Year 2

Under this grant awarded TR1-01232 titled ‘Mouse Models for Stem Cell Therapeutic Development’ we proposed to eliminate bottlenecks to the translation of stem cell research by a) developing a comprehensive collection of standardized mouse models on the appropriate immune backgrounds, and b) establishing production-scale processes to ensure their efficient availability to California researchers.

Year 1 milestones included the release of the streptozotocin induced type 1 diabetes model and this achievement was reported in our year 1 progress report. Year 1 milestones also included the development of a model of Multiple Sclerosis (MS).

At the end of year 1 we reported that we had optimized the induction protocol for both the proteiolipid protein (PLP) and myelin-oligodendrocyte (MOG) induced MS models. We have been able to reproducibly induce disease in both male and female NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG) mice. In the PLP model we are able to consistently achieve a pattern of relapsing and remitting disease although the remission is only about 50% of the peak disease severity. We have assessed the efficacy of glatiramer acetate (GA; Copaxone) as a reference compound in the PLP model and determined its effectiveness in reducing the severity of disease in female NSG mice but not in males. In the MOG model of MS we show disease relapse and remission although the changes in disease severity are not as discrete as that observed in the PLP model and may be more correctly labeled a vacillation in disease severity. Although the standard test used in these MS models is disease activity index scoring we sought to develop a less subjective and more rapid test for higher throughput screening of test compounds. We have looked at grip strength and found a reproducible reduction in this parameter in mice induced to disease compared to naïve animals. We also observed a reproducible improvement in grip strength in GA treated animals. Although higher throughput. We may, if time permits, look at locomotor function in these models. We are currently awaiting the pathology report on these models. We have completed the scale up study and the shipping assessment and we are currently finalizing the release of these models.

Year 2 milestones include the release of our chronic (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) induced Parkinson’s disease model and the release of our stroke model.

We have completed the optimization of our chronic MPTP model and identified that although significant death is observed following twice weekly dosing of MPTP at 27.5mg/kg for 5 weeks (50%) this dose of MPTP provides the most reproducible behavioral changes. On locomotor testing MPTP treated mice showed a consistent increase in clinical signs of anxiety. This is important as this is a feature of the disease observed in human patients and is associated with pathology rather than the trauma of the disease alone. We also showed a reproducible decrease in motor function although interestingly this was seen after an initial observation of improved performance measured by rotarod and open field testing. The initial improvement is likely a phenomenon associated with a compensatory change in gait, such as decreased stride length and increased stride frequency, which masks clinical signs of disease during the early milder phase. Immobility testing by tail suspension showed a significant decrease in movement on tail suspension in MPTP treated male NSG mice but this finding was not reproducible in females. We are awaiting dopaminergic cell counts to see if females have less reproducible cells loss following MPTP treatment. Interestingly, reduced risk of Parkinson’s disease and disease severity is associated with females. The chronic MPTP model of Parkinson’s disease is available to researchers for both shipment to their facility and for execution of studies through our internal service.

Although initially a year 3 milestone we reported at the end of our first year of the award that we had moved TBI to year 2 and stroke to year 3. We have initiated the development of the traumatic brain injury (TBI) protocol. The TBI surgical procedure has been optimized and we have standardized the injury protocol. Using the Precision Instruments automated TBI system we have established mild, moderate, and severe injury profiles.

Earlier than scheduled, the development and standardization of the surgical procedure for stroke induction has been completed. Due to variability in the vasculature of the NSG mouse reproducibility of injury is proving difficult.

We have also initiated the SCI model development and significant progress is being made.

I can report that the adjustment to our proposed milestones has allowed us to remain on schedule for the release of all models proposed in the original grant within the 3 years of the grant award.