Objective: This study investigates the role of fluid shear stress, induced by lymph flow within lymphatic vessels, in regulating mitochondrial fusion in lymphatic endothelial cells. We hypothesize laminar shear stress promotes mitochondrial fusion through the regulation of mitochondrial fusion-related genes, with corresponding morphological changes reflecting fusion events. Methods: Human Dermal Lymphatic Endothelial Cells were cultured in MV2 medium on μ-slides. To apply different types of fluid shear stress, laminar shear stress was generated under unidirectional flow (UF), and oscillatory shear stress under disturbed flow (DF), using the ibidi pump system (laminar shear stress: 4 dyn/cm 2 ; oscillatory shear stress: 4 dyn/cm 2 , 0.25 Hz). After 24 hours of shear stress application, RNA was extracted, and real-time PCR was performed to evaluate the expression of key mitochondrial fusion-related genes MFN1, MFN2, and OPA1. In a separate set of experiments, cells were stained with 100 nM MitoTracker Green FM in MV2 medium for 30 minutes following shear stress. Mitochondrial morphology was evaluated by mitochondrial fluorescence staining. Cells were then visualized under a fluorescence microscope in live cell imaging solution. Fluorescence images were analyzed using ImageJ to quantify mitochondrial form factor, aspect ratio, number, size, and total area. Results: Mitochondrial fusion-related gene expression tended to be increased in UF, with only MFN1 significantly increased (MFN1: ~2.1-fold increase vs DF, p< 0.05). Under UF, cells exhibited an elongated morphology, whereas cell shape was irregular under DF. Both form factor and aspect ratio were elevated under UF compared to DF, with only aspect ratio showing a statistically significant difference (aspect ratio: UF 4.00±0.10 vs DF 3.45±0.07, p< 0.05; mean ± SEM). The total mitochondrial area was significantly higher under UF (mitochondrial area: UF 2776±183 vs DF 2227±92, p< 0.05; mean ± SEM), while mitochondrial size and number did not differ significantly between the two shear stress conditions. Conclusions: In this study, differential expression of mitochondrial fusion–related genes was observed between UF and DF, with only MFN1 showing a significant change. Although gene expression alone is insufficient to conclude increased mitochondrial fusion, morphological changes under different flow conditions indicate lymphatic endothelial cells respond to flow patterns. In UF, increased aspect ratio and total mitochondrial area, despite similar mitochondrial numbers, suggest fusion may contribute to these morphological adaptations. Further investigation of upstream signaling genes or post-translational modifications of fusion proteins will clarify molecular mechanisms underlying shear stress–induced mitochondrial fusion. Funding sources: N.A. This abstract was presented at the American Physiology Summit 2026 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.
Jeom et al. (Fri,) studied this question.