Background: Adipose tissue-derived mesenchymal stem cells (AD-MSCs) were proved to differentiate into insulin-producing cells (IPCs), but the amount of insulin secreted was relatively low compared to the insulin secreted by mature pancreatic islets. Enrichment of MSCs culture with melatonin (MT) was detected to promote cartilage matrix synthesis, osteogenic and neuronal differentiation. Therefore, the present study was conducted to evaluate the potential role of MT pre-treated AD-MSCs in enhancing the treatment and regeneration of the islet cells of Langerhans in diabetic rats induced by streptozotocin (STZ). Materials and Methods: Forty adult male Sprague Dawley albino rats were divided equally into groups; group I (control group), group II (STZ group), group III (STZ+AD-MSCs) and group IV (STZ+MT pre-treated AD-MSCs). Biochemical studies were implemented including measurements of the body weight, fasting blood glucose and serum insulin levels, Interleukin 17 (IL-17) and IL-10. Samples of the pancreas were taken and prepared for light, fluorescent microscopic examination, PCNA and caspase-3 immunohistochemical studies and histomorphometric analysis.  Results: The present study confirmed the regenerative and therapeutic effects of AD-MSCs on the pancreatic cells. Concomitant supply of MT to the culture of AD-MSCs, in group IV, was shown to retain the normal architecture of the islet cells of Langerhans. They appeared well-defined and lightly stained, surrounded by classical pancreatic acini and contained a large number of islet cells with vesicular nuclei and prominent nucleoli. Improvements of all the biochemical parameters, in the same group, was demonstrated by increased body weight and serum insulin levels with a decrease in the fasting blood glucose levels. Significant decrease in the pro-inflammatory cytokine; IL-17 and increase in the anti-inflammatory cytokine; IL-10, compared to the STZ group, were also discovered.  Significant increase in the PCNA proliferation index, decrease in caspase-3 and increase in PKH26 labeled MSCs area percent was recorded in the group of AD-MSCs enriched with MT compared to the group of AD-MSCs without MT.   Conclusions: The present study confirmed the potential therapeutic and protective role of MT pre-treated AD-MSCs against the STZ-induced pancreatic islet cells’ damage. Further studies were recommended to investigate the efficacy of MT and AD-MSCs over longer experimental durations.