Inhibition of Heat Shock Protein 90 Reduces Inflammatory Signal Transduction in Murine J774 Macrophage Cells and Lessens Disease in Autoimmune MRL/lpr Mice: What in vitro, in vivo, and in silico Models Reveal
| dc.contributor.author | Shimp, Samuel Kline | en |
| dc.contributor.committeechair | Rylander, M. Nichole | en |
| dc.contributor.committeecochair | Reilly, Christopher M. | en |
| dc.contributor.committeemember | Caudell, David L. | en |
| dc.contributor.committeemember | Achenie, Luke E. K. | en |
| dc.contributor.committeemember | Lee, Yong Woo | en |
| dc.contributor.department | Biomedical Engineering | en |
| dc.date.accessioned | 2017-04-06T15:42:30Z | en |
| dc.date.adate | 2012-05-30 | en |
| dc.date.available | 2017-04-06T15:42:30Z | en |
| dc.date.issued | 2012-04-30 | en |
| dc.date.rdate | 2016-10-18 | en |
| dc.date.sdate | 2012-05-14 | en |
| dc.description.abstract | Heat shock protein 90 (HSP90) is a molecular chaperone protein that protects proteins from degradation, repairs damaged proteins, and assists proteins in carrying out their functions. HSP90 has hundreds of clients, many of which are inflammatory signaling kinases. The mechanism by which HSP90 enables inflammatory pathways is an active area of investigation. The HSP90 inhibitors such as geldanamycin (GA) and its derivative 17-dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG) have been shown to reduce inflammation. It was hypothesized that inhibiting HSP90 would reduce inflammatory signal cascade levels. To test this, J774 mouse macrophage cells were treated with 17-DMAG and immune-stimulated with lipopolysaccharide (LPS). 17-DMAG treatment reduced nitric oxide (NO) production and the expression of pro-inflammatory cytokines interleukin (IL)-6, IL-12, and TNF-α. Inhibition of HSP90 also prevented nuclear translocation of NF-κB. To investigate the anti-inflammatory effects of HSP90 inhibition in vivo, MRL/lpr lupus mice were administered 5 mg/kg 17-DMAG for six weeks via intraperitoneal injection. Mice treated with 17-DMAG were found to have reduced proteinuria and reduced splenomegaly. Flow cytometric analysis of splenocytes showed that 17-DMAG decreased double negative T (DNT) cells. Renal expression of HSP90 was also measured and found to be increased in MRL/lpr mice that did not receive 17-DMAG. The mechanistic interactions between HSP90 and the pro-inflammatory nuclear factor-κB (NF-κB) pathway were studied and a computational model was developed. The model predicts cellular response of inhibitor of κB kinase (IKK) activation and NF-κB activation to LPS stimulation. Model parameters were fit to IKK activation data. Parameter sensitivity was assessed through simulation studies and showed a strong dependence on IKK-HSP90 binding. The model also accounts for the effect of a general HSP90 inhibitor to disrupt the IKK-HSP90 interaction for reduced activation of NF-κB. Model simulations were validated with experimental data. In conclusion, HSP90 facilitates inflammation through multiple signal pathways including Akt and IKK. Inhibition of HSP90 by 17-DMAG reduced disease in the MRL/lpr lupus mouse model. A computational model supported the hypothesis that HSP90 is required for IKK to activate the NF-κB pathway. Taken together, HSP90 is a prime target for therapeutic regulation of many inflammatory processes and warrants further study to understand its mechanism of regulating cell signaling cascades. | en |
| dc.description.degree | Ph. D. | en |
| dc.identifier.other | etd-05142012-164452 | en |
| dc.identifier.sourceurl | http://scholar.lib.vt.edu/theses/available/etd-05142012-164452/ | en |
| dc.identifier.uri | http://hdl.handle.net/10919/77080 | en |
| dc.language.iso | en_US | en |
| dc.publisher | Virginia Tech | en |
| dc.rights | In Copyright | en |
| dc.rights.uri | http://rightsstatements.org/vocab/InC/1.0/ | en |
| dc.subject | inflammation | en |
| dc.subject | heat shock protein 90 | en |
| dc.subject | computational models | en |
| dc.subject | geldanamycin | en |
| dc.subject | 17-DMAG | en |
| dc.subject | nuclear factor-κB | en |
| dc.title | Inhibition of Heat Shock Protein 90 Reduces Inflammatory Signal Transduction in Murine J774 Macrophage Cells and Lessens Disease in Autoimmune MRL/lpr Mice: What in vitro, in vivo, and in silico Models Reveal | en |
| dc.type | Dissertation | en |
| dc.type.dcmitype | Text | en |
| thesis.degree.discipline | Biomedical Engineering | en |
| thesis.degree.grantor | Virginia Polytechnic Institute and State University | en |
| thesis.degree.level | doctoral | en |
| thesis.degree.name | Ph. D. | en |