Polyalthia Longifolia Induced Apoptosis via miR-484 Downregulation: A Multimodal In Situ Microscopy, In Vitro, and In Vivo Investigation.

Polyalthia longifolia is a promising botanical source of anticancer compounds; yet, the microstructural mechanisms underlying its activity remain insufficiently defined. This study examines the role of miR-484 in P. longifolia methanolic leaf extract (PLME)-induced apoptosis in HeLa cells using a multimodal framework integrating in situ microscopy, in vitro assays, and in vivo validation. In in vitro study, PLME markedly reduced miR-484 expression, corresponding with enhanced apoptotic morphology visualized through in situ microscopic techniques, including scanning electron microscopy (SEM). These structural alterations coincided with reduced cell viability in MTT assays (IC50 value = 21.41 μg/mL) and increased active caspase-3 activity. miR-484 overexpression attenuated PLME-induced apoptosis, showing increased membrane integrity and proliferation-associated morphology on SEM. In vivo xenograft models supported these findings: miR-484 overexpression promoted tumor growth, whereas PLME induced significant tumor suppression. PLME alone reduced tumor weight (0.45 ± 0.08 g) compared with PLME + miR-484 mimic (0.92 ± 0.18 g) and 5% DMSO (1.74 ± 0.18 g). Histopathology revealed apoptotic signatures consistent with miR-484 downregulation, and qRT-PCR confirmed the inverse relationship between PLME exposure and miR-484 levels across models (PLME reduced miR-484 expression by 26%, versus 43% with PLME + miR-484 mimic). Collectively, these results show that PLME induces apoptosis through miR-484 downregulation, with multimodal microscopy providing critical in situ evidence of structural and biological changes. This study underscores the value of integrated microscopy-driven analysis in natural product-based anticancer research.