HPLC‐ESI‐MS analysis of oral human fluids reveals that gingival crevicular fluid is the main source of oral thymosins β4 and β10

Thymosin β₄ (Tβ₄), its sulfoxide, and thymosin β₁₀ (Tβ₁₀) were detected in human saliva and identified by different strategies based on RP HPLC coupled to electrospray multidimensional IT MS. Tβ₄ was almost always detected in whole saliva, its sulfoxide sporadically, Tβ₁₀ rarely. Tβ₄ was undetectable in parotid saliva and less concentrated in submandibular/sublingual saliva than in whole saliva. Analysis of gingival crevicular fluid revealed high relative amounts of Tβ₄, Tβ₄ sulfoxide, and Tβ₁₀ in all the samples. Tβ₄ mean concentration was 200 times higher in crevicular fluid (20 μmol/L, N = 9) than in whole saliva (0.1 μmol/L, N = 9). Crevicular fluid concentration of Tβ₄ (ca. 5% represented by its sulfoxide) and β₁₀ significantly correlated (r = 0.856; N = 9), and their ratio was about 5. A significant correlation was also observed between Tβ₄ concentrations in whole saliva and gingival crevicular fluid (r = 0.738; N = 9). Immunohistochemical analysis of the major salivary glands showed that immunoreactivity for Tβ₄ is restricted to ductal cells, with minor degree of focal positivity in some acinar cells. On the whole, results indicate that gingival sulcus is a main, although not the sole, source for oral Tβ₄ and Tβ₁₀.