Low levels of high-density cholesterol (HDLc) accompany chronic kidney disease, but the association between HDLc and the estimated glomerular filtration rate (eGFR) in the general population is unclear. We investigated the HDLc-eGFR association in nondiabetic Han Chinese (HC, n = 1100), West Africans (WA, n = 1497), and African Americans (AA, n = 1539). There were significant differences by ancestry: HDLc was positively associated with eGFR in HC (β = 0.13, P < 0.0001), but negatively associated among African ancestry populations (WA: -0.19, P < 0.0001; AA: -0.09, P = 0.02). These differences were also seen in nationally-representative NHANES data (among European Americans: 0.09, P = 0.005; among African Americans -0.14, P = 0.03). To further explore the findings in African ancestry populations, we investigated the role of an African ancestry-specific nephropathy risk variant, rs73885319, in the gene encoding HDL-associated APOL1. Among AA, an inverse HDLc-eGFR association was observed only with the risk genotype (-0.38 versus 0.001; P = 0.03). This interaction was not seen in WA. In summary, counter to expectation, an inverse HDLc-eGFR association was observed among those of African ancestry. Given the APOL1 × HDLc interaction among AA, genetic factors may contribute to this paradoxical association. Notably, these findings suggest that the unexplained mechanism by which APOL1 affects kidney-disease risk may involve HDLc.