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The prevalence of obesity, characterized by an excessive accumulation of adipose tissue and adipocyte hypertrophy, presents a major public health challenge. This study investigates the therapeutic potential of two probiotic strains, <i>Lactobacillus sakei</i> Probio65 and <i>Lactobacillus plantarum</i> Probio-093, in the context of obesity. Utilizing 3T3-L1 cell-derived human adipocytes, we assessed Probio65’s and Probio-093’s capacity to mitigate triglyceride accumulation and influence adipocytokine production in vitro. Subsequently, an in vivo trial with male C57BL/6J mice examined the effects of both probiotic strains on adipose tissue characteristics, body weight, fat mass, and obesity-related gene expression. This study employed both live and ethanol-extracted bacterial cells. The results demonstrated significant reductions in the triglyceride deposition, body weight, and adipose tissue mass in the treated groups (<i>p</i> < 0.05). Furthermore, both strains modulated adipokine profiles by downregulating proinflammatory markers such as PAI-1, leptin, TNF-α, STAMP2, F4/80, resistin, and MCP-1, and upregulating the insulin-sensitive transporter GLUT4 and the anti-inflammatory adiponectin (<i>p</i> < 0.05). Our findings suggest that <i>Lactobacillus sakei</i> Probio65 and <i>Lactobacillus plantarum</i> Probio-093 are promising agents for microbiome-targeted anti-obesity therapies, offering the effective mitigation of obesity and improvement in adipocyte function in a murine model.