To the Editor: Sleep is a vital physiological process essential for cognitive function, memory consolidation, metabolic homeostasis, and immune regulation. Insomnia and chronic sleep deprivation (SD) have emerged as major public health concerns, inducing systemic inflammation, multiorgan dysfunction, and, in extreme cases, fatal outcomes.1 However, the pathological mechanisms underlying SD and effective interventions to mitigate its life-threatening consequences remain poorly understood. Mesenchymal stem cells (MSCs) exert therapeutic effects primarily through paracrine secretion of trophic factors, conferring anti-inflammatory, pro-repair, and tissue-protective functions. Extracellular vesicles derived from human umbilical cord MSCs have been shown to alleviate SD-induced anxiety and cognitive impairment by modulating neuroinflammation.2 Immunity- and matrix-regulatory cells (IMRCs), a novel MSC-like population derived from human embryonic stem cells, exhibit enhanced immunomodulatory and tissue-repair capacities. IMRC infusion suppresses proinflammatory cytokines (e.g., tumor necrosis factor-α TNF-α and interleukin 6 IL-6), enhances anti-inflammatory mediators (e.g., IL-10), and protects against organ damage through macrophage polarization and T cell regulation.3,4 Given that SD is closely associated with oxidative stress and inflammation, this study investigated whether IMRC infusion could attenuate SD-induced pathology and improve survival. The clinical-grade hESC line (Q-CTS-hESC-2, established and provided by National Stem Cell Resource Center, Beijing, China) for IMRC preparation was obtained, and IMRCs were generated according to previous protocols Figure 1A, 1B and Supplementary Figure 1A, https://links.lww.com/CM9/C796.3,4 To further explore the immunomodulatory properties of IMRCs, a coculture system was established with M1-polarized macrophages. Flow cytometric analysis revealed a marked reduction in CD80 surface expression (71.50% vs. 41.22%) when M1 macrophages were exposed to IMRC-conditioned medium Supplementary Figure 1B, https://links.lww.com/CM9/C796. These findings demonstrated that IMRCs effectively suppress inflammation.Figure 1: IMRC infusion as a protective strategy against sudden death triggered by severe SD. (A) Preparation workflow and representative images of IMRCs. (B) Flow cytometric analysis of cell surface marker expression in IMRCs. (C) Survival curves for C57BL/6J mice subjected to the control (Ctrl; n = 30 mice), SDsaline (n = 28 mice), and SDIMRC (n = 30 mice) treatments. The data are presented as the mean ± standard error of the mean (SEM). Statistical analysis was performed using nonlinear regression with curve comparison to determine significance (* P <0.05). (D) Representative staining of heart (Masson’s), liver (H CK-MB: MB isoenzyme of CK; GO: Gene ontology; HBDH: α-Hydroxybutyrate dehydrogenase; H IMRC: Immunity- and matrix-regulatory cells; LDH: Lactate dehydrogenase; ns: Not significant; SD: Sleep deprivation; SEM: Standard error of mean.All animal experiments were approved by the Animal Care and Use Committees of the Institute of Zoology, Chinese Academy of Sciences (No. IOZ-IACUC-2024-220). C57BL/6J mice were purchased from SPF Biotechnology Co., Ltd (Beijing, China). Mice were housed in the Laboratory Animal Center of the Institute of Zoology under identical conditions (12-hour light-dark cycle, five mice housed per cage, and free access to food and water). Previous studies have shown that SD in mice not only leads to premature death but also triggers systemic inflammation and reactive oxygen species (ROS) production.5 To evaluate whether IMRCs prevent premature death induced by severe SD, 8-week-old C57/BL6J mice were administered IMRCs (SDIMRC group) and normal saline (SDsaline group) through the tail vein, while the control group was untreated. Subsequently, both experimental groups were subjected to SD, and the control group was not subjected to SD Supplementary Figure 1C, https://links.lww.com/CM9/C796. The median survival time in the SDIMRC group was markedly extended compared with the SDsaline group, indicating the protective effect of IMRC therapy against sudden death induced by severe SD Figure 1C. In clinical practice, most sudden deaths are of cardiac origin. To assess the severity of myocardial injury, the plasma levels of four key myocardial enzymes—creatine kinase (CK), MB isoenzyme of CK (CK-MB), lactate dehydrogenase (LDH), and α-hydroxybutyrate dehydrogenase (HBDH)—were determined. Compared with the SDsaline group, the SDIMRC group exhibited a significant reduction in plasma levels of CK and HBDH, as well as a decreasing trend in plasma CK-MB and LDH levels (without statistical significance) Supplementary Figure 1D, https://links.lww.com/CM9/C796. These findings suggested that IMRC treatment effectively mitigates SD-induced myocardial injury. Given that severe SD increases the risk of cardiac events and induces neurological dysfunction and multisystem damage, the potential mechanisms by which IMRC treatment prolongs the survival time of SD mice were investigated. Immunofluorescence and hematoxylin and eosin (H the enriched pathways were involved in neurogenesis and memory formation, indicating enhanced brain plasticity and cognitive function Supplementary Figure 2E, https://links.lww.com/CM9/C796. In contrast, the SDsaline group exhibited enrichment of pathways related to inflammatory and oxidative stress pathways Supplementary Figure 2F, https://links.lww.com/CM9/C796. Quantitative polymerase chain reaction (qPCR) and bulk ribonucleic acid (RNA) sequencing (RNA-seq) analyses of peripheral tissues revealed that IMRC treatment conferred multiorgan protective effects under pathological conditions. In the liver, IMRC treatment improved metabolic homeostasis, restored the expression of genes related to lipid metabolism, and significantly reduced inflammatory cytokine levels Supplementary Figure 2G–L, https://links.lww.com/CM9/C796. In the lungs, IMRC treatment facilitated epithelial regeneration, attenuated pulmonary inflammation, and promoted tissue remodeling, contributing to the preservation of alveolar structure and function Supplementary Figure 3A–G, https://links.lww.com/CM9/C796. In the kidneys, IMRC treatment maintained renal function by alleviating tubular injury, reducing renal fibrosis, and upregulating the expression of reparative genes associated with tissue remodeling and anti-inflammation Supplementary Figure 3H–M, https://links.lww.com/CM9/C796. In the spleen, IMRC treatment effectively restored immune homeostasis by balancing proinflammatory and anti-inflammatory cytokines, reducing splenic immune cell activation, and preventing splenic atrophy Supplementary Figure 3N and O, https://links.lww.com/CM9/C796. Weighted gene coexpression network analysis (WGCNA) of bulk RNA-seq data derived from six tissues (with three biological replicates per group) identified 66 distinct gene modules. As a key analytical tool, WGCNA clusters genes with similar expression patterns, which helps elucidate functionally relevant biological pathways in complex transcriptomic datasets. The cluster dendrogram shown in Supplementary Figure 4A https://links.lww.com/CM9/C796 displays the gene modules based on coexpression patterns, while the eigengene adjacency heatmap shown in Supplementary Figure 4B https://links.lww.com/CM9/C796 illustrates the correlations among module eigengenes. Among these 66 modules, the dark olive green module was specifically enriched in the SDsaline group, whereas the violet module was preferentially enriched in the SDIMRC group Supplementary Figure 4C, https://links.lww.com/CM9/C796. Gene Ontology (GO) enrichment analysis of the module revealed activation of immune and inflammatory pathways, such as regulation of innate immune response and cytokine-mediated signaling Figure 1F, indicating that SD induces systemic inflammation and potential organ dysfunction. Conversely, the violet module in the SDIMRC group was enriched in tissue repair pathways, including angiogenesis regulation, mesenchyme development, and Wingless/Integrated (Wnt) signaling Supplementary Figure 4D, https://links.lww.com/CM9/C796, suggesting that IMRC treatment promotes regeneration and tissue recovery following SD-induced damage. Collectively, these results highlighted the therapeutic role of IMRCs in restoring tissue homeostasis and counteracting SD-induced stress. In conclusion, a single pre-SD dose of IMRCs markedly reduced inflammation and ROS accumulation, prevented multiorgan damage (including brain, liver, and spleen), and extended survival by nearly one-third. These findings support systemic IMRC-based anti-inflammatory intervention as a promising strategy to mitigate SD-induced organ injury and sudden death risk. Funding This work was financially supported by the Strategic Priority Research Program of Chinese Academy of Sciences (No. XDA 046205, XDB 1480201), the National Key Research and Development Program (Nos. 2021YFA1101400, 2020YFA0804000, 2024YFA1108302, and 2022YFA1103603), and the National Natural Science Foundation of China (No. 32370851). Conflicts of interest None.
Hou et al. (Fri,) studied this question.