Cysteine proteinase inhibitor in eccrine sweat is derived from sweat gland

Although cysteine proteinases have been reported to be present in human eccrine sweat, their endogenous inhibitors, cysteine proteinase inhibitors (CPIs), have remained unstudied. We now present evidence that CPIs are indeed a true ingredient of human eccrine sweat. Sweat induced in sauna was collected over a Vaseline barrier placed on the skin to minimize epidermal contamination. The absence of major epidermal contamination of the sweat was further ensured by monitoring an epidermal marker, high-molecular-mass aminopeptidase. Sweat CPI was purified sequentially by chromatography with Sephacryl S-200, carboxymethylated papain-Sepharose, and anion-exchange Mono Q fast-protein liquid chromatography columns. Sweat CPI has a molecular mass of approximately 15 kDa, is stable for temperature (up to 80 degrees C) and pH (from 3 to 10), and inhibits papain, ficin, and sweat cathepsin B- and H-like enzymes. Sweat CPI may be of sweat gland origin because 1) the rate of CPI output in sweat (CPI concentration x sweat rate) is constant over 45 min; 2) antibody against epidermal CPI, which cross-reacts with sweat CPI, localized immunoreactivity in the sweat duct; 3) CPI activity was present in the glandular extracts of control and methacholine-stimulated (for 1 h in vitro) human sweat glands; and 4) the peaks of CPI activity in the glandular extract and sweat CPI were both eluted (by high-pressure liquid chromatography) at around 15 kDa. Sweat CPI may be very similar to epidermal CPI (which belongs to the stefin family of CPIs) because of many shared characteristics. The identity and function of sweat CPI remain to be studied.