bioRxiv Science⌕ Search
Home/Archive/bioRxiv

bioRxiv · 10.1101/2025.11.28.691223

Development of Long-Term Human Adipocyte Organoids Manifesting Aging in Response to Intermittent Hypoxia

Dong, Y.·Wang, J.·Feng, F.·Ali, I. H.·Karim, S.·Bock, J. M.·Somers, V. K.
View original source ↗Export metadata ↓

Abstract

Obstructive sleep apnea (OSA) (and consequent intermittent hypoxia (IH)) is increasingly recognized as a driver of adipose tissue dysfunction, insulin resistance, and aging. However, current in vitro experimental models inadequately capture the long-term effects of IH on human adipocytes. Here, we report the development and optimization of a robust long-term human adipocyte organoid culture system that faithfully recapitulates IH-induced adipocyte aging in vitro. Human stromal vascular fraction (SVF) cells, isolated from subcutaneous abdominal fat biopsies, were embedded in Matrigel and seeded into Biofloat U-bottom 96-well plates. Using a 1:1 Matrigel-cell mixture and optimized seeding volumes (5-20 {micro}L), we established adipocyte organoids that formed within 10-12 days and remained viable with stable morphology for up to 90 days or more. Matrigel was essential for organoid integrity, while alternative matrices such as gelatin and low-melting agarose failed to support proper organoid formation. Subcutaneous preadipocyte medium with 10% FBS from ZenBio was superior to "Advanced/F12K" medium for adipogenic differentiation and long-term maintenance. To model OSA-related hypoxic stress, we exposed organoids to intermittent hypoxia using a programmable hypoxia chamber. IH treatment suppressed adipogenesis, as shown by reduced lipid accumulation, downregulation of adipogenic markers (e.g., PPAR{gamma}, adiponectin, FABP4), and smaller intracellular lipid droplets. Transmission electron microscopy (TEM) revealed IH-induced structural abnormalities, including ER fragmentation, mitochondrial disruption, nuclear enlargement, and heterochromatin formation, all of which are hallmarks of cellular aging. Furthermore, IH upregulated HIF1, H2AX, and aging-associated histone methylation markers (H3K9me3, H3K79me3, H4K20me3), as well as extracellular matrix remodeling proteins such as fibronectin and LOX. Insulin signaling was also impaired, evidenced by decreased phosphorylation of PI3K and AKT. Collectively, these results establish a reliable platform for long-term human adipocyte organoid culture and demonstrate its utility in modeling IH-induced adipocyte dysfunction and aging. This system offers a physiologically relevant tool for mechanistic studies and preclinical therapeutic screening targeting hypoxia-related metabolic disorders.

Explore related subjects

cell biologyPreprint

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Dong, Y., Wang, J., Feng, F., Ali, I. H., Karim, S., Bock, J. M., Somers, V. K.. 2025-12-01. Development of Long-Term Human Adipocyte Organoids Manifesting Aging in Response to Intermittent Hypoxia. https://doi.org/10.1101/2025.11.28.691223

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

Nucleosome Core Allostery Governs Chromatin Recognition and Cell Fate

Nucleosomes regulate chromatin folding, accessibility, and factor recruitment. Current models primarily attribute these functions to histone tail modifications, while the core is largely viewed as a structural scaffold. Yet subtle changes within the nucleosome core can produce profound functional consequences, and the mechanisms underlying these effects remain unclear. Here, we describe nucleosome core allostery as a fundamental principle of chromatin regulation that amplifies the impact of minimal nucleosome variations. Leveraging natural differences between H2A.Z variants, we show that the nucleosome core encodes distinct conformational dynamics that propagate allosterically, thereby controlling nucleosome accessibility and recognition by chromatin factors. As a result, a single buried amino acid substitution alone is sufficient to reprogram nucleosome dynamics and bias cell identity. Our findings establish the nucleosome core as an allosteric regulatory module and provide a generalizable framework for how subtle variation within nucleosomes is amplified into diverse biological outcomes in development and disease.

cell biology↗
bioRxiv2026

SOX4 Reprograms Adipose Stromal Cells into a Cancer-Associated Fibroblast-like State to Drive Metabolic Disease

Pathogenic adipose tissue remodeling promotes metabolic disease in obesity, but the mechanisms that establish this unhealthy tissue state remain poorly understood. Here, we show that obesity drives SOX4-dependent reprogramming of mesenchymal stromal cells (MSCs) into cancer-associated fibroblast-like (CAF-like) cells that promote adipose tissue dysfunction. TGF{beta} signaling is elevated in obesity and activates SOX4 in mouse and human MSCs, inducing their conversion to a CAF-like state. In mice, MSC-specific SOX4 activation induces the CAF-like program and exacerbates adipose tissue inflammation and glucose intolerance, whereas Sox4 deletion attenuates inflammation and improves glucose homeostasis during obesity. We further identify the growth factor Midkine (MDK) as a SOX4-regulated paracrine effector produced by CAF-like cells. MDK inhibition in obese mice reduces adipose tissue inflammation and improves metabolic function. Together, these findings define a TGF{beta}-SOX4-MDK stromal signaling axis that drives pathological adipose tissue remodeling in obesity and highlight this pathway as a potential therapeutic target for improving metabolic health.

cell biology↗
bioRxiv2026

PDLIM5 Modulates YAP1 Localisation and Fibrogenic Gene Expression in Hepatic Stellate Cells

Hepatic stellate cells (HSCs) are the key cellular drivers of liver fibrosis. During liver injury and chronic inflammation HSCs adopt an activated phenotype and secrete fibrotic extracellular matrix (ECM) components such as collagen 1. Mechanical cues derived from the fibrotic ECM drive and support the activation of HSCs, via mechanisms that involve integrins and the mechano-sensitive transcriptional regulator YAP1. It is not yet well understood how external mechanical cues are translated into a molecular response that alters YAP1 nuclear shuttling. There is evidence that suggests the PDZ and LIM domain protein (PDLIM) 5 can regulate YAP1 shuttling in human epithelial cells. We therefore investigated whether PDLIM5 is expressed in HSCs and contributes to YAP1 associated HSC mechano-activation. PDLIM5 protein was detected in HSCs in fibrotic human and mouse liver. PDLIM5 transcript and protein were expressed by primary human and mouse HSCs and by the immortalised HSC LX-2 cell line. PDLIM5 localised with actin stress fibres suggesting a role in HSC adhesion. Co-immunoprecipitation and proximity ligation in LX-2 cells support an association between PDLIM5 and YAP1. We used pharmacological (paclitaxel) and genetic (siRNA and CRISPRi) approaches to inhibit PDLIM5 in HSCs. Inhibiting PDLIM5 reduced YAP1 nuclear localisation and fibrotic gene (COL1A1, ACTA2) expression in LX-2 cells. Overall, these data support a role for PDLIM5 in regulating YAP1 localisation and fibrogenic gene expression in HSCs.

cell biology↗