Skeletal muscle is an important site for nutrient metabolism. The capillaries in the skeletal muscle and the blood flowing through these capillaries, known as muscle microvascular blood flow (MBF), enhance glucose delivery and disposal in the muscle in response to insulin infusion (1) or ingestion of a mixed nutrient meal (2). Polyunsaturated fatty acids (PUFA), particularly omega-3 (n-3) PUFA, have demonstrated a variety of cardiometabolic benefits, including inhibiting inflammation, improving lipid profiles, endothelial function, insulin sensitivity, and reducing the risk of metabolic and chronic diseases (3). However, the impact of PUFA on muscle microvascular health and function remains unknown. The aim of this systematic review is to determine the effects of PUFA interventions on muscle microvascular health and function in animals and humans. A systematic search of Medline Complete, PubMed, Embase, and Cochrane Library was conducted. Studies were considered eligible if they examined the effect of PUFA interventions on microvascular measures including capillary density or MBF in skeletal muscle of animals or humans. Quality assessment was conducted through the Systematic Review Centre for Laboratory Animal Experimentation (SYRCLE's) risk of bias tool for animal studies and the revised Cochrane risk of bias tool for randomized trials (RoB) for human studies. Twelve eligible studies were included in this systematic review, out of which nine involved animal models, and three studies were conducted in humans. Four animal studies showed that PUFA increased capillary density in skeletal muscle in various models. Out of these four studies, three studies used n-3 PUFA-rich fish oil with doses ranging from 5% to 20% weight (w)/w of diet and 0.64 g/100 g of body weight (BW)/day and duration of 14 to 28 days and one study used n-6 PUFA-rich primrose oil (10% w/w of diet for 2 months). In terms of muscle MBF, three animal studies showed n-3 PUFA-rich fish oil (5% to 14.5% w/w of diet and 0.64 g/100 g of BW/day and duration of 14 days to 6 weeks) prevented the impairment in muscle MBF caused by various experimental conditions such as ischemia, endotoxin infusion or during vasoactive agent infusion. None of the animal studies using n-6 PUFA oils showed an effect on muscle MBF. In humans, neither n-3 nor n-6 PUFA affected muscle MBF. While there seems to be a wide range of intervention dose and duration used in the animal studies, they do suggest n-3 PUFA may be beneficial for microvascular health and function; however, further investigation in humans is warranted.