Drug Discovery : Disease models


Adapted Rhinovirus C for Maximum Virus Yield

Building on their work, the researchers have now developed a mutated RV-C strain that induces strong cytopathic effect and replicates vigorously in the HeLa-E8 cells, yielding more than a log higher level of infectious rhinovirus particles compared to the parental clinical isolate.

Photoreceptor Scaffold for In Vitro Modeling and Transplantation Therapy

Using state-of-the-art microfabrication techniques, UW–Madison researchers have developed microstructured scaffold systems that can guide the growth of photoreceptor cells and mimic polarized outer retinal tissue. The scaffolds also may be used for transplantation of organized photoreceptor tissue with or without RPE.

Transplantation of photoreceptor-seeded scaffolds may improve grafted cell retention, survival, integration and functional visual rescue as compared to simple bolus injections. By recapitulating in vivo outer retinal architecture, these uniquely fabricated scaffolds also can be used for in vitro developmental and disease studies as well as drug screening.

The microfabrication process for scaffold production is fully compatible with numerous biomaterials, including biodegradable and non-biodegradable materials, thus allowing the scaffolds to be tailored to both in vitro and in vivo applications. The scaffolds feature biocompatible support layers (e.g., PDMS film) patterned with an array of unique through-holes having a curvilinear cell receiver and cell guide channels. The structure enables photoreceptors to be grown in a polarized orientation that mimics their development in vivo.

New Viral Propagation Method Yields Insight Into Childhood Asthma

UW–Madison researchers have developed an efficient, cost-effective method of propagating RV-C. They discovered that human cadherin-related family member 3 (CDHR3) is the receptor for RV-C and allows cell lines normally unsusceptible to HRV-C infection to support virus binding and replication.

To create cell lines capable of efficiently growing RV-C, the researchers modify the host cell so it expresses an effective amount of the CDHR3 receptor. This method enables high-throughput, large-scale production of RV-C, which in turn enables critical basic and applied research regarding this understudied pathogen.

Understanding and Treating Nervous System Dysfunction Using Modified Fly Models

UW–Madison researchers have developed new methods to study such time-dependent neurological mechanisms and to screen for potentially therapeutic small molecules using extended third instar stage (ETI) Drosophila larvae. These flies have been genetically modified to remain in the larval period for up to 30 days but are otherwise normal. Given the longer time window, these ETI larvae can be utilized to identify agents that stimulate nerve regeneration, confer neuroprotection or prevent synaptic degeneration.

For such studies, the ETI larvae are fed test compounds (e.g., from a chemical library) and the functional consequences of the test agent on processes such as neuronal survival, axonal regrowth, and synaptic maintenance are assessed in appropriate assays.

Enhanced Blood-Brain Barrier Model Outperforms All Others

UW–Madison researchers have developed a more realistic, reproducible in vitro model of the BBB. The model uses either human embryonic or induced pluripotent stem cells as a source of BMECs.

These cells are treated with retinoic acid to produce further BBB maturation and improve barrier properties. They are purified and co-cultured with other types of neurovascular cells, including pericytes, astrocytes and differentiated neural progenitor cells (NPCs).

Blood-Brain Barrier Model

UW-Madison researchers have developed a simpler, more reproducible in vitro model of the blood-brain barrier. Most existing models include primary brain microvasculature endothelial cells (BMECs), which form the blood-brain barrier in vivo, and the corresponding primary astrocytes, which affect the barrier. The improved model consists of primary BMECs on a permeable membrane support. Embryonic neural progenitor cells (NPCs), which can be stimulated to differentiate into each of the major brain lineages that help govern the blood-brain barrier, are co-cultured with the BMECs.

Adding differentiating NPCs to the BMEC model results in more realistic and in vivo-like properties, including increased trans-endothelial electrical resistance, reduced permeability and rearranged tight junctions. Alternatively, NPCs that have differentiated into a mixture of astrocytes, neurons and oligodendrocytes can be co-cultured with the BMECs to “tune” the model for specific applications.

Improved Method to Produce Brain Microvascular Endothelial Cells for a Robust Human Blood-Brain Barrier Model

UW–Madison researchers have devised a novel method for reproducibly and efficiently growing human BMECs from human pluripotent stem cells (hPSCs), which include both induced pluripotent stem cells (IPSCs) and embryonic stem cells (ESCs). The BMECs have characteristic BBB properties, such as the expression of well-organized tight junctions and high transendothelial resistance. They can be used to create high-fidelity in vitro human BBB models.

The hPSCs are grown on a suitable matrix, such as Matrigel-coated plates, and subjected to an unconditioned medium. After several days, endothelial cell medium is added. Finally, other signals needed for further maturation of the BMECs are provided, and the cells are tested for BBB properties. Flow cytometry may be used to quantify cell development.

Methods for Identifying Neurodegeneration Mutants and Screening Neuroprotective Agents

UW-Madison researchers have developed neurodegeneration mutants of Drosophila and a method for obtaining them. The method starts with a collection of temperature-sensitive paralytic or bang-sensitive paralytic mutants, among which are relatively high numbers of neurodegeneration mutants with altered neural function. The neural function mutants are then screened for age-dependent neurodegeneration.

The researchers used this method to identify several neurodegeneration mutants in Drosophila. Several of the identified mutants have marked neuropathology throughout their entire central nervous systems, which leads to a shortened lifespan. These mutant animals are also particularly well suited for screening assays aimed at identifying neuroprotective agents.

Method for Identifying Genetic Markers of Biological Age and Metabolism

UW-Madison researchers have developed a method of identifying biomarkers of aging. The method involves comparing gene expression profiles from a young subject; a chronologically and biologically older subject; and a biologically younger, calorically restricted subject that is the same chronological age as the older subject. Oligonucleotide arrays may be used to compare the gene expression profiles. Alterations in gene expression that are observed when comparing the young subject to the older one, but that are not observed, or are reduced, when comparing the young subject to the biologically younger, calorically restricted subject are then identified. These gene expression alterations likely represent markers of biological, rather than chronological, age.

A Model for Familial Combined Hyperlipidemia and Type II Diabetes Mellitus

UW-Madison researchers have developed a population of California mice that is an excellent model for familial combined hyperlipidemia and related disorders in humans. They discovered a colony of California mice (Peromyscus californicus) that, like humans with FCH, becomes hyperlipidemic and eventually diabetic when fed a high fat diet. Some California mice also show elevated levels of triglycerides and cholesterol, and develop hyperinsulinemia and insulin resistance after 12 weeks on this diet. Susceptibility to these metabolic abnormalities varies considerably within the population and is passed on to offspring. The researchers plan to selectively breed California mice that show either high or low triglyceride and cholesterol responses to dietary fat.

Genetically Modified Mouse Model Displaying Abnormalities in Circadian Rhythms

UW-Madison researchers have developed a mouse model generated by targeted mutation of a key locus in circadian regulation, the mop3 locus. Mop3 mutant mice display characteristics indicative of complete loss of circadian clock control, including abnormal sleep patterns under conditions of 12:12 hour light:dark cycles, loss of circadian control of activity under conditions of constant darkness and reduced total activity. Other physiological consequences of the mutation include enlarged heart and liver, premature death, abnormal bone growth and decreased fertility in females.

The mop3 mutant mouse is the first to possess a single gene mutation resulting in complete loss of the circadian clock. Analyses of this mouse model also indicate that mop3 lies at the top of the hierarchy for molecular control of circadian rhythm. A surprising outgrowth of this research is that this mouse has also been shown to be a model of a common human arthropathy, known as diffuse skeletal hyperossification (DISH).

Collection of Drosophila Mutants with Neurodegenerative Effects

UW-Madison researchers have compiled a collection of mutant Drosophila with obvious and distinct neurodegeneration.  Each of these mutants has a defect in a single gene.  They could be used to identify novel drug targets for neurodegenerative diseases.

Immortalized Cells from the Mouse Urogenital Sinus and Adult Mouse Prostate

A UW-Madison researcher has developed biological material that includes immortalized murine mesenchymal cells from the urogenital sinus and stromal cells from the lobes of the adult prostate. The cells were isolated from the INK4A transgenic mouse, which lacks genes that normally enforce senescence. A construct that expresses lacZ was included in these cells to allow their identification when they are grafted into mouse tissues.

Mouse Model of Diabetes

Based on this observation, UW-Madison researchers developed a line of TSP1 negative mice that provide an animal model for diabetic retinopathy. They crossed Akita/+ mice that develop diabetes between three and four weeks of age with TSP1 -/- mice. The resulting Akita/+ TSP1 -/- mice develop diabetes-associated vasculopathies of greater severity and at a much earlier stage of diabetes than the Akita/+ mice. These mice also show a dramatic increase in acellular capillaries and saccular microaneurysms, which are some of the early signs of diabetic retinopathy.

Methods and Materials for Assaying Non-SCD1 Isoforms

UW-Madison researchers have developed useful tools for the discovery of SCD inhibitors. The invention consists of cDNAs for murine SCD2 and SCD3 and human SCD5, along with a stable mammalian cell line and yeast strain that express human SCD5. In addition, the invention includes SCD2, and SCD3 knockout and transgenic mice, as well as the targeting constructs used to generate the SCD1 and SCD3 transgenic mice.

Tsp1 -/- Stz Mice, A Model For Diabetes

Based on this observation, UW-Madison researchers have developed a line of TSP1-negative mice that provides an improved animal model for the study of diabetic retinopathy. They induced diabetes in TSP1-/- mice by injecting them with a single dose of streptozotocin to destroy their pancreatic beta cells. The resulting TSP1 -/- mice develop diabetes-associated early vasculopathies of similar severity to those observed in a previous mouse model developed by the inventors, but after a shorter duration of diabetes.

Mouse Model for Mania

A UW–Madison researcher has developed a novel line of mice which exhibit at least four characteristics of mania: hyperactivity, elevated aggression, increased risk taking and decreased sleep.

The line of mice originally was derived from outbred hsd:ICR mice as part of a study of high wheel-running. The researcher now has found that the mice exhibit mania behaviors like decreased daytime sleep and cage hyperactivity. Furthermore, some of these behaviors were tempered following treatment with the common anti-mania drugs valproate and olanzapine, as well as lithium.

Wisconsin Miniature SwineTM for Biomedical Research

UW–Madison researchers have developed a novel line called “Wisconsin Miniature Swine,” or WMS, that is well suited to biomedical research and genetic modeling of human diseases. The animals’ body weight, size and composition are similar to humans and can be manipulated easily. For example, on an unrestricted feeding regimen, WMS become obese and appear to develop the hallmarks of metabolic syndrome.

Ringworm Fungal Strain for Infection Studies

UW–Madison researchers have established that a strain of M. canis, called UW-8, is particularly useful in experimental infection studies. The strain produces a uniquely bright and consistent fluorescence that makes ringworm lesions much easier to identify and measure.

The researchers have used UW-8 for infection studies since 1993, and the attributes of the strain are supported by a body of peer-reviewed literature. UW-8 was selected from natural field strains and can be cultured readily.

Wisconsin Miniature SwineTM for Cardiovascular Research

UW–Madison researchers have developed a novel line called Wisconsin Miniature Swine – Familial Hypercholesterolemia (WMS-FH). The animals are hypercholesterolemic and develop atherosclerotic vascular disease that is remarkably similar to that of humans.

The body weight, size and composition of WMS-FH are similar to humans and can be manipulated easily. They are ideally suited for studying the mechanisms of cardiovascular disease, and for developing/validating diagnostic and therapeutic technologies in the field of cardiology.

Animal Model of Brittle Bone Disease

A UW–Madison researcher and collaborator have developed a mouse strain that is conditionally null for BMP1 and mTLL1. The model exhibits the hallmark features of OI and provides a new means to screen drugs and study mechanism.

The mice exhibit dramatically weakened and brittle bone with spontaneous fractures – defining features of OI. Additional skeletal features include osteomalacia, thinned/porous cortical bone, reduced processing of procollagen and dentin matrix protein 1, remarkably high bone turnover and defective osteocyte maturation.

WARF also owns and can license one of the parental lines (a mouse strain with a floxed TLL1 allele).

Collection of Temperature-Sensitive Paralytic Mutants of Drosophila

UW-Madison researchers have compiled several temperature-sensitive paralytic mutants of Drosophila. The collection consists of more than 100 different neurological mutants that are behaviorally normal at 25 degrees C, but display severe locomotor defects, including uncoordinated movement, ataxia, seizures or complete paralysis, within five minutes of exposure to 38 degrees C. The phenotypes are reversible upon return to 25 degrees. Kinetics of paralysis and recovery vary from strain to strain, but are characteristic to any particular mutant.

Genes affected by these mutations include those encoding ion channel subunits, ion channel regulators, components of the synaptic release machinery and other proteins required for proper neuronal signaling, viability and development. Most of the mutations were induced by ethylmethane sulfonate, a chemical mutagen; others were generated by transposable-element mutagenesis.