The impact of genetic background on neurodegeneration and behavior in seizured mice

We used pilocarpine-induced seizures in mice to determine the impact of genetic background on the vulnerability of hippocampal neurons and associated changes of behavioral performance. The susceptibility of hippocampal neurons to seizure-induced cell death paralleled the severity of the seizures and depended on genetic background. Hippocampal neurons in C57BL/6 mice were most resistant to cell death, whereas they were highly vulnerable in FVB/N mice. The degree of neuronal degeneration in F1 hybrid mice obtained by crossing the two strains was at an intermediate level between the parent strains. Two weeks after the severe seizures, performance in a water-maze place navigation task showed a bimodal distribution. Seventeen of 19 (90%) F1 mice were completely unable to learn while the other two learned reasonably well. Of 28 C57BL/6 mice with similarly severe seizures, six were as strongly impaired as their F1 counterparts (22%). The remaining 22 performed normally, indicating a much lower probability of C57BL/6 mice to be affected. Treated mice showed a deficit of open-field exploration which was strongly correlated with the impairment in the place navigation task and was again more severe in F1 mice. Our results show that the vulnerability of hippocampal neurons to pilocarpine-induced seizures, as well as the associated behavioral changes, depended on genetic background. Furthermore, they confirm and extend our earlier finding that a relatively modest reduction of hippocampal cell death can be associated with dramatic changes of behavioral performance and emphasize the importance of tightly-controlled genetic backgrounds in biological studies

In vitro investigations of Cynara scolymus L. extract on cell physiology of HepG2 liver cells

The objective of this study was the investigation of a potential influence of artichoke leaf extract (ALE) on the cell physiology and gene expression of phase I/II enzymes of human liver cells HepG2 and investigation on potential cell protective effects against ethanol-induced cell toxicity against HepG2 cells. Cell biological assays under in vitro conditions using HepG2 liver cells and investigation of mitochondrial activity (MTT test), proliferation assay (BrdU incorporation ELISA), LDH as toxicity marker, gene expression analysis by RT-PCR and enzyme activity of glutationtransferase. Artichocke extract, containing 27% caffeoylquinic acids and 7% flavonoids induced mitochondrial activity, proliferation and total protein content under in vitro conditions in human liver cells HepG2. These effects could not be correlated to the well-known artichoke secondary compounds cynarin, caffeic acid, chlorogenic acid, luteolin and luteolin-7-O-glucoside. The flavones luteolin and luteolin-7-O-glucoside had inhibitory effects at 100 µg/mL level on HepG2 cells, with luteolin being a significant stronger inhibitor compared to the respective glucoside. Artichoke leaf extract had minor stimulating effect on gene expression of CYP1A2, while CYP3A4, GGT, GPX2, GSR and GST were slightly inhibited. GST inhibition under in vitro conditions was also shown by quantification of GST enzyme activity. Induction of gene expression of CYP1A2 was shown to be supraadditive after simultaneous application of ethanol plus artichoke extract. Artichoke leaf extract exhibited cell protective effects against ethanol-induced toxicity within cotreatment under in vitro conditions. Also H2O2 damage was significantly inhibited by simultaneous artichoke incubation. Pre- and posttreatments did not exert protective effects. DMSO-induced toxicity was significantly reduced by pre-, post- and cotreatment with artichoke extract and especially with luteolin-7-O-glucoside, indicating a direct interaction with the toxifying agent and an induction of repair mechanisms.<br>O objetivo deste estudo foi a investigação de uma potente influência do extrato das folhas da alcachofra (ALE) na fisiologia celular e na expressão gênica de enzimas de fase I/II de células hepáticas humanas HepG2 e investigação no potencial efeito protetor celular em células HepG2 contra toxicidade celular induzida por etanol. Ensaios biológicos de células em condições in vitro usando células de fígado HepG2 e investigação da atividade mitocondrial (teste MTT), ensaio de proliferação, LDH como marcador de toxicidade, análise de expressão gênica por RT-PCR e atividade da enzima glutationa transferase. O extrato da alcachofra, contendo 27% de ácidos cafeoilquínico e 7% de flavonóides, induzem a atividade mitocondrial, proliferação e o teor de proteína total em condições in vitro em células hepáticas humanas HepG2. Estes efeitos não podem ser correlacionados aos compostos secundários conhecidos da alcachofra, cinarina, ácido cafeico, ácido clorogênico, luteolina e luteolin-7-O-glicosídeo. As flavonas luteolina e luteolin-7-O-glicosídeo possuem efeitos inibitórios em nível de 100 µg/mL em células HepG2, com a luteonina sendo uma inibidora significativamente mais forte comparada com o respectivo glicosídeo. O extrato das folhas de alcachofra possui um efeito mínimo da estimulação na expressão gênica de CYP1A2, enquanto CYP3A4, GGT, GPX2, GSR e GST foram sutilmente inibidos. A inibição de GST em condições in vitro também foi mostrada pela quantificação da atividade da enzima GST. Indução da expressão gênica de CYP1A2 mostrou-se supraaditiva após aplicação simultânea do etanol mais o extrato de alcachofra. O extrato das folhas de alcachofra exibiu efeitos protetores celulares frente à toxicidade induzida por etanol em co-tratamento em condições in vitro. Além disso, danos por H2O2 foram significativamente inibidos pela incubação simultânea do extrato de alcachofra. Pré e pós-tratamento não exerceram efeitos protetores. Toxicidade induzida por DMSO foi significativamente reduzida por pré, pós e co-tratamento com extrato de alcachofra e especialmente com luteína-7-O-glicosídeo, indicando uma interação direta com o agente toxicante e a indução dos mecanismos de reparo