Background & Aims Interferon-free direct-acting antiviral (DAA) therapies are effective in patients with hepatitis C virus-induced cryoglobulinemia vasculitis (HCV-CV). We analyzed blood samples from patients with HCV-CV before and after DAA therapy to determine mechanisms of these drugs and their effects on cellular immunity. Methods We performed a prospective study of 27 consecutive patients with HCV-CV (median age 59 years) treated with DAA therapy (21 patients received sofosbuvir plus ribavirin for 24 weeks, 4 patients received sofosbuvir plus daclatasvir for 12 weeks, and 2 patients received sofosbuvir plus simeprevir for 12 weeks) in Paris, France. Blood samples were collected from these patients before and after DAA therapy, and also from 12 healthy donors and 12 individuals with HCV infection without CV. HCV load, cryoglobulins, and cytokines were quantified by flow cytometry, cytokine multiplex assays, and ELISAs. Results Twenty-four patients (88.9%) had a complete clinical response of CV to DAA therapy at week 24, defined by improvement of all the affected organs and the absence of relapse. Compared to healthy donors and patients with HCV infection without CV, patients with HCV-CV, before DAA therapy, had a lower percentage of CD4+CD25hiFoxP3+ regulatory T cells (P<.01) but higher proportions of IgM+CD21–/low memory B cells (P<.05), CD4+IFNγ+ cells (P<.01), CD4+IL17A+ cells (P<.01), and CD4+CXCR5+IL21+ follicular T-helper (Tfh) cells (P<.01). In patients with HCV-CV, there was a negative correlation between numbers of IgM+CD21–/low memory B cells and T-regulatory cells (P=.03), and positive correlations with numbers of Tfh cells (P=.03) and serum levels of cryoglobulin (P=.01). DAA therapy increased patients’ numbers of T-regulatory cells (1.5%±0.18% before therapy vs 2.1%±0.18% after therapy), decreased percentages of IgM+CD21–/low memory B cells (35.7%±6.1% before therapy vs 14.9% ± 3.8% after therapy), and decreased numbers of Tfh cells (12%±1.3% before therapy vs 8%±0.9% after therapy). Expression levels of B lymphocyte stimulator receptor 3 and programmed cell death 1 on B cells increased in patients with HCV-CV after DAA-based therapy (mean fluorescence units, 37 ± 2.4 before therapy vs 47 ± 2.6 after therapy; P<.01 and 29 ± 7.3 before therapy vs 48 ± 9.3 after therapy; P<.05, respectively). Conclusions In a prospective clinical trial of patients with HCV-CV, DAA-based therapy restored disturbances in peripheral B- and T-cell homeostasis.