Modeling cognition and disease using human glial chimeric mice.

Human glial progenitors can colonize and fully myelinate the hypomyelinated mouse CNSA–B. Sagittal images of an immmunodeficient shiverer mouse brain (MBP shi/shi x rag2−/−), sacrificed at 1 year after neonatal transplant of hGPCs. Each image in A and B represents a montage of 50 to 100 individual 10x photomicrographs; each series begins 750 μm lateral to the midline and continues at 600 μm intervals. A. Human donor cells, immunolabeled in 14 μm cryosections using an anti-human nuclear antibody (hN; red). B Myelin basic protein (MBP; green) in sections adjacent or nearly so to their matched sections in A. C. Donor-derived MBP (green) in the spinal cord at 1 year, at the level of the conus medullaris. D shows a Kaplan-Meier plot of the effect of neonatal hGPC engraftment upon the survival of these mice. Shiverer x rag2−/− mice were either engrafted at birth with hGPCs (n=26, red), injected with saline (n = 29, green), or not treated (n=59, blue). Whereas all control mice died between 18 and 21 weeks of age, a fraction of engrafted mice lived substantially longer than any controls; some were frankly rescued, with concomitant recovery of neurological phenotype as well. The experiment was terminated at 13 months, and rescued mice were sacrificed for observation through 2 years of age. Scale: 200 μm.A, B and D from (Windrem et al. 2008).

Human GPCs out-compete and ultimately replace resident mouse GPCsA. Human glial progenitor cells (hGPCs), neonatally transplanted into either congenitally hypomyelinated shiverer x rag2−/− (left column) or normally myelinated rag2−/− (right column) mouse brain, disperse and expand broadly throughout the brain as a function of age. hGPCs reach higher density in white matter than gray matter of the hypomyelinated shiverer (left), in contrast to their relatively uniform distribution in normally myelinated brain (right). Red dots indicate individual human donor GPCs, as labeled by human nuclear antigen (left and right columns). B. Progressive replacement of host mouse GPCs, by donor hGPCs, as identified by species-specific antibodies against NG2, compared to an unengrafted control mouse (left-most column). By one year, hGPCs have replaced mouse GPCs throughout the entire depth of the cortex. C. EGFP-tagged human astrocytes in the corpus callosum, cortex, and fimbria in a 2-year old rag2−/− mouse. The hGPCs were labeled with an EGFP-expressing lentivirus (green) in vitro, prior to transplantation. Dorsal is toward the top and rostral to the left in this sagittal section. (EGFP, green; human-specific GFAP, red). Scale: 100 μm.A and B, from (Windrem et al. 2014).

Human glial chimeras manifest enhanced long-term potentiation and learn more rapidlyA. Induction of LTP by two trains of high-frequency stimulation (each train consisted of 100 pulses at 100 Hz, with 30 s between bursts) in human chimeric mice, but not in unengrafted littermates or mice allografted with conspecific mouse GPCs (n=7 mice/group, *p<0.05; t-test compares fEPSP slopes before and 60 min after stimulation, for each group). B. Object-Location Memory Task (OLT) in chimeric mice and their unengrafted rag1 null littermate controls demonstrated a learning advantage in chimeric mice, via enhanced recognition of and preference for the novel displaced object (n=7, **p<0.01, one-way ANOVA). C. Auditory fear conditioning assessed in a cohort of human glial chimeric, mouse allografted, and unengrafted rag2−/− control mice. The human glial chimeric mice exhibit prolonged freezing behavior in test chamber 2 during exposure to the tonal conditioned stimulus, when compared to their unengrafted mice and allografted controls (n=5–20, *p<.05, **p<0.01, 2-way repeated-measures ANOVA with Bonferroni t-tests). D. Barnes maze testing in chimeric and unengrafted immunodeficient littermate controls. Chimeric mice demonstrated a significant learning advantage, as reflected in a shorted latency and fewer errors in solving the maze (n=6, *p<0.05, **p,0.01, 2-way ANOVA with Bonferroni t-tests).From (Han et al. 2013).

Neonatal engraftment with hiPSC GPCs produces patient-specific human glial chimerasConfocal images of the callosal and capsular white matter of shiverer mice neonatally engrafted with hiPSC OPCs produced from hiPSCs derived from 3 independent patient-derived lines, designated C27 (A–D); K04 (E–H); and C14 (I–K). A, G, and J show abundant, donor-derived myelin basic protein expression (MBP, green) by C27, K04, and C14 hiPSC hGPCs (hNA, red), respectively. Representative z stacks of individual hNA+ oligodendroglia are shown as asterisks in A and E. By 19 weeks, hiPSC oligodendroglia derived from all 3 lines (B, C27; E–F, K04; J, C14) robustly myelinated axons (neurofilament, red). hiPSC-derived oligodendroglial morphologies are exemplified in panels F (K04) and I (C14); F in particular shows multi-axon myelination by single oligodendroglia in the striatum. hiPSC hGPCs also generated astroglia (C, C27; H, K04), which exhibited the complex fibrous morphologies typical of human astrocytes (human-specific GFAP, green). Many cells also remained as progenitors, immunostaining for NG2 (D, C27) or PDGFαR (K, C14). Scale: 50 μm (A–C, G, J); 20 μm (C–F, H, K); and 10 μm (I, insets to A and E).From (Wang et al. 2013).

Human glial chimeric mice can develop a prototypic human viral encephalitisJCV infection and spread in vivo was tracked by immunostaining human glial-engrafted shiverer brains for the late viral VP1 antigen, as a function of time after infection. In these chimeric shiverers, essentially all myelinating oligodendrocytes, as well as most GPCs and large numbers of astrocytes, are human. A–B, By 4 weeks after infection by the polyomavirus JCV, focal regions of demyelination (A, arrows) and infection-associated astrogliosis (B, arrow) are noted in the forebrain white matter of human glial chimeric mice, typically abutting the callosal wall of the lateral ventricle. C, By 11 weeks after infection, diffuse hypomyelination of the corpus callosa and capsules of infected chimeric mice was noted. D, Sagittal sections of 3 different infected chimeras at each of 3 time-points are shown; individual VP1+ cells are dot-mapped onto the schematic. VP1+ human cells became progressively more widespread with time, with JCV infection progressing from the site of viral injection to include much of the forebrain white matter by 12 weeks post-infection, with marked cortical spread by that point as well. Scale: 200 μm (A, C); 100 μm (B).From (Kondo et al. 2014).