Hypothesis

Aging neurons convert dangerous misfolded proteins into chaperone‑rich aggregates that serve as reversible storage depots for select RNA‑binding proteins (RBPs). These RBPs remain functionally silent but intact within the aggregate matrix and can be liberated by disaggregase activity to restart translation during recovery phases.

Mechanistic Basis

1. Phase‑separation seeding – Stress‑induced low‑complexity regions of RBPs drive transient liquid droplets that mature into amyloid‑like fibrils, a process observed in both physiological stress granules and pathological aggregates 1.
2. Chaperone coating – Hsp70/Hsp90 and small Hsps bind the fibril surface, creating a protective shell that prevents further seeding while keeping incorporated RBPs in a near‑native conformation 2.
3. Post‑translational gating – Phosphorylation of RBPs lowers their affinity for the fibril core, acting as a molecular switch that favors release when kinase activity drops 3.
4. Disaggregase extraction – The HSP110‑HSP70‑HSP40 complex can thread polypeptides out of the fibril without dissolving the core, delivering RBPs to the cytosol in a refold‑competent state 4.

Testable Predictions

• P1: Elevating disaggregase levels in aged brains will decrease the fraction of chaperone‑rich aggregates w

Hypothesis

Aged skeletal muscle loses eosinophils not because of deficient IL‑5 production but because resident eosinophils fail to sustain an autocrine IL‑33 loop that normally maintains their survival and IL‑4‑secreting capacity. Restoring this IL‑33→IL‑5 axis within muscle will rescue eosinophil numbers, improve regeneration, and attenuate age‑related fibrosis.

Background

• Eosinophils support muscle repair by secreting IL‑4, which expands fibro/adipogenic progenitors (FAPs) while blocking adipogenic differentiation [1].
• IL‑5 drives eosinophil differentiation, survival, and tissue homing, but IL‑33 acts upstream, expanding IL‑5Rα⁺ precursors and inducing systemic IL‑5 [3].
• With age, eosinophils decline in visceral adipose tissue, correlating with elevated CCL11/CCL2 and systemic inflammation [4]; young eosinophil transfer reverses these effects in an IL‑4‑dependent manner.
• Direct eosinophil‑satellite cell interactions in muscle remain poorly defined, raising the possibility that eosinophil‑derived factors beyond IL‑4 influence myogenesis.

Mechanistic insight

We propose that, in addition to the circulating IL‑33→IL‑5 axis, muscle‑resident eosinophils produce IL‑33 in response to damage‑associated molecular patterns (DAMPs) released during injury. This locally secreted IL‑33 engages the IL‑33 receptor (ST2) on eosinophils themselves, activating NF‑κB and u

Hypothesis

Age-related loss of MSC positional identity is not solely an intrinsic epigenetic clock but is amplified by microbiome-derived signals that travel the gut‑brain‑vagus axis to alter HOX locus chromatin states.

Mechanistic Link

Aged microbiota increase production of medium‑chain fatty acids (MCFAs) such as isovaleric acid, which activate gut myeloid cells to release inflammatory mediators. These mediators activate vagal afferents, altering neurotransmitter release in the bone marrow niche. Norepinephrine and acetylcholine fluctuations influence MSC histone acetyltransferase and deacetylase activity, leading to hyper‑methylation of HOXB1, HOXA3 and HOXC‑AS3 enhancers and reduced ATAC‑seq signal [2,3,4]. The resulting positional memory loss impairs myogenic differentiation, linking gut inflammation to stem cell functional decline.

Predictions & Experimental Design

1. Block vagal signaling – Perform subdiaphragmatic vagotomy in aged mice or administer a vagal antagonist (e.g., atropine locally to the gut). Prediction: MSCs isolated from bone marrow will show restored HOX enhancer accessibility (ATAC‑seq) and reduced methylation at HOX loci compared with sham‑operated controls.
2. Neutralize MCFA – Treat aged mice with an antibody or small molecule that binds isovaleric acid, or feed them a diet low in precursors for MCFA. Prediction: Decrease in circulating isovaleric acid will lower my

Hypothesis

Senescent immune cells create a localized NAD+ sink through SASP‑induced CD38 overexpression, which depletes NAD+ in stromal and parenchymal cells, suppresses SIRT1‑mediated deacetylation, triggers mitochondrial dysfunction, and propagates senescence beyond the immune compartment. Restoring NAD+ specifically within immune niches breaks this feed‑forward loop and delays organismal aging.

Mechanistic Model

• Step 1: Immune senescence – Aging lymphoid and myeloid compartments accumulate senescent cells that secrete IL‑6, IL‑8, TNF‑α and upregulate CD38 (3).
• Step 2: NAD+ consumption – CD38 hydrolyzes NAD+ to generate ADPR and cADPR, lowering intracellular NAD+ levels in nearby non‑immune cells.
• Step 3: SIRT1 inhibition – Reduced NAD+ diminishes SIRT1 deacetylase activity, leading to hyperacetylation of PGC‑1α, FOXO, and NF‑κB subunits, which impairs mitochondrial biogenesis and increases ROS production.
• Step 4: Mitochondrial damage & secondary senescence – Dysfunctional mitochondria release mtDNA and ROS, activating the cGAS‑STING pathway and reinforcing SASP expression in parenchymal cells, thereby spreading senescence.
• Step 5: Feedback amplification – Newly senescent parenchymal cells additionally secrete cytokines that further stimulate CD38 expression in immune cells, tightening the loop.

This model places immune‑derived NAD+ depletion upstream of tissue‑specific aging, extending the observation that transplanting aged splenocytes induces senescence in young tissues ([1](https://doi

Hypothesis

Aging colonic fibroblasts develop a senescence-associated secretory phenotype (SASP) that alters mucin O‑glycosylation, reducing the binding affinity of mucin for butyrate‑producing Firmicutes (e.g., Roseburia, Faecalibacterium) while preserving or enhancing adhesion for mucin‑degrading Bacteroidetes. This shift in the mucosal niche explains the biphasic F/B ratio: early‑life increases reflect expansion of Firmicutes as the mucosal surface matures; after ~70 years, fibroblast‑driven mucin remodeling creates a permissive environment for Bacteroidetes, lowering the ratio despite continued loss of butyrate genes[1].

Mechanistic Rationale

1. Senescent fibroblasts secrete MMP‑9 and TGF‑β, which activate colonic epithelial glycosyltransferases (e.g., C1GALT1) leading to increased sialylation and fucosylation of MUC2[2].
2. Altered glycan motifs diminish lectin‑like interactions with Firmicutes surface adhesins that preferentially bind terminal galactose/N‑acetylglucosamine residues, whereas Bacteroidetes utilize polysaccharide‑utilization loci (PULs) that recognize sialylated/fucosylated glycans[3].
3. Consequently, butyrate‑producing Firmicutes lose mucosal residence, drop luminal concentrations, and their metabolic output (butyrate) falls, triggering colonocyte energy stress, AMPK activation, and barrier leak as previously described[4].
4. The resulting LPS influx fuels inflammaging, which further reinforces fibroblast senescence via NF‑κB signaling, establishing a positive feedback loop[5].

Testable Predictions

• Prediction 1: Colonic fibroblast isola

Hypothesis

A twelve‑month regimen that combines daily time‑restricted feeding (TRF), moderate‑intensity aerobic exercise, and a specific prebiotic fiber blend will produce greater improvements in thymic output and systemic inflammation than either rapamycin or anti‑IL‑6 monoclonal antibody monotherapy in adults aged 65‑80.

Mechanistic Rationale

• TRF and exercise raise NAD⁺ levels, activating SIRT1 in thymic epithelial cells (TECs). SIRT1 deacetylates FOXN1, driving its transcription and promoting TEC differentiation, which expands naïve T‑cell output【https://pmc.ncbi.nlm.nih.gov/articles/PMC12711513/】.
• Exercise‑induced irisin shifts macrophage polarization toward M2 and upregulates Foxp3 in regulatory T cells, while the accompanying IL‑6 transient differs from chronic inflammatory IL‑6 signaling【https://pmc.ncbi.nlm.nih.gov/articles/PMC12784853/】.
• Prebiotic fiber fermentation yields butyrate, which binds GPR109A on hematopoietic stem cells (HSCs), biasing lymphoid lineage commitment and limiting expansion of pro‑inflammatory myeloid cells【https://pmc.ncbi.nlm.nih.gov/articles/PMC12711513/】.
• The three interventions concurrently hit SASP‑driven inflammation, HSC lineage skewing, thymic involution, and metabolic dysregulation, breaking the self‑reinforcing loop where chronic inflammation impairs immunity and weakened immunity fails to clear senescent cells.

Predictions

1. Thymic output (measured by circulating T‑cell receptor excision circles) will increase ≥30 % in the combination arm, versus <10 % in each monotherapy arm.
2. **Plasma inflamm

Hypothesis

Rapamycin extends lifespan by imposing a famine‑like epigenetic state that enhances stress resistance but simultaneously dampens tissue‑regenerative pathways; intermittent withdrawal permits re‑activation of these pathways, allowing damage clearance when combined with youthful circulating factors.

Mechanistic Basis

Caloric restriction and rapamycin both inhibit mTORC1, yet transcriptomic profiles diverge [2]. We propose that the shared mTORC1 inhibition triggers a conserved chromatin response: increased histone deacetylase activity and reduced H3K27ac at promoters of growth‑related genes, mimicking the chromatin landscape observed during natural famine [1]. This epigenetic shift boosts autophagy and FOXO‑mediated stress resistance, explaining the healthspan gains without removing existing lesions [3]. Importantly, the same repressive marks also silence stem‑cell niche regulators (e.g., Sox9, Pax7) and senescence‑clearance mediators, accounting for the limited reversal of damage seen with rapamycin alone.

Prediction

If the epigenetic famine state is reversible, then scheduled rapamycin pulses followed by drug‑free intervals will allow transient re‑opening of regenerative chromatin, enabling senescent cell clearance and DNA‑repair upregulation. Adding young plasma heterochronic exposure during the drug‑free window should synergize, producing measurable damage reduction beyond what either intervention achieves alone.

Experimental Design

1. Treatment groups (mice, 20 months old): (a) continuous rapamycin; (b) intermittent rapamycin (5 days on/2 day

Background

The hypothalamus regulates systemic aging via IKKβ/NF‑κB–mediated repression of GnRH. Microglial‑derived TNF‑α activates IKKβ/NF‑κB in neighboring neurons, establishing a feed‑forward loop that suppresses GnRH promoter activity by ~50 % [1]. Chronic metabolic overload and insulin/IGF‑1 signaling further amplify NF‑κB through ER stress and PI‑3K/AKT/mTOR pathways [2] [3]. Downstream, NF‑κB induces c‑Fos/c‑Jun and PKC isoforms that repress GnRH transcription, linking hypothalamic inflammation to reduced adult neurogenesis [1]. A critical gap remains: how mitochondrial dysfunction in microglia translates into neuronal NF‑κB activation beyond TNF‑α signaling.

Hypothesis

Aged microglia release extracellular vesicles enriched in oxidized mitochondrial DNA (mtDNA) and cardiolipin. These vesicles act as damage‑associated molecular patterns (DAMPs) that are taken up by hypothalamic neurons, where mtDNA engages endosomal TLR9 and cardiolipin activates the NLRP3 inflammasome. The resulting caspase‑1‑dependent release of IL‑1β and reactive oxygen species (ROS) amplifies IKKβ/NF‑κB signaling in a TNF‑α–independent manner, further suppressing GnRH synthesis.

Mechanistic Rationale

1. Mitochondrial stress in microglia – Age‑related ROS production damages mitochondrial membranes, causing release of mtDNA and oxidation of cardiolipin into the cytosol.
2. **Packag

Hypothesis

Berberine’s low-dose activation of lysosomal AMPK drives TFEB-dependent lysosomal biogenesis and autophagic flux, leading to post-translational clearance of PCSK9 protein. This mechanism operates alongside its transcriptional inhibition of PCSK9 and explains the dose- and formulation-dependent inconsistencies observed across models. When combined with metformin, which activates cytosolic AMPK via a PEN2-dependent pathway, the two drugs achieve complementary spatial AMPK signaling (lysosomal vs cytosolic), resulting in synergistic metabolic improvements.

Testable Predictions

• Prediction 1: In HepG2 cells treated with berberine (0.1–2.5 µM), lysosomal AMPK phosphorylation (p-AMPKα-Lys) will increase, accompanied by TFEB nuclear translocation and up-regulation of lysosomal genes (LAMP1, CTSB). siRNA-mediated TFEB knock-down will block berberine-induced reduction of PCSK9 protein without altering PCSK9 mRNA levels.
• Prediction 2: Metformin will increase cytosolic AMPK phosphorylation (p-AMPKα-Cyt) but not lysosomal AMPK or TFEB activation. The combination of berberine + metformin will produce additive increases in both lysosomal and cytosolic p-AMPK, correlating with greater LDLR upregulation and LDL-C clearance.
• Prediction 3: In vivo, high-fat diet mice given berberine will show elevated hepatic autophagic flux (LC3-II/I ratio, p62 degradation) and reduced PCSK9 protein; autophagy inhibition (chloroquine or ATG5 KO) will abolish the PCSK9-lowering effect while preserving mRNA suppression.

Mechanistic Rationale

Berberine’s AXIN1-depend

Hypothesis\nWe hypothesize that integrating spatial proteomics, metabolomics, and single-cell transcriptomics from serial liquid biopsies can identify pre-resistant niches in the tumor microenvironment that activate bypass signaling pathways (e.g., MET/HER2, VEGF) weeks before radiographic progression. This early detection will allow preemptive therapeutic adaptation, improving progression‑free survival in patients with EGFR‑mutant NSCLC undergoing TKI therapy.\n\n## Rationale\nMulti‑omics reveals functional layers missed by genomics, such as post‑translational activation of bypass receptors and TME-derived metabolites that drive resistance [1][2]. Spatial technologies now map where these active proteins and metabolites coexist with immune and stromal cells, revealing signaling hotspots that bulk assays miss. The TuPro trial showed serial biopsies altered therapy in 50% of cases, proving resistance evolves temporally [3]. By coupling serial liquid biopsies (ctDNA, exosomes) with spatial omics of circulating tumor‑derived microvesicles, we can capture the evolving TME without invasive repeats.\n\n## Predictions\n1. Patients whose baseline liquid‑biopsy spatial atlas shows co-localization of phosphorylated MET/HER2 with lactate-rich stromal zones will develop TKI resistance within 8 weeks, whereas those lacking this signature will remain progress

Core Hypothesis

Certain bioactive peptides may contribute to increased longevity by enhancing cellular repair, reducing chronic inflammation, and supporting metabolic efficiency — positioning peptides as viable targets for healthspan-extending interventions.

Scientific Rationale

Peptides function as endogenous signaling molecules with the capacity to modulate several hallmarks of aging:

1. Autophagy Enhancement

Specific peptide sequences have been shown to upregulate autophagy — the cellular quality-control process responsible for clearing damaged proteins and organelles. By optimizing autophagic flux, cells maintain homeostasis longer, potentially delaying senescence and age-related tissue deterioration.

2. Systemic Anti-Inflammatory Action

Chronic low-grade inflammation ("inflammaging") is a major contributor to age-related pathology. Certain bioactive peptides derived from food proteins or endogenous sources demonstrate inhibition of NF-κB signaling and reduction in pro-inflammatory cytokines (IL-6, TNF-α), potentially attenuating the inflammatory cascade underlying cardiovascular disease, neurodegeneration, and metabolic syndrome.

We hypothesize that coupling brief OSK expression cycles with timed senolytic interventions will prevent senescence rebound and lock in youthful DNA methylation states, thereby sustaining lifespan extension beyond factor withdrawal.

Recent work shows that systemic OSK gene therapy extends median remaining lifespan in old mice by 109% and reverses epigenetic age without teratoma formation [https://pmc.ncbi.nlm.nih.gov/articles/PMC10909732/]. However, rejuvenation effects decay once OSK expression stops, with methylation age rebounding after treatment ends [https://www.fightingaging.org/archives/2025/10/a-review-of-the-present-state-of-epigenetic-reprogramming-to-treat-aging/]. This durability loss coincides with a senescence rebound that occurs when OSKM exposure persists beyond day 13‑15, triggering a p16^INK4a‑dependent secretory phenotype that undermines the epigenetically youthful state [https://pmc.ncbi.nlm.nih.gov/articles/PMC6351826/].

We propose that the senescence burst induced by transient OSK is not merely a toxic side‑effect but a signal that can be harnessed. OSK activates a stress response that upregulates p16^INK4a and SASP factors in a subset of cells; removing those senescent cells shortly after each OSK pulse should interrupt the inflammatory feedback loop that drives remethylation of aging‑associated loci. Senolytic agents such as dasatinib plus quercetin efficiently clear p16^INK4a‑positive cells and have been shown to improve healthspan in aged mice [https://doi.org/10.1016/j.cell.2015.05.031]. By administering senolytics 24 h after each 3‑day OSK ON p

Hypothesis

Telomere length in colonic epithelium operates as an entropy‑sensing clock that predicts the velocity of tight‑junction (TJ) protein loss and downstream inflammaging, independent of mitotic history.

Rationale

Recent work shows age‑dependent decline of ZO‑1, occludin and JAM‑A in the primate gut, driving permeability‑mediated endotoxemia and inflammaging [1][2]. Oxidative stress and NLRP3 activation directly damage TJ complexes [5]. If telomeres quantify informational entropy rather than division count, then accelerated telomere attrition should mirror rising molecular disorder in epithelial cells, precipitating TJ gene dysregulation before any detectable change in proliferation markers.

Mechanistic Extension

1. Entropy‑driven chromatin destabilization – Shortened telomeres increase local heterochromatin loss, releasing repressive marks from TJ gene promoters (ZO‑1, occludin, JAM‑A). This transcriptional silencing occurs even when Ki‑67 indices are low, linking telomere entropy directly to barrier gene expression.
2. Telomere damage‑induced SASP – Persistent telomere‑associated DNA foci activate ATM‑p53 pathways, provoking a senescence‑associated secretory phenotype rich in IL‑1α, IL‑6 and MMPs. These factors phosphorylate myosin light chain kinase (MLCK), augmenting actomyosin contractility and physically pulling TJ strands apart [5] [[6](http

Hypothesis

We propose that glucosepane buildup in stromal collagen impairs integrin‑β1 mechanotransduction, triggering keratocyte senescence via p16^INK4a upregulation and a secretory phenotype that further disrupts collagen organization.

Mechanistic Basis

• It's known that glucosepane forms non‑reducible cross‑links that stiffen fibrils, altering the nanoscale rigidity sensed by keratocyte integrins (Human corneal keratocyte density declines linearly at approximately 0.3–0.45% per year).
• Increased matrix rigidity shifts focal adhesion kinase (FAK) signaling toward a pro‑senescent state, raising nuclear p16^INK4a and lowering proliferative capacity (The aging stroma exhibits increased collagen cross‑linking, fibril disorganization, thinning, and nerve fiber loss, disrupting transparency and biomechanics).
• Senescent keratocytes release matrix metalloproteinases and TGF‑β isoforms that degrade ordered lamellae and encourage aberrant fibril bundling, creating a feed‑forward loop of disorganization.
• This mechanistic link explains why metabolic improvements (e.g., after bariatric surgery) can partially restore keratocyte density (Keratocyte density reductions in severe obesity can partially reverse following bariatric surgery): reduced systemic glucose flux lowers glycation and glucosepane formation.

Testable Predictions

1. In donor corneas grouped by age, glucosepane levels will positively correlate with p16^INK4

Hypothesis

We hypothesize that a regimen of pulsed low‑dose IGF‑1 combined with oral BPC‑157 and intermittent TB‑500 will produce clinically meaningful improvements in pain and tendon structure in humans with chronic tendinopathy, while keeping mTORC1 activity (as measured by phospho‑S6K in peripheral blood mononuclear cells) at baseline levels.

Rationale

• BPC‑157 accelerates tendon and bone healing in rodents via ERK1/2 signaling and nitric‑oxide‑mediated vasodilation[1]. These pathways promote fibroblast proliferation and collagen synthesis without directly stimulating mTORC1.
• TB‑500 (thymosin β4) sequesters G‑actin, facilitating actin polymerization and cell migration, which supports tenocyte infiltration into injured matrix. No standalone human data exist, but its mechanism is conserved across species[1].
• IGF‑1 activates the PI3K‑Akt pathway, which can branch toward both mTORC1‑dependent growth and ERK‑mediated survival. Pulsed low‑dose exposure favors transient Akt activation and downstream ERK signaling, limiting sustained mTORC1 drive that is linked to oncogenic risk[5].
• Nitric oxide from BPC‑157‑induced eNOS activation can S‑nitrosylate mTOR complex components, transiently inhibiting mTORC1 signaling[1].
• Combining these agents may therefore create a synergistic milieu: enhanced microvascular perfusion (BPC‑157), incr

Aggregates as Redox Sinks

Protein aggregation in aging may represent a controlled redox‑buffering system rather than mere waste. Hyperoxidation of cysteine residues (to sulfinic/sulfonic acids) promotes liquid‑liquid phase separation (LLPS) that sequesters these reactive thiols into insoluble deposits, lowering cytosolic oxidative pressure. When the ubiquitin‑proteasome system is overwhelmed, the cell actively shuttles hyperoxidized proteins to microtubule‑organized inclusions, converting a dangerous soluble pool into a thermodynamically stable solid phase.

Mechanistic Extension

We hypothesize that aggregates function as redox sinks: they store oxidizing equivalents derived from cysteine hyperoxidation, and their reversible persulfidation can release reducing equivalents back to the cytosol on demand. Persulfidation (‑SSH) decreases with age, tipping the balance toward hyperoxidation and aggregation. Restoring persulfidation should dissolve aggregates without releasing toxic oligomers, because the sequestered cysteines are already in their highest oxidation state and cannot regenerate harmful soluble intermediates.

• Step 1: Cysteine thiols become hyperoxidized (Cys‑SO₂H/Cys‑SO₃H) under chronic oxidative stress.
• Step 2: Hyperoxidized cysteines promote LLPS via increased polarity and hydrogen‑bonding capacity, nucleating chaperone‑rich hydrogels.
• Step 3: Hydrogels mature into amyloid‑like aggregates that sequester the oxidized species, preventing aberrant disulfide‑mediated cross‑links with vital proteins.
• Step 4: Persulfidating enzymes (e.

Hypothesis

Transient mTOR inhibition before cyclic expression of Yamanaka factors enhances chromatin accessibility and TET‑mediated DNA demethylation, thereby increasing the efficiency and durability of epigenetic rejuvenation without compromising cellular identity.

Mechanistic Rationale

mTOR activity drives anabolic programs that favor chromatin condensation through increased histone acetylation and methylation, limiting the plasticity needed for epigenetic resetting. When mTOR is briefly suppressed, cells shift toward a catabolic state marked by upregulated autophagy and NAD+‑dependent sirtuin activity. Autophagy clears damaged histones and chromatin‑associated proteins, while sirtuins deacetylate histones H3K9 and H3K56, creating a more open nucleosome landscape. This metabolic milieu coincides with the early window (days 3‑7) when partial reprogramming initiates steady epigenetic age decline. In this state, TET enzymes gain better access to 5‑mC sites, accelerating hydroxylation and subsequent base‑excision repair–driven demethylation. The result is a deeper, more stable epigenetic reset that retains lineage‑specific transcription factors, avoiding dedifferentiation.

Experimental Design

1. Animal cohorts – Aged (20‑month) C57BL/6 mice divided into four groups (n=20 per group):
   • Control (vehicle)
   • Rapamycin pretreatment (2 mg/kg i.p. 3×/week for 2 weeks) followed by vehicle
   • Cyclic OSK (AAV‑OSK, 2 days on/5 days off) for 8 weeks
   • Rapamycin pretreatment + cyclic OSK (same regimens)
2. Readouts
   • Epigenetic age via multi‑tissue DNA met

Hypothesis

Senescent stromal cells function as physical scaffolds that preserve topologically associating domain (TAD) integrity in aged muscle stem cells through juxtacrine interactions involving lamin‑associated domain (LAD) proteins.

Mechanistic basis

• Senescent fibroblasts and mesenchymal stromal cells retain a distinct nuclear lamina composition enriched in lamin A/C and LAD‑binding proteins such as BAF and LBR[1].
• These lamin‑rich nuclei can form stable cadherin‑mediated adhesions with neighboring muscle stem cells (MuSCs), creating a juxtacrine niche that transmits mechanical cues to the MuSC nucleus.
• Mechanical tension transmitted via N‑cadherin complexes promotes chromatin decondensation at specific LADs, preserving boundary elements and preventing ectopic enhancer‑promoter contacts that would otherwise drive inflammatory gene programs.
• In aged tissue, senescent cells accumulate and maintain this scaffold, while their SASP concurrently suppresses proliferation; the net effect is a trade‑off where architectural support offsets some SASP‑driven dysfunction.
• When senescent cells are cleared by senolytics, the loss of juxtacrine LAD‑lamin contacts leads to rapid erosion of TAD insulation in MuSCs, manifesting as increased inter‑TAD contacts and loss of age‑specific loops[2].

Testable predictions

1. Proximity dependency – MuSCs that are physically isolated from senescent stromal cells (e.g., by transwell separation) will show TAD deteriorat