor cells is necessary for immune surveillance. A, Ifngr1 KO of both proliferating and senescent NSP cells validated by flow cytometry. B, Tumor regression phenotype of Ifngr1 KO or control sgRNA–transfected tumor cells orthotopically injected into Bl/6N mice upon p53 restoration. A control sgRNA targeting a gene desert located on Chr8 (Ctrl KO) serves as a control. C, Tumor regression phenotype of parental NSP tumor cells orthotopically injected into WT or Ifng KO mice upon p53 restoration. D, Representative macroscopic images of tumor collected at day 21 after p53 restoration from C. E, Flow cytometry analysis of CD45 abundance in tumor from indicated groups. F, Representative immunofluorescence in p53-suppressed (proliferating) and p53-restored (senescent, 7 days after p53 restoration) tumor from the indicated host. NSP tumor cells were transduced with a GFP-expressing vector for visualization. Scale bars, 50 μm. Data are presented as mean ± SEM. Two-tailed Student t-test was used. **, P < 0.01; ***, P < 0.001.


Daim duab 7. IFN signaling hauv senescent qog hlwb yog tsim nyog rau kev tiv thaiv kab mob. A, Ifngr1 KO ntawm ob qho tib si proliferating thiab senescent NSP hlwb validated los ntawm ntws cytometry. B, Tumor regression phenotype ntawm Ifngr1 KO los yog tswj sgRNA-transfected qog hlwb orthotopically txhaj rau hauv Bl/6N nas thaum p53 kho. Kev tswj sgRNA tsom rau cov noob suab puam nyob ntawm Chr8 (Ctrl KO) ua haujlwm raws li kev tswj hwm. C, Tumor regression phenotype ntawm niam txiv NSP qog hlwb orthotopically txhaj rau hauv WT lossis Ifng KO nas thaum p53 kho. D, Cov neeg sawv cev macroscopic dluab ntawm cov qog tau sau nyob rau hnub 21 tom qab p53 rov qab los ntawm C. E, Flow cytometry tsom xam ntawm CD45 abundance nyob rau hauv cov qog los ntawm pawg qhia. F, Tus Neeg Sawv Cev immunofluorescence hauv p{{10}}suppressed (proliferating) thiab p53-rov qab (senescent, 7 hnub tom qab p53 kho) qog los ntawm tus tswv tsev qhia. NSP qog hlwb tau hloov nrog GFP-expressing vector rau kev pom. Scale bars, 50 μm. Cov ntaub ntawv raug nthuav tawm raws li txhais tau tias ± SEM. Ob-tailed Student t-test tau siv. **, P <0.01; ***, P <0.001.

Tej zaum qhov zoo tshaj plaws tsim tawm ntawm hom II IFN signaling cuam tshuam nrog nws lub peev xwm los ntxias cov antigen-kev nthuav qhia machinery. Tseeb, IFN induced cell-surface nthuav qhia ntawm MHC-I (los yog HLA nyob rau hauv tib neeg hlwb) nyob rau hauv peb cov qauv nyob rau hauv ob qho tib si proliferating thiab senescence tej yam kev mob. Txawm li cas los xij, IFN - induced MHC-I upregulation tau tshaj tawm nyob rau hauv cov hlwb senescent, ib qho kev cuam tshuam nrog kev nthuav qhia ntawm kev thauj mus los cuam tshuam nrog kev ua cov tshuaj antigen, lwm yam kev ua haujlwm antigen, thiab cov qauv ntawm MHC-I. Ib qho kev hnov ​​​​mob zoo sib xws rau IFN hauv kev ua rau MHC-I / HLA tau pom nyob rau hauv tib neeg lub siab thiab lub ntsws qog nqaij hlav hlwb ua rau muaj kev ntxhov siab. Cov txiaj ntsig no txhais tau hais tias qhov kev pab cuam senescence tuaj yeem txhim kho qhov kev nthuav qhia antigen hauv cov hlwb uas tsis muaj lub cev tiv thaiv kab mob, yog li ua kom cov qog tiv thaiv kab mob. Peb cov txiaj ntsig tau txhawb nqa tus qauv uas qhov kawg cuam tshuam ntawm cov hlwb senescent ntawm cov ntaub so ntswg biology yog txiav txim los ntawm kev sib xyaw ua ke ntawm qhov lawv xa thiab tau txais cov cim ib puag ncig. Tsis tsuas yog ua cov hlwb senescent induce lub SASP, uas ua rau cov ntaub so ntswg hloov kho thiab hloov lub xeev ntawm tes thiab muaj pes tsawg leeg ntawm lub cev tiv thaiv kab mob nyob rau hauv ib puag ncig, tab sis lawv kuj hloov pauv lawv cov surfaceome, uas ua rau muaj peev xwm sib txawv rau kev nkag siab ib puag ncig yam, ntawm no ua piv txwv los ntawm IFN . Qhov tseem ceeb, kev cuam tshuam ntawm IFN signaling tsis muaj kev cuam tshuam rau senescence induction lossis SASP hauv peb lub cev, tseem ua rau cov qog nqaij hlav tom qab, qhia tias kev hloov pauv ib puag ncig kev hnov ​​​​lus hauv kev hais kwv txhiaj nrog SASP los txiav txim siab qhov txiaj ntsig kawg ntawm qhov kev pab cuam senescence-hauv qhov no, tiv thaiv kab mob. Cov teebmeem no tshwm sim los ua ib feem ntawm kev sib koom ua ke ntawm epigenetic program, vim tias ob qho tib si SASP thiab cov kev pabcuam pom tau pom muaj kev vam khom tseem ceeb ntawm chromatin remodeling factor BRD4.

Txawm hais tias cov txheej txheem ntawm kev tiv thaiv kab mob hauv peb cov qauv yog nyob ntawm kev sib koom tes ntawm CD8 T-cell thiab macrophage cov neeg muaj kev cuam tshuam ntawm kev hloov pauv ntawm "kev tiv thaiv kab mob khaub thuas" mus rau "lub cev tsis muaj zog" qog microenvironment, lwm yam innate lossis adaptive immune cell hom yuav paub. thiab ntshiab senescent hlwb nyob rau hauv ntau lub ntsiab lus, los yog, hloov, kev tiv thaiv kab mob yuav tsis tshwm sim txhua (18, 19). Undoubtedly, qee qhov kev sib txawv no muaj kev cuam tshuam txog heterogeneity hauv SASP yam zais cia (13, 14), txawm hais tias peb cov txiaj ntsig tau nce qhov muaj peev xwm hais tias qhov ntau thiab qhov xwm txheej ntawm kev hloov pauv ib puag ncig kev hnov ​​​​lus kuj tseem cuam tshuam li cas senescent hlwb cuam tshuam rau cov ntaub so ntswg biology. Txawm hais tias knockout ntawm IFN sensing (ntawm Ifngr1 KO) thiab tshem tawm ntawm MHC-I (B2M KO) nyob rau hauv cov qog nqaij hlav hlwb ua rau lawv lub cev tiv thaiv kab mob hauv vivo, nws tsis tau tshem tawm cov qog regression tom qab senescence induction, qhia tias IFN sensing nyob rau hauv senescent hlwb yog. tsis yog tib txoj kev uas ua rau cov qog regression. Txawm li cas los xij, qhov tseeb tias cov hlwb muaj peev xwm teb tau txawv ntawm ib puag ncig cov cim qhia tau hais tias lawv qhov kawg ntawm lub xeev molecular hauv cov ntaub so ntswg yuav txawv dua li hauv kab lis kev cai ntawm tes, qhia txog qhov yuav tsum tau ua kom zoo dua cov txheej txheem hauv vivo.

Peb cov txiaj ntsig tuaj yeem pab piav qhia txog qhov cuam tshuam tsis zoo ntawm senescence biology hauv physiology thiab kab mob thiab muaj feem cuam tshuam rau kev siv cov tshuaj kho senescence-modulating zoo. Piv txwv li, nyob rau hauv peb cov qauv, qhov sib txawv ntawm cov qog senescent cell clearance thiab persistence tau txiav txim, tsawg kawg yog ib feem, los ntawm lub xub ntiag ntawm ib puag ncig IFN thiab kev ncaj ncees ntawm hom II IFN signaling nyob rau hauv lub senescent hlwb. Qhov no qhia tau hais tias kev hloov pauv ntawm lub peev xwm ntawm cov hlwb senescent los nrhiav thiab paub txog IFN -secreting immune cells lossis lwm yam kab mob hauv lub cev tuaj yeem cuam tshuam rau kev tshem tawm ntawm tes, xws li kev txo qis ib puag ncig IFN lossis txo qis hom II IFN signaling tuaj yeem ua rau senescent cell persistence hauv cov ntaub so ntswg. Nyob rau hauv cov ntsiab lus ntawm kev mob qog noj ntshav, kev kho mob uas ua rau cov qog cell senescence-ib txoj haujlwm cytostatic- tuaj yeem ua rau lub cev tiv thaiv kab mob qog noj ntshav lossis rov ua kom cov qog ua kom lub cev tiv thaiv kab mob, tab sis cov no tsis yog qhov tshwm sim thoob ntiaj teb. Raws li xws li, heterogeneity hauv SASP (uas tuaj yeem sib txawv ntawm hom qog cell thiab senescence inducers) lossis IFN sensing thiab tso zis [tej zaum cuam tshuam los ntawm kev tshem tawm lossis hloov pauv ntawm IFN txoj hauv kev lossis HLA Cheebtsam (60) lossis cov txheej txheem hloov pauv hloov pauv tsis pom ntawm no] tuaj yeem cuam tshuam. kev ua tau zoo ntawm cov kev kho mob zoo li no hauv cov neeg mob. Raws li qhov kev xav no, kev kho-tsav induction ntawm SASP profile tshwj xeeb kwv yees cov txiaj ntsig ntawm tus neeg mob hauv pawg neeg mob qog noj ntshav ntawm zes qe menyuam (61). Los ntawm qhov sib txawv, cov tswv yim los txhim kho lub cev tiv thaiv kab mob ntawm cov hlwb senescent los ntawm kev ua kom lawv rhiab heev rau IFN (piv txwv li, nrog PTPN2 inhibitors) tuaj yeem pab txhawb kev tsis ncaj ncees rau cov qog cell rejection. Peb xav tias kev tshawb xyuas qhov no thiab lwm yam kev kho cov ntaub so ntswg thiab kev soj ntsuam cov kev pab cuam nyob rau hauv pre- thiab posttreatment qog nqaij hlav biopsies (xws li, los ntawm transcriptomic los yog proteomic profiles) yuav nthuav tawm cov lus teb tshiab biomarkers thiab / los yog cov tswv yim sib xyaw ua ke los txhim kho kev tswj xyuas kev mob qog noj ntshav.

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