acase@tulane.edu / The goal of the following experiments was to determine the cellular mechanisms through which estrogen receptor activity is maintained in hippocampal cells following termination of ovarian function. Aim 1 determined that kinase signaling contributes to the maintenance of estrogen receptor activity in the hippocampus of ovariectomized mice in addition to local synthesis of brain derived “neuroestrogens”. Inhibition of both the mitogen activated protein kinase (MAPK) and phosphoinositide-3 kinase (PI3K) cascades with intracerebroventricular infusion of specific kinase inhibitors reduced estrogen response element (ERE)-dependent gene expression in the hippocampus of ovariectomized mice. Aim 2 determined that neuroestrogen synthesis, MAPK signaling, and PI3K signaling interact to regulate the transcriptional output of estrogen receptors in response to insulin like growth factor-1 receptor (IGF-1R) activation in the Neuro-2A cell culture model. Rapid IGF-1R-dependent MAPK signaling promotes, while PI3K signaling inhibits, IGF-1R-dependent activation of endogenous estrogen receptors in Neuro-2A cells. Long-term IGF-1R stimulation reduces ERE-dependent gene expression in part through phosphorylation of estrogen receptor alpha (ERα). Rapid IGF-1R-dependent activation but not long-term repression of estrogen receptor activity in Neuro-2A cells requires neuroestrogen synthesis. Aim 3 determined that exposure to 40 days of continuous unopposed estradiol at the time of ovariectomy results in lasting enhancement of estrogen receptor activity in the hippocampus and lasting enhancement of hippocampus dependent memory in female mice beyond the period of short-term estradiol exposure. Together these three aims determine that neuroestrogen synthesis and kinase signaling interact to actively maintain estrogen receptor signaling in neuronal cells and these autonomous neuronal mechanisms of estrogen receptor activation have functional consequences on cognition long after cessation of ovarian function. / 1 / Kevin J Pollard

1 / Sarah Elizabeth Galvez

Tendon plays an important role in regulating body locomotion and providing additional stability to the body. However, tendon is susceptible to injuries and the healing process could be devastating along with the several issues, namely adhesion formations, slow healing and failure at fixation sites, which have deferred the success of proper tendon healing via tendon tissue engineering. This dissertation thus aims to create a sustained dual-drug therapy to address these issues. For adhesion formation, naproxen sodium (NPS) has been shown to be able to avoid this symptom through inhibiting inflammation process.

Life-history trade-offs have been well-documented within the literature through correlational and experimental studies. However, the physiological mechanisms underlying these trade-offs are less understood. Currently, there is great interest in shared mechanisms, specifically endocrine mechanisms, that might underlie the variation in life-history traits. Insulin-like growth factor-1 (IGF-1) may be one shared mechanism that is particularly important. IGF-1 is a metabolic hormone that is part of a highly conserved insulin-signaling pathway known to influence multiple life-history traits including growth and longevity across taxa, however, little is known about these trade-offs outside of laboratory populations. This dissertation focuses on the role of IGF-1 as a hormonal mechanism underlying the life-history trade-off between growth and aging in wild birds. While the causes of aging are not fully understood, telomere dynamics (length and change in length) are a potentially important mechanism underlying lifespan. To investigate the role of IGF-1 as a hormonal mechanism underlying the life-history trade-off between growth and aging in Franklin’s gulls (Leucophaeus pipixcan) and house sparrows (Passer domesticus). In Franklin’s gulls, dietary restriction reduced growth rate and IGF-1 levels but did not impact telomere dynamics. However, there was a significant negative correlation between IGF-1 levels and telomere length at the end of the post-natal growth period. In house sparrows, we found that nestling growth rates varied with respect to year, but IGF-1 levels did not. Telomere dynamics were not related to growth rates or IGF-1 levels, suggesting that during post-natal growth nestlings may be able to mitigate or even delay costs to later life stages. Finally, when exogenous IGF-1 was administered to house sparrow nestlings during the post-natal growth period, nestling growth was impacted but only in some years. Exogenous IGF-1 increased growth and final mass in 2016 and final mass in 2018. There was a trend suggesting experimental birds had shorter telomeres in 2016. Similarly, in 2018, experimental birds had significantly shorter telomeres than control birds. These effects were not observed in 2017, suggesting that trade-offs between growth an aging might only be visible under certain environmental conditions, which may vary with respect to year.

Astronauts at are risk of losing 1.0-1.5% of their bone mass for every month they spend in space despite their adherence to high impact exercise training programs and diets high in nutrients, potassium, calcium, and vitamin D, all designed to preserve the skeletal system. This article reviews the basics of bone formation and resorption and details how exposure to microgravity or simulated microgravity affects the structure and function of osteoblasts, osteocytes, osteoclasts, and their mesenchymal and hematologic stem cell precursors. It details the critical roles that insulin-like growth factor-1 and its receptor insulin-like growth factor-1 receptor (GFR1) play in maintaining bone homeostasis and how exposure of bone cells to microgravity affects the function of these growth factors. Lastly, it discusses the potential of tumor necrosis factor-related apoptosis-inducing ligand, syncytin-A, sclerostin inhibitors and recombinant IGF-1 as a bone-saving treatment for astronauts in space and during their colonization of the Moon.

Traumatic brain injury (TBI) is a prevalent CNS neurodegenerative condition that results in lasting neurological dysfunction, including potentially debilitating cognitive impairments. Despite the advancements in understanding the complex damage that can culminate in cellular dysfunction and loss, no therapeutic treatment has been effective in clinical trials, highlighting that new approaches are desperately needed. A therapy that limits cell death while simultaneously promoting reparative mechanisms, including post-traumatic neurogenesis, in the injured brain may have maximum effectiveness in improving recovery of function after TBI. Insulin-like growth factor-1 (IGF-1) is a potent growth factor that has previously been shown to promote recovery of function after TBI, but no studies have evaluated the efficacy of IGF-1 to promote cell survival and modulate neurogenesis following brain injury. Systemic infusion of IGF-1 resulted in undetectable levels of IGF-1 in the brain, but did promote increased cortical activation of Akt, a pro-survival downstream mediator of IGF-1 signaling, in mice subjected to controlled cortical impact (CCI), a well-established model of contusion TBI. However, systemic infusion of IGF-1 did not promote recovery of motor function in mice after CCI. A one week central infusion of IGF-1 elevated brain levels of IGF-1, increased Akt activation and improved motor and cognitive function after CCI. Central infusion of IGF-1 also significantly increased immature neuron density at 7 d post-injury for a range of doses and when administered with a clinically relevant delayed onset of 6 hr post-injury. To mitigate potential side effects of central infusion, an alternative conditional astrocyte-specific IGF-1 overexpressing mouse model was utilized to evaluate the efficacy of IGF-1 to promote post-traumatic neurogenesis. Overexpression of IGF-1 did not protect against acute immature neuron loss, but did increase immature neuron density above uninjured levels at 10 d post-injury. The increase in immature neuron density appeared to be driven by enhanced neuronal differentiation. In wildtype mice, immature neurons exhibited injury-induced reductions in dendritic arbor complexity following severe CCI, a previously unknown pathological phenomenon. Overexpression of IGF-1 in brain-injured mice promoted the restoration of dendritic arbor complexity to the dendritic morphology observed in uninjured mice. Together, these findings provide strong evidence that treatment with IGF-1 promotes the recovery of neurobehavioral function and enhances post-traumatic neurogenesis in a mouse model of contusion TBI.

Background and objectives Evidence suggests that circulating IGF-I levels increase mammographic density (a breast cancer risk factor) and breast cancer risk in premenopausal women. The objective of this thesis was to examine the association of genetic variation at the IGF1 gene with IGF-I concentration, mammographic density, breast cancer risk, and related anthropometric measures in premenopausal women. Methods Three IGF1 CA repeat polymorphisms (at the 5′ and 3′ ends, and in intron 2) were genotyped. A cross-sectional design was used to investigate their associations with IGF-I levels, mammographic density, BMI, weight, and height. Families from registries in Ontario and Australia were used to investigate associations with breast cancer risk and also BMI, weight and height. Results In the cross-sectional study, greater number of copies of the 5′ 19 allele were associated with lower circulating IGF-I levels. Greater number of 3′ 185 alleles were associated with greater percentage breast density, smaller amount of non-dense tissue, and lower BMI. Including BMI in regression models removed the association of the 3′ 185 allele with percentage breast density. In the family based study, nominally significant associations (5′ 21 allele, intron 2 212 allele, intron 2 216 allele) with breast cancer risk were observed, but significance was lost after multiple comparison adjustment. There was a stronger association between the intron 2 216 allele and risk under a recessive model, and 5′ allele groupings of length 18 to 20 and 20 or more repeats produced significant positive and negative associations respectively. These associations were not strongly supported in analyses stratified by registry. Results from the family based study did not support an association between genetic variation at IGF1 with BMI, weight or height. Conclusions No specific IGF1 variant influenced each of circulating IGF-I levels, mammographic density, and breast cancer risk. The failure to replicate the association of the 3′ 185 allele with BMI in the family based study suggests that the association of the 3′ 185 allele with percentage breast density is spurious, since this association was mediated through the relationship with BMI (suggesting IGF-I action on body fat). Evidence for an association between IGF1 and breast cancer risk was limited.

The purpose of this study was to look at the genotoxic and cytotoxic effects of benzo-á-pyrene (BáP), a chemical mutagen that is present in cigarette smoke, on the insulin-like growth factor-I (IGF-I) gene. Women who smoke during pregnancy are more likely to have a growth-restricted baby. We hypothesized that BáP exerts its effects through genotoxic and cytotoxic avenues. The cytotoxicity is manifested by chromosomal abnormalities and a decrease in the rate of cell division. The genotoxicity is manifested by changes in certain genes known to be important in mammalian fetal development such as IGF-I. IGF-I is implicated in intrauterine growth restriction (IUGR), a problem that greatly increases the risk of perinatal morbidity and mortality. To futher understand the mechanism by which BáP influences the normal growth and development of human placental cells, human placental trophoblast cells from an established immortalized cell line were utilized. Cells were cultured in appropriate media, starved (using starvation "Serum Free Medium"), and treated with two doses of BáP, 1µM (dose 1) and 5µM (dose 2). Chromosomes were prepared for cytogenetic analysis and visualized using light microscopy after Giemsa staining. Chromosomal aberrations were identified and the rate of cell division was determined through the analysis of the mitotic index for treated cells compared to a control group. To further understand the influence of BáP on the IGF-I gene expression level, RNA was extracted from control and treated cells, from which cDNA was synthesized and used for further analysis using polymerized chain reaction (PCR). The PCR results were used to better understand the genotoxicity of BáP, while chromosomal aberration analysis was used to determine the cytotoxic effects of BáP on human placental cells. Our results indicate that many chromosomal abnormalities were present in the treated groups compared to the control group. In addition, there was a significant decrease in the mitotic index of the BáP-treated cells (MI=0.3%) verses the control group (MI=0.93%), p value 0.0447. Through the PCR assay, we speculate that there is a dose-related response to BáP of the IGF-I RNA expression level, with low levels in the treated groups compared to the control group. We conclude from these results that BáP influences placental cells at both the gene and chromosome level. It also affects the cell cycle of human placental cells. It is known that smoking is deleterious for fetal development. We believe that the current study brings us closer to understanding the mechanism by which smoking can lead to fetal growth restriction.

Thesis (Ph. D.)--Cornell University, January, 2007. / Vita. Includes bibliographical references.

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