Optimizing the function and proliferative capacity of stem cells is essential to maximize their therapeutic benefits. High glucose concentrations are known to have detrimental effects on many cell types. We hypothesized that human mesenchymal stem cells (hMSCs) cultured in high glucose-containing media would exhibit diminished proliferation and attenuated production of VEGF, hepatocyte growth factor (HGF), and FGF2 in response to treatment with TNF-alpha, LPS, or hypoxia. hMSCs were plated in medium containing low (5.5 mM) and high (20 mM or 30 mM) glucose concentrations and treated with TNF-alpha, LPS, or hypoxia. Supernatants were collected at 24 and 48 h and assayed via ELISA for VEGF, HGF, and FGF2. In addition, hMSCs were cultured on 96-well plates at the above glucose concentrations, and proliferation at 48 h was determined via bromo-2'-deoxy-uridine (BrdU) incorporation. At 24 and 48 h, TNF-alpha, LPS, and hypoxia-treated hMSCs produced significantly higher VEGF, HGF, and FGF2 compared with control. Hypoxia-induced VEGF production by hMSCs was the most pronounced change over baseline. At both 24 and 48 h, glucose concentration did not affect production of VEGF, HGF, or FGF2 by untreated hMSCs and those treated with TNF-alpha, LPS, or hypoxia. Proliferation of hMSCs as determined via BrdU incorporation was unaffected by glucose concentration of the media. Contrary to what has been observed with other cells, hMSCs may be resistant to the short-term effects of high glucose. Ongoing efforts to characterize and optimize ex vivo and in vivo conditions are critical if the therapeutic benefits of MSCs are to be maximized.