Terpenoids and Phenylpropanoids in Ligularia duciformis, L. kongkalingensis, L. nelumbifolia, and L. limprichtii.

The diversity in root chemicals and evolutionally neutral DNA regions in the complex of Ligularia duciformis, L. kongkalingensis, and L. nelumbifolia (the d/k/n complex) was studied using eight samples collected in central and northern Sichuan Province of China. Cacalol (14) and epicacalone (15), rearranged eremophilanes, were isolated from the complex for the first time. Two new phenylpropanoids were also obtained. Seven of the eight samples produced phenylpropanoids and the other produced lupeol alone. Two of the seven samples also produced furanoeremophilanes or their derivatives and one produced oplopanes. The geographical distribution of the sesquiterpene-producing populations suggests that the production of sesquiterpenes evolved independently in separate regions. L. limprichtii collected in northern Sichuan was also analyzed and its chemical composition and the sequence of internal transcribed spacers (ITSs) in the ribosomal RNA gene cluster were found to be similar to that in the d/k/n complex and L. yunnanensis, which are morphologically similar.

The photocarcinogenesis of antibiotic lomefloxacin and UVA radiation is enhanced in xeroderma pigmentosum group A gene-deficient mice.

Lomefloxacin (LFLX) is phototoxic and phototumorigenic, but the mechanisms of phototumorigenesis of quinolone drugs have not been fully elucidated. Formation of cyclobutane pyrimidine dimers (CPD) by UVB radiation is primarily involved in the carcinogenesis of ultraviolet (UV) radiation. On the other hand, UVA region is responsible to photobiologic reactions of quinolone drugs. To know if CPD can be formed by UVA radiation in the presence of LFLX and is involved in the phototumorigenesis, we used xeroderma pigmentosum (XP) group A gene-deficient (XPA-/-) mouse, which is defective in nucleotide excision repair. XPA-/- and XPA+/+ mice were irradiated to 5 J per cm2-UVA with or without the administration of LFLX. In XPA-/- mice treated with LFLX, the first skin tumor appeared after exposures to 75 J per cm2 in 5 wk. In XPA+/+ mice treated with LFLX, the first tumor appeared after exposures to 345 J per cm2 in 23 wk. Immunohistochemically, CPD formation was observed after UVA-exposure in the skin of XPA+/+ as well as XPA-/- mice which had been given LFLX. The CPD disappeared, however, earlier from XPA+/+ mice than from XPA-/- mice. The acute inflammatory reaction after LFLX administration and exposure to UVA were greatly enhanced in XPA-/- mice. These results indicate that UVA exposure induces DNA damage in the form of CPD in the presence of LFLX, which exerts phototoxicity and phototumorigenesis.

Effects of calcium concentration on the SOD activity and UVB-induced cytotoxicity in cultured human keratinocytes.

BACKGROUND/PURPOSE: Cellular differentiation due to the extracellular calcium (Ca(2+)) concentration affects the level of several antioxidant enzymes in cultured human keratinocytes. Because the epidermis includes well- and un-differentiated keratinocytes, we expected that keratinocytes possess different antioxidant capacity and sensitivity to damaging effects of ultraviolet-B (UVB) depending on the differentiation. We examined the effects of Ca(2+) concentration of culture medium (DMEM (Dulbecco's modified Eagle's medium)) on the superoxide dismutase (SOD) activity and UVB-induced cytotoxicity in cultured human keratinocytes in order to investigate the relationship between cell differentiation and antioxidant defense. METHODS: Human keratinocytes (HaCaT cells) were incubated in high Ca(2+) (>1 mM) or low Ca(2+) (<0.1 mM) concentration DMEM for 24 h at 37 degrees C in 5% CO(2). Then, we measured total SOD activity and also individual Cu,Zn- and Mn-SOD activities in keratinocytes. Furthermore, after incubation in high or low Ca(2+) concentration DMEM, human keratinocytes were irradiated with 10, 20 or 30 mJ/cm(2) UVB. The quantity of lactate dehydrogenase (LDH) leaked in the supernatant from damaged keratinocytes, cell viability and TdT-mediated dUTP nick end labelings (TUNEL) positive keratinocytes were measured at 24 h after UVB irradiation. RESULTS: Total SOD activity and Cu,Zn-SOD activity in human keratinocytes cultured in low Ca(2+) were significantly lower than in keratinocytes cultured in high Ca(2+) concentration DMEM. In contrast, Mn-SOD activity was not affected. LDH leakage in the supernatant from keratinocytes cultured in low Ca(2+) concentration was significantly higher than that from keratinocytes cultured in high Ca(2+) concentration DMEM after UVB irradiation. The cell viability of keratinocytes cultured in low Ca(2+) concentration DMEM was significantly decreased compared to that of keratinocytes cultured in high Ca(2+) concentration DMEM after UVB irradiation. Furthermore, UVB-induced apoptosis was increased in keratinocytes cultured in low Ca(2+) concentration DMEM by the TUNEL method. CONCLUSIONS: These results suggest that cellular differentiation due to the change of Ca(2+) concentration of culture medium affects the Cu,Zn-SOD activity and UVB-induced cytotoxicity in cultured human keratinocytes.

XPA gene-deficient, SCF-transgenic mice with epidermal melanin are resistant to UV-induced carcinogenesis.

Photobiologic investigations have been performed using animals without epidermal melanocytes. We developed xeroderma pigmentosum group A gene-deficient (XPA (-/-)), stem cell factor transgenic (SCF-Tg) mice, which one defective in nucleotide excision repair and have epidermal melanocytes, and investigated protective effects of epidermal melanin against UV-induced injuries. When irradiated to UVB, XPA (-/-) mice developed greatly enhanced responses including acute inflammation, cyclobutane pyrimidine dimer (CPD) formation, keratinocyte apoptosis, depletion of Langerhans cells and immunosuppression of contact hypersensitivity, but XPA (-/-), SCF-Tg mice showed much less responses to the same dose of UVB. XPA (-/-), SCF-Tg mice did not develop skin cancers after repeated exposures to UVB for 30 wk at a total dose of 72 J per cm(2), which induced a significant number of tumors even in wild-type, XPA (+/+) mice, and was lethal dose for XPA (-/-) mice. Dimethylbenz (alpha) anthracence (DMBA) induces DNA damages, which require XPA protein to be repaired. Topical application of DMBA produced a significant inflammation, CPD formation, apoptosis, immunosuppression, and skin cancers in XPA (-/-), SCF-Tg mice as well as XPA (-/-) mice. These findings indicate that epidermal melanin has a high ability to protect DNA damage by UVB radiation, and thereby, prevent UV-induced inflammation, immunosuppression, and carcinogenesis.