Adiponectin reduces the stimulatory effect of leptin on clonogenicity, anchorage-independent growth migration and invasion potential of breast carcinoma cells. (A) Breast cancer cells (MCF7 and MDA-MB-231) were treated with 100 ng/ml leptin (L) and 10 µg/ml adiponectin (Adn) alone and in combination (L + Adn) and subjected to clonogenicity assay. Colonies containing >50 normal-appearing cells were counted. Adiponectin inhibited leptin-induced clonogenicity. (B) Breast cancer cells were subjected to soft-agar colony formation assay in the presence of leptin (L) and/or adiponectin (Adn) as in A for 3 weeks. Results are expressed as average number of colonies counted (in six microfields). *P < .005, compared with untreated controls; **P < .001, compared with untreated controls; #P < .005, compared with L treatment. Adiponectin inhibited leptin-induced anchorage-independent growth. (C) Breast cancer (MCF7, T47D, MDA-MB-231, and MDA-MB-468) cells were subjected to scratch migration assay in the presence of leptin and adiponectin treatments as described in A. The histogram shows the fold change in migration. *P < .01 and **P < .005, compared to untreated controls; #P < 0.01, compared to leptin (L)-treated cells. Adiponectin inhibited migration of breast cancer cells even in the presence of leptin. (D) MCF7 and MDA-MB-231 cells were cultured in Matrigel invasion chambers followed by treatment as in A for 24 hours. The number of cells that invaded through the matrigel was counted in five different regions. The slides were blinded to remove counting bias. The result shows mean of three independent experiments performed in triplicates. *P < .005, compared with untreated controls; #P < .001, compared to leptin-treated cells. Adiponectin treatment significantly reduced leptin-induced matrigel invasion.