Background: A current, albeit unproven, hypothesis is that an acceleration of cellular senescence is involved in impaired renal repair and progression of glomerular diseases. Focal segmental glomerulosclerosis (FSGS) is a glomerular disease with a substantial risk for progression to ESRD. However, if and to what extent cell senescence predicts a negative outcome in FSGS is still unknown. Methods: The hypothesis that cell senescence represents a proximate mechanism by which the kidney is damaged in FSGS (NOS phenotype) was investigated in 26 consecutive kidney biopsies from adult FSGS cases (eGFR 72 ± 4 mL/min, proteinuria 2.3 ± 0.6 g/day) who were incident for 2 years in a Northern Italian nephrology center and had a 6-year clinical follow-up. Results: Cell senescence (p16<sup>INK4A</sup>, SA-β-galactosidase [SA-β-Gal]) was upregulated by ∼3- to 4-fold in both glomerular and tubular cells in kidney biopsies of FSGS as compared to age-matched controls (p < 0.05–0.01). Tubular SA-β-Gal correlated with proteinuria and glomerulosclerosis, while only as a trend, tubular p16<sup>INK4A</sup> was directly associated with interstitial fibrosis. At univariate analysis, basal eGFR, proteinuria, and tubular expression of SA-β-Gal and p16<sup>INK4A</sup> were significantly directly related to the annual loss of eGFR. No correlation was observed between glomerular p16<sup>INK4A</sup> and eGFR loss. However, at multivariate analysis, eGFR, proteinuria, and tubular p16<sup>INK4A</sup>, but not SA-β-Gal, contributed significantly to the prediction of eGFR loss. Conclusions: The results indicate that an elevated cell senescence rate, expressed by an upregulation of p16<sup>INK4A</sup> in tubules at the time of initial biopsy, represents an independent predictor of progression to ESRD in adult patients with FSGS.