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Background: Several studies reveal an inverse relation between serum 25-hydroxyvitamin D [25(OH)D] concentration and the risk of acute ischemic stroke (AIS). The aim of this study was to determine relationships between 25(OH)D concentration and the course and outcomes of AIS treatment and the level of indices of inflammatory response to brain injury. Patients and Methods: Retrospective analysis of medical documentation of 1381 real-world AIS patients hospitalized in a single center between 1 January 2020 and 31 May 2025. Serum 25(OH)D level, several inflammatory indices, and clinical data were assessed. Results: Compared to patients in the lowest quartile of 25(OH)D concentration, those in the highest quartile had a shorter length of in-hospital stay, a lower risk of all-cause death, and a lower score for disability on a modified Rankin scale (mRS). Along with an increase in 25(OH)D quartiles, we found: a decrease in neutrophil count; a decrease in glucose, HbA1c, albumin, C-reactive protein (CRP), and CRP-to-albumin, -lymphocyte, -neutrophil, and -platelet ratios; lower neutrophil-to-lymphocyte and -albumin ratios, and lower systemic immune inflammation, and systemic inflammation response indices. In multifactorial logistic regression, the quartile of 25(OH)D (OR, 95% CI: 1.52, 1.09-2.12; p = 0.012) was the only variable to have a positive association with a mRS score ≤ 2 at discharge from hospital, and neutrophil-to-lymphocyte ratio, age, diabetes, and treatment with endovascular mechanical thrombectomy were biomarkers of poor functional status at discharge. Conclusions: Higher 25(OH)D concentration in AIS patients is related to better survival and a lower level of inflammatory response indices and disability at discharge.