Stem cells transplanted into the brain are immunogenic and are actively rejected by the host [abstract]

Abstract only availableStem cell-based therapies have shown great promise in the treatment of neurodegenerative diseases such as Batten's Disease and Parkinson's Disease. Intracranial stem cell transplantation has the potential to restore function and compensate for neural cells lost due to injury or disease. However, rejection of donor cells by the host immune system may limit the effectiveness of stem cell therapies. Recent research has suggested that some stem cells may be immunoprivileged, able to avoid rejection by the host's immune system in both allogeneic and xenogeneic settings. This can occur regardless of an MHC mismatch. However, several aspects of these studies complicate interpretations of their results. Transplant recipients are often irradiated before the transplant (Drukker et al. 2006) or otherwise immunocompromised and studies often culminate in a time period insufficient for immune rejection to have occurred (under a week) (Li et al. 2004). To evaluate whether stem cells are immunoprivileged when transplanted into the brain, GFP-expressing neural stem cells (NSCs) were transplanted into the brains of immunocompetent, immunologically mismatched mice. Mice were then sacrificed at time points of one, three, five, seven and nine days posttransplantation. The brains were fixed, freeze-embedded with OCT and sectioned. Graft survival was evaluated by observing the amount of GFP-expressing donor cells in the sections. Sections were also immunolabeled for cells expressing CD4, CD8 and CD11b, all of which are markers for infiltrating immune cells. Presence of such cells indicates immune detection and rejection and can be used to quantify the immune response to foreign stem cells. In future studies, the use of Regulatory T Cells may help to alleviate this rejection by suppressing the activity of CD8 (cytotoxic) and CD4 (helper) T Cells.National Institutes of Healt

The effect of Camgaroo-2 incorporation on the differentiation potential of embryonic stem cells

Abstract only availableEmbryonic stem (ES) cells are capable of differentiating into any cell type in the body and are a promising therapeutic agent. Our research focuses on the differentiation of ES cells into functional neurons and/or glial that can nurture host cells of the nervous system that are damaged due to disease. Cells must express the appropriate phenotype and perform the proper function after transplantation. Camgaroo-2 is a fluorescence protein that provides a basal fluorescence and responds to a rise in intracellular calcium by producing an increase in fluorescence emission. Our lab transfected a mouse ES cell line (GSI-1) with the Camgaroo-2 gene and is testing this fluorescence indicator to determine the physiological function of cells grown in vitro. There is concern that the incorporation of the Camgaroo-2 gene could alter the cell phenotype, potentially decreasing their differentiation potential. GSI-1 cells were plated on culture slides following a neural induction protocol that uses retinoic acid and allowed to proliferate. Immunohistochemisty of slides was performed to label for neural precursors, immature and mature neurons, astrocytes, and oligodendrocytes (anti-O4). GSI-1 labeling was compared to corresponding immunohistochemistry performed on another ES cell line that had also been 'neuralized' to determine if the differentiation potential of the GSI-1 cells was similar to that of the other ES cell line. Similar labeling was seen for all markers except O4 which did not label for the GSI-1 cells, indicating the GSI-1 cells have the potential to differentiate into all cells of neural lineage except possibly oligodendrocytes. GSI-1 cells retained the ability to differentiate post-transfection with the Camgaroo-2 gene. Because of their unique ability to respond to an influx of intracellular calcium, GSI-1 cells expressing Camgaroo-2 can be transplanted into rodent models for human disease, and can be tested post transplantation for their ability to function as neural cells.Life Sciences Undergraduate Research Opportunity Progra

Camgaroo-2 as an indicator of function in embryonic and neuralized stem cells

Abstract only availableThe transplantation of stem cells to replace cells that have been lost or damaged due to disease or injury is quickly becoming a conceivable treatment method. Embryonic stem (ES) cells have the capacity to become any cell in the body, so the therapeutic possibilities are vast. The ultimate goal of our research on ES cells is to induce them to differentiate into functioning neurons to replace those that are lost in patients suffering from neurodegenerative disorders. However, it is important that the differentiated cells possess the appropriate phenotype and are able to perform the correct function after transplantation. In the past, it was common to accept a differentiated cell's fate based solely on its morphology and the presence of specific membrane markers. Now, it is becoming increasingly important to determine a donor stem cell's fate based on its function, especially if the cell is to be transplanted into a subject as a means of therapy. This study used the calcium-sensitive protein Camgaroo-2 to test the function of embryonic stem cells and cells directed toward a neural lineage. Camgaroo-2 is a fusion protein that consists of calmodulin in between two halves of yellow fluorescent protein. When calcium is present, it binds to the calmodulin portion of the Camgaroo-2, inducing a conformational change that results in increased fluorescence. After mouse embryonic stem cells were transfected with Camgaroo-2, we used reagents such as potassium chloride and ionomycin, known to elevate intracellular calcium, to confirm that the ES cells were stably transfected with the plasmid, and that Camgaroo-2 was functioning correctly. Potassium chloride causes the cell to depolarize while ionomycin (a calcium ionophore) creates large pores in the cell membrane. Both reagents allow for an influx of calcium into the cell, leading to increased fluorescence. The Camgaroo-2 transfected ES cells showed the appropriate responses to KCl and ionomycin by depolarizing and showing visible increases in fluorescence. This confirms that our Camgaroo-2 construct is functioning in the ES cells. We are in the process of testing the responses of neuralized ES cells using appropriate neurotransmitters, the presence of which should induce unique fluorescent signatures in a cell specific manner. Confirming neuronal function from differentiated Camgaroo-2 ES cells is an important step toward neuron transplantation in a neurodegenerative disease model.Arts & Sciences Undergraduate Research Mentorship Progra

Potential Role for Programmed Cell Death in the Formation of an In Vitro Neural Stem Cell Niche [abstract]

Abstract only availableFaculty Mentor: Dr. Mark Kirk, Biological SciencesStem cell therapies have the potential to treat neurodegenerative diseases, such as Batten Disease. Batten disease, a rare inherited disease in children, causes severe neurodegeneratoin, which results in blindness, seizures, and eventual death. In Batten disease, the transplantation of stem cells into a patient may replace lost cells or to prevent cell loss due to the disease. In one form of Batten Disease, transplantation of stem cells into the retinas of mutant model mice have shown signs of neuroprotection; including enhanced survival of photoreceptors (Meyer et al. 2006). One possible method to increase the efficiency of this treatment is the transplantation of a functional unit capable of producing its own neural precursors "on demand". Such a structure, known as a neural stem cell (NSC) niche, can be found in two small areas in the brain of mammals, and is the center for adult neurogenesis throughout the lives of these mammals. In our lab, we have developed a way to produce a NSC niche in vitro from neuralized mouse embryonic stem cells. To test how this structure is formed and maintained, I am investigating cell death within this in vitro NSC niche-like structure. I performed two different tests for apoptosis, or programmed cell death, Trypan Blue exclusion and TUNEL. Trypan Blue shows membrane permeability; if cells turn blue it is indicative of a late stage in the apoptotic process. In the TUNEL assay, nicked ends of DNA are labeled, an indication of early stage apoptosis. I also tested the affects of induced cell death on the advancement of in vitro niche formation. We hope that this information will lead to a better understanding of how the in vitro niche forms.College of Arts and Science Undergraduate Research Mentorship Progra

Characterization of gene and protein marker expression by human dental pulp stem cells

Abstract only availableNeurodegenerative diseases result from deterioration of neurons or their myelin sheath that over time leads to brain dysfunction and premature death. Cells of the brain and spinal cord do not readily regenerate therefore excessive damage can be devastating. Our lab focuses on stem cell-based therapies for brain disorders. Previous studies indicate the potential of stem cells for use in therapies to treat neurodegenerative disorders. In particular, my lab project deals with dental pulp mesenchymal stem cells (DPMSCs) that are currently being investigated due to their ability to differentiate into multiple cell types, including neural cells. Our DPMSCs are composed of populations of mesenchymal stem cells harvested from normal human third molars (wisdom teeth). The initial goal of my research is to assess the variation of marker expression by the dental pulp mesenchymal stem cells to describe their developmental potentials, particularly neuronal development since neurons are the functional unit of the brain. Our results identified expression of neuronal-specific markers (indicative of neuronal precursors and mature neurons) at the gene and protein level by the DPMSCs specifically, we observed expression of nestin, β-III tubulin, and GFAP as well as the MSC markers CD 90, CD 73 and CD 44. Based on these findings, we propose that human DPMSCs may possess the capabilities necessary for therapeutic treatment of neurodegenerative disorders. In future experiments, we plan to perform cell transplantations into mouse models with neurodegenerative disorders. Our results are very significant because they could lead to cures for serious CNS disorders.NSF-REU Program in Biological Sciences & Biochemistr

Near-universal marriage, early childbearing, and low fertility: India's alternative fertility transition

Objective: To compare fertility in India to both low-to-middle-income and high-income countries (LMICs and HICs) and describe the patterns that have accompanied India's transition to low fertility. Methods: We use data from the Demographic and Health Surveys (DHS), the United Nations (UN), and the Organisation for Economic Co-operation and Development (OECD) to observe factors associated with fertility decline in 36 Indian states and 76 countries. Results: Although fertility in India has declined to levels similar to HICs, women's entry into marriage and initiation of childbearing are more in line with patterns found in LMICs. The vast majority of women in India (97Š) are married by age 30, and their average age at first birth is only 21.3 years old. In spite of these patterns, average fertility has declined in India as a result of earlier termination of childbearing. Among more recent cohorts, fewer women progressed to higher-order births and about half of women obtained a sterilization by age 35. Conclusions: India has reached low fertility by mechanisms outside the traditional indicators of fertility decline. In contrast to countries that have achieved low fertility through delayed age at first birth, women in India have continued to enter unions and bear children early, lowered their age at last birth, and increasingly ended their fertility via sterilization following the birth of two children. Contribution: Evidence from India reveals an alternative pathway to low fertility, highlighting the limitations of traditional socioeconomic indicators for explaining fertility decline

Immunogenic properties of neuralized embryonic stem cells in a model of allogenic intracranial transplantation

Title from PDF of title page (University of Missouri--Columbia, viewed on May 31, 2012).The entire thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file; a non-technical public abstract appears in the public.pdf file.Dissertation advisor: Dr. Mark D. KirkVita.Includes bibliographical references.Ph. D. University of Missouri-Columbia 2011."December 2011"[ACCESS RESTRICTED TO THE UNIVERSITY OF MISSOURI AT REQUEST OF AUTHOR.] Intracranial stem cell transplantation has may restore central nervous system (CNS) function and compensate for neural cell loss. However, immunological rejection of stem cells can limit the effectiveness of such therapy. Neuralized embryonic stem cells (nESCs) were grafted into the striatum of murine hosts and monitored from 3 days to 6 weeks post-transplantation. Allogeneic nESCs were rapidly rejected brain grafts over the course of 7 days, while syngeneic grafts persisted for greater than 14 days. Our results indicate that rejection of nESCs occurs within the CNS, and demonstrates that intraparenchymal antigens can recruit a systemic immune response; the adaptive immune system is strongly implicated in the destruction of graft tissue. We also describe a novel NSC culture system with the potential for intracranial transplantation, derived from ES cells, that maintains and expands a population of neural stem cells similar to those in the developing and adult brain. FLOW cytometry, qRT-PCR, and immunocytochemistry were used to examine the unique structures formed by embryonic stem cells that exhibit key properties of a developing neural stem cell niche. We believe this system to have promising applications in stem cell transplantation, emphasizing the importance of understanding graft rejection in the brain in order to effectively introduce and preserve this complex microenvironment in intracranial transplantation therapy

Native American Food Practice and Provisioning

Join our panelists for a discussion of the history, roles, and uses of food in maintaining indigenous cultures. This panel considers the Native food sovereignty movement and the creative ways that some museums have connected with Native food production; cooking traditions and the perpetuation of culture; and the continuities from past into present in approaches to Native American cuisine. Moderator: Kathleen Barker, Assistant Director of Education & Public Programs, Massachusetts Historical Society Presenters: Elizabeth Hoover, Assistant Professor of American Studies and Ethnic Studies, Brown University Rachel Sayet, Mashpee Archives Loren Spears, Executive Director, Tomaquag Museu