School of Medicine, Department of Genetics
Institute for Diabetes, Obesity and Metabolism
Dr. Kaestner’s lab is employing modern genetic, genomic and epigenomic
approaches (ChIP-Seq, RNA-Seq, gene targeting, tissue-specific and
inducible gene ablation) to understand the molecular mechanisms of
organogenesis and physiology of the liver, pancreas and gastrointestinal
tract. Disease areas targeted by our research include diabetes and
Epigenomic rejuvenation of human pancreatic beta-cells.
The prevalence of Diabetes Mellitus has reached epidemic proportions world-wide, and is predicted to increase rapidly in the years to come, putting a tremendous strain on health care budgets in both developed and developing countries. There are two major forms of diabetes and both are associated with decreased beta-cell mass. No treatments have been devised that increase beta-cell mass in vivo in humans, and transplantation of beta-cells is extremely limited due to lack of appropriate donors. For these reasons, increasing functional beta-cell mass in vitro, or in vivo prior to or after transplantation, has become a “Holy Grail” of diabetes research. Our previous studies clearly show that adult human beta-cells can be induced to replicate, and – importantly - that cells can maintain normal glucose responsiveness after cell division. However, the replication rate achieved was still low, likely due in part to the known age-related decline in the ability of the beta-cell to replicate. We propose to build on our previous findings and to develop more efficacious methods to increase functional beta-cell mass by inducing replication of adult beta-cells, and by restoring juvenile functional properties to aged beta-cells. We will focus on mechanisms derived from studies of non-neoplastic human disease as well as age-related phenotypic changes in human beta-cells.
1, we will target the genes altered in patients with marked beta-cell
hyperplasia, such as those suffering from Beckwith-Wiedemann Syndrome or
Multiple Endocrine Neoplasia. Expression of these genes will be altered
in human beta-cells via shRNA-mediated gene suppression and
locus-specific epigenetic targeting. Success will be assessed in
transplanted human islets by determination of beta-cell replication rate
and retention of function. In Aim 2, we will determine the mechanisms
of age-related decline in beta-cell function and replicative capacity,
by mapping the changes in the beta-cell epigenome that occur with age.
Selected genes will then be targeted as in Aim 1 to improve human
beta-cell function, as assessed by glucose responsiveness. To accomplish
these aims, we will use cutting-edge and emerging technologies that are
already established or are being developed in our laboratories. The
research team combines clinical experience with expertise in molecular
biology and extensive experience in genomic modification aimed at
enhancing beta-cell replication. By basing interventions on changes
found in human disease and normal aging, this approach will increase the
chances that discoveries made can be translated more rapidly into
clinically relevant protocols.
Regulatory cascades in differentiation and proliferation of the gastrointestinal epithelium.
The mammalian gut epithelium is a highly organized and dynamic system which requires continuous controlled proliferation and differentiation throughout life. Proliferation, cell migration and cell adhesion all must be tightly controlled in order to prevent either inflammatory diseases or epithelial cancers. As with many other vertebrate organs, the digestive tract develops from heterogeneous embryonic origins. While the musculature and the connective tissue are derived from lateral plate mesoderm, the epithelium is derived from the endoderm. We have identified a novel member of the winged helix gene family termed Foxl1 which is expressed in the gut mesoderm and have begun its functional analysis in vivo through targeted mutagenesis in mice. Null mutations in the mesodermal transcription factor Foxl1 result in dramatic alterations in endoderm development, including epithelial hyperproliferation. We have now identified APC/Min and GKLF as downstream targets of Foxl1 and have begun the analysis of these genes in gastrointestinal differentiation by tissue-specific gene ablation.
Innovative Genetic Approaches for Hepatic Repopulation
A better understanding of the liver’s response to toxic injury, which includes hepatocyte proliferation, activation and differentiation of facultative hepatic stem cells (“oval cells”), and – unfortunately – an increased risk for hepatocellular carcinoma, is a prerequisite for the development of novel clinical treatments for chronic liver disease and improved cancer prevention. Likewise, cell replacement therapy, either through direct hepatocyte transplantation or in bio-artificial liver devices, needs to be improved in order to become a reliable alternative to liver transplantation. To date, investigations of hepatocyte proliferation have frequently focused on the partial hepatectomy paradigm, a “non¬injury” model that is not reflective of liver injury in humans and which has therefore failed to identify specific targets for either improved regeneration following toxic injury or for limiting proliferation in HCC in humans.
In Specific Aim 1, we will determine
which genes and gene combinations promote or repress hepatocyte
repopulation following toxic liver injury using an innovative genetic
approach. In Specific Aim 2, we will employ expression of key hepatic
transcription factors to improve the differentiation of hepatic
progenitor cells to functional hepatocytes. Together, these approaches
will provide an improved understanding of the liver’s response to toxic
injury, and facilitate the discovery of new cell replacement therapies
to treat chronic liver disease and liver failure.