Preclinical Models Core
Intellectual and developmental disabilities (IDD) and neurodevelopmental disorders (NDD) are complex conditions with wide-ranging impact on affected individuals and caretakers. It is now possible to replicate many IDDs in the laboratory using animal, cellular, or genetic modeling. This core makes rapidly advancing techniques for generating cellular and organismal models of IDD available and accessible to KIDDRC scientists.
The PMC provides the following services to KIDDRC scientists and projects:
The Model Development program produces genetically modified mice and patient-specific and modified pluripotent stem cells for IDD-related studies.
Genome editing. Inherited or acquired gene mutations are believed to cause a majority of NDDs. Accordingly, techniques are required to introduce suspected causal mutations into cells or organisms to catalogue the consequences. The core utilizes CRISPR/Cas9 methods to modify the mouse genome or for transfection of human and mouse pluripotent stem cells for making genetic alterations in vitro.
Rodent modeling services. Humans and mice share significant portions of their genomes, and large numbers of orthologous genes can be edited to faithfully recreate human mutations. While simpler, higher throughput organismal models exist, common genes are fewer and findings typically must be confirmed in rodents, where phenotypes can be better ascertained. The rapidity with which mice can be generated with gene editing technologies minimizes throughput time and expense.
Cellular modeling services. Understanding how mutations lead to developmental disorders requires study of their impact at the cellular level. Cells obtained by skin biopsy or blood draw can be reprogrammed to take on stem cell properties and subsequently differentiated into neurons that are essentially indistinguishable from native cells. Patient-specific induced pluripotent stem (iPS) cell models allow us to capture genetic material of patients with developmental disorders, permitting in vitro study of the genetic basis for these conditions.
Once an IDD-relevant model is generated, it must be characterized. Moreover, when an intervention is delivered, it is necessary to assess its impact at an organismal and/or cellular level.
Behavioral, Metabolic and Biomarker Analysis. If a genetic variant or environmental insult is responsible for neurological dysfunction, then a rodent model incorporating the variant or exposed to the insult should display behavioral changes that recapitulate the human scenario. A variety of behavioral services and equipment are available in conjuction with fully trained staff to interrogate behavioral manifestations. Similarly, metabolic disorders commonly co-present in IDD patients. An assortment of metabolic equipment and services is available for training or assisted services as warranted by KIDDRC projects. Biomarkers can be analyzed on demand from human or rodent serum, plasma or other potential matrices by expert staff.
Histology. Histological analysis is a mainstay for understanding neuroanatomical bases of developmental disorders. The PMC provides full-service histology facilities to all KIDDRC members. The facility has large dedicated space, multiple microtomes and cryostats, paraffin embedding, automated staining and labeling, and associated services. Staff perform an array of immunohistochemical and immunofluorescence procedures and provide training on all facility equipment. Investigators can engage our staff beginning with fixation and dissection, through embedding, sectioning, staining and mounting of their sectioned material, and have it placed in a queue for our imaging services.
Microscopic Imaging and Cellular Phenotyping. The PMC maintains a variety of microscope platforms including three Nikon confocal microscopes in inverted and upright configurations for fixed and live cell imaging, spectral separation, and confocal stereology. Additionally, the core maintains a a Nikon High Content Analysis System, allowing rapid automated fluorescent and brightfield imaging of slides and culture plates.
Image Analysis and Graphics. The PMC has dedicated workstations for standard analysis needs and provides training on all software. Imaging software (MetaMorph, Elements) is installed on multiple shared computers for off-line analysis, together with a full software spectrum for data management and statistical analysis. The core leverages the 1Pb shared storage the university provides without charge to house and manage the large image files generated during service provision. The PMC offers 3D printing, and printed images have included cranial reconstructions and novel objects for behavioral phenotyping services. Graphics staff oversees 2D and 3D image output and graphic and illustrative design.
Genomics. Gene mutations can result in transcriptional changes leading to IDD. Identifying genetic changes is necessary for understanding IDD etiology and identifying potential therapeutic targets. KIDDRC provides genomics services including microarray profiling and a variety of next-generation sequencing techniques. Sequencing is performed on investigator-provided animal or human tissue, cells, or isolated genomic material, and the facility generates libraries, prepares samples for sequencing, performs all QC, and conducts sequencing (RNA, DNA, exome analysis, Methyl-seq, ChIP-seq, etc.) to the PI’s specifications. To complement the global-based sequencing applications, single-cell sequencing is available and is further enhanced with the multi-omic capabilities of a Mission Bio Tapestry system. The combined interrogative power of single-cell and global sequencing applications, paired with the versatile capabilities of the NovaSeq, provides greater ability to understanding genetic underpinnings of IDD. To ensure rigor and validity of these complex experiments, the genomics facility dovetails with the KIDDRC Bioinfomatics team providing a free consultation for all KIDDRC investigators to facilitate optimal experimental design. Upon generation, sequencing data are transferred automatically to bioinformatics for analysis.
Mass Spectrometry Proteomics. Like transcriptional changes, altered protein translation and processing occurs within target cells and contributes to IDD. Mass spectrometry proteomics can identify these changes and facilitate understanding of targets and etiology of IDD.
Additional available core technologies
While the PMC consolidates a wide range of core services that are integral to developing and characterizing IDD models, additional technologies are sometimes required on an ad hoc basis. KU is fortunate in having an abundance of cores offering cutting-edge technologies that the PMC can access. Frequently used, informative key core technologies for IDD modeling include:
- Flow Cytometry (used in stem cell selection and single-cell seq preparation)
- Hoglund Biomedical Imaging Center (9.4T magnet for small animal imaging)
- In Vivo Imaging Lab (IVIS non-invasive fluorescence small animal imaging)
- Electron Microscopy Lab (subcellular relationships)
- Small molecule mass spectrometry Analytical Core
- The Biospecimen Repository (for human samples)
- High Throughput Screening Lab (screening large numbers of molecules using cell-based assays)
Because many of these cores are underwritten by the CTSA, COBREs or the Kansas INBRE, they provide cost-effective access for KIDDRC investigators.