The mechanisms that underlie valvular inflammation in streptococci-induced infective endocarditis (IE) remain unclear. We previously demonstrated that streptococcal glucosyltransferases (GTFs) can activate human heart valvular interstitial cells (VICs) to secrete IL-6, a cytokine involved in T helper (Th) 17 cell differentiation. Here, we tested the hypothesis that activated VICs can enhance neutrophil infiltration through sustained IL-17 production, leading to valvular damage. To monitor cytokine and chemokine production, leukocyte recruitment, and the induction or expansion of CD4+CD45RA–CD25–CCR6+ Th17 cells, primary human VICs were cultured in vitro and activated by GTFs. Serum cytokine levels were measured using ELISA, and neutrophils and Th17 cells were detected by immunohistochemistry in infected valves from patients with IE. The expression of IL-21, IL-23, IL-17, and retinoic acid receptor-related orphan receptor C (Rorc) were up-regulated in GTF-activated VICs, which may enhance the proliferation of memory Th17 cells in an IL-6 dependent manner. Many chemokines, including chemokine (C-X-C motif) ligand 1 (CXCL1), were up-regulated in GTF-activated VICs, which might recruit neutrophils and CD4+ T cells. Moreover, CXCL1 production in VICs was induced dose-dependently by IL-17 to enhance neutrophil chemotaxis. CXCL1-expressing VICs and infiltrating neutrophils could be detected in infected valves, and serum concentrations of IL-17, IL-21, and IL-23 were increased in patients with IE compared to healthy donors. Furthermore, elevated serum IL-21 levels have been significantly associated with severe valvular damage, including rupture of chordate tendineae, in IE patients. Our findings suggest that VICs are activated by bacterial modulins to recruit neutrophils, and such activities might be further enhanced by the production of Th17-associated cytokines. Together, these factors can amplify the release of neutrophilic contents in situ, which might lead to severe valvular damage.