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2 protocols using nafamostat mesylate

1

Apoptosis Signaling Pathway Inhibitors

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Staurosporine, etoposide, phenylmethanesulfonyl fluoride (PMSF), calpain inhibitor, tumor necrosis factor (TNF)-α, cycloheximide, and enterokinase were purchased from Sigma-Aldrich (St. Louis, MO, USA). Z-VAD-FMK, Z-DEVD-FMK, Z-VEID-FMK, Z-VDVAD-FMK, Z-IETD-FMK, and Z-LEHD-FMK were purchased from R&D systems (Minneapolis, MN, USA). Cathepsin inhibitor I and Omi/HtrA2 inhibitor were purchased from Millipore (Billerica, MA, USA). 3,4-Dichloroisocoumarin and nafamostat mesylate (NFM) were purchased from Santa Cruz (Santa Cruz, CA, USA). Bio-Gel HTP hydroxyapatite was purchased from Bio-Rad (Hercules, CA, USA). Anti-histone H4 and histone H4 modification antibodies were purchased from Abcam (Cambridge, MA, USA). Caspase-3, -8, -9, and poly ADP-ribose polymerase (PARP) antibodies were purchased from Cell Signaling (Danvers, MA, USA). Anti-glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was purchased from Santa Cruz.
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2

Protease Inhibitor Characterization

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Recombinant human prostasin, matriptase and HAT, expressed as soluble catalytic domains, were obtained from R&D Systems (Minneapolis, MN). Human lung (β) tryptase was from EMD-Millipore (Billerica, MA). The tryptic peptidase substrates t-butyloxycarbonyl-L-Gln-L-Ala-L-Arg-4-nitroanilide (QAR) and N-(p-tosyl)-Gly-L-Pro-L-Lys-4-nitroanilide (GPK) were from Bachem Americas (Torrance, CA) and Sigma-Aldrich (St. Louis, MO), respectively. Protease inhibitors camostat mesylate and nafamostat mesylate were from Santa Cruz Biotechnology (Dallas, TX), as was nafamostat’s cleavage product 6-amidino-2-naphthol. Benzamidine, aprotinin, p-nitrophenyl-p’-guanidinobenzoate and bovine trypsin were from Sigma-Aldrich. BABIM was provided by Dr. Richard Tidwell as described previously [34 (link)]. Structures of chemical inhibitors used in this study are given in Fig 1.
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