Similar to those in HDL2 patients, the RNA foci were rarely colocalized with NIs and were colocalized with Mbnl1 (Figure S2B). Selleckchem GSK1349572 Thus, we concluded that BAC-HDL2 mice also recapitulate the phenotype of CUG RNA foci, another

molecular pathological marker for HDL2. One intriguing finding in HDL2 neuropathology is the immunoreactivity of NIs with 1C2, a monoclonal antibody that has relatively high specificity to all expanded neuropathogenic polyQ proteins (Trottier et al., 1995), but can also recognize some normal long polyQ proteins such as TBP as well as some other amino acid stretches such as polyleucine (Dorsman et al., 2002). Because of this latter possibility, the precise molecular nature of the 1C2 immunoreactivity within NIs in HDL2 remains to be clarified. We next asked whether the NIs in BAC-HDL2 mice, like those in HDL2 patients, could be immunostained with 1C2. By using a sensitive antigen retrieval technique (Osmand et al., 2006) we were able to detect 1C2-immunoreactive NIs in 12-month-old BAC-HLD2

brains that are unlike the faint diffuse nuclear staining found in the Volasertib mw wild-type controls (Figure S3A). Such 1C2 (+) NIs were not detected at 1 month old, but could be detected at 3 months old and became progressively enlarged at 6 and 12 months old (Figures S3A and S3C). Finally, double immunofluorescent staining revealed that 1C2-immunoreactive NIs colocalized with ubiquitin-positive NIs (Figure S3B), suggesting that the composition of NIs in BAC-HDL2 mice is quite similar to those described in HDL2 patients. To provide further evidence that BAC-HDL2 NIs contain an expanded polyQ protein, we used another monoclonal antibody, 3B5H10, which has been shown to be specific to the expanded crotamiton polyQ epitope in all known polyQ disorders (Brooks et al., 2004). Immunostaining with 3B5H10 after antigen retrieval revealed that NIs in 12-month-old BAC-HDL2 cortices and striatum were prominently stained with this expanded polyQ-specific antibody (Figure 3). No such 3B5H10 (+) NIs were detected in the brains of wild-type control littermates at 12 months old (Figure 3). Importantly, the distribution of

3B5H10-immunoreactive NIs in BAC-HDL2 brains is strikingly similar to that of patients, with prominent levels of NIs in the cortex (the upper cortical layers more than the deep cortical layers), hippocampus, and amygdala, decreased abundance in the striatum, and very few if any NIs detected in the cerebellum, thalamus, and brain stem (Figure 4 and data not shown). Taken together, our neuropathological studies with both 1C2 and 3B5H10 antibodies demonstrated that the NIs found in BAC-HDL2 brains recapitulate the patterns seen in HDL2 patients. Furthermore, an expanded polyQ protein is probably a component of such NIs. Because pathogenesis of HDL2 has been linked to the expansion of CTG/CAG repeats at the human JPH3 locus ( Holmes et al.