Polymorphisms in genes coding for pro-inflammatory molecules represent important factors for the pathogenesis and outcome of stroke. The aim of this study was to evaluate the relationship between the tumor necrosis factor beta (TNF-β) NcoI (rs909253) polymorphism with inflammatory and metabolic markers in acute ischemic stroke. Ninety-three patients and 134 controls were included. The TNF-β polymorphism was determined using PCR-RFLP with NcoI restriction enzyme. Stroke subtypes and neurological deficit score were evaluated. White blood cell counts, erythrocyte sedimentation rate (ESR), plasma levels of IL-6 and TNF-α, serum high sensitivity C-reactive Protein (hsCRP), serum lipid profile, plasma levels of glucose and insulin, and homeostatic model assessment of insulin resistance (HOMA-IR) were determined. Stroke patients presented higher white blood cell counts, hsCRP, ESR, glucose, insulin, and HOMA-IR, and lower HDL cholesterol than controls (p < 0.01). There was no difference in genotypic and allelic frequency of TNF-β NcoI polymorphism among patients and controls (p > 0.05). However, stroke patients carrying the TNFB2/B2 genotype presented higher levels of TNF-α, white blood cell counts, total cholesterol, LDL cholesterol, glucose, insulin, and HOMA-IR than those with other genotypes (p < 0.05). White blood cells, IL-6, hsCRP, and ESR were positively correlated with the neurological deficit of the patients (p < 0.05). Taken together, TNF-β NcoI polymorphism, by itself, was not associated with increased susceptibility for stroke development. However, the homozygous genotype for the allele TNFB2 was associated with higher expression of classical inflammatory and metabolic markers of development and outcome of stroke than other genotypes. The identification of variant alleles might allow both better prediction of susceptibility for stroke as well the identification of novel stroke mechanisms that could be target to new therapeutic approaches. Stroke patients carrying the TNFB2 variant allele could have a beneficial effect with the anti-inflammatory therapies in the early inflammatory phase of stroke.