MSUD is a serious liver-based metabolic disorder caused by a deficiency in the branched-chain alpha-ketoacid dehydrogenase (BCKDH) complex. Resulting branched-chain amino acid (BCAA) accretion in the body mainly affects the brain, which in most cases results in permanent neurological dysfunction or death without life-long attentive care. Recently it was shown liver transplantation alone restored BCKDH to a level sufficient to correct MSUD. To test novel therapies, a mouse model of intermediate MSUD (iMSUD) was created (Homanics et al., 2006), which mimicked human iMSUD. Therefore, this dissertation focused on the investigation of liver-directed therapeutic approaches to correct MSUD.

In the first aim, iMSUD mice were further characterized and determined to closely mirror the human disease phenotype. iMSUD mice suffered from developmental delay, seizures, and altered brain amino acid and neurotransmitter concentration. iMSUD brains also displayed histological abnormalities while liver morphology was normal.

In the second aim, adeno-associated viral (AAV) vectors were used to deliver E2 to the liver. However, no significant improvement was determined in AAV-treated iMSUD mice compared to controls. The most likely reasons this study was unsuccessful were low treatment dose, a weak albumin promoter, and possible competition and interaction between AAV-derived and iMSUD transgene-derived E2.

The third aim focused on hepatocyte transplantation (HTx). iMSUD-HTx mice had a 75% reduction in BCAA/alanine levels compared to iMSUD controls. BCKDH activity was increased, and Real Time qPCR detected donor-derived E2 in the liver. Dopamine and serotonin, along with several related metabolites, were corrected to control levels. Body weight at weaning and survival were also significantly improved in iMSUD-HTx mice.

The fourth aim focused on differentiated embryonic stem cell (ESC) transplantation. Differentiated ESCs expressed liver-specific markers after 3 days in culture and BCKDH activity was significantly increased over undifferentiated ESC populations. Liver-like ESC engraftment was verified up to 1 month following transplantation into wildtype mouse liver.

In summary, iMSUD mice were determined to be a superior model to test novel liver-directed therapies. Our findings of partial metabolic correction of iMSUD in a mouse model by HTx were very encouraging. Therefore, liver-directed therapeutic intervention for human MSUD should be investigated further.