Cell179, October 31, 2019 a 2019?
Cellular Senescence: Defining a Path Forward
Definition and Characteristics of Cellular Senescence?
Cellular senescence is a cell state?
triggered by ：stressful insults and certain physiological processes,
characterized by ：aprolongedand generally irreversible cell-cycle arrest withsecretory features, macromolecular damage, andaltered metabolism?
These features can be inter-dependent?
Cell-Cycle Arrest?
mammalian cells?
retinoblastoma(RB) familyandp53 proteinsare important for establishing senescent cell-cycle arrest?
RB1 and its family members p107 (RBL1) and p130 (RBL2) are phosphorylated by specific cyclin-dependent kinases (CDKs; CDK4, CDK6, CDK2).?
This phosphorylation reduces the ability of the RB family members torepressE2F family transcription factor activity, which is required for cell-cycle progression?
CDK2 inhibitor p21WAF1/Cip1 (CDKN1A) and CDK4/6 inhibitor p16INK4A (CDKN2A) accumulate.?
Secretion?
SASP的功能
the SASP reinforces and spreads senescence in autocrineandparacrinefashions?
activates immune responses that eliminate senescent cells?
SASP factors mediate developmental senescence?
tissue plasticity?
?contribute to persistent chronic inflammation (known as inflammaging)?
the SASP can recruit immature immune-suppressive myeloid cells to prostate and liver tumors （肝臟腫瘤！＜鼐铡茴她！看一下Distinct Functions of Senescence-Associated Immune Responses in Liver Tumor Surveillance and Tumor Progression?
）
控制SASP的相關(guān)通路
enhancer remodeling?
activation of transcription factors, such as NF-kB, C/EBPb, GATA4?
mammalian target of rapamycin(mTOR)?
p38MAPK signaling pathways?
上游因子控制SASP的方式不同并且和老化誘導(dǎo)的途徑有關(guān)
type 1 interferon response：DNA damage, cytoplasmic chromatin fragments (CCFs)?
inflammasome ：damage-associated molecular patterns (DAMPs)?
SASP的成分和強度不同決定于：duration of senescence, origin of the prosenescence stimulus, and cell type?
SASP可以和外界的微環(huán)境通過：juxtacrine NOTCH/JAG1 signaling猫缭；release of ROS誊薄；cytoplasmic bridges；extracellular vesicles （egexosomes）
）
Macromolecular Damage?
DNA Damage ：The first molecular feature associated with senescence wastelomere shortening, a result of the DNA end-replication problem, during serial passages?
端粒導(dǎo)致DDR的過程
Type1
Telomere shortening during proliferation?
telomeric DNA loop destabilization &telomere uncapping?
generating telomere dysfunction-induced foci (TIFs)?
ctivate the DDR?
cell-cycle arrest?
（This response can also be elicited by inhibiting or altering genes involved in telomere maintenance ）
Type2
oxidative DNA damage at telomeric G-reach repeats?
telomere-associated foci (TAFs), can exist at telomeres （irrespective of telomere length or shelterin loss ）
DNA損傷種類
half of the persistent DNA damage foci in senescent cells localize to telomeres
other stressful subcytotoxic insults can trigger senescence by inducing irreparable DNA damage?
genotoxic agents： radiation (ionizing and UV),
?? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? pharmacological agents (e.g., certain chemotherapeutics),
?? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? oxidative stress?
activated oncogenes can induce senescence (known as OIS) as a tumor-suppressive response, restricting the uncontrolled proliferation of potentially oncogenic cells.?
——OIS is often mediated by the tumor suppressorsp16INK4A andARF, both encoded by theCDKN2A locus, imposing a cell-cycle arrest?
DDR也可以產(chǎn)生OIS的激活
oncogene-driven hyperproliferation→damage signal originates at collapsed replication forks?
DNA-SCARSs (DNA segments with chromatin alter- ations reinforcing senescence) ：Senescent cells harbor persistent nuclear DNA damage foci?
特點：associate with promyelocytic leukemian (PML) nuclear bodies （？？很泊？）
?? ? ? ? ? lack theDNA repair proteinsRPAandRAD51as well as single- stranded DNA (ssDNA)?
?? ? ? ? ? contain activated forms of the DDR mediatorsCHK2andp53?
DNA-SCARSs 可以調(diào)節(jié)細(xì)胞生長停滯和SASP，但是并不是一個老化細(xì)胞的特征
Protein Damage?
Proteotoxicity is a hallmark of aging and cellular senescence?
damaged proteins help identify senescent cells?
蛋白質(zhì)損傷的途徑
ROS?
↓
oxidize both methionine and cysteine residues?
↓
alter protein folding and function?
eg
（蛋氨酸和半胱氨酸沾谓，很多絡(luò)氨酸磷酸酶都含有cys委造，因此他們所去磷酸化的部位就會被inactivate）PTPs：protein tyrosine phosphatases?
↓
hyperactivating ERK signaling （similar to the effect of activated oncogenes ）
↓
trigger senescence?
在腫瘤前期的病變處可以發(fā)現(xiàn)高磷酸化的ERK
Most protein oxidative damage：degradation by the ubiquitin proteasome system(UPS) orautophagy因此這兩個特點可以作為檢測老化情況的指標(biāo)
promyelocytic leukemian (PML) 小體可以作為ROS和氧化損傷的sensor；同樣也可以作為細(xì)胞老化的一個非特異性指標(biāo)
Lipid Damage?
老化細(xì)胞的清除可以減少老年小鼠肝臟和大腦中脂肪的沉積
（特定的脂代謝和老化相關(guān)知之甚少）
雖然很多方法可以檢測脂肪在組織和細(xì)胞之前的變化均驶，但是作為老化的biomarker還是未知的昏兆，因為不同的老化相關(guān)的脂肪十分不同。
比如在原癌基因激活的老化和復(fù)制性老化中脂肪代謝非常不同
Deregulated Metabolic Profile?
Mitochondria?
Mitochondria in senescent cells are less functional, showingdecreased membrane potential,increased proton leak, reduced fusion and fission rates, increased mass, andabundance of tricarboxylic acid (TCA) cycle metabolites?
Altered AMP:ATP and ADP:ATP ratios during senescence?
↓
activating AMPK (AMP-activated protein kinase), a main sensor of energy deprivation?
↓
contribute to cell-cycle withdrawal?
線粒體的功能失調(diào)和SASP的調(diào)節(jié)也相關(guān)
Mitophagy (mitochondrial clearance) in senescent cells appears tosuppressthe SASP?
genetic or pharmacological inhibition of the ETC（electron transport chain ） can induce senescence even though cells lack expression of key pro-inflammatory SASP factors
?NAD+/NADH ratios are reduced n senescent cells →alter the activity of poly-ADP ribose polymerase (PARP) and sirtuins （乙醺狙ǎ化酶）——both involved in activation of the SASP regulatorNF-kB?
lysosomes
?溶酶體的代謝是有細(xì)胞的代謝或者降解需要決定的
當(dāng)AA在溶酶體的腔內(nèi)含量比較高的時候爬虱，mTOR1 is recruited and activated?
并且溶酶體可以和線粒相互關(guān)聯(lián)
老化的細(xì)胞中溶酶體的數(shù)量和大小都有所增加，但是活性并沒有增加
↓
lysosome-mitochondrial axis degrades?
↓
decreased mitochondrial?
↓
ncreases ROS production?
↓
ROS targets cellular structures, including lysosomes, which forms a vicious feedback loop that induces more damage
溶酶體的數(shù)量和SA-b-gal 的活性有關(guān)
從治療角度來講腾它，溶酶體增大的三部分可以為捕捉藥物增加空間跑筝，比如CDK4/6 in- hibitors palbociclib哌柏西利, ribociclib, and abemaciclib?
Senescence-Associated (Epi-)genetic and Gene Expression Changes
上述的表型變化大多是由于基因的表達(dá)所導(dǎo)致，這里我們解釋下相關(guān)基因的情況
老化相關(guān)基因查詢網(wǎng)址(http://Senequest.net)?
Chromatin Landscape?
表關(guān)遺傳的變化大多和環(huán)境相關(guān)瞒滴。replicative senescence has been correlated with global loss of DNA methylation at CpG sites 继蜡；Cells undergoing OIS fail to show such alterations in DNA methylation?
老化的細(xì)胞呈現(xiàn)普遍的increase in chromatin accessibility, but the genome-wide profilevariesdepending on the stimulus?
大多是組蛋白的變化individual histone modifications and variants
H4K16ac is often enriched at active pro- moters in senescent, but not proliferating, cells?
N terminus proteolytic cleavage of H3.3 correlates with gene repression in a different subset of genes during senescence?
Certain histone modifications are crucial for senescence, such as elevatedH4K20me3andH3K9me3,which contribute to theproliferation arrest ；elevatedH3K27acat gene enhancers promotes aSASP?
Senescence is also associated with chromatinmorphological changes.?
Senescence-associated heterochromatin foci (SAHFs), visualized as DAPI-dense foci, are enriched in heterochromatin protein (HP) 1.?
?SAHFs derive from chromatin factors—
?including RB,
? histone variant macroH2A,?
?high mobility group A proteins,?
?the HIRA/UBN1/CABIN1, a
?ASF1a chaperones—and increased nuclear pore density?
SAHF主要組成復(fù)制后基因的poor eterochromatic 部分，因此和細(xì)胞老化的相關(guān)性很小
細(xì)胞的老化同時也和大面積的H1丟失有關(guān)
enescence-associated distension of satellites (SADSs) ——先于SAHF稀并，并且和細(xì)胞老化相關(guān)
Retrotransposable elements
repressed LINE-1 (L1) retrotransposons are activated, stimulating the cGAS-STING pathway that elicits a type 1 interferon response 仅颇，從而和SASP相關(guān)
Downregulation of lamin B1（a major component of the nuclear lamina,）——也是老化細(xì)胞的一個主要特點
Lamin B1 和表觀基因、enescence-associated chromatin structures (SAHFs and SADSs) 相關(guān)
主要發(fā)生在H3K9me3-rich regions：liberate H3K9me3 from the nuclear lamina promoting spatial rearrangement of H3K9me3 heterochromatin to form SAHFs 尤其是在OIS中發(fā)揮作用碘举，然而復(fù)制性老化并沒有什么相互作用
lamin B1 loss and reduced nuclear integrity 也可以促進(jìn)SASP的形成
Transcriptional Signatures?
genes linked to thecell-cycle arrestand SASPare frequently interrogated in combination with other biomarkers to validate the senescence phenotype or type of senescence?
eg：CDKN1A (p21WAF1/Cip1), CDKN2A (p16INK4A), and CDK2B (p15INK4B), and a subset of SASP genes, along with decreased expression of cyclinsCCNA2andCCNE2andLMNB1 should be determined.
為了更好研究轉(zhuǎn)錄情況——Whole-transcriptome studies忘瓦，可以預(yù)測藥物靶向；
但是現(xiàn)在的轉(zhuǎn)錄因子數(shù)據(jù)還是不足引颈，因此相關(guān)轉(zhuǎn)錄因子的發(fā)現(xiàn)還是很重要的
?miRNAs and Non-coding RNAs （）
尤其是miRNA和細(xì)胞老化的關(guān)聯(lián)更大耕皮；可以直接或者間接的調(diào)節(jié)老化關(guān)鍵因子，
「p53, p21WAF1/Cip1, and SIRT1 」
negative：miR-504 targets the p53 30UTR, reducing p53 abundance and activity?
? ? ? ? ? ? ? ? ? Gld2-mediated stabilization of miR-122 enables its binding to the CBEP 30 UTR, resulting in decreased p53 mRNA polyadenylation and translation?
? ? ? ? ? ? ? ? ? multiple miRNAs downregu- late p21WAF1/Cip1, including 28 miRNAs that block OIS?
? ? ? ? ? ? ? ? ? miR-24 suppresses p16INK4a?
positive：miR-605 targets MDM2, triggering p53-mediated senescence?
miRNA feedback loops can modulate senescence programs.?
——For example, ap53/ miRNA/CCNA2 pathwaydrives senescence independently of the p53/p21WAF1/Cip1 axis 蝙场；p53- dependent upregulation ofmiR-34a/b/cdownregulates cell proliferation and survival factors?
「regulate the SASP 」
MiR-146a/b ：dampens a proinflammatory arm of the SAS?
miRNAs also downregulate repressors of senescence, including polycomb group(PcG)members CBX7, EED, EZH2, and SUZ12 (miR-26b, 181a, 210, and 424), leading top16INK4a derepressionand senescence initiation?
「the role of miRNAs in senes- cence extends beyond their classical functions」
Argonaute 2 (AGO2) binds let-7f in the nucleus, forming a complex with RB1(pRB), resulting inrepressivechromatin atCDC2andCDCA8promoters . Silencing theseE2Ftarget genes is required for senescence initiation.?
Long non-coding RNAs (lncRNAs) (>200 nt) canbind RNA, DNA, or proteinsto regulate senescence.?
ANRIL, a 30-40kb antisense transcript encoded by the CDKN2A locus, binds CBX7 torepress INK4b/ARF/INK4a expression?
lncRNA PANDA recruits PcG com- plexes, suppressing senescence-promoting genes?
silencing ofGUARDIN, a p53-responsive lncRNA, causes senescence or apoptosis?
following OIS induced by RAF, the lncRNA VADpreserves senescence by decreasing repressive H2A.Z deposition at INK promoters?
lncRNA UCA1 disrupts association of the RNA-binding protein hnRNP A1 with p16INK4A
Immune-Regulation and Anti-apoptotic Proteins?
雖然細(xì)胞老化的誘導(dǎo)環(huán)境可以激活一些炎癥反應(yīng)因子凌停，尤其是一些細(xì)胞表面的marker，可以作為研究從單個細(xì)胞到組織的方式售滤，但是DCR2 and NKG2D ligands are not conserved among species, making mouse-to-human comparisons not possible.?
Notch1 in OIS and DPP4 in replicative and OIS：upregulated cell surface markers, 并且對于調(diào)節(jié)SASP有著一定的作用
increased expression of anti-apoptotic BCL-2 family members 也可以罚拟，因為老化的細(xì)胞是有凋亡抵抗的。
in Vivo Models to Study Cellular Senescence?
Senescence Reporter Mice
通過報告基因estimatep16Ink4a expression?
Murine Models of Accelerated Senescence and Aging?
（各種早衰小鼠模型的建立方法）
Identification of Cellular Senescence In Vivo
A Simplified Algorithm for Detecting Senescent Cells In Situ
實驗室免疫染色/WB+用顯微鏡觀察下
不同方法聯(lián)合檢測
Challenges to Detect Senescent Cells in Humans?
將細(xì)胞的老化和人類的疾病聯(lián)系起來的
neurodegenerative disorders, glaucoma青光眼, cataract白內(nèi)障完箩， atherosclerosis and cardiovascular disease, diabetes, osteoarthritis, pulmonary, and renal and liver fibrosis?
組織的檢測可以：fresh samplesby SA-b-gal staining?
?or indirect markers in formalin-fixed tissues?
histochemical dye SBB interacts with lipofuscin, another hallmark of senescent cells?
Lipofuscin is preserved in fixed material?
reagent (GL13) is amenable to immunohistochemistry （identified senescent Hodgkin and Reed-Sternberg (HRS) cells in Hodgkin lymphomas (cHL) where they predicted poor prognosis ）
Another method for identi- fying and quantifying senescent cellsin vivo is SA-b-gal staining combined with ImageStream X analysis?
We recommendcombining cytoplasmic (e.g., SA-b-gal, lipofuscin), nuclear (e.g., p16INK4A, p21WAF1/Cip1, Ki67) and SASP, context and/or cell-type-specific markers?
老化相關(guān)仍然存在的問題
不可逆的細(xì)胞周期停滯并不一定存在所有的老化細(xì)胞中赐俗！
遺傳學(xué)和表觀遺傳學(xué)在細(xì)胞老化中的關(guān)聯(lián)？
在不同的老化誘導(dǎo)條件下細(xì)胞的修復(fù)方式又是什么弊知？
抗衰老治療的應(yīng)用阻逮？
（我個人覺得mirNA&Non-coding RNAs是不是也挺熱門的）
