rifampin and Neuroblastoma

Mitochondrial and autophagic dysfunction, as well as neuroinflammation, are associated with the pathophysiology of Parkinson's disease (PD). Rotenone, an inhibitor of mitochondrial complex I, has been associated as an environmental neurotoxin related to PD. Our previous studies reported that rifampicin inhibited microglia activation and production of proinflammatory mediators induced by rotenone, but the precise mechanism has not been completely elucidated. BV2 cells were pretreated for 2h with rifampicin followed by 0.1μM rotenone, alone or in combination with chloroquine. Here, we demonstrate that rifampicin pretreatment alleviated rotenone induced release of IL-1β and IL-6, and its effects were suppressed when autophagy was inhibited by chloroquine. Moreover, preconditioning with 50μM rifampicin significantly increased viability of SH-SY5Y cells cocultured with rotenone-treated BV2 cells in the transwell coculture system. Chloroquine partially abolished the neuroprotective effects of rifampicin pretreatment. Rifampicin pretreatment significantly reversed rotenone-induced mitochondrial membrane potential reduction and reactive oxygen species accumulation. We suggest that the mechanism for rifampicin-mediated anti-inflammatory and antioxidant effects is the enhancement of autophagy. Indeed, the ratio of LC3-II/LC3-I in rifampicin-pretreated BV2 cells was significantly higher than that in cells without pretreatment. Fluorescence and electron microscopy analyses indicate an increase of lysosomes colocalized with mitochondria in cells pretreated with rifampicin, which confirms that the damaged mitochondria were cleared through autophagy (mitophagy). Taken together, the data provide further evidence that rifampicin exerts neuroprotection against rotenone-induced microglia inflammation, partially through the autophagy pathway. Modulation of autophagy by rifampicin is a novel therapeutic strategy for PD.

Topics: Analysis of Variance; Antirheumatic Agents; Autophagy; Cell Line, Tumor; Chloroquine; Coculture Techniques; Humans; Insecticides; Interleukin-1beta; Interleukin-6; Membrane Potential, Mitochondrial; Microglia; Microscopy, Electron, Transmission; Mitochondria; Neuroblastoma; Neuroprotective Agents; Reactive Oxygen Species; Rifampin; Rotenone

Pregnane X receptor (PXR) is an important transcriptional regulator that plays important roles in the cell metabolism and cell growth by regulating the transcriptional of a sort of metabolizing enzymes.. To investigate whether rifampicin effected HepG2 cells growth and the inhibition was due to the G0/G1 phase arrest.. PXR-knockdown experiments using RNAi showed that the cell cycle phase arrest mediated by rifampicin based on activation of PXR. The results also indicated that cell phase arrest by rifampicin could protect cells form UVB-induced DNA damage. Retinoid X receptor alpha (RXRα) expression level in cells is another key factor for cell cycle phase arrest mediated by rifampicin. Both over expression and lacking expression of RXRα in cell reduced the cell arrest efficiency mediated by rifampicin. In the study, we found that rifampicin inhibited HepG2 cells growth and demonstrated that the inhibition is due to the G0/G1 phase arrest through flow cytometry analysis.. The results showed that RXRα promote cell cycle phase transition rate of HepG2. Competitive bind of rifampicin-activated PXR with RXRα is one main reason to arrest cell cycle phase through inhibiting combination of RXRα with other partners. Rifampicin could promote cell growth rate when RXRα expressed more excessively than PXR in cells.

Topics: Binding, Competitive; Cell Division; Cell Line, Tumor; DNA Damage; G1 Phase; G1 Phase Cell Cycle Checkpoints; Hep G2 Cells; Humans; Neuroblastoma; Pregnane X Receptor; Real-Time Polymerase Chain Reaction; Receptors, Steroid; Recombinant Fusion Proteins; Resting Phase, Cell Cycle; Retinoid X Receptor alpha; Rifampin; RNA Interference; RNA, Small Interfering; Transcription, Genetic; Ultraviolet Rays

The cellular uptake of microcystin-LR, a cyclic heptapeptide hepatotoxin from the cyanobacterium Microcystis aeruginosa, was studied by means of a radiolabelled derivative of the toxin. 3H-dihydromicrocystin-LR. The uptake of 3H-dihydromicrocystin-LR was shown to be specific for freshly isolated rat hepatocytes whereas the uptake in the human hepatocarcinoma cell line Hep G2 as well as the mouse fibroblast cell line NIH-3T3, and the human neuroblastoma cell line SH-SY5Y, was negligible. By means of a surface barostat technique it was shown that the membrane penetrating capacity (surface activity) of microcystin-LR was low, indicating that the toxin requires an active uptake mechanism. The hepatocellular uptake of microcystin-LR could be inhibited in the presence of bile acid transport inhibitors such as antamanide (5 microM), sulfobromophthalein (50 microM) and rifampicin (30 microM). The uptake was also reduced in a concentration dependent manner when the hepatocytes were incubated in the presence the bile salts cholate and taurocholate. A complete inhibition of the hepatocellular uptake was achieved by 100 microM of either bile salt. The overall results indicate that the uptake of microcystin-LR is through the multispecific transport system for bile acids. This mechanism of cell entry would explain the previously observed cell specificity and organotropism of microcystin-LR.

Topics: Amino Acid Sequence; Animals; Bile Acids and Salts; Biological Transport; Cell Membrane; Humans; Kinetics; Liver; Liver Neoplasms; Liver Neoplasms, Experimental; Mice; Mice, Inbred Strains; Molecular Sequence Data; Neuroblastoma; Peptides, Cyclic; Rifampin; Sulfobromophthalein; Tumor Cells, Cultured