The original upstream model selleckchem Sunitinib in, which includes IKK cycling among three states and feedback from A20, was unable to adequately fit either the rapid activation or deactivation of micro glial activation. Therefore, we examined ways in which the model could be modified consistent with the biology to better correspond with the data. Activation of the IKK complex at the biomolecular level involves the recruitment and assembly of a signal ing complex following TNFa binding to its receptor, as well as numerous post translational modifications to the complex subunits before IKK is activated by phosphory lation at two residues within its kinase domain. Although other studies have attempted to model the upstream pathway in a greater level of detail, many of the details are still being resolved and we opted to retain the basic IKK cycling description from.

The activation reaction rate was changed from a linear function to a nonlinear Hill equation as a coarse approximation to the many intermediate steps involved in IKK activation. The quick attenuation of IKK activity following its induction is essential to proper signaling and the result ing biphasic NF B activity. IKK reportedly undergoes hyperphosphorylation at 9 or 10 residues in the C terminal, which was found to significantly decrease kinase activity in cells. We posited that potential cooperativity in IKK inactivation due to autophosphory lation may lead to nonlinearites in the inactivation rate equation of the model. Accordingly the linear reaction rate was changed to a nonlinear Hill equation.

Feedback from A20 in the published model was pro posed to inhibit the transition of inactivated IKK back to its native state. Because we were unaware of any bio logical basis for such a mechanism, we adopted two mechanisms of A20 interaction that had been identified in the literature and had also been included in prior models. The first is direct inactivation of the IKK complex by A20 protein, a mechanism reported in and previously modeled in. We used the identical mathematical description of this interaction from in our model. Secondly A20 is known to inhibit activation indirectly through its ubiquitin editing activities of upstream signaling components. This mechanism has been included in previous models that have a more detailed description of the upstream signaling pathway.

We adapted this second interaction to our model by assuming that A20 attenuates the rate of TNF induced IKK activation in a concentration dependent manner. Parameter estimation was performed using the newly developed upstream model with fixed nuclear NF B as the model input. Parameters were found for which the model produced excellent agreement with Batimastat microglial IKK activation, decreasing the fitting error by more than an order of magnitude compared to the best fit achieved with the original upstream model.