Li Lab Linheng Li, Ph.D. Investigator lil@stowers-institute.org

     Stem cells are the key subset of cells in the body functioning as ancestor cells to produce a variety of types of functionally specialized mature (differentiated) cells in a given tissue, while at the same time maintaining the capacity to continuously divide and reproduce themselves (self-renewal). This self-renewal process is controlled by intrinsic genetic pathways that are subject to regulation by extrinsic signals from the microenvironment in which stem cells reside. Stem cells play essential roles ranging from embryonic development and organogenesis (fetal stem cells and embryonic stem cells) to tissue homeostasis and regeneration (adult stem cells). Stem cell development is a complex process, and a precise balance is maintained among different cell events including self-renewal, differentiation, apoptosis (cell death), and migration. Loss of this balance tends to lead to uncontrolled cell growth or cell death, thereby developing into a variety of diseases including cancer or tissue defects.

     We mainly focus on two systems to study stem cell development: hematopoietic and intestinal stem cell compartments. The hematopoietic system facilitates functional characterization of stem cells, as bone marrow transplantation experiments can be readily performed. The intestinal system has a well-organized developmental architecture in which stem cell marking and lineage tracing can be used to investigate how stem cells are maintained by their microenvironment (niche), how stem cells undergo asymmetric division to keep the balance between self-renewal and lineage commitment, and what molecular signals are involved in this regulation. To investigate the molecular mechanisms that control stem cell properties, we use the combined approaches described as follows:

• A global view of the changes in the gene expression patterns during hematopoietic stem cell development to reveal important pathways regulating hematopoietic stem cell self-renewal and lineage commitment. The Notch, Wnt, BMP, and PTEN signal pathways have been well documented to be involved in developmental regulation and tumorigenesis. Expression of Notch, BMP4, and ß-Catenin (a transcription factor) in hematopoietic stem cells suggests that these pathways may play important roles in the regulation of hematopoietic stem cell proliferation and differentiation.
• To further characterize the functions of these pathways, we use genetic approaches such as transgenic or gene targeting animal models to examine their influence on stem cell development. Our goal is to understand how these signal pathways or mechanisms regulate normal development in the hematopoietic and intestinal systems. This information should reveal how they may malfunction or be altered in association with human diseases such as leukemia and colon cancer.

1. Molecular basis of multipotentiality of stem cells
     What determines the multipotentiality of stem cells is a fundamental question. Analysis of the gene expression profiles during early hematopoietic stem cell (HSC) development revealed that the step-wise decrease in promiscuity (diversity) for multiple lineage-affiliated genes correlates with a progressive restriction of developmental potential in early hematopoiesis. These results support the hypothesis that stem cells maintain their multipotentiality via a wide-open chromatin structure (Blood 2003).

2. Identification of the hematopoietic stem cell niche
     Schofield first proposed the hypothesis in 1978 that the niche (the cellular components of the microenvironment) plays an essential role in the maintenance of HSCs; however, the HSC niche has been a mystery since then. Using the Bmpr1a conditional knock-out (KO) model, we have identified that spindle-shaped N-cadherin+ osteoblastic cells are a key component of the HSC niche. This is the first stem cell niche in the mammalian system to be identified at the cellular level. As an increase in the HSC number resulted from an increase in the size of the HSC niche in Bmpr1a mutant mice, BMP signaling controls the HSC number via regulation of the niche size (Nature 2003). The clinical implication of this discovery is the potential of maintaining and expanding hematopoietic stem cells in vitro.

3. BMP and Wnt signaling yin-yang controls intestinal stem cell (ISC) properties
     The molecular mechanism underlying juvenile polyposis syndrome (JPS) caused by defects in bone morphogenetic protein receptor-IA (BMPR1A) in humans remains largely unknown. Inactivation of Bmpr1a in mice resulted in JPS. We found that the BMP and Wnt signal pathways are antagonistic, ensuring the balanced (yin-yang) control of stem cell self-renewal and proliferation versus differentiation. We further propose that BMP signaling inhibits Wnt signaling through either Smad-dependent transcriptional repression or inhibiting β-catenin activity via a PTEN-PI3K/Akt pathway (Nature Genetics 2004).

4. Normal stem cells versus cancer stem cells
      In the studies of the Bmpr1a mutant mouse model, we found that the PTEN-controlled PI3K-Akt pathway may mediate a cross talk between BMP and Wnt signaling pathways. We therefore examined the consequences of inactivation of PTEN (a tumor suppressor) in both hematopoietic and intestinal systems. Loss of PTEN leads to enhanced proliferation and mobilization of stem cells, which in turn result in acute myeloid/lymphoid leukemia and intestinal polyposis, respectively. Mechanistic studies of the PTEN deficient mouse model revealed that PTEN plays a critical role in maintaining normal hematopoietic stem cells and preventing leukemia development (Nature 2006); and loss of PTEN results in conversion of normal stem cells into cancer initiating/stem cells. We documented the process of how cancer stem cells initiate tumorigenesis in intestinal polyposis (Nature Genetics 2007).

5. Discovery of two subpopulations of reserved and primed hematopoietic stem cells
     To sustain blood production over a lifetime, primitive hematopoietic stem cells (HSCs) must support daily regeneration of lost cells and must also avoid becoming depleted or exhausted. We have shown how HSCs may balance distinct and sometimes opposing needs. We found that the primitive HSC pool contains two subpopulations distinguished by the expression level of N-cadherin, an adhesion molecule thought to mediate HSC-niche interaction. Low levels of N-cadherin identify HSCs in a “primed” state ready to support active blood regeneration. Cells with intermediate levels form a larger pool of “reserved” HSCs adapted to a maintenance role.

Future directions

      We are extending our finding of the HSC niche, attempting to further dissect the niche signals, and investigating how these signals coordinate to regulate stem cell self-renewal. We also are attempting to systematically study the distinct and complementary functions of different types of niches in regulation of HSCs. We will map out the different microenvironments required for different stages of hematopoietic lineage commitment and maturation.

     As cancer is derived from cancer stem cells, a key to completely curing cancer may depend on whether or not we can successfully target cancer stem cells (not just the bulk of the tumor mass). To this end, identification and characterization of cancer stem cells will be the essential step. We are engaged in identifying cancer stem cells in hematopoietic and intestinal tissues, followed by characterization of cancer stem cells at cellular and molecular levels.

     Academic Appointment: Associate Professor, Department of Pathology & Laboratory Medicine, The University of Kansas School of Medicine