Intermittent hypobaric pressure induces selective senescent cell death and alleviates age-related osteoporosis

Meng, Bowen; Qu, Yan; Yang, Benyi; Fu, Chaoran; He, Yifan; Li, Jing; Wan, Rentao; Li, Xin; Xue, Zhulin; Cao, Zeyuan; Hao, Meng; Zhang, Xiao; An, Zhe; Chen, Fen; Ren, Ruibao; Mao, Xueli; Cao, Yang; Shi, Songtao

doi:10.1038/s41551-025-01584-5

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Published: 14 January 2026

Intermittent hypobaric pressure induces selective senescent cell death and alleviates age-related osteoporosis

Bowen Meng¹^na1,
Yan Qu¹^na1,
Benyi Yang¹,
Chaoran Fu¹,
Yifan He¹,
Jing Li²,
Rentao Wan²,
Xin Li³,
Zhulin Xue^1,4,
Zeyuan Cao¹,
Meng Hao¹,
Xiao Zhang^1,5,
Zhe An¹,
Fen Chen¹,
Ruibao Ren^6,7,
Xueli Mao¹,
Yang Cao¹ &
…
Songtao Shi ORCID: orcid.org/0000-0003-1516-7500^1,6

Nature Biomedical Engineering (2026)Cite this article

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Abstract

Senescent cell accumulation contributes to aging, and their clearance represents an effective anti-aging strategy. Current senolytic strategies focus on drug-mediated senescent cell clearance, but it is unknown whether a hypobaric condition can induce senescent cell death. Here we show that hypobaric pressure (HP) at −375 mmHg without hypoxia induces cells to undergo lysosome-dependent cell death (LDCD). Mechanistically, we unveil that HP activates transmembrane protein 59 (TMEM59) to induce cellular Ca²⁺ influx, which triggers calpain 2 to cleave lysosomal associated membrane protein 2 (LAMP2), leading to lysosomal membrane permeabilization and subsequent LDCD. Furthermore, given that senescent cells contain elevated numbers of lysosomes, we report intermittent HP treatment to specifically induce senescent cells to undergo LDCD and reduce the senescence-associated secretory phenotype. Eventually, we report that intermittent HP treatment can substantially extend the lifespan and rescue the osteoporosis phenotype in aged mice. This study reveals a previously unknown role of HP as a natural senolytic to eliminate senescent cells, and identifies TMEM59 as a new HP-activated ion channel protein.

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Main

Aging is a complex process that affects various tissues and organs, along with senescent cell accumulation¹. Senescent cells exhibit irreversible arrest in proliferation and resistance to undergo programmed cell death¹. Importantly, senescent cells can secrete multiple pro-inflammatory cytokines, chemokines and tissue-remodelling proteins that are collectively termed the senescence-associated secretory phenotype (SASP)¹. SASP has been implicated in the pathogenesis of aging-associated degenerative diseases. Accumulated evidence demonstrated that clearance of senescent cells using pharmacological or genetic approaches is an effective anti-aging approach termed senolytics^2,3. Senolytic agents such as dasatinib (a chemotherapy drug for leukaemia), plant extract quercetin, natural flavonoids fisetin, procyanidin C1, piperlongumine, curcumin and taurine in foods, Bcl2 family inhibitors navitoclax and venetoclax^{2,3,4,5,6,7,8} mainly target anti-apoptotic pathways in senescent cells (SCAPs). Recently, newly developed senolytics have been shown to specifically target senescent cells with reduced side effects^9,10,11. So far, it is unknown whether hypobaric pressure is capable of inducing death in senescent cells.

Mechanical force plays important biological roles in physiological development and pathological processes¹². There are various forms of mechanical forces inside the body, such as compressive force, tensile force, fluid shear force and hypobaric pressure¹². The body can transduce mechanical stimulation signals into biochemical signals, thereby activating a series of response reactions within the cell, affecting normal physiological behaviours such as cell migration, proliferation and differentiation, aging, as well as pathological processes such as cancer and age-related diseases^{13,14,15,16,17,18}. Normal mechanical force contributes to cell proliferation, differentiation and development^13,14; abnormal mechanical force can lead to cell death and aggravate diseases such as osteoarthritis, cancer, hypertension and atherosclerosis^15,16,17,18. However, the detailed molecular mechanism by which mechanical force regulates cell death is not fully understood. Hypobaric pressure (HP), a state of relative pressure lower than standard air pressure, is a natural condition at high altitudes and exists inside the body. The distribution of HP within the body is an important manifestation of biological evolution and adaptation. HP is necessary for maintaining basic life activities in the heart, lung, abdominal cavity, gastrointestinal tract and joint, helping to distribute oxygen, CO₂, energy substances and metabolic products to the desired organs^{19,20,21,22,23}. Moderate HP (−125 mmHg) has been used to promote wound healing in clinics for many years^24,25,26,27. Its therapeutic mechanism has been suggested to be associated with deformation of the tissues and drainage of extracellular inflammatory fluids²⁸. In addition, hypobaric hypoxia is used for training pilots and athletes to improve their tolerance to high-altitude hypoxic environments and physical fitness^29,30. However, the molecular mechanisms and biological cues of HP-induced cellular and molecular responses without hypoxia are still not fully understood.

Living organisms depend on their ability to receive and respond to chemical and physical signals from the environment³¹. Mechanosensitive ion channels exhibit channel gating in response to alterations in physical stress within their microenvironment. These changes may arise from various stimuli, including tactile contact with the skin, gravitational forces, proprioceptive feedback, auditory vibrations, variations in food texture, muscle elongation, airflow and fluctuations in atmospheric pressure³¹. The activation of mechanosensitive ion channels is essential for numerous fundamental physiological processes that necessitate the detection of mechanical forces³¹. It is well known that mechanosensitive ion channels include Piezo1, TREK/TRAAK and the transient receptor potential (TRP) family^32,33,34. Transmembrane proteins can conduct mechanical force-sensitive ion channel activities, including those of TMEM63, TMC-1 and TACAN/TMEM120 (refs. ^31,35,36,37). However, it is unknown whether there is any HP-specific ion channel protein. In this study, we found that transmembrane protein 59 (TMEM59) is an ion channel protein that is specifically activated by HP, and intermittent HP (−375 mmHg) treatment can eliminate senescent cells via lysosome-dependent cell death (LDCD).

Results

HP can induce cells to undergo LDCD

Mechanical force can be divided into two categories: mechanical positive pressure (PP) and hypobaric pressure (HP), due to positive and negative loading, respectively. We used mesenchymal stem cells (MSCs) as a cell source to study the mechanism of mechanical force-induced cell death. Surprisingly, HP treatment induced MSCs to undergo a ‘rosette’ blebbing change. However, PP treatment induced MSCs to show a typical apoptotic morphological change, including cell membrane and cytoplasm shrinkage, cell membrane blebbing and nuclear shrinkage, as assessed by immunofluorescence staining (Fig. 1a). HP treatment failed to activate cleaved caspase 3, whereas PP induced substantial activation of cleaved caspase 3, as assessed by western blotting analysis and immunofluorescence staining (Fig. 1b,c and Supplementary Fig. 1a). The cell death rate of MSCs in the HP group was higher than that in the PP group, as assessed by flow cytometry analysis (Fig. 1d). After treatment with the apoptosis inhibitor Z-VAD, the cell death rate of HP-induced MSCs failed to change, but the cell death rate in PP-induced MSCs was substantially reduced, adding to the evidence that HP-induced MSC death is not associated with caspase 3 activation (Fig. 1d and Supplementary Fig. 1b,c).

**Fig. 1: Hypobaric pressure induces MSCs to undergo non-apoptotic cell death.**

To examine the molecular mechanism of mechanical force-induced MSC death, we used the data-independent acquisition (DIA) protein profiling method for high-throughput analysis of protein expression. A total of 6,274, 6,061 and 6,263 proteins were identified in untreated MSCs, PP- and HP-treated MSCs, respectively (Fig. 1e). Furthermore, 3,547 proteins were differentially expressed between the PP and HP groups (fold-change > 1.2, Q-value < 0.05), which included 2,219 notably upregulated proteins and 1,328 notably downregulated proteins in the HP group (Fig. 1f,g and Supplementary Table 1). KEGG enrichment analysis showed that 3,547 differentially expressed proteins were mainly concentrated in the ‘lysosome’, ‘protein digestion and absorption’ and ‘other glycan degradation’ categories (Supplementary Fig. 1d). Meanwhile, volcano map and clustering heat map analysis showed that 4,508 proteins were differentially expressed between the untreated MSC group and the HP group (fold-change > 1.2, Q-value < 0.05), which included 1,795 notably upregulated proteins and 2,263 notably downregulated proteins in the HP group (Supplementary Fig. 1e,f and Supplementary Table 2). These data indicate that HP-induced MSC death is distinguishable from untreated MSCs and PP-induced apoptotic MSCs.

We next focused on the differentially expressed proteins that further distinguish the HP from the PP group, and found 4,357 lysosome-related proteins³⁸ in the mass spectrometry library (Supplementary Table 3). Gene Ontology (GO) molecular function analysis showed that 199 hydrolase activity proteins were highly expressed in HP-treated MSCs (Fig. 2a,b and Supplementary Table 4), of which 169 belonged to lysosomal proteins. Further GO biological process analysis indicated that 59 proteolysis proteins were highly expressed in HP-treated MSCs (Fig. 2c,d and Supplementary Table 5), of which 53 belonged to the category of lysosomal proteins. Meanwhile, we show that hydrolase function proteins and proteolytic process proteins were highly expressed in the HP group compared with the untreated MSC group (Supplementary Fig. 2a–d, and Supplementary Tables 6 and 7). GO molecular function enrichment analysis of the 199 hydrolase activity proteins and 59 proteolysis proteins unveiled enrichment of hydrolase activity, peptidase activity, metallopeptidase activity and metabolic processes (Supplementary Fig. 2f). These data indicate that HP-induced MSC death may be related to lysosome-mediated hydrolysis.

**Fig. 2: HP induces MSC death in a lysosome-dependent manner.**

Lysosomal membrane permeabilization (LMP) may result in the liberation of hydrolytic enzymes that are implicated in LDCD³⁹. We showed that calpain 2 was notably activated after HP treatment and the lysosomal membrane protein LAMP2 was massively cleaved, indicating that HP treatment may increase LMP in MSCs (Fig. 2e). PP treatment cannot activate the LDCD-related proteins, including calpain 1, calpain 2, LAMP1, LAMP2, cathepsin B and cathepsin D (Supplementary Fig. 2e). Further analysis showed that hydrolase proteins including cathepsin B, cathepsin D, ADAM9, ADAM10, MMP-1, MMP2, MMP-14 and ADAMTS4 were highly expressed in HP-treated MSCs (Fig. 2e). Immunofluorescence analysis confirmed that calpain 2 and LAMP2 were activated after HP treatment, and their co-localization area was notably increased (Fig. 2f,g). HP treatment elevated cathepsin B enzyme activity and the expression of ADAM9, MMP2 and ADAMTS4, as assessed by immunofluorescence analysis and western blotting, respectively (Fig. 2e and Supplementary Fig. 2g). Meanwhile, HP treatment caused lysosomal volume enlargement (Fig. 2f–h). After small interfering (si)RNA knockdown of calpain 2 expression, its co-localization area with LAMP2 was notably decreased (Fig. 2f,g and Supplementary Fig. 2h). Under HP treatment, the fluorescence signal of lysotracker gradually increased in MSCs, while calpain 2 siRNA treatment abolished this effect (Fig. 2h). Acridine orange (AO), a lysomotropic metachromatic fluorescent dye, shows red fluorescence (emission wavelength: 617 nm) in the lysosomal body, but green fluorescence (emission wavelength: 528 nm) in permeability-increased lysosomal membrane⁴⁰. AO staining showed that HP treatment caused LMP as evidenced by reduced red and increased green fluorescence intensity, which was reversed by calpain 2 siRNA treatment (Fig. 2i). Higher HP notably increased the expression of cleaved LAMP2. Mild PP increased the expression of cleaved caspase 3, but excessive PP increased the expression of RIP3 (Supplementary Fig. 2i). These results indicate that HP can induce MSC LDCD, and mild PP can induce MSC apoptosis, but excessive PP can induce MSC necrosis. In addition, cycle pull force notably increased the expression of cleaved caspase 3 and decreased the expression of LAMP2 (Supplementary Fig. 2j). These results indicate that cycle pull force induces MSC apoptosis, but not LDCD. These data suggest that HP induces cells to undergo LDCD.

We performed additional RNA sequencing (RNA-seq) analysis to visualize homo/heterogeneity of the observed effects at selected HP/PP levels. The results showed that a total of 15,245, 16,216 and 15,307 genes were identified in untreated MSCs, PP- and HP-treated MSCs, respectively (Supplementary Fig. 3a). Furthermore, 7,871 genes were differentially expressed between the PP and HP groups (log₂fold-change > 0.5, Q-value < 0.05), which included 4,265 notably upregulated genes and 3,606 notably downregulated genes in the HP group (Supplementary Fig. 3b,c and Supplementary Table 8). We next focused on the differentially expressed genes (DEGs) that further distinguished the HP group from the PP group and found 7,129 lysosome-related genes (LAG) in the mass spectrometry library (Supplementary Table 9). GO molecular function analysis showed that 183 hydrolase activity genes were highly expressed in HP-treated MSCs (Supplementary Fig. 3d,e and Supplementary Table 10), of which 101 belonged to lysosomal genes. Further GO biological process analysis indicated that 106 proteolysis genes were highly expressed in HP-treated MSCs (Supplementary Fig. 3f,g and Supplementary Table 11), of which 55 belonged to the category of lysosomal genes. GO molecular function enrichment analysis of the 183 hydrolase activity genes and 106 proteolysis genes unveiled enrichment of hydrolase activity, peptidase activity, metallopeptidase activity and metabolic processes (Supplementary Fig. 3h,i). These data indicate that HP can induce LDCD in MSCs.

TMEM59 is an HP-responsive mechanosensitive ion channel protein

Calcium influx is the key initiator of calpain 2 activation⁴¹. Therefore, we screened highly expressed calcium ion-related channel proteins in the HP group by DIA protein profiling. The result showed that the HP group highly expressed 12 calcium-related channel proteins, including TRPV2, Piezo1, MCU, PKD2, TMEM63B, ANXA2, CLU, SEC61A1, ITPR3, PANX1, ITPR2 and TRPM4 (Supplementary Fig. 4a,b). Western blotting results showed that TRPV2 and Piezo1 notably increased in the PP treatment group, but their expression showed no significant change in the early or middle stage of HP treatment, suggesting that TRPV2 and Piezo1 may be not the main HP-responsive mechanosensitive ion channels (Supplementary Fig. 4c).

We next found that the TMEM family was highly expressed in the HP group but not in the PP group (Fig. 3a,b and Supplementary Table 12). We used western blotting to confirm that the expression levels of TMEM59, TMEM263 and TMEM127 were notably increased in the HP group but not in the PP group (Fig. 3c). Immunofluorescence results showed that HP treatment increased the expression of TMEM59 and calpain 2, while TMEM59 siRNA treatment and TMEM59-KO inhibited the expression of calpain 2 (Supplementary Fig. 4g). However, HP treatment increased the expression levels of TMEM263, TMEM127 and calpain 2, but TMEM263 or TMEM127 siRNA treatment failed to inhibit the expression of calpain 2 (Supplementary Fig. 4h,i). These data suggest that the expression of TMEM59 is critical for the activation of calpain 2. To examine whether TMEM59 is an HP-sensitive ion channel, we used siRNA technology to knock down TMEM59 to examine its effect on intracellular calcium ion levels. Amlodipine, an L-type calcium ion channel inhibitor, was used to treat MSCs to exclude potential off-target interference. Calcium ion fluorescence staining results showed that HP treatment failed to increase the intracellular Ca²⁺ levels in the TMEM59 siRNA knockdown and TMEM59-KO groups, but knockdown of TMEM263, TMEM127, TRPV2 and Piezo1 by siRNA has no effect on Ca²⁺ influx (Fig. 3d,e and Supplementary Fig. 4e,f). These results suggest that TMEM59 is a major HP-responsive ion channel protein.

**Fig. 3: HP specifically activates TMEM59-mediated calcium influx.**

We next used cell-attached recordings to assess whether TMEM59 affects mechanically evoked currents. To exclude potential interference of TRPV2 and Piezo1, we used siRNA to knockdown both TRPV2 and Piezo1 in MSCs for subsequent experiments (Supplementary Fig. 4d). We applied brief pulses of HP through the recording electrode and found the presence of endogenous mechanosensitive ion channels in MSCs. However, siRNA knockdown of TMEM59 and TMEM59-KO notably decreased mechanically evoked currents in MSCs under different HP intensities (Fig. 3f,g). These data confirm that TMEM59 is an HP-responsive mechanosensitive calcium ion channel. Flow cytometry analysis showed that after treatment with a calcium ion chelator BAPTA-AM/EGTA-AM, calpain inhibitor PD150606 and broad-spectrum protease inhibitors E64D//Leupeptin hemisulfate, the HP-induced MSC death rate was notably reduced (Fig. 3h). However, the above inhibitor treatments had no effect on the PP-induced MSC death rate (Supplementary Fig. 4j). These results suggested that HP-induced MSC death is calcium influx-activated LDCD.

HP treatment specifically eliminates senescent cells via TMEM59-mediated LDCD

Previous studies have shown that the number of lysosomes increases in senescent cells, along with enlarged size and enhanced membrane permeability⁴². To further examine the alterations of lysosomal function in senescent MSCs, we used iTRAQ protein profiling to identify a total of 6,189 lysosome-related proteins in young and senescent MSC lysosomes. A volcano map and clustering heat map showed that 2,301 proteins were differentially expressed in lysosomes between young and aged mice (fold-change > 1.1, Q-value < 0.05), including 1,286 upregulated proteins and 1,015 downregulated proteins in the senescent lysosome group (Fig. 4a,b and Supplementary Table 13). Subsequently, we directed our attention to the differentially expressed proteins and performed a functional analysis. We analysed 1,522 senescence-associated secretory phenotype (SASP) proteins that were previously shown to be expressed in human cell lines⁴³ and found 1,057 proteins in the mass spectrometry library (Supplementary Table 14). Further analysis showed that senescent MSC-derived lysosomes contained 265 notably upregulated SASP proteins and 177 notably downregulated SASP proteins compared with young MSC-derived lysosomes, suggesting that senescent lysosomes have higher levels of SASP proteins (Fig. 4c and Supplementary Table 15). GO molecular function analysis showed that 118 hydrolase activity proteins were highly expressed in the senescent MSC lysosome group, of which 27 were more highly expressed in the HP group than in the PP group (Fig. 4d and Supplementary Table 16). Further GO biological process analysis indicated that 43 proteolysis proteins were highly expressed in the senescent MSC lysosome group, of which 6 were more highly expressed in the HP than in the PP group (Fig. 4e and Supplementary Table 17). GO molecular function and biological process enrichment analysis of 118 hydrolase activity proteins and 43 proteolysis proteins unveiled enrichment of hydrolase activity, peptidase activity, metallopeptidase activity and metabolic processes, indicating that these proteins are involved in the hydrolysis of various metabolites (Supplementary Fig. 5a,b).

**Fig. 4: High-throughput analysis of protein profiling of lysosomes from senescent and young bone marrow MSCs.**

Senescent MSCs showed a higher number of and enlarged lysosomes than young MSCs (Fig. 4f). After HP treatment, the number of lysosomes increased more notably in senescent MSCs than in young MSCs (Fig. 4f). LAMP2 immunofluorescence analysis further confirmed that HP treatment can increase the expression of lysosomal membrane protein LAMP2 in senescent MSCs vs in young MSCs (Fig. 4f). AO staining showed that the red fluorescence of senescent MSCs decreased more notably than that of young MSCs with the extension of HP treatment time, suggesting that the permeability of lysosomes was enhanced in senescent MSCs (Fig. 4g and Supplementary Fig. 5c). We also used ratiometric fluorescence imaging of the pH-sensitive dye RatioWorks PDMPO to perform more quantitative measurement of lysosomal pH, as previously reported⁴⁴. The results of immunofluorescence showed that the pH of lysosomes in senescent MSCs was markedly increased (Fig. 4h). These results indicate that senescent MSCs are more sensitive to HP-induced LDCD.

Combined with our finding that HP treatment increased the expression of lysosome-related proteolytic enzymes, we speculate that HP treatment may be able to eliminate senescent MSCs. We used two types of senescent MSC, namely, bone marrow MSCs from aged mice and senescent human umbilical MSCs obtained by serial passage to passage 25, to examine whether HP treatment can eliminate senescent MSCs. Western blot and cellular immunofluorescence analysis showed that senescent bone marrow MSCs and senescent umbilical MSCs highly expressed senescence marker proteins P16, P21 and γH2AX (Supplementary Fig. 6a–c). SA-β-gal staining showed that HP treatment eliminated SA-β-gal-positive MSCs from senescent bone marrow MSCs and umbilical MSCs (Fig. 5a,b and Supplementary Fig. 6d,e). Only a very limited number of SA-β-gal-positive MSCs were still detectable after HP treatment in young bone marrow MSCs and umbilical MSCs (Fig. 5a,b and Supplementary Fig. 6d,e). However, HP treatment failed to eliminate senescent MSCs from TMEM59 knockout or TMEM59 siRNA knockdown MSCs (Fig. 5a,b and Supplementary Fig. 6d,e). In contrast, HP treatment eliminated senescent MSCs from TRPV2, Piezo1, TMEM263 and TMEM127 siRNA knockdown MSCs (Fig. 5a,b and Supplementary Fig. 6d,e). In addition, we showed that HP of −50 kPa (−375 mmHg) could effectively remove senescent MSCs (Supplementary Fig. 6g,h). D + Q treatment showed similar senolytic effect to HP treatment (Supplementary Fig. 6i,j). Flow cytometry analysis showed that the cell death rates of senescent bone marrow MSCs and umbilical MSCs gradually increased after HP treatment for 3–9 h. The cell death rate of young bone marrow MSCs and umbilical MSCs did not increase notably after HP treatment (Fig. 5c and Supplementary Fig. 6f). ELISA showed that the senescent MSCs secreted higher levels of SASP-related factors IL-1, IL-6, IL-8, monocyte chemoattractant protein-1 (MCP-1), tumour necrosis factor-α (TNF-α), MMP-1 and MMP-14 than the young MSCs. HP treatment notably reduced the levels of the above SASP-related factors (Fig. 5d). Immunofluorescence results showed that HP treatment reduced the fluorescence level of P16 and increased the fluorescence level of LAMP2 in senescent MSCs when compared to untreated control MSCs, but HP failed to alter P16 and LAMP2 expression in young MSCs (Fig. 5e). In addition, long-term continuous HP failed to increase the aging marker P16 in young cells (Supplementary Fig. 6k), suggesting that HP may not initially induce senescence for subsequent selective elimination. The above results suggest that HP treatment can efficiently eliminate senescent MSCs by TMEM59-mediated LDCD.

**Fig. 5: HP specifically eliminates senescent cells via TMEM59-mediated LDCD.**

Intermittent HP treatment eliminates senescent cells in middle-aged mice

To further explore whether HP treatment can eliminate senescent cells in vivo, 15-month-old middle-aged mice were placed in an HP chamber for 2 h a day over the course of 4 weeks. H&E staining showed that HP treatment resulted in a variety of histological changes in these middle-aged mice: reductions in the numbers of vacuoles in the liver and renal tubular oedema in the kidney, the thickness of the red pulp in the spleen, the amount of heterochromatin around the nuclei in the brain and inflammatory cells around the alveoli in the lung; and increases in the number of glomeruli in the kidney and the thickness of white pulp in the spleen (Fig. 6a). After HP treatment, SA-β-gal-positive cells were decreased in multiple organs including the liver, kidney, spleen, brain and lung in middle-aged mice (Fig. 6b,c). Immunofluorescence and SA-β-gal staining showed that HP treatment reduced the P16 fluorescence intensity of peripheral blood mononuclear cells (PBMCs) and SA-β-gal-positive PBMCs in middle-aged mice (Supplementary Fig. 7a–c). In vivo animal imaging results showed that HP treatment reduced the fluorescence levels of various organs in P16 fluorescently labelled mice (Fig. 6d).

**Fig. 6: HP eliminates senescent cells in various organs of middle-aged mice.**

We next showed that TMEM59-KO mice exhibited premature aging in various organs when compared to control C57BL/6 mice (Supplementary Fig. 7d). A previous study showed that the expression of TMEM59 in various organs of older mice is notably reduced relative to that of young mice⁴⁵. We also found that the expression of TMEM59 was reduced in tissues and organs of aged mice, including the liver, kidney, spleen, brain and lung (Fig. 6e). Therefore, we speculated that TMEM59 may be associated with clearance of senescent cells in vivo. HP treatment failed to alleviate the histological changes seen in the aged organs and did not have a significant effect on the number of SA-β-gal-positive cells in aged TMEM59-KO mice (Fig. 6a–c). We showed that HP treatment reduced P16 fluorescence intensity and increased TMEM59 fluorescence intensity in P16 fluorescently labelled middle-aged mice, but not in TMEM59-KO middle-aged mice (Fig. 6f). However, TMEM59 fluorescence intensity in aged mice was lower than in young mice after treatment with HP (Fig. 6e,f). The above results suggest that HP treatment eliminates senescent cells in various tissues and organs in a TMEM59-dependent manner.

To further confirm the molecular mechanism by which HP stimulates removal of senescent cells in middle-aged mice, we examined changes in the senescence marker P21 and the lysosomal marker LAMP2 after HP treatment. The results showed that HP treatment decreased the fluorescence intensity of P21 while increasing the fluorescence intensity of LAMP2 in the liver, kidney, spleen, brain and lung in middle-aged mice (Supplementary Fig. 7e,f). In addition, long-term treatment with intermittent HP improved MSC viability and increased the number of PCNA⁺ cells in the bone marrow of 15-month-old C57BL/6J mice (Supplementary Fig. 7g,h). The above results suggest that HP promotes the clearance of senescent cells by activating lysosomal function (Supplementary Fig. 9).

Intermittent HP treatment extends lifespan

We next assessed the feasibility of using intermittent HP treatment to extend life. We put 15-month-old mice into an HP chamber (without hypoxia) for 1 h every other day until they died. Intermittently HP-treated mice survived longer than control mice (Fig. 7a–c). In the control group, all untreated mice died before 27.1 months of age. In contrast, intermittently HP-treated mice showed a better survival rate (Fig. 7a–c). Intermittent HP treatment notably extended the median lifespan of aged C57BL/6J mice from 24.16 months to 27.3 months and showed a 20.93% longer median post-treatment lifespan (female mice from 25.98 months to 29.52 months and male mice from 23.55 months to 26.34 months, P < 0.0001) (Fig. 7a–c). The average lifespan increased from 24.25 ± 1.94 months to 27.86 ± 2.24 months and showed a 24.07% longer average post-treatment lifespan (female mice from 25.30 ± 1.79 months to 29.52 ± 1.66 months (P < 0.0001) and male mice from 23.20 ± 1.52 months to 26.20 ± 1.30 months (P < 0.001)) (Fig. 7d–f). Intermittent HP treatment also reduced body weight gain in aged mice (Fig. 7g). Moreover, intermittent HP treatment reduced the amount of grey hair and hair loss in the aged group (Fig. 7h). ELISA results showed that intermittent HP treatment reduced the levels of SASP factors in the serum of aged mice, including IL-1, IL-6, IL-17, MCP-1, TNF-α and interferon-gamma (IFNγ) (Fig. 7i).

**Fig. 7: HP extends lifespan and reduces age-related hallmarks in aged mice.**

To explore whether HP can treat age-related osteoporosis, aged mice were placed in the HP chamber for 2 h a day over the course of 6 weeks, after which the femurs of the mice were isolated for micro-computed tomography (micro-CT) scanning. The results showed that HP treatment rescued the osteoporosis phenotype of aged mice, as evidenced by increased bone volume/tissue volume (BV/TV), trabecular number (Tb.n) and trabecular thickness (Tb.Th) (Fig. 7j,k). Alizarin red, oil red O and TRAP staining showed that HP treatment enhanced osteogenesis and reduced adipogenesis of bone marrow MSCs, along with decreased osteoclast differentiation (Fig. 7l–n). These results confirmed that HP can rescue osteoporosis phenotypes in aged mice.

Since intermittent HP treatment reduced systematic chronic inflammation, we examined physical function in HP-treated aged mice. HP treatment for 9 months notably increased the maximal rotarod time (Supplementary Fig. 8a), hanging endurance (Supplementary Fig. 8b), grip strength (Supplementary Fig. 8c) and treadmill endurance (Supplementary Fig. 8d) in aged mice. HP treatment showed no obvious systemic toxicity according to serum biochemical test and routine blood analysis (Supplementary Fig. 8e–p). Notably, HP treatment for 9 months notably reduced the levels of ALT, AST, TG, FFA, LDL-C, UA, CRE, BUN and WBC, but increased the level of HDL-C (Supplementary Fig. 8e–p).

Discussion

The biological activities of organisms at all levels, from organs and tissues to cells, are carried out in a certain mechanical environment⁴⁶. Therefore, understanding the molecular mechanism of how natural life makes adaptive changes by sensing the surrounding environment is crucial. The internal body cavities are connected to each other and to the external air pressure^{47,48,49,50,51}. Body cavities maintain a specific air pressure and its dynamic balance is crucial to maintaining organ homeostasis. In local HP application, such as capping and negative pressure wound therapy, unbalanced internal and external pressure may cause a local continuous pull and push⁵². In contrast, in whole-body HP treatment, the internal pressure of the body will quickly adapt to external pressure and reach a newly established pressure balance. The whole process from life to death is also accompanied by changes in the internal mechanical environment of an organism⁵³, including physiological and pathological processes. However, the molecular mechanisms by which different forms of forces, such as PP and HP, exert their impact on cell death are still largely unknown. Moreover, it is also unclear whether senescent cells can be eliminated by adjusting the mechanical environment. It was previously reported that moderate mechanical PP may induce apoptosis⁵⁴. Local HP, in forms such as HP wound therapy⁵⁵ and cupping therapy^56,57, has been widely used in the treatment of various diseases. However, the relationships between HP and cell death as well as aging have not been elucidated. In this study, we discovered that HP treatment can induce LDCD through the HP-activated mechanosensitive ion channel TMEM59. Importantly, we further show that intermittent HP can remove senescent cells and is capable of extending the lifespan and alleviating age-related osteoporosis. This study provides a nature-powered senolytic therapeutic approach.

The transmembrane protein family has recently been known to conduct mechanical force-sensitive ion channel activities. OSCA/TMEM63 was first screened for osmotic stress response defects in plants⁵⁸ and was also shown to be heterologously expressed in Xenopus laevis oocytes to respond to osmotic stress⁵⁹. Compared with the low pressure response threshold (P₅₀ = −31.2 ± 3.5 mmHg) of PIEZO1 (ref. ³²), OSCA/TMEM63 is a mechanosensitive ion channel with a higher pressure response (P₅₀ = −215.9 ± 10.44 mmHg)³⁵. Transmembrane channel-like protein-1 (TMC-1) initially attracted attention because of its hearing-related ion channel activity^60,61. Recently, the TMC-1 complex was confirmed to be a mechanosensitive ion channel³⁶. In addition, TMEM16 is evolutionarily related to TMC family proteins and is a calcium ion-activated chloride channel protein⁶². TACAN/TMEM120A was originally discovered in a proteomic screen for membrane proteins involved in mechanotransduction in smooth muscle cells⁶³ and was later identified as an ion channel involved in sensing mechanical pain³⁷. Although relatively insensitive to PP stimulation, TACAN is more sensitive to HP stimulation³⁷. Interestingly, TMEM59 is also on the list of membrane proteins involved in smooth muscle cell force transduction identified in previous proteomic screening⁶³. Recently, whole-exome sequencing analysis found that the mutations of TMEM59 are closely related to hereditary deafness⁶³, and overexpression of TMEM59 throughout the nervous system improves deficits in nerve electrical signal conduction in a Drosophila model of Parkinson’s disease⁶⁴. The above results suggest that TMEM59 may be involved in mechanoreception and ion channel conduction. Our proteomic screening found that TMEM59 was the most highly expressed transmembrane protein after HP stimulation. Meanwhile, the expression of TMEM59 increased under HP but not under PP stimulation. Patch clamp results further confirmed that TMEM59 is more sensitive to HP than to PP.

Lysosomes, the main degradative organelles of mammalian cells, have emerged as the centre of metabolic regulation⁶⁵. When LMP is altered and not repaired, the massive release of lysosomal hydrolase and extensive cytoplasmic acidification caused LDCD⁶⁶. Limited LMP can activate caspase-independent death^67,68. Our study found that HP-induced MSC death is independent of caspases. It is mainly due to the activation of calpain 2 by TMEM59-mediated calcium ion influx, which further cleaves the lysosomal membrane, resulting in enhanced LMP and hydrolase cleavage that in turn cause LDCD.

Organelle dysfunction is one of the important characteristics of senescent cells, among which lysosome changes are the most prominent⁶⁹. Senescent lysosomes have higher mass and pH, increased LMP and reduced proteolytic capacity⁴². However, the reduced degradation function of lysosomes in senescent cells may be compensated to some extent by their increased number^70,71,72. This study found that intermittent HP treatment can eliminate senescent MSCs by enhancing LMP and increasing the number of lysosomes. In addition, lysosome-mediated exocytosis may play an important role in SASP in senescent cells⁷⁰. This study also found that senescent MSCs have elevated SASP secretion potential, which may be the main reason why SASP levels decreased after HP treatment. Therefore, targeting the functional status of lysosomes has the potential to become a new senolytic strategy^71,73,74,75.

Aging is accompanied by the gradual accumulation of senescent cells, leading to a high incidence of age-related diseases⁶⁹. Aging can be reversed to a certain extent by removing senescent cells, restoring body health and treating age-related diseases⁶⁹. Senescent cells display the development of SASP and resistance to apoptosis. Therefore, senotherapeutics are mainly divided into two categories: (1) senolytics, developed to target the senescent cell anti-apoptotic pathway (SCAP), which selectively kills senescent cells or induces senolysis; and (2) senomorphics, which attenuate the pathological SASPs to cause senostasis⁶⁹. However, clinical trials of senotherapeutic drugs are still under evaluation. Long-term use of anti-tumour drugs dasatinib and quercetin (D + Q), the most widely studied senolytics, causes side effects^76,77. Therefore, intermittent administration is currently used to reduce the side effects of senotherapeutic drugs. In addition, many new multidimensional intervention strategies for delaying aging are being explored, including CAR-T therapy^9,10, blocking the reactivation and spread of ERV ancient viruses⁷⁸, restoring the integrity of the epigenome⁷⁹, a synthetic genetic oscillator⁸⁰, dietary restriction⁸¹, nutrient supplementation^82,83, replenishing longevity-associated molecules with young blood⁸⁴ and transferring longevity genes⁸⁵. Interestingly, a study of the life expectancy of 1,494 retired airline pilots showed that they appeared to enjoy a life expectancy of more than 5 years longer than the 1980 US general population of 60-year-old white males⁸⁶. Pilot’s flight and landing processes are accompanied by alternating air pressure changes, with intermittent HP and normal pressure. In addition, it has been reported that elderly people of the Tibetan Plateau have a longer lifespan than elderly people in other regions of China, and the proportion of Tibetan men over 100 years old is about twice as high as that of Han Chinese⁸⁷, despite the fact that high-altitude areas are accompanied by low oxygen, cold temperatures and higher exposure to solar radiation. However, the relationship between hypoxia and aging is elusive^88,89. In this study, we developed an HP chamber with a continual supply of oxygen to ensure no hypoxia. We found that intermittent HP treatment can remove senescent cells from various tissues and organs, improve age-related osteoporosis and hair phenotypes, reduce SASP levels and extend lifespan. Gender difference is a factor that affects life expectancy of C57BL/6 mice after HP treatment. Females showed a 23.6% median post-treatment lifespan increase, while males showed an 18.6% median post-treatment lifespan increase. Previous studies have reported that overexpression of TMEM59 can improve the impaired motor behaviour in flies that model Parkinson’s disease and extend their lifespan⁶⁴. Interestingly, in our study, intermittent HP could not clear senescent cells from aged TMEM59-KO mice. This study paves the way for a new, nature-powered senolytic therapy that uses naturally existing hypobaric pressure to remove senescent cells and extend lifespan via TMEM59-mediated LDCD.

Limitations of the study

This study showed that different gradients of HP could remove senescent cells, and provided the most appropriate HP intensity of −50 kPa. We also verified the safety and effectiveness of intermittent HP (−50 kPa, once every 2 days) in removing senescent cells from multiple tissues and organs of mice. However, more interventional doses need to be further explored in future work to provide safer and more efficient anti-aging strategies for potential clinical translational applications. Further clinical experiments are required to evaluate the safety and effectiveness of more interventional doses of HP on human lifespan and age-related diseases.

Methods

Animals

C57BL/6J mice, aged 6 weeks and 15 months with equal numbers of males and females, were acquired from the Model Animal Research Center at Nanjing University and Gempharmatech. Cdkn2a-Luc-2A-tdTomato-2A-CreERT2 mice (designated as NM-KI-18039) with equal numbers of males and females were acquired from the Shanghai Model Organisms Center. In addition, TMEM59 conditional knockout (KO) mice (Tmem59^flox/flox, C57BL/6J background, identified as S-CKO-12059) with equal numbers of males and females were obtained from Cyagen Biosciences. Homozygous Tmem59^−/− (KO) Cdkn2a-Luc-2A-tdTomato mice were produced by crossing Tmem59^flox/flox mice with Cdkn2a-Luc-2A-tdTomato-2A-CreERT2 mice. All animal experiments were conducted in accordance with the guidelines of the Institutional Animal Care and Use Committee under an approved protocol at Sun Yat-Sen University (SYSU-IACUC-2023-B1163).

Antibodies and reagents

A comprehensive list of all antibodies, cytokines, kits and other resources utilized in this study can be found in Supplementary Table 18.

Cell culture and induction of senescent cells

Human umbilical cord mesenchymal stem cells (UMSCs) and mouse bone marrow mesenchymal stem cells (mBMMSCs) were isolated and cultured in accordance with the methodologies outlined in our previous research^90,91. In summary, UMSCs were procured from full-term caesarean section procedures, with informed consent of the donors. These cells were subsequently cultured in alpha minimum essential medium (α-MEM, Invitrogen), supplemented with 15% fetal bovine serum (FBS, Gibco), L-ascorbic acid phosphate at a concentration of 0.1 × 10⁻³ mol (Wako), and 1% penicillin/streptomycin (Invitrogen). Senescent UMSCs were obtained by continuously passaging beyond 25 generations.

mBMMSCs, either from young or aged specimens, were extracted from the bone marrow of the femurs and tibias of young or aged C57BL/6J mice, as well as from TMEM59-KO mice. Subsequently, mBMMSCs were maintained in α-MEM medium (Invitrogen) supplemented with 20% FBS (Gibco), 2 mM L-glutamine (Invitrogen), 55 mM 2-mercaptoethanol (Invitrogen) and 1% penicillin/streptomycin (Invitrogen). All cellular materials utilized in this research and associated experiments received approval from the Medical Ethics Committee at the Hospital of Stomatology, Sun Yat-Sen University (KQEC-2021-59-01).

Design and use of hypobaric pressure chambers

The hypobaric pressure chamber used in this experiment was independently designed and mainly consists of a cabin sealing structure, an intelligent control system, a vacuum pump, an oxygen supply machine and a pipeline structure (patent application number: CN202310638877.2; PCT/CN2023/097692; Gaoqishou Inc.). Hypobaric pressure chambers ensure maintenance of the required temperature, humidity and normal O₂ and CO₂ concentration inside the cabin.

Mice were randomly divided into HP treatment and control groups. HP group mice (whole body) were kept in the HP chamber (pressure: −50 kPa, O₂ partial pressure: 20–22 kPa, 22 ± 1 °C) for the indicated times, and were kept in normal housing conditions the rest of the time. Control group mice (whole body) were kept in the HP chamber (pressure: 101 kPa, O₂ partial pressure: 20–22 kPa, 22 ± 1 °C) for the indicated times, and were kept in normal housing conditions the rest of the time. All mice were housed in groups under a 12-h light/dark cycle, with illumination commencing at 7:00, and maintained at a temperature range of 20–22 °C and humidity levels between 30–60%. They had ad libitum access to food and water; however, the mice were transitioned to individual housing 1 week before the commencement of the experimental procedures. Tested cells were seeded on 6-well plates and placed in the HP chamber (pressure: −50 kPa, CO₂ partial pressure: 5 kPa, 37 °C) for the indicated times.

Physical function measurements and blood analysis

Rotarod evaluations were conducted in accordance with previously established methodologies⁹². Assessment of exercise capacity was conducted following established methodology⁹³, as was assessment of grip strength⁹⁴. Gross haematological parameters were evaluated through the analysis of EDTA-treated whole blood, utilizing a blood cell analyser (Innovent, HB-7021V). For serum biochemical analysis, blood samples were collected and allowed to clot for a duration of 2 h at room temperature or overnight at a temperature of 4 °C. Subsequently, the samples were subjected to centrifugation at 1,000 × g for 10 min to isolate the serum. An aliquot of 200 μl of serum was then utilized for the assessment of liver and renal function, as well as blood lipid levels, employing a test kit.

Postmortem pathological examination

Postmortem pathological examination was conducted as previously reported^92,94. Briefly, mice corpses were preserved in 4% paraformaldehyde individually within 24 h, and the corpses were subjected to a blinded examination by pathologists after undergoing a systematic assessment protocol. In summary, the evaluation included quantification of tumour burden, defined as the aggregate of various tumour types present in each mouse, as well as disease burden, which encompassed the totality of distinct histopathological alterations observed in the major organs of each mouse. In addition, the severity of each lesion and the extent of inflammation, characterized by lymphocytic infiltrate, were assessed.

Implementation of mechanical positive pressure

The implementation of mechanical positive pressure adhered to the methodology previously outlined in our research⁹⁵. In summary, when cell confluence reached ~80%, a continuous compressive force of 4 g cm⁻² was exerted on the MSCs, utilizing class layers and 50 ml plastic tube caps that contained weighted metal spheres.

Flow cytometric analysis

To assess the rate of cell death, cells were incubated with Annexin V and 7AAD (BD Pharmingen) in a 1× Annexin V binding buffer (BD Pharmingen) for 15 min at room temperature. The analysis of cell death levels was conducted within a 30-min timeframe using FlowJo (v.10.0).

Cathepsin B detection with Rhodamine 110

Live cells were cultured on a confocal dish and subsequently stained with 1× Rhodamine 110-(RR)2 substrate for 15 min, in accordance with manufacturer guidelines provided in the Cathepsin B Assay kit. Red fluorescence intensities, indicative of cathepsin B activity within the cells, were observed using the Zeiss Elyra 7 system equipped with Lattice SIM technology.

LysoTracker staining

Live cells were seeded on a confocal dish and incubated in LysoTracker Green according to manufacturer protocol. Images were taken using the Zeiss Elyra 7 system with Lattice SIM at 37 °C in 5% CO₂.

LMP measurement with AO staining

AO is a metachromatic fluorescent dye that specifically targets lysosomes, displaying red fluorescence within intact lysosomal structures and green fluorescence in the cytoplasmic region. Acridine orange enters lysosomes and is redistributed under stress stimulation, which is used to analyse lysosomal membrane permeability through absorption and intracellular redistribution⁴⁰. Following manufacturer protocol, live cells were incubated with AO and rinsed with Hank’s Balanced Salt Solution (HBSS) with 3% FBS. Images were taken using the Zeiss Elyra 7 system with Lattice SIM at 37 °C in 5% CO₂.

Measurement of lysosomal pH utilizing RatioWorks PDMPO Dextran

The measurement of lysosomal pH was conducted using ratiometric fluorescence imaging with the pH-sensitive dye RatioWorks PDMPO Dextran, following the methodology outlined in previous studies⁴⁴. In accordance with manufacturer guidelines, cells were incubated with RatioWorks PDMPO Dextran for 15 min, followed by washing with HBSS supplemented with 3% FBS. Fluorescence was then read at Ex/Em= 360/540 and 360/450 nm using the Zeiss Elyra 7 with Lattice SIM at 37 °C in 5% CO₂. The fluorescence emission intensity ratios of individual cells, measured following each pH treatment, were analysed as a function of pH and fitted to a Boltzmann sigmoid equation. This equation was subsequently employed as a standard curve to determine the experimental lysosomal pH for each cell.

Assessment of cytosolic free calcium ion concentrations

Cytosolic calcium ion (Ca²⁺) concentrations were quantified utilizing Fluo-4 AM. In accordance with manufacturer instructions, cells were incubated with Fluo-4 AM and subsequently imaged using the Zeiss Elyra 7 equipped with Lattice SIM at a temperature of 37 °C in a 5% CO₂ environment. A total of 10–22 cells were examined across a minimum of 3 distinct experimental trials, and the data were analysed using ImageJ software.

Cell-attached recordings

In cell-attached patch mode recordings, the bath solution utilized comprised the following concentrations (in mM): 140 KCl, 10 D-glucose, 10 HEPES and 1 MgCl₂, with the pH adjusted to 7.4 using KOH. The solution utilized in the patch pipette was composed of the following concentrations (in mM): 130 NaCl, 5 KCl, 10 TEA (tetraethylammonium) chloride, 8 D-glucose, 10 HEPES, 1.2 MgCl₂ and 1.5 CaCl₂, with the pH adjusted to 7.4 using sodium hydroxide. Electrodes were fabricated utilizing fire-polished glass electrodes (AM Systems, Glass Borosilicate, 1.5 mm outer diameter, 0.86 mm inner diameter) exhibiting resistances ranging from 1.5 to 2 MOhms. Electrical signals were enhanced and documented utilizing an Axopatch 1D amplifier in conjunction with pCLAMP 10 software (Molecular Devices). The data underwent filtering at a frequency of 1 kHz and were digitally sampled at a rate of 3 kHz through a Digidata 1322A analogue-to-digital converter (Molecular Devices). The analysis was conducted offline utilizing Clampfit 10.2 (Molecular Devices) and Origin 7.5 software (OriginLab). Data recordings were carried out at ambient room temperature. Hypobaric pressure was induced by administering short pulses of hypobaric pressure via the recording electrode, utilizing a Clampex-controlled pressure clamp HSPC-1 device (ALA Scientific). Furthermore, stretch-activated channels were documented at a holding potential of −80 mV, utilizing pressure steps ranging from 0 to −200 mmHg in increments of −10 mmHg. For analytical purposes, 3–10 recording traces were averaged per cell.

Immunofluorescence staining

Frozen sections and cellular samples were subjected to fixation in 4% paraformaldehyde (Sigma-Aldrich) for 20 min at ambient temperature. Subsequently, the samples underwent staining with primary antibodies, followed by the application of secondary antibodies. Ultimately, the samples were preserved using DAPI mounting medium and affixed to adhesive microscope slides.

Western blotting

Total protein was extracted using a protein extraction kit (Thermo Fisher) following manufacturer guidelines. A total of 20 μg of protein from each sample was applied to SDS–polyacrylamide electrophoresis gels and subsequently transferred to PVDF membranes (Millipore). Following this, the membranes underwent a blocking procedure for 1 h, after which they were incubated with primary antibodies at 4 °C overnight. This was succeeded by a 1-h incubation with HRP-conjugated secondary antibodies at room temperature. Immunoreactive proteins were visualized using the SuperSignal West Pico Chemiluminescent Substrate (Thermo Fisher) and subsequently scanned through gel electrophoresis. Image Studio Software (v.5.2) was used for western blot scanning and analysis.

ELISA

In accordance with manufacturer guidelines, the concentrations of human interleukins IL-1, IL-6 and IL-8, TNF-α, MCP-1, matrix metalloproteinase-1 (MMP-1) and matrix metalloproteinase-14 (MMP-14) were quantified using the human ELISA Quantikine Immunoassay kit (CUSABIO). In addition, the concentrations of mouse IL-1, IL-6, IL-8, IL-17, TNF-α, MCP-1, MMP-1, MMP-14 and IFNγ were assessed using the human ELISA Quantikine Immunoassay kit (Maisha).

SA-β-galactosidase staining

The cellular senescence of frozen sections and MSCs was evaluated following manufacturer instructions for the β-galactosidase staining kit (Solarbio). The proportion of senescent cells was determined by calculating the ratio of positively stained cells to the total number of cells counted across five distinct fields of view. Each experiment was conducted at least three times to ensure reliability of the results.

H&E staining

The samples that were subjected to dissection were subsequently fixed in a 4% paraformaldehyde solution (Sigma-Aldrich) and then embedded in paraffin. The paraffin sections, measuring 5 μm in thickness, were stained using H&E.

Micro-CT and analysis

The femurs were preserved in a 4% paraformaldehyde solution and subsequently examined using a Venus Micro CT (PINGSENG Healthcare). Scanning was conducted under the following parameters: tube voltage of 90 kV, tube current of 70 μA and a voxel size of 13 μm. The resulting data were visualized and analysed with the aid of Avatar software (PINGSENG Healthcare).

Bioluminescence imaging

Mice were administered an intraperitoneal injection of 3 mg d-luciferin in 200 μl of phosphate-buffered saline. Anaesthesia was induced in the mice using isoflurane, and bioluminescence imaging was conducted using a Xenogen IVIS 100 system (PerkinElmer) following manufacturer guidelines. Fluorescence intensity was subsequently quantified using Living Image software (PerkinElmer).

Lysosomal protein extraction

Lysosomal protein was enriched using superparamagnetic iron oxide nanoparticles (SPIONs), as described in a previous report⁶⁵. In summary, mBMMSCs were plated at a density of 2 × 10⁶ cells per 10-cm dish. The cells were treated with a 10% (v/v) solution of SPIONs and incubated for 24 h. Following this incubation, the cells were washed and subsequently cultured in fresh DMEM for an additional 24 h. After this period, the cells were washed again, collected in 2 ml of isolation buffer and then homogenized using a 15 ml Dounce homogenizer. Then the above sample was centrifuged to obtain combined post-nuclear supernatant and the resuspended pellet. Finally, the combined sample was enriched using LS columns in combination with a QuadroMACS magnet (both Miltenyi Biotec) to elute lysosomes.

DIA analysis

Proteins were subjected to analysis using the Q-Exactive HF X mass spectrometer (Thermo Fisher). The extraction of proteins was performed using a 1× cocktail containing the requisite concentrations of SDS L3 and EDTA. Subsequently, the samples were centrifuged for 15 min at 25,000 × g at a temperature of 4 °C. The resulting supernatants were then collected for subsequent Bradford quantification and SDS–PAGE analysis. The proteins underwent enzymatic hydrolysis utilizing the Trypsin enzyme, followed by separation through High pH Reverse Phase chromatography to yield freeze-dried peptide samples. For data-dependent acquisition (DDA) analysis, the peptides, once separated by liquid chromatography (LC), were ionized using nano-electrospray ionization (nanoESI) and subsequently introduced into a Q-Exactive HF X tandem mass spectrometer (Thermo Fisher) operating in DDA detection mode. In the context of DIA analysis, peptides that were separated via LC underwent ionization through nanoESI before being introduced into a Q-Exactive HF X tandem mass spectrometer (Thermo Fisher) operating in DIA detection mode. The sample data obtained from the high-resolution mass spectrometer facilitated the identification of data-dependent acquisition (DDA) data using the Andromeda search engine integrated within MaxQuant v.1.5.3.30. The results of these identifications were subsequently utilized for the construction of a spectral library. In the context of extensive data derived from DIA, the mProphet algorithm was employed to conduct quality control measures. For the differential analysis utilizing MSstats, the criteria for identifying differentially expressed proteins were established as a fold-change greater than 1.5 and a P value of less than 0.05, which were deemed significant. The identified proteins were systematically annotated and categorized into specific pathways utilizing the GO and the Kyoto Encyclopedia of Genes and Genomes (KEGG) databases. Following the quantitative analysis, differential proteins between the compared groups were discerned for functional enrichment and subcellular localization assessment.

iTRAQ analysis

Lysosomal protein levels were measured using a protein concentration assay (Bio-Rad). Subsequently, proteolysis and iTRAQ labelling of the resulting peptides were performed. The fractionation of peptides was carried out using the Shimadzu LC-20AB liquid chromatography system. The samples were amalgamated in accordance with the chromatographic elution peak profile. The separation process was conducted using the Thermo UltiMate 3000 Ultra High-Performance Liquid Chromatography system. The peptides that were separated through liquid phase chromatography underwent ionization via a nanoESI source and subsequently directed to a tandem mass spectrometer (Q-Exactive HF X, Thermo Fisher) for analysis utilizing the DDA mode. The raw MS/MS data were transformed into MGF format and the resulting MGF files were subsequently analysed using the local Mascot server for database comparison. Furthermore, protein quantification was conducted using IQuant, accompanied by quality control measures. Proteins exhibiting a false-discovery rate (FDR) of less than 1% were utilized as the input for GO analysis. In addition, a comprehensive examination of differentially expressed proteins was conducted, encompassing both GO enrichment analysis and KEGG pathway enrichment analysis.

siRNA transfection

In the process of siRNA transfection, human UMSCs were subjected to transfection with siRNAs targeting TRPV2, Piezo1, calpain 2, TMEM59, TMEM263 and TMEM127 (Ribo, China) utilizing the Lipofectamine RNAiMAX transfection reagent (Thermo Fisher), in accordance with manufacturer guidelines. Non-targeting control siRNAs (Ribo) served as negative controls. The efficiency of transfection was assessed through western blot analysis.

Osteogenic differentiation assay

mBMMSCs were cultured in 6-well plates. Upon reaching 100% confluence, the growth medium was substituted with an osteogenic medium comprising 2 mM β-glycerophosphate (Sigma), 100 μM L-ascorbic acid 2-phosphate (Sigma) and 10 nM dexamethasone (Sigma). Following a 4-week period of osteogenic induction, the cultures were subjected to staining with 1% alizarin red-S (Sigma). The area positive for alizarin red was quantified using ImageJ software and expressed as a percentage of the total area.

Adipogenic differentiation

mBMMSCs were cultured in a 6-well plate. Upon reaching 100% confluence, the culture medium was substituted with an adipogenic medium comprising 500 nM isobutylmethylxanthine (Sigma-Aldrich), 60 μM indomethacin (Sigma-Aldrich), 500 nM hydrocortisone (Sigma-Aldrich), 10 μg ml⁻¹ insulin (Sigma-Aldrich) and 100 nM L-ascorbic acid phosphate. After a 4-week induction period, the adipocytes were subjected to oil red O staining (Sigma-Aldrich), and the number of positively stained cells was quantified using microscopy, expressed as a ratio of the total cell count.

TRAP staining

Frozen sections were subjected to staining using the TRAP Stain kit (Wako), following the protocols outlined by the manufacturer⁹⁶. Multinucleated cells that exhibited red staining (TRAP-positive) were identified as osteoclasts. The resulting images were analysed using an optical microscope (Zeiss).

Statistical analysis

All experiments were conducted in biological triplicate, and the data are presented as mean values accompanied by standard deviation. Statistical analyses and graphical representations were conducted using GraphPad Prism 7 v.7.1.0. Comparisons among multiple groups were evaluated using one-way analysis of variance (ANOVA), followed by Tukey’s test. For comparisons between two groups, statistical significance was assessed utilizing independent unpaired two-tailed Student’s t-tests. P values less than 0.05 were deemed statistically significant.

Reporting summary

Further information on research design is available in the Nature Portfolio Reporting Summary linked to this article.

Data availability

The data supporting the results in this study are available in the main text or the supplementary materials. The iTRAQ analysis datasets generated during the study are available from the PRIDE database (https://www.ebi.ac.uk/pride/archive/projects/PXD065406)⁹⁷. The DIA analysis datasets generated during the study are available from the iProX database (https://www.iprox.cn/page/project.html?id=IPX0012428000)⁹⁸. The RNA-seq datasets generated during the study are available from the Sequence Read Archive (SRA) (https://www.ncbi.nlm.nih.gov/Traces/study/?acc=PRJNA1281710&o=acc_s%3Aa)⁹⁹. All data generated and analysed during the study are available from the corresponding authors on reasonable request. Source data are provided with this paper.

References

López-Otín, C., Blasco, M. A., Partridge, L., Serrano, M. & Kroemer, G. Hallmarks of aging: an expanding universe. Cell 186, 243–278 (2023).
Article PubMed Google Scholar
Lelarge, V., Capelle, R., Oger, F., Mathieu, T. & Le Calvé, B. Senolytics: from pharmacological inhibitors to immunotherapies, a promising future for patients’ treatment. npj Aging 10, 12 (2024).
Article PubMed PubMed Central CAS Google Scholar
Kirkland, J. L. & Tchkonia, T. Senolytic drugs: from discovery to translation. J. Intern. Med. 288, 518–536 (2020).
Article PubMed PubMed Central CAS Google Scholar
Zhu, Y. et al. The Achilles’ heel of senescent cells: from transcriptome to senolytic drugs. Aging Cell 14, 644–658 (2015).
Article PubMed PubMed Central CAS Google Scholar
Xu, M. et al. Senolytics improve physical function and increase lifespan in old age. Nat. Med. 24, 1246–1256 (2018).
Article PubMed PubMed Central CAS Google Scholar
Zhang, P. et al. Senolytic therapy alleviates Aβ-associated oligodendrocyte progenitor cell senescence and cognitive deficits in an Alzheimer’s disease model. Nat. Neurosci. 22, 719–728 (2019).
Article PubMed PubMed Central CAS Google Scholar
Novais, E. J. et al. Long-term treatment with senolytic drugs Dasatinib and Quercetin ameliorates age-dependent intervertebral disc degeneration in mice. Nat. Commun. 12, 5213 (2021).
Article PubMed PubMed Central CAS Google Scholar
Li, W. et al. Emerging senolytic agents derived from natural products. Mech. Ageing Dev. 181, 1–6 (2019).
Article PubMed CAS Google Scholar
Hasegawa, T. et al. Cytotoxic CD4⁺ T cells eliminate senescent cells by targeting cytomegalovirus antigen. Cell 186, 1417–1431.e20 (2023).
Article PubMed CAS Google Scholar
Amor, C. et al. Senolytic CAR T cells reverse senescence-associated pathologies. Nature 583, 127–132 (2020).
Article PubMed PubMed Central CAS Google Scholar
Suda, M. et al. Senolytic vaccination improves normal and pathological age-related phenotypes and increases lifespan in progeroid mice. Nat. Aging 1, 1117–1126 (2021).
Article PubMed Google Scholar
Heisenberg, C. P. & Bellaiche, Y. Forces in tissue morphogenesis and patterning. Cell 153, 948–962 (2013).
Article PubMed CAS Google Scholar
Bergert, M. et al. Cell surface mechanics gate embryonic stem cell differentiation. Cell Stem Cell 28, 209–216.e4 (2021).
Article PubMed PubMed Central CAS Google Scholar
Sun, Y., Chen, C. S. & Fu, J. Forcing stem cells to behave: a biophysical perspective of the cellular microenvironment. Annu. Rev. Biophys. 41, 519–542 (2012).
Article PubMed PubMed Central Google Scholar
Hodgkinson, T., Kelly, D. C., Curtin, C. M. & O’Brien, F. J. Mechanosignalling in cartilage: an emerging target for the treatment of osteoarthritis. Nat. Rev. Rheumatol. 18, 67–84 (2022).
Article PubMed Google Scholar
An, J. et al. Mechanical stimuli-driven cancer therapeutics. Chem. Soc. Rev. 52, 30–46 (2023).
Article PubMed CAS Google Scholar
Hilscher, M. B. et al. Mechanical stretch increases expression of CXCL1 in liver sinusoidal endothelial cells to recruit neutrophils, generate sinusoidal microthombi, and promote portal hypertension. Gastroenterology 157, 193–209.e9 (2019).
Article PubMed CAS Google Scholar
Mehta, V. et al. Mechanical forces regulate endothelial-to-mesenchymal transition and atherosclerosis via an Alk5-Shc mechanotransduction pathway. Sci. Adv. 7, eabg5060 (2021).
Fogarty, M. J. & Sieck, G. C. Evolution and functional differentiation of the diaphragm muscle of mammals. Compr. Physiol. 9, 715–766 (2019).
Article PubMed PubMed Central Google Scholar
Arnoldi, C. C. The venous return from the lower leg in health and in chronic venous insufficiency: a synthesis. Acta Orthop. 35, 3–75 (1964).
Ludbrook, J. The musculovenous pumps of the human lower limb. Am. Heart J. 71, 635–641 (1966).
Article PubMed CAS Google Scholar
Knox, P., Levick, J. R. & McDonald, J. N. Synovial fluid—its mass, macromolecular content and pressure in major limb joints of the rabbit. Q. J. Exp. Physiol. 73, 33–45 (1988).
Article PubMed CAS Google Scholar
Scott, D., Levick, J. R. & Miserocchi, G. Non-linear dependence of interstitial fluid pressure on joint cavity pressure and implications for interstitial resistance in rabbit knee. Acta Physiol. Scand. 179, 93–101 (2003).
Article PubMed CAS Google Scholar
Costa, M. L. et al. Effect of negative pressure wound therapy vs standard wound management on 12-month disability among adults with severe open fracture of the lower limb: the WOLLF Randomized Clinical Trial. JAMA 319, 2280–2288 (2018).
Article PubMed PubMed Central Google Scholar
Costa, M. L. et al. Effect of incisional negative pressure wound therapy vs standard wound dressing on deep surgical site infection after surgery for lower limb fractures associated with major trauma: the WHIST Randomized Clinical Trial. JAMA 323, 519–526 (2020).
Article PubMed PubMed Central Google Scholar
Gillespie, B. M. et al. Closed incision negative pressure wound therapy versus standard dressings in obese women undergoing caesarean section: multicentre parallel group randomised controlled trial. BMJ 373, n893 (2021).
Article PubMed PubMed Central Google Scholar
Seidel, D. et al. Negative pressure wound therapy vs conventional wound treatment in subcutaneous abdominal wound healing impairment: the SAWHI Randomized Clinical Trial. JAMA Surg. 155, 469–478 (2020).
Article PubMed PubMed Central Google Scholar
Powers, J. G., Higham, C., Broussard, K. & Phillips, T. J. Wound healing and treating wounds: chronic wound care and management. J. Am. Acad. Dermatol. 74, 607–625; quiz 625–626 (2016).
Neuhaus, C. & Hinkelbein, J. Cognitive responses to hypobaric hypoxia: implications for aviation training. Psychol. Res. Behav. Manage. 7, 297–302 (2014).
Article Google Scholar
Mujika, I., Sharma, A. P. & Stellingwerff, T. Contemporary periodization of altitude training for elite endurance athletes: a narrative review. Sports Med. 49, 1651–1669 (2019).
Article PubMed Google Scholar
Jin, P., Jan, L. Y. & Jan, Y. N. Mechanosensitive ion channels: structural features relevant to mechanotransduction mechanisms. Annu. Rev. Neurosci. 43, 207–229 (2020).
Article PubMed CAS Google Scholar
Coste, B. et al. Piezo1 and Piezo2 are essential components of distinct mechanically activated cation channels. Science 330, 55–60 (2010).
Article PubMed PubMed Central CAS Google Scholar
Muraki, K. et al. TRPV2 is a component of osmotically sensitive cation channels in murine aortic myocytes. Circ. Res. 93, 829–838 (2003).
Article PubMed CAS Google Scholar
Bang, H., Kim, Y. & Kim, D. TREK-2, a new member of the mechanosensitive tandem-pore K⁺ channel family. J. Biol. Chem. 275, 17412–17419 (2000).
Article PubMed CAS Google Scholar
Zhang, M. et al. Structure of the mechanosensitive OSCA channels. Nat. Struct. Mol. Biol. 25, 850–858 (2018).
Article PubMed CAS Google Scholar
Jeong, H. et al. Structures of the TMC-1 complex illuminate mechanosensory transduction. Nature 610, 796–803 (2022).
Article PubMed PubMed Central CAS Google Scholar
Beaulieu-Laroche, L. et al. TACAN is an ion channel involved in sensing mechanical pain. Cell 180, 956–967.e17 (2020).
Article PubMed CAS Google Scholar
Akter, F. et al. Multi-cell line analysis of lysosomal proteomes reveals unique features and novel lysosomal proteins. Mol. Cell. Proteom. 22, 100509 (2023).
Article CAS Google Scholar
Wang, F., Gómez-Sintes, R. & Boya, P. Lysosomal membrane permeabilization and cell death. Traffic 19, 918–931 (2018).
Article PubMed CAS Google Scholar
Wang, L. H. et al. KPT330 promotes the sensitivity of glioblastoma to olaparib by retaining SQSTM1 in the nucleus and disrupting lysosomal function. Autophagy 20, 295–310 (2024).
Article PubMed CAS Google Scholar
Goll, D. E., Thompson, V. F., Li, H., Wei, W. & Cong, J. The calpain system. Physiol. Rev. 83, 731–801 (2003).
Article PubMed CAS Google Scholar
Gómez-Sintes, R., Ledesma, M. D. & Boya, P. Lysosomal cell death mechanisms in aging. Ageing Res. Rev. 32, 150–168 (2016).
Article PubMed Google Scholar
Basisty, N. et al. A proteomic atlas of senescence-associated secretomes for aging biomarker development. PLoS Biol. 18, e3000599 (2020).
Article PubMed PubMed Central Google Scholar
Mo, S. et al. Mycobacterium tuberculosis utilizes host histamine receptor H1 to modulate reactive oxygen species production and phagosome maturation via the p38MAPK-NOX2 axis. mBio 13, e0200422 (2022).
Article PubMed Google Scholar
Ma, Z. et al. TMEM59 ablation leads to loss of olfactory sensory neurons and impairs olfactory functions via interaction with inflammation. Brain Behav. Immun. 111, 151–168 (2023).
Article PubMed CAS Google Scholar
Kefauver, J. M., Ward, A. B. & Patapoutian, A. Discoveries in structure and physiology of mechanically activated ion channels. Nature 587, 567–576 (2020).
Article PubMed PubMed Central CAS Google Scholar
Kamine, T. H., Elmadhun, N. Y., Kasper, E. M., Papavassiliou, E. & Schneider, B. E. Abdominal insufflation for laparoscopy increases intracranial and intrathoracic pressure in human subjects. Surg. Endosc. 30, 4029–4032 (2016).
Article PubMed Google Scholar
Valenza, F. et al. Effects of continuous negative extra-abdominal pressure on cardiorespiratory function during abdominal hypertension: an experimental study. Intensive Care Med. 31, 105–111 (2005).
Article PubMed Google Scholar
Scalea, T. M. et al. Increased intra-abdominal, intrathoracic, and intracranial pressure after severe brain injury: multiple compartment syndrome. J. Trauma 62, 647–656 (2007).
PubMed Google Scholar
Saggi, B. H. et al. Treatment of intracranial hypertension using nonsurgical abdominal decompression. J. Trauma 46, 646–651 (1999).
Article PubMed CAS Google Scholar
Nagpal, S. et al. Decompressive laparotomy to treat intractable cerebral hypoxia. J. Trauma 67, E152–E155 (2009).
PubMed Google Scholar
Kairinos, N., Solomons, M. & Hudson, D. A. The paradox of negative pressure wound therapy—in vitro studies. J. Plast. Reconstr. Aesthet. Surg. 63, 174–179 (2010).
Article PubMed Google Scholar
Evers, T. M. J., Holt, L. J., Alberti, S. & Mashaghi, A. Reciprocal regulation of cellular mechanics and metabolism. Nat. Metab. 3, 456–468 (2021).
Article PubMed PubMed Central Google Scholar
Gudipaty, S. A. et al. Mechanical stretch triggers rapid epithelial cell division through Piezo1. Nature 543, 118–121 (2017).
Article PubMed PubMed Central CAS Google Scholar
Yang, L. et al. Diabetic foot wound ulcers management by vacuum sealing drainage: a meta-analysis. Int. Wound J. 21, e14390 (2024).
Article PubMed Google Scholar
Choi, T. Y., Ang, L., Ku, B., Jun, J. H. & Lee, M. S. Evidence map of cupping therapy. J. Clin. Med. 10, 1750 (2021).
Vaccaro, M., Coppola, M., Ceccarelli, M., Montopoli, M. & Guarneri, C. The good and the bad of cupping therapy: case report and review of the literature. Eur. Rev. Med. Pharmacol. Sci. 25, 2327–2330 (2021).
PubMed CAS Google Scholar
Yuan, F. et al. OSCA1 mediates osmotic-stress-evoked Ca²⁺ increases vital for osmosensing in Arabidopsis. Nature 514, 367–371 (2014).
Article PubMed CAS Google Scholar
Hou, C. et al. DUF221 proteins are a family of osmosensitive calcium-permeable cation channels conserved across eukaryotes. Cell Res. 24, 632–635 (2014).
Article PubMed PubMed Central CAS Google Scholar
Gao, X. et al. Treatment of autosomal dominant hearing loss by in vivo delivery of genome editing agents. Nature 553, 217–221 (2018).
Article PubMed CAS Google Scholar
Chatzigeorgiou, M., Bang, S., Hwang, S. W. & Schafer, W. R. tmc-1 encodes a sodium-sensitive channel required for salt chemosensation in C. elegans. Nature 494, 95–99 (2013).
Article PubMed PubMed Central CAS Google Scholar
Schroeder, B. C., Cheng, T., Jan, Y. N. & Jan, L. Y. Expression cloning of TMEM16A as a calcium-activated chloride channel subunit. Cell 134, 1019–1029 (2008).
Article PubMed PubMed Central CAS Google Scholar
Sharif-Naeini, R. et al. Polycystin-1 and -2 dosage regulates pressure sensing. Cell 139, 587–596 (2009).
Article PubMed CAS Google Scholar
Zhang, S. et al. Degradation of alpha-synuclein by dendritic cell factor 1 delays neurodegeneration and extends lifespan in Drosophila. Neurobiol. Aging 67, 67–74 (2018).
Article PubMed CAS Google Scholar
Ebner, M. et al. Nutrient-regulated control of lysosome function by signaling lipid conversion. Cell 186, 5328–5346.e6 (2023).
Article PubMed CAS Google Scholar
Serrano-Puebla, A. & Boya, P. Lysosomal membrane permeabilization in cell death: new evidence and implications for health and disease. Ann. N. Y. Acad. Sci. 1371, 30–44 (2016).
Article PubMed Google Scholar
Ballabio, A. & Bonifacino, J. S. Lysosomes as dynamic regulators of cell and organismal homeostasis. Nat. Rev. Mol. Cell Biol. 21, 101–118 (2020).
Article PubMed CAS Google Scholar
Aits, S. & Jäättelä, M. Lysosomal cell death at a glance. J. Cell Sci. 126, 1905–1912 (2013).
Article PubMed CAS Google Scholar
Chaib, S., Tchkonia, T. & Kirkland, J. L. Cellular senescence and senolytics: the path to the clinic. Nat. Med. 28, 1556–1568 (2022).
Article PubMed PubMed Central CAS Google Scholar
Rovira, M. et al. The lysosomal proteome of senescent cells contributes to the senescence secretome. Aging Cell 21, e13707 (2022).
Article PubMed PubMed Central CAS Google Scholar
Johmura, Y. et al. Senolysis by glutaminolysis inhibition ameliorates various age-associated disorders. Science 371, 265–270 (2021).
Article PubMed CAS Google Scholar
Curnock, R. et al. TFEB-dependent lysosome biogenesis is required for senescence. EMBO J. 42, e111241 (2023).
Article PubMed PubMed Central CAS Google Scholar
Guerrero, A. et al. Galactose-modified duocarmycin prodrugs as senolytics. Aging Cell 19, e13133 (2020).
Article PubMed PubMed Central CAS Google Scholar
Munoz-Espin, D. et al. A versatile drug delivery system targeting senescent cells. EMBO Mol. Med. 10, EMMM201809355 (2018).
Fuhrmann-Stroissnigg, H. et al. Identification of HSP90 inhibitors as a novel class of senolytics. Nat. Commun. 8, 422 (2017).
Article PubMed PubMed Central Google Scholar
Naqvi, K. et al. Long-term follow-up of lower dose dasatinib (50 mg daily) as frontline therapy in newly diagnosed chronic-phase chronic myeloid leukemia. Cancer 126, 67–75 (2020).
Article PubMed CAS Google Scholar
Ottmann, O. et al. Long-term efficacy and safety of dasatinib in patients with chronic myeloid leukemia in accelerated phase who are resistant to or intolerant of imatinib. Blood Cancer J. 8, 88 (2018).
Article PubMed PubMed Central Google Scholar
Liu, X. et al. Resurrection of endogenous retroviruses during aging reinforces senescence. Cell 186, 287–304.e26 (2023).
Article PubMed CAS Google Scholar
Yang, J. H. et al. Loss of epigenetic information as a cause of mammalian aging. Cell 186, 305–326.e27 (2023).
Article PubMed PubMed Central CAS Google Scholar
Zhou, Z. et al. Engineering longevity-design of a synthetic gene oscillator to slow cellular aging. Science 380, 376–381 (2023).
Article PubMed PubMed Central CAS Google Scholar
Weaver, K. J., Holt, R. A., Henry, E., Lyu, Y. & Pletcher, S. D. Effects of hunger on neuronal histone modifications slow aging in Drosophila. Science 380, 625–632 (2023).
Article PubMed PubMed Central CAS Google Scholar
Singh, P. et al. Taurine deficiency as a driver of aging. Science 380, eabn9257 (2023).
Article PubMed PubMed Central CAS Google Scholar
Sun, S. et al. CHIT1-positive microglia drive motor neuron ageing in the primate spinal cord. Nature 624, 611–620 (2023).
Article PubMed CAS Google Scholar
Schroer, A. B. et al. Platelet factors attenuate inflammation and rescue cognition in ageing. Nature 620, 1071–1079 (2023).
Article PubMed PubMed Central CAS Google Scholar
Zhang, Z. et al. Increased hyaluronan by naked mole-rat Has2 improves healthspan in mice. Nature 621, 196–205 (2023).
Article PubMed PubMed Central CAS Google Scholar
Besco, R. O., Sangal, S. P., Nesthus, T. E. & Veronneau, S. J. H. Study of life expectancy for a sample of retired airline pilots. Proc. Hum. Factors Ergon. Soc. Annu. Meet. 1, 128–132 (1994).
Article Google Scholar
Li, Y. et al. Hypoxia potentially promotes Tibetan longevity. Cell Res. 27, 302–305 (2017).
Article PubMed Google Scholar
Burtscher, J., Mallet, R. T., Burtscher, M. & Millet, G. P. Hypoxia and brain aging: neurodegeneration or neuroprotection?. Ageing Res. Rev. 68, 101343 (2021).
Article PubMed CAS Google Scholar
Yeo, E. J. Hypoxia and aging. Exp. Mol. Med. 51, 1–15 (2019).
PubMed CAS Google Scholar
Qu, Y. et al. Apoptotic vesicles inherit SOX2 from pluripotent stem cells to accelerate wound healing by energizing mesenchymal stem cells. Acta Biomater. 149, 258–272 (2022).
Article PubMed CAS Google Scholar
Liu, D. et al. Circulating apoptotic bodies maintain mesenchymal stem cell homeostasis and ameliorate osteopenia via transferring multiple cellular factors. Cell Res. 28, 918–933 (2018).
Article PubMed PubMed Central CAS Google Scholar
Baker, D. J. et al. Naturally occurring p16(Ink4a)-positive cells shorten healthy lifespan. Nature 530, 184–189 (2016).
Article PubMed PubMed Central CAS Google Scholar
Cai, Y. et al. Elimination of senescent cells by β-galactosidase-targeted prodrug attenuates inflammation and restores physical function in aged mice. Cell Res. 30, 574–589 (2020).
Article PubMed PubMed Central CAS Google Scholar
Xu, Q. et al. The flavonoid procyanidin C1 has senotherapeutic activity and increases lifespan in mice. Nat. Metab. 3, 1706–1726 (2021).
Article PubMed PubMed Central CAS Google Scholar
He, D. et al. Mechanical load-induced H₂S production by periodontal ligament stem cells activates M1 macrophages to promote bone remodeling and tooth movement via STAT1. Stem Cell Res. Ther. 11, 112 (2020).
Article PubMed PubMed Central CAS Google Scholar
Saita, M. et al. Novel antioxidative nanotherapeutics in a rat periodontitis model: reactive oxygen species scavenging by redox injectable gel suppresses alveolar bone resorption. Biomaterials 76, 292–301 (2016).
Article PubMed CAS Google Scholar
Meng, B. et al. Intermittent hypobaric pressure induces selective senescent cells death and alleviate age-related osteoporosis. Pride https://www.ebi.ac.uk/pride/archive/projects/PXD065406 (2025).
Meng, B. et al. Intermittent hypobaric pressure induces selective senescent cells death and alleviate age-related osteoporosis. iProX https://www.iprox.cn/page/project.html?id=IPX0012428000 (2025).
Meng, B. et al. Intermittent hypobaric pressure induces selective senescent cells death and alleviate age-related osteoporosis. SRA https://www.ncbi.nlm.nih.gov/Traces/study/?acc=PRJNA1281710&o=acc_s%3Aa (2025).

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Acknowledgements

This work was supported by grants from the National Key R&D Program of China (2021YFA1100600 to S.S.), the National Natural Science Foundation of China (82401162 to B.M., 82301123 to Y.Q.), the Guangdong Financial Fund for High-Caliber Hospital Construction (174-2018-XMZC-0001-03-0125, D-07 to S.S.), the Natural Science Foundation of Guangdong Province (2023A1515010626, 2024A1515012820 to Y.Q.; 2023A1515111127, 2025A1515010453 to B.M.), the Pearl River Talent Recruitment Program (2019ZT08Y485, 2019QN01Y138, 2019JC01Y182), the Guangzhou Basic and Applied Basic Research Scheme (SL2024A04J02205 to Y.Q.), and the nationally Funded Postdoctoral Researcher Program (2024M753795, GZC20233267 to B.M.).

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These authors contributed equally: Bowen Meng, Yan Qu.

Authors and Affiliations

Hospital of Stomatology, Guanghua School of Stomatology, Sun Yat-sen University, South China Center of Craniofacial Stem Cell Research, Guangdong Provincial Key Laboratory of Stomatology, Guangzhou, China
Bowen Meng, Yan Qu, Benyi Yang, Chaoran Fu, Yifan He, Zhulin Xue, Zeyuan Cao, Meng Hao, Xiao Zhang, Zhe An, Fen Chen, Xueli Mao, Yang Cao & Songtao Shi
Lingnan Medical Research Center, The first affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou, China
Jing Li & Rentao Wan
School and Hospital of Stomatology, Guangdong Engineering Research Center of Oral Restoration and Reconstruction, Guangzhou Medical University, Guangzhou, China
Xin Li
Department of Oral and Maxillofacial Surgery, Peking University School and Hospital of Stomatology, and National Center for Stomatology, and National Clinical Research Center for Oral Diseases, and National Engineering Research Center of Oral Biomaterials and Digital Medical Devices, Beijing, China
Zhulin Xue
Department of Prosthodontics, Peking University School and Hospital of Stomatology, Beijing, China
Xiao Zhang
International Center for Aging and Cancer, Hainan Medical University, Haikou, China
Ruibao Ren & Songtao Shi
Shanghai Institute of Hematology, State Key Laboratory for Medical Genomics, National Research Center for Translational Medicine, Ruijin Hospital affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, China
Ruibao Ren

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Bowen Meng
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Yan Qu
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Benyi Yang
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Chaoran Fu
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Yifan He
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Jing Li
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Rentao Wan
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Xin Li
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Zhulin Xue
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Zeyuan Cao
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Meng Hao
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Xiao Zhang
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Zhe An
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Fen Chen
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Ruibao Ren
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Xueli Mao
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Yang Cao
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Songtao Shi
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Contributions

B.M. designed the study plan, performed experimental procedures and drafted the manuscript. Y.Q. contributed to data acquisition and performed experimental procedures. B.Y. and C.F. contributed to data analysis and interpretation. Y.H. and X.L. contributed to data analysis and visualization. J.L. and R.W. performed electrophysiological experiments. Z.X. analysed mouse bone marrow. Z.C. and M.H. extracted lysosomal protein. X.Z. analysed protein profiles. Z.A. cultured cells. F.C. used the hypobaric pressure chamber. R.R., X.M. and Y.C. contributed to experimental design. S.S. conceived and supervised the project, designed experiments and wrote the manuscript. All authors approved the final version of the manuscript.

Corresponding author

Correspondence to Songtao Shi.

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The authors declare no competing interests.

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Nature Biomedical Engineering thanks Binsheng Wang and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

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Supplementary information

Supplementary Information

Supplementary Figs. 1–9.

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Supplementary Tables 1–18.

Supplementary Data 1

Unprocessed western blots for Supplementary Figs. 1, 2, 4 and 6.

Supplementary Data 2

Statistical source data for Supplementary Figs. 1, 2, 4 and 6–8.

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Statistical source data.

Source Data Figs. 1–3

Unmodified blots.

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Meng, B., Qu, Y., Yang, B. et al. Intermittent hypobaric pressure induces selective senescent cell death and alleviates age-related osteoporosis. Nat. Biomed. Eng (2026). https://doi.org/10.1038/s41551-025-01584-5

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Received: 12 June 2024
Accepted: 10 November 2025
Published: 14 January 2026
Version of record: 14 January 2026
DOI: https://doi.org/10.1038/s41551-025-01584-5