Myocardial infarction is the most common cause of heart failure and remains one of the leading causes of morbidity and mortality in humans. Stem cells are important in the maintenance and repair of adult tissues. Hoechst effluxing cells, termed side population cells are a rare subset of cells found in adult tissues that are highly enriched for stem and progenitor cell activity. Recent studies have suggested that Sca-1+/CD31- cardiac side population cells are capable of differentiation into cardiomyocytes in vitro. However, the response of cardiac side population cells to myocardial injury remains unknown in vivo. In this study, we directly transplanted Sca-1+/CD31- cardiac side population cells into an acutely infarcted mouse heart. After two weeks, the transplanted cells were found to express cardiomyocyte or endothelial cell markers. Importantly, when these cells were transplanted into a remote nonischemic part of the heart after MI, they were able to migrate to the damaged myocardium. Consistent with these cells homing property, we found that SDF-1α, a chemotactic chemokine and its receptor, CXCR4 were up-regulated in the damaged myocardium and on Sca-1+/CD31- cardiac SP cells respectively following an acute myocardial infarction. We further showed that SDF-1α was able to induce migration of Sca-1+/CD31- cardiac side population cells in vitro. Our results have therefore suggested that Sca-1+/CD31- cardiac side population cells are able to migrate to damaged myocardium from non-ischemic myocardium and differentiate into cardiomyocytes as well as endothelial cells in the acutely infarcted mouse heart. We postulate that the SDF-1α/CXCR4 interaction may play an important role in the migration of these cells. Understanding and enhancing these processes may hold enormous potential possibilities for therapeutic myocardial regeneration for the treatment of cardiovascular disease.

Thesis (M. D.)--University of Hong Kong, 2006. / Title proper from title frame. Also available in printed format.

Unlike the central nervous system, axonal regeneration does occur in the peripheral nervous system, however, despite this, functional recovery from nerve transection injury remains dismal. This has been attributed to factors intrinsic to the motor or sensory cell body and to elements in the local site of injury including nerve gaps, scar and a limited time frame in which supportive growth factors and extracellular matrix molecules are expressed. The aim of this thesis is to review the mechanisms behind axonal damage and regeneration in the peripheral nervous system and investigate surgical, pharmacological and biological approaches to overcome limitations in regeneration and functional recovery. By taking a broad approach to the topic, I hoped to gain a greater understanding of the inhibitory and regenerative processes at play and provide a contribution to the understanding in the field of peripheral nerve surgery. / Experimental Surgery

Identifying the mechanisms that regulate neural precursor cell (NPC) proliferation and differentiation is important for understanding CNS development among different vertebrates. My work has focused specifically on understanding how mitogenic factors, like basic fibroblast growth factor (FGF2), regulate the NPC cell cycle. Mitogenic factors and serum are thought to drive cell cycle and therefore proliferation mainly by activating G1-type cyclin-dependent kinases (CDKs). The general hypothesis being addressed here is that FGF2 also promotes cell cycle progression of NPCs through the degradation of the cell cycle inhibitor p27Kip1. I show that, in the presence of FGF2 in vitro, embryonic rat cortical NPCs express high protein levels of the CDC28 protein kinase regulatory subunit 1b (Cks1b), a component of the SCFSkp2 E3 ubiquitin ligase complex that targets p27Kip1 for proteasomal degradation. I also show that NPCs maintained in FGF2 express undetectable levels of p27KIP1, while removal of FGF2 results in increased p27Kip1 protein expression and decreased protein expression of Cks1b. RNA expression data shows that Cks1b mRNA is reduced in non-dividing NPCs but is present in dividing NPCs, suggesting that Cks1b is being regulated at the transcriptional level. Analysis of the putative promoter of Cks1b reveals numerous conserved transcription factor consensus sites that could potentially play a role in regulation of Cks1b transcription, including consensus sites for E2F and the cell cycle-dependent element (CDE) cell cycle genes homology region (CHR) tandem repressor element. I use chromatin immunoprecipitation and luciferase assays to identify which E2Fs occupy and regulate the transcription of Cks1b under different conditions of mitogen stimulation. The data show that E2F4 occupies the promoter of Cks1b in non-dividing NPCs while E2F1 binds exclusively in proliferating NPCs. Mutation of either the E2F or CDE/CHR consensus sites independently de-represses the activity of a Cks1b promoter reporter in NPCs in G0/G1, while mutation of both sites delays induction of promoter activity. Finally, I use in ovo electroporation to determine if p27Kip1 has an additional role in neuronal differentiation during early spinal cord development. I show that ectopic expression of p27Kip1 is insufficient to induce neuronal differentiation in spinal cord progenitors.

The purpose of this study was to develop a model describing the relationship of inquiry-based teaching elements on middle school students’ science, technology, engineering, and mathematics (STEM) interests and belief changes. The study utilized pretest/posttest, correlational, and longitudinal designs. Classroom inquiry data (N = 139) and middle school students’ attitudinal data (N = 1779) were collected in middle school classrooms within a 40 mile radius of Texas A&M University during the 2004-2005 and 2005-2006 school years. Results indicated 24% of the variation in middle school students’ change in science, technology, engineering, and mathematics (STEM) interests was explained by the inquiry-learning element “teacher as listener” was very characteristic of this classroom.” STEM interest change explained 55% of the variation in middle school students’ STEM belief change. Analyses indicated NSF Fellows and teachers affected the rate at which middle school students’ STEM beliefs and interests changed. Middle school students’ STEM interests and beliefs remained significantly unchanged from pre- to post-NSF Fellow each year of the study. Classroom inquiry levels did significantly increase from beginning of school-year to end of school-year each year of the project. NSF Fellows had a positive relationship with the one inquiry element “teacher as listener” was very characteristic of the classroom; which explained middle school students’ change in STEM interests. NSF Fellows had negative relationships with the inquiry elements, lessons involved fundamental concepts of the subject; lessons were designed to engage students as members of a learning community; lessons promoted strong conceptual understanding; and elements of abstraction were encouraged when it was important to do so. No inquiry elements were associated with middle school students’ change in STEM beliefs. Middle school students’ change in STEM interests were positively associated with three inquiry elements, “teacher as listener” was very characteristic of the classroom; students were involved in the communication of their ideas to others using a variety of means and media; and student questions and comments often determine the focus and direction of classroom discourse. The inquiry element, instructional strategies and activities respected students’ prior knowledge and the preconceptions inherent therein, was negatively associated with changes in middle school students’ STEM interests.

In recent years, research in the area of cancer stem cells has spiked tremendously. Numerous investigators have found that several types of cancers contain a subpopulation of tumor cells that display many defining characteristics of normal tissue stem cells, including multipotent differentiation potential, long-term self-renewal capacity, and expression of molecular markers of stemness. Most importantly, these cancer stem cells (CSCs) have very high tumor initiating potential, a finding that has led to the development of the cancer stem cell model for tumor progression. This model suggests that tumors are organized in a developmental hierarchy (similar to healthy tissue), with long-term tumor progression being driven by self-renewing CSCs at the top of the hierarchy. The CSC model represents a significant shift in our understanding of tumor progression, and as such, it may be possible to expand our knowledge of other aspects of tumor biology by re-examining them in the context of the CSC model. My work focuses on investigating interactions between CSCs and the tumor vasculature. Previous work has demonstrated heterogeneity in the proangiogenic potential of cells in a tumor. Considering the possibility that angiogenesis may be driven by specific subsets of tumor cells, I investigated the contribution of the CSC fraction to tumor angiogenesis. Comparing tumors with low or high CSC fractions, I have found that CSCs contribute to tumor vascular development through promotion of endothelial cell activity and recruitment of bone marrow-derived proangiogenic cells, mediated in part by vascular endothelial growth factor (VEGF) and stromal-derived factor 1 (SDF1). Since some tissue stem cells are known to reside in a vascular niche, I investigated the possibility that CSCs may also be supported by blood vessels in the tumor microenvironment, and that consequently CSCs may be targeted by disruption of tumor vasculature with antiangiogenic therapy. By testing multiple antiangiogenic therapeutic strategies, I have found that antiangiogenic therapy sensitizes CSCs to the effects of cytotoxic chemotherapy. Taken together, my work demonstrates a bi-directional relationship in which CSCs promote tumor vascular development, and tumor vasculature supports and protects CSCs. This work has implications for our understanding of CSC biology, tumor angiogenesis and antiangiogenic therapy, and provides insight into strategies for targeting the critical CSC population.

Skin-derived precursors (SKPs) are multipotent dermal stem cells that reside within a hair follicle niche and that share properties with embryonic neural crest precursors. Here, we have asked whether SKPs and their endogenous dermal precursors originate from the neural crest or whether, like the dermis itself, they originate from multiple developmental origins. To do this, we used two different mouse Cre lines that allow us to perform lineage tracing: Wnt1-cre, which targets cells deriving from the neural crest, and Myf5-cre, which targets cells of a somite origin. By crossing these Cre lines to reporter mice, we show that the endogenous follicle-associated dermal precursors in the face derive from the neural crest, and those in the dorsal trunk derive from the somites, as do the SKPs they generate. Despite these different developmental origins, SKPs from these two locations are functionally similar, even with regard to their ability to differentiate into Schwann cells, a cell type only thought to be generated from the neural crest. Analysis of global gene expression using microarrays confirmed that facial and dorsal SKPs exhibit a very high degree of similarity, and that they are also very similar to SKPs derived from ventral dermis, which has a lateral plate origin. However, these developmentally distinct SKPs also retain differential expression of a small number of genes that reflect their developmental origins. Thus, an adult neural crest-like dermal precursor can be generated from a non-neural crest origin, a finding with broad implications for the many neuroendocrine cells in the body.

In recent years, research in the area of cancer stem cells has spiked tremendously. Numerous investigators have found that several types of cancers contain a subpopulation of tumor cells that display many defining characteristics of normal tissue stem cells, including multipotent differentiation potential, long-term self-renewal capacity, and expression of molecular markers of stemness. Most importantly, these cancer stem cells (CSCs) have very high tumor initiating potential, a finding that has led to the development of the cancer stem cell model for tumor progression. This model suggests that tumors are organized in a developmental hierarchy (similar to healthy tissue), with long-term tumor progression being driven by self-renewing CSCs at the top of the hierarchy. The CSC model represents a significant shift in our understanding of tumor progression, and as such, it may be possible to expand our knowledge of other aspects of tumor biology by re-examining them in the context of the CSC model. My work focuses on investigating interactions between CSCs and the tumor vasculature. Previous work has demonstrated heterogeneity in the proangiogenic potential of cells in a tumor. Considering the possibility that angiogenesis may be driven by specific subsets of tumor cells, I investigated the contribution of the CSC fraction to tumor angiogenesis. Comparing tumors with low or high CSC fractions, I have found that CSCs contribute to tumor vascular development through promotion of endothelial cell activity and recruitment of bone marrow-derived proangiogenic cells, mediated in part by vascular endothelial growth factor (VEGF) and stromal-derived factor 1 (SDF1). Since some tissue stem cells are known to reside in a vascular niche, I investigated the possibility that CSCs may also be supported by blood vessels in the tumor microenvironment, and that consequently CSCs may be targeted by disruption of tumor vasculature with antiangiogenic therapy. By testing multiple antiangiogenic therapeutic strategies, I have found that antiangiogenic therapy sensitizes CSCs to the effects of cytotoxic chemotherapy. Taken together, my work demonstrates a bi-directional relationship in which CSCs promote tumor vascular development, and tumor vasculature supports and protects CSCs. This work has implications for our understanding of CSC biology, tumor angiogenesis and antiangiogenic therapy, and provides insight into strategies for targeting the critical CSC population.

The purpose of this study was to develop a model describing the relationship of inquiry-based teaching elements on middle school students’ science, technology, engineering, and mathematics (STEM) interests and belief changes. The study utilized pretest/posttest, correlational, and longitudinal designs. Classroom inquiry data (N = 139) and middle school students’ attitudinal data (N = 1779) were collected in middle school classrooms within a 40 mile radius of Texas A&M University during the 2004-2005 and 2005-2006 school years. Results indicated 24% of the variation in middle school students’ change in science, technology, engineering, and mathematics (STEM) interests was explained by the inquiry-learning element “teacher as listener” was very characteristic of this classroom.” STEM interest change explained 55% of the variation in middle school students’ STEM belief change. Analyses indicated NSF Fellows and teachers affected the rate at which middle school students’ STEM beliefs and interests changed. Middle school students’ STEM interests and beliefs remained significantly unchanged from pre- to post-NSF Fellow each year of the study. Classroom inquiry levels did significantly increase from beginning of school-year to end of school-year each year of the project. NSF Fellows had a positive relationship with the one inquiry element “teacher as listener” was very characteristic of the classroom; which explained middle school students’ change in STEM interests. NSF Fellows had negative relationships with the inquiry elements, lessons involved fundamental concepts of the subject; lessons were designed to engage students as members of a learning community; lessons promoted strong conceptual understanding; and elements of abstraction were encouraged when it was important to do so. No inquiry elements were associated with middle school students’ change in STEM beliefs. Middle school students’ change in STEM interests were positively associated with three inquiry elements, “teacher as listener” was very characteristic of the classroom; students were involved in the communication of their ideas to others using a variety of means and media; and student questions and comments often determine the focus and direction of classroom discourse. The inquiry element, instructional strategies and activities respected students’ prior knowledge and the preconceptions inherent therein, was negatively associated with changes in middle school students’ STEM interests.

Abstract Recent studies demonstrated that LPS treated Sprague-Dawley rats induced a reduction (Phase I), followed by an augmentation (Phase II) and a secondary decrease (Phase III) in the power density of vasomotor components (0-0.8 Hz) in systemic arterial pressure signals. The molecular mechanisms underlie the progression toward death in the brain stem is unclear. In order to find out the differentially expressed genes between LPS-treated RVLM and saline-treated RVLM, we used suppression subtractive hybridization,a method commonly used to search differentially expressed genes, and subtractive cDNA library construction. At present, we have found some differentially expressed genes and these genes are up-regulation expression. These genes may be involved in the progression toward death in the rat brain stem.