TNF-α is a pro-inflammatory cytokine in insulin resistance. Cytokine-induced increase in TNF-α production by the iNOS pathway plays an important role in diabetes. This study was designed to determine the effect of cow's milk kefir as an antidiabetic and expression of the iNOS gene. Twenty-five Wistar rats were divided into 5 groups. Group cow’s milk kefir dose variation (50%, 75%, and 100%). This study showed that TNF-α levels in the DM control rat group were higher compared to other DM rat groups that were given cow’s milk kefir treatment. The cow's milk kefir treatment group (50%) had a higher average TNF-α content (146.3± 6.68) compared to the other two treatment groups given 75% cow's milk kefir (169.9± 6.39) and 100% cow's milk kefir (175.3± 4.66). There were differences in TNF-α levels between treatment groups (p=0.0013). The expression of the iNOS gene in the healthy control group was highest found in the M rat groupgiven 50% milk kefir and the lowest in the DM rat group given 100% milk kefir. There weresignificant differences in iNOS gene expression between groups (p = 0.0019). There was a significant difference between the DM rat group that was treated in the form of 50% group with DM Rats (positive controls) (p= 0.0048) and DM cow’s milk Kefir rats 75% milk with DM Rats (Positive controls) (p=0.0128). This study shows that there is an effect of cow's milk kefir as an antidiabetic on TNF-α and iNOS gene expression.