title: Activation of prodeath Bcl-2 family proteins and mitochondrial apoptosis pathway by sanguinarine in immortalized human HaCaT keratinocytes.
abstract: Sanguinarine, derived from the root of Sanguinaria canadensis and other poppy fumaria species, possesses strong antimicrobial, anti-inflammatory, and antioxidant properties. We earlier showed that sanguinarine kills human epidermoid carcinoma A431 cells via an induction of apoptosis [N. Ahmad et al., Clin. Cancer Res., 6: 1524-1528, 2000]. In this study, using immortalized human keratinocytes (HaCaT cells), we provide information about mechanism of the antiproliferative effect of sanguinarine. Sanguinarine [0.1 (M-2 (M)] treatment to HaCaT cells was found to inhibit in a dose-dependent manner the cell proliferation and induce apoptosis, as measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and ELISA, respectively. Sanguinarine treatment also resulted in a significant cleavage of poly(ADP-ribose) polymerase in HaCaT cells. Because mitochondrial pathway is critical for the regulation of apoptosis, we studied the involvement and regulation of mitochondrial events in sanguinarine-mediated apoptosis of HaCaT cells. As shown by the immunoblot analysis, our data clearly demonstrated that sanguinarine treatment to HaCaT cells resulted in a dose-dependent (a) increase in the level of Bax with a concomitant decrease in Bcl-2 levels and (b) increase in Bax/Bcl-2 ratio. Sanguinarine also resulted in significant increases in the proapoptotic members of Bcl-2 family proteins, i.e., Bak and Bid. This was accompanied by increase in (a) protein expression of cytochrome c and apoptotic protease-activating factor-1 and (b) activity and protein expression of caspase-3, caspase-7, caspase-8, and caspase-9. Taken together, our data showed the involvement of mitochondrial pathway and Bcl-2 family proteins during sanguinarine-mediated apoptosis of immortalized keratinocytes. We suggest that sanguinarine could be developed as a drug for the management of hyperproliferative skin disorders, including skin cancer.
authors.affiliation: Department of Dermatology, University of Wisconsin, Madison, Wisconsin 53706, USA.
authors.forename: Vaqar Mustafa
authors.initials: VM
authors.lastname: Adhami
authors.forename: Moammir Hasan
authors.initials: MH
authors.lastname: Aziz
authors.forename: Hasan
authors.initials: H
authors.lastname: Mukhtar
authors.forename: Nihal
authors.initials: N
authors.lastname: Ahmad
mesh_terms: D000470:Alkaloids / Q000627:therapeutic use*; D000975:Antioxidants / Q000494:pharmacology; D017209:Apoptosis; D053119:Benzophenanthridines; D053148:Caspase 3; D053179:Caspase 7; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases / Q000096:biosynthesis; D002455:Cell Division; D045744:Cell Line, Tumor; D002470:Cell Survival; D045304:Cytochromes c / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000627:therapeutic use*; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D015151:Immunoblotting; D007546:Isoquinolines; D015603:Keratinocytes / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism; D008954:Models, Biological; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism*; D051028:bcl-2-Associated X Protein
chemical_list: D000470:Alkaloids; D000975:Antioxidants; C494826:BAX protein, human; D053119:Benzophenanthridines; D004791:Enzyme Inhibitors; D007546:Isoquinolines; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D045304:Cytochromes c; C005705:sanguinarine; D011065:Poly(ADP-ribose) Polymerases; C505407:CASP3 protein, human; C505520:CASP7 protein, human; C505530:CASP8 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053179:Caspase 7; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases