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The metabolism of the insecticide momfluorothrin (1), 2,3,5,6-tetrafluoro-4-(methoxymethyl)benzyl (EZ)-(1R,3R)-3-(2-cyanoprop-1-enyl)-2,2-dimethylcyclopropanecarboxylate, 14C-labeled at the benzyl or cyclopropyl carbon, was investigated in lettuce. The metabolic profiles were similar between the two active ingredients, 1-R-trans-Z and 1-R-trans-E. On the leaf surface, 1 gradually volatilized and penetrated into the plant with concomitant degradation to form aldehyde/carboxylic acid derivatives via oxidative cleavage of the propenyl double bond. No isomerization of 1 proceeded at any chiral carbon. In the leaf tissues, 1 underwent ester hydrolysis to give the corresponding alcohol and chrysanthemic acid moieties, followed by glucose conjugation and successive malonic acid or ribose modification. Assuming O3 or 1O2 as the major reactant for the degradation on the plant, the reactivity with the alkenyl group in the substructure methyl (1R,3R)-3-[(Z)-2-cyanoprop-1-enyl]-2,2-dimethylcyclopropane-1-carboxylate was estimated from the HOMO/LUMO energy at the B3LYP/6-311+G** level, which indicated a lower potential of 1 than analogous pyrethroids due to its electron-withdrawing cyano group.

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A mass spectrometry (MS)-based proteomic methodology was employed to monitor oxidative modifications in keratins, the main constituents of human skin ("Non-invasive proteomic analysis of human skin keratins: screening of methionine oxidation in keratins by mass spectrometry" [1], "UV irradiation-induced methionine oxidation in human skin keratins: mass spectrometry-based non-invasive proteomic analysis" [2]). Human skin proteins were obtained non-invasively by tape stripping and solubilized in sodium dodecyl sulfate (SDS) buffer, followed by purification and digestion using the filter-aided sample preparation method. The tryptic peptides were then analyzed by liquid chromatography (LC)/electrospray ionization (ESI)-MS, tandem MS (MS/MS), and LC/ESI-selected reaction monitoring (SRM)/MS. The MS/MS data were generated to confirm amino acid sequences and oxidation sites of tryptic peptides D(290)VDGAYMTK(298) (P1) and N(258)MQDMVEDYR(267) (P2), which contain the most susceptible oxidation sites (Met(259), Met(262), and Met(296) in K1 keratin) upon UVA irradiation [2]. Subsequently, quantitative determination of the relative oxidation levels of P1 and P1 [2] was achieved by LC/ESI-SRM/MS analyses of P1 and P2 together with their oxidized forms after exposure to UVA radiation or treatment with hydrogen peroxide (H2O2).

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Ultraviolet (UV) radiation is the major environmental factor that causes oxidative skin damage. Keratins are the main constituents of human skin and have been identified as oxidative target proteins. We have recently developed a mass spectrometry (MS)-based non-invasive proteomic methodology to screen oxidative modifications in human skin keratins. Using this methodology, UV effects on methionine (Met) oxidation in human skin keratins were investigated. The initial screening revealed that Met(259), Met(262), and Met(296) in K1 keratin were the most susceptible oxidation sites upon UVA (or UVB) irradiation of human tape-stripped skin. Subsequent liquid chromatography/electrospray ionization-MS and tandem MS analyses confirmed amino acid sequences and oxidation sites of tryptic peptides D(290)VDGAYMTK(298) (P1) and N(258)MQDMVEDYR(267) (P2). The relative oxidation levels of P1 and P2 increased in a time-dependent manner upon UVA irradiation. Butylated hydroxytoluene was the most effective antioxidant for artifactual oxidation of Met residues. The relative oxidation levels of P1 and P2 after UVA irradiation for 48 h corresponded to treatment with 100mM hydrogen peroxide for 15 min. In addition, Met(259) was oxidized by only UVA irradiation. The Met sites identified in conjunction with the current proteomic methodology can be used to evaluate skin damage under various conditions of oxidative stress. BIOLOGICAL SIGNIFICANCE: We demonstrated that the relative Met oxidation levels in keratins directly reflected UV-induced damages to human tape-stripped skin. Human skin proteins isolated by tape stripping were analyzed by MS-based non-invasive proteomic methodology. Met(259), Met(262), and Met(296) in K1 keratin were the most susceptible oxidation sites upon UV irradiation. Met(259) was oxidized by only UVA irradiation. Quantitative LC/ESI-SRM/MS analyses confirmed a time-dependent increase in the relative oxidation of target peptides (P1 and P2) containing these Met residues, upon UVA irradiation of isolated human skin. The relative oxidation levels of P1 and P2 along with the current proteomic methodology could be applied to the assessment of oxidative stress levels in skin after exposure to sunlight.

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This study was designed to evaluate changes in auditory brainstem response (ABR) in the course of auditory disturbance in rats induced by Kanamycin (KM). KM was administered subcutaneously to 12 CD (SD) male rats aged 6 weeks for 10 days at a dose of 800 mg/kg. Death was observed in one male on day 8 and 2 males on day 10. It was thought that kidney damage was the cause of death from histopathological findings. ABR was recorded before KM treatment and on days 4, 8, 10 and 11 after KM treatment. The ABR changes after KM treatment in rats were as follows. On day 4, 6 rats showed an increase in amplitude of waves I and/or II and on day 8, among those, 4 rats still showed a high amplitude of waves I and/or II. On day 8, 2 rats showed an elevation of ABR threshold (15-40 dB SPL) and a decrease in amplitude of wave I and increase in amplitude of wave II at the same time. On day 11, 7 rats showed a decrease in amplitude of wave I. In addition, ABR threshold shifts (10-70 dB SPL) were observed in those rats. In ABR recording, KM-induced auditory disturbance model rats showed an increase in amplitude of waves I and/or II earlier than an ABR threshold shift. By analyzing temporal alteration of amplitude of the ABR components, we could detect precursory phenomenon of the auditory disturbance at an early phase of treatment. By following the pathway of click-ABR and tone pip-ABR examination, the auditory disturbance of low- frequency to high-frequency range could be analyzed at an early date in detail.

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Extraskeletal osteosarcoma is a very rare tumor in humans and animals including rats. This paper describes a case of extraskeletal osteosarcoma observed in the glandular stomach of an aged female Fischer 344 rat. Grossly, a whitish solid mass was observed at the greater curvature of the glandular stomach. Histologically, the tumor consisted of both atypical polygonal and pleomorphic spindle-shaped cells, with pleomorphic nuclei, and it contained variable amounts of osteoids and small clumps of mature bone tissue. In addition, mitotic figures were frequently observed. Neither invasion of the muscle layer or vessels in the stomach nor metastasis to distant organs was detected. There were no skeletal tumors in the body. Immunohistochemically, the tumor cells were positive for osteocalcin, osteonectin, vimentin and S-100 protein. Judging from these results, the present tumor was diagnosed as extraskeletal osteosarcoma. This is the first report of spontaneous extraskeletal osteosarcoma arising from the stomach in a rat.

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We investigated a case of spontaneous malignant T-cell lymphoma observed in a 19-week-old male Crl:CD (SD) rat. The rat showed paralysis beginning 1 week before euthanasia. Hematological examination revealed marked lymphocytosis without distinct atypia. Macroscopically, hepatosplenomegaly and partial atrophy of the thoracic spinal cord were observed. Microscopically, neoplastic cells infiltrated into the liver, splenic red pulp, bone marrow and epidural space of the thoracic spinal cord, while no neoplastic cells were observed in the thymus and lymph nodes. Moreover, the spinal cord showed focal degeneration due to compression by marked infiltration of neoplastic cells in the subdural space. The neoplastic cells were generally small-sized round cells that had a round nucleus with/without a single nucleolus and scanty cytoplasm. Immunohistochemically, the neoplastic cells were positive for CD3 and CD8 and negative for CD79α. Judging from these results, the present tumor in this young adult rat was diagnosed as malignant T-cell lymphoma.