Chemotherapy-Induced Damage of the Bone Marrow Microenvironment

The bone marrow microenvironment is characterized as the anatomical site including specialized niches that support stem cells. In addition, these niches also provide both soluble and physical cues leading to the differentiation of stem cells into all the cells of the blood. The studies in this dissertation focus on two supportive niches in the bone marrow microenvironment, osteoblasts (HOB) and bone marrow stromal cells (BMSC), in the setting of high dose chemotherapy and the potential damage that chemotherapy treatment causes to the cells of the bone marrow microenvironment.;In the first study, we investigated the effects of melphalan and Etoposide (VP-16) treatment on osteoblasts. Previous studies from our lab had shown that chemotherapy treatment increased the amount of active TGF-beta secreted from BMSC, leading to decreases in the ability to support pro-B cells. Here we describe the novel observation that osteoblasts pre-treated with chemotherapy have increased active TGF-beta and a decreased capacity to support human embryonic stem cells (hESC), CD34+ bone marrow derived cells and pro-B cells. We also evaluated the effects of adding recombinant TGF-beta (rTGF-beta) to osteoblasts to mimic the autocrine and paracrine TGF-beta in the microenvironment during chemotherapy treatment. rTGF- beta treatment of osteoblasts increased TGF-beta secretion and also led to a decreased ability to support hESC, CD34+ bone marrow derived cells and pro-B cells. Microarray analysis of the cells treated directly with chemotherapy or rTGF-beta or conditioned media from BMSC that were treated with chemotherapy suggested that many genes are changing in response to all of these treatment groups, indicating that osteoblasts are a vulnerable cell population that can be affected by high dose chemotherapy, potentially resulting in decreased hematopoietic support.;We also investigated Interleukin-6 (IL-6), a known hematopoietic factor important in both myeloid and lymphoid differentiation, acute and inflammatory immune responses and bone metabolism. Neuroendocrine modulation of the bone marrow microenvironment is thought to be important in both hematopoiesis and immune regulation. We investigated the roles of neurotrophins in the bone marrow and their effects on BMSC. We show that BMSC express functional neurotrophin receptors and that treatment of BMSC with two neurotrophins, NGF or BDNF, led to an increase in IL-6 expression. Increased IL-6 is associated with a number of inflammatory diseases and our data support the idea that increased neurotrophins in the bone marrow microenvironment could lead to dysregulated hematopoiesis.;Additionally, we also evaluated the effects of chemotherapy treatment of BMSC and HOB, focusing on IL-6. Previous data has suggested IL-6 to be involved in graft versus host disease and we investigated the effects of melphalan on IL-6 expression in BMSC and HOB. Interestingly, we determined that melphalan treatment led to a decrease in IL-6 mRNA and protein, and compared to other chemotherapeutic agents used in our studies, melphalan had the most pronounced effect. We also evaluated the effects of recombinant IL-6 (rIL-6) in combination with melphalan and determined that the addition of rIL-6 restored both IL-6 mRNA and protein expression, suggesting that pathways associated with IL-6 expression may be disrupted. The decrease in IL-6 could potentially affect hematopoiesis and further studies in vivo need to be completed. Additionally, melphalan is used as first-line therapy in the treatment of multiple myeloma (MM). IL-6 is a proliferative factor in MM, allowing for disease progression. The melphalan-induced decrease in IL-6 observed in our studies may, in part, contribute eradicating the tumor population by decreasing the potent proliferative factor, IL-6.;Collectively, these data contribute to our understanding of alterations to the bone marrow microenvironment that occur during high dose chemotherapy and emphasize the importance of understanding the mechanisms that underlie the potential damage leading to altered ability to support normal hematopoiesis

Effectiveness of LISTEN on Loneliness, Neuroimmunological Stress Response, Psychosocial Functioning, Quality of Life, and Physical Health Measures of Chronic Illness

Objectives: Loneliness is a biopsychosocial determinant of health and contributes to physical and psychological chronic illnesses, functional decline, and mortality in older adults. This paper presents the results of the first randomized trial of LISTEN, which is anew cognitive behavioral intervention for loneliness, on loneliness, neuroimmunological stress response, psychosocial functioning, quality of life, and measures of physical health. Methods: The effectiveness of LISTEN was evaluated in a sample population comprising 27lonely, chronically ill, older adults living in Appalachia. Participants were randomized intoLISTEN or educational attention control groups. Outcome measures included salivary cortisol and DHEA, interleukin-6, interleukin-2, depressive symptoms, loneliness, perceived social support, functional ability, quality of life, fasting glucose, blood pressure, and body mass index. Results: At 12 weeks after the last intervention session, participants of the LISTEN group reported reduced loneliness (p¼0.03), enhanced overall social support(p¼0.05), and decreased systolic blood pressure (p¼0.02). The attention control group reported decreased functional ability (p¼0.10) and reduced quality of life (p¼0.13). Conclusions: LISTEN can effectively diminish loneliness and decrease the systolic blood pressure in community-dwelling, chronically ill, older adults. Results indicate that this population, if left with untreated loneliness, may experience functional impairment over a period as short as 4 months. Further studies on LISTEN are needed with larger samples, in varied populations, and over longer periods of time to assess the long-term effects of diminishing loneliness in multiple chronic condition

Extracellular Vesicles Secreted in Response to Cytokine Exposure Increase Mitochondrial Oxygen Consumption in Recipient Cells

Extracellular vesicles (EVs) are small, membrane-bound nanoparticles released from most, if not all cells, and can carry functionally active cargo (proteins, nucleic acids) which can be taken up by neighboring cells and mediate physiologically relevant effects. In this capacity, EVs are being regarded as novel cell-to-cell communicators, which may play important roles in the progression of neurodegenerative diseases, like Alzheimer’s disease (AD). Aside from the canonical physical hallmarks of this disease [amyloid β (Aβ) plaques, neurofibrillary tangles, and widespread cell death], AD is characterized by chronic neuroinflammation and mitochondrial dysfunction. In the current study, we sought to better understand the role of tumor necrosis factor-alpha (TNF-α), known to be involved in inflammation, in mediating alterations in mitochondrial function and EV secretion. Using an immortalized hippocampal cell line, we observed significant reductions in several parameters of mitochondrial oxygen consumption after a 24-h exposure period to TNF-α. In addition, after TNF-α exposure we also observed significant upregulation of two microRNAs (miRNAs; miR-34a and miR-146a) associated with mitochondrial dysfunction in secreted EVs. Despite this, when naïve cells are exposed to EVs isolated from TNF-α treated cells, mitochondrial respiration, proton leak, and reactive oxygen species (ROS) production are all significantly increased. Collectively these data indicate that a potent proinflammatory cytokine, TNF-α, induces significant mitochondrial dysfunction in a neuronal cell type, in part via the secretion of EVs, which significantly alter mitochondrial activity in recipient cells

Neurotrophins Regulate Bone Marrow Stromal Cell IL-6 Expression through the MAPK Pathway

The host's response to infection is characterized by altered levels of neurotrophins and an influx of inflammatory cells to sites of injured tissue. Progenitor cells that give rise to the differentiated cellular mediators of inflammation are derived from bone marrow progenitor cells where their development is regulated, in part, by cues from bone marrow stromal cells (BMSC). As such, alteration of BMSC function in response to elevated systemic mediators has the potential to alter their function in biologically relevant ways, including downstream alteration of cytokine production that influences hematopoietic development.In the current study we investigated BMSC neurotrophin receptor expression by flow cytometric analysis to determine differences in expression as well as potential to respond to NGF or BDNF. Intracellular signaling subsequent to neurotrophin stimulation of BMSC was analyzed by western blot, microarray analysis, confocal microscopy and real-time PCR. Analysis of BMSC Interleukin-6 (IL-6) expression was completed using ELISA and real-time PCR.BMSC established from different individuals had distinct expression profiles of the neurotrophin receptors, TrkA, TrkB, TrkC, and p75(NTR). These receptors were functional, demonstrated by an increase in Akt-phosphorylation following BMSC exposure to recombinant NGF or BDNF. Neurotrophin stimulation of BMSC resulted in increased IL-6 gene and protein expression which required activation of ERK and p38 MAPK signaling, but was not mediated by the NFkappaB pathway. BMSC response to neurotrophins, including the up-regulation of IL-6, may alter their support of hematopoiesis and regulate the availability of inflammatory cells for migration to sites of injury or infection. As such, these studies are relevant to the growing appreciation of the interplay between neurotropic mediators and the regulation of hematopoiesis

Extracellular Vesicles Secreted in Response to Cytokine Exposure Increase Mitochondrial Oxygen Consumption in Recipient Cells

Extracellular vesicles (EVs) are small, membrane-bound nanoparticles released from most, if not all cells, and can carry functionally active cargo (proteins, nucleic acids) which can be taken up by neighboring cells and mediate physiologically relevant effects. In this capacity, EVs are being regarded as novel cell-to-cell communicators, which may play important roles in the progression of neurodegenerative diseases, like Alzheimer’s disease (AD). Aside from the canonical physical hallmarks of this disease [amyloid β (Aβ) plaques, neurofibrillary tangles, and widespread cell death], AD is characterized by chronic neuroinflammation and mitochondrial dysfunction. In the current study, we sought to better understand the role of tumor necrosis factor-alpha (TNF-α), known to be involved in inflammation, in mediating alterations in mitochondrial function and EV secretion. Using an immortalized hippocampal cell line, we observed significant reductions in several parameters of mitochondrial oxygen consumption after a 24-h exposure period to TNF-α. In addition, after TNF-α exposure we also observed significant upregulation of two microRNAs (miRNAs; miR-34a and miR-146a) associated with mitochondrial dysfunction in secreted EVs. Despite this, when naïve cells are exposed to EVs isolated from TNF-α treated cells, mitochondrial respiration, proton leak, and reactive oxygen species (ROS) production are all significantly increased. Collectively these data indicate that a potent proinflammatory cytokine, TNF-α, induces significant mitochondrial dysfunction in a neuronal cell type, in part via the secretion of EVs, which significantly alter mitochondrial activity in recipient cells

Fibrous Monolithic Ceramics

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/66419/1/j.1151-2916.1997.tb03148.x.pd

Bone Marrow Osteoblast Damage by Chemotherapeutic Agents

Hematopoietic reconstitution, following bone marrow or stem cell transplantation, requires a microenvironment niche capable of supporting both immature progenitors and stem cells with the capacity to differentiate and expand. Osteoblasts comprise one important component of this niche. We determined that treatment of human primary osteoblasts (HOB) with melphalan or VP-16 resulted in increased phospho-Smad2, consistent with increased TGF-β1 activity. This increase was coincident with reduced HOB capacity to support immature B lineage cell chemotaxis and adherence. The supportive deficit was not limited to committed progenitor cells, as human embryonic stem cells (hESC) or human CD34+ bone marrow cells co-cultured with HOB pre-exposed to melphalan, VP-16 or rTGF-β1 had profiles distinct from the same populations co-cultured with untreated HOB. Functional support deficits were downstream of changes in HOB gene expression profiles following chemotherapy exposure. Melphalan and VP-16 induced damage of HOB suggests vulnerability of this critical niche to therapeutic agents frequently utilized in pre-transplant regimens and suggests that dose escalated chemotherapy may contribute to post-transplantation hematopoietic deficits by damaging structural components of this supportive niche