# Extracellular Vesicles in Osteogenesis: Comparative Analysis of Stem Cell Sources, Conditioning Strategies, and In Vitro Models Toward Advanced Bone Regeneration

## TL;DR

This paper reviews how extracellular vesicles from different stem cells can be used to promote bone regeneration, focusing on sources, conditioning methods, and in vitro models.

## Contribution

The paper uniquely integrates a comparative analysis of extracellular vesicles from multiple stem cell sources with engineering strategies and clinical perspectives.

## Key findings

- Different stem cell sources produce extracellular vesicles with distinct therapeutic potentials for bone regeneration.
- Conditioning strategies like osteogenic induction and hypoxia modulate EV cargo and enhance their regenerative effects.
- Advanced in vitro models, including 3D spheroids and organoids, are critical for evaluating EV-mediated bone regeneration.

## Abstract

Extracellular vesicles (EVs) derived from stem cells have emerged as promising mediators of osteogenesis, suggesting cell-free alternatives for bone tissue engineering and regenerative medicine. This review provides a comprehensive analysis of the main stem cell sources used for EV production, including bone marrow mesenchymal stem cells (BM-MSCs), adipose-derived stem cells (ADSCs), umbilical cord MSCs (UC-MSCs), induced pluripotent stem cells (iPSCs), and alternative stromal populations. Particular attention is given to the ways in which different conditioning and differentiation strategies, such as osteogenic induction, hypoxia, and mechanical stimulation, modulate EV cargo composition and enhance their therapeutic potential. We further discuss the in vitro models employed to evaluate EV-mediated bone regeneration, ranging from 2D cultures to complex 3D spheroids, scaffold-based systems, and bone organoids. Overall, this review emphasizes the current challenges related to standardization, scalable production, and clinical translation. It also outlines future directions, including bioengineering approaches, advanced preclinical models, and the integration of multi-omics approaches and artificial intelligence to optimize EV-based therapies. By integrating current knowledge, this work aims to guide researchers toward more consistent and physiologically relevant strategies to harness EVs for effective bone regeneration. Finally, this work uniquely integrates a comparative analysis of EVs from multiple stem cell sources with engineering strategies and emerging clinical perspectives, thereby providing an updated and translational framework for their application in bone regeneration.