MSCAGR (Animal Science) / Department of Animal Science / The use of in vitro culture media and assisted hatching techniques remain a challenging obstacle to hatching of blastocyst-stage embryos. Mechanical, chemical, enzymatic thinning and laser assisted techniques have been used previously, but there is still a lack of information on its application and implication in livestock. The aim of this study was to compare the effect of two in vitro culture media ((Ham’s F10 and Tissue Culture Medium 199 (TCM-199)) and four assisted hatching techniques (mechanical, chemical, enzymatic and laser) on blastocyst formation and hatching rate using murine embryos as a model. The C57BL/6 and BALB/c mouse breeds were bred and raised until they reach maturity and then bred naturally to produce a hybrid F1 generation. The light in the breeder house was controlled at 14 hours light and 10 hours darkness. Feed and water were provided ad libitum for the mice. Mature female mice were super-ovulated using equine chorionic gonadotropin (eCG) and human chorionic gonadotropin (hCG). A total of 400 blastocysts were collected from the F1 generation and these were allocated equally for the four assisted hatching techniques (laser, mechanical, chemical and enzymatic) as well as a non-treated control group. The blastocysts were paired into a group of 10 and replicated 4-four times for each assisted hatching techniques and control group. The embryos were then cultured for 24 hours and the hatching of the embryos were observed. Hatched embryos were stained for blastomere counting. The general linear model (GLM) of statistical analysis software (SAS) version 9.4 was used to analyze the data. Assisted hatching techniques (laser, mechanical, enzymatic and chemical) yielded 46.86±37.12; 51.07±40.19; 39.05±35.83 and 33.32±37.50% of hatching, respectively under in vitro culture in Ham’s F10. There was a significant difference (p<0.05) observed between assisted hatching techniques using Ham’s F10 as culture medium. In the TCM-199, laser, mechanical, enzymatic and chemical assisted hatching techniques yielded 56.25±43.30; 52.55±35.50; 49.16±37.50 and 33.85±35.50%, respectively, with significant differences (p<0.05). However, the hatching rate of embryos for all techniques was higher when in vitro cultured in TCM-199 compared to those cultured in Ham’s F10, and statistically higher than the control group. In conclusion, laser assisted hatching technique is the best of the techniques to use to assist the hatching of murine embryos and TCM-199 is the best of the two in vitro culture media for the hatching percentage.

Recent studies have revealed that crosstalk between glioma cells and the brain microenvironment is a crucial regulator of cancer initiation and progression. A vast majority of glioma patients suffer from seizures, and this pathological neuronal activity has been proposed to contribute to increased glioma cell proliferation. Glioma patients also suffer from additional neurological symptoms, including deficits in attention, concentration, memory, and language. These neurological effects of gliomas along with the limited therapeutic options underscore the need for novel therapies. This thesis investigates the neuronal alterations at the margins of glioma which contribute to the neurological symptoms (Chapter 2), and on the effect of sensory stimulation on the glioma cells and microglia in the glioma microenvironment (Chapter 3). The work describes the development of new mouse models in which glioma cells are infiltrating the somatosensory cortex in mice that express GCaMP in neurons or microglia. Methodologies include a combination of in vivo two-photon calcium imaging and tissue-based analysis to determine the glioma-induced alteration on whisker stimulation-evoked responses of these different cell types. This work also tests the effects of pharmacologically inhibiting mTOR signaling on neuronal responses (Chapter 2) or purinergic signaling on microglial responses (Chapter 3). Together these studies demonstrate that glioma infiltration induces local effects in functionally-responsive cortex, and that many of these glioma-induced effects on neurons and microglia are ameliorated by pharmacological inhibition of mTOR or purinergic signaling. This reveals a highly dynamic and plastic nature of the glioma-induced alterations, and points towards new strategies to treat glioma-associated neurological symptoms while potentially slowing tumor progression.

Adult neurogenesis is affected in neurodegenerative diseases and also represents an important therapeutic target. The goal of this dissertation research was to test the hypothesis that regeneration of neurons and glia in the adult brain can be manipulated by neurotrophic drugs in the context of two mouse models of neurodegenerative disease : Parkinson's disease and Huntington's disease.... These findings have implications for the pathophysiology of Huntington's disease and neurodegeneration in general. Specific alterations to the SVZ neurogenic niche parallel some of the pre-motor symptoms of Parkinson's disease and Huntington's disease. This dissertation research contributes to the growing body of literature concerning the pharmacological modulation of SVZ-derived neurogenesis designed to attenuate the progressive loss of neurons in neurodegenerative diseases and perhaps delay the onset of symptoms. / by Mark Harvey McCollum. / Vita. / Thesis (Ph.D.)--Florida Atlantic University, 2012. / Includes bibliography. / Mode of access: World Wide Web. / System requirements: Adobe Reader.

Sjogren's Syndrome (SS) is characterized by lymphocytic infiltration, destruction and dysfunction of the lacrimal and salivary glands and the presence of serum autoantibodies. Although, approximately 0.5% of the population suffers from SS, there is a female predominance of 9:1 compared with males. Most women with SS are postmenopausal; however, not all women who are post-menopausal develop SS. Therefore, we postulate that a decrease in the circulating levels of hormones creates an environment favorable to the development of SS in a predisposed genetic background. In order to carry out our studies, we used the NOD.B10.H2b mouse model of SS, and ovariectomized (OVX) them as a model for the post-menopausal condition. We removed the lacrimal glands and measured the gene expression and protein levels of several cytokines and chemokines known to be upregulated in patients with SS such as : lL-1B, IL-10, INF-y, TNFa, CCL9 and CXCL13. / We also stained for markers of B cells (B220+) and T cells (CD4+ and CD8+), and counted positively stained cleaved caspase-3 cells as an indication of apoptosis. These experiments were done 3, 7 and 21 days post-OVX and compared to sham operated animals. In order to determine whether the changes observed with OVX were triggered mainly by a genetic pre-disposition, a non-prediposed OVX and sham operated mouse (C57BL/10) was used as control. We found that gene expression of IL-1B, IL-10 and IF-y were upregulated in the lacrimal glands of the OVX NOD.B10.H2b mice at 3 days post-OVX compared with sham operated animals. Gene expression of IL-1B, IL-10, IFN-y, TNF-a, CCL9 and CXCL13, and protein levels of IL-1B, IL-10 and CCL9 were upregulated in the OVX NOD.B10.H2b mice at 7 days post-OVX compared to sham operated animals. / Also, at 7 days, an increase in B220+ B cells and an increase in cleaved caspase-3 were also observed in the OVX NOD.B10.H2b mice lacrimal glands compared to sham operated animals. At 21 days, protein levels of IL-10 were also highly upregulated in the OVX NOD.B10.H2b mice, together with an increase of B220+ B cells, a slight increase in the CD4/CD8 ratio and an increase on the number of caspase-3 positive cells. No changes were observed in any of the above parameters measured in the OVX C57BL/10 mice compared to the sham operated group, supporting our hypothesis that both, genetics and a decrease in the levels of hormones are necessary for SS to occur. / by Vanessa Seamon. / Thesis (Ph.D.)--Florida Atlantic University, 2009. / Includes bibliography. / Electronic reproduction. Boca Raton, Fla., 2009. Mode of access: World Wide Web.

Metastasis is the primary cause of mortality in women with breast cancer. Recently, elevated serum levels of a glycoprotein known as chitinase-3 likeprotein- 1 (CHI3L1) has been correlated with poor prognosis and shorter survival of patients with cancer and inflammatory diseases. The biological and physiological functions of CHI3L1 in tumor progression have not yet been elucidated. In this document, we describe the role of CHI3L1 in tumor growth and metastasis and its relationship with inflammation. Using well-established models of breast cancer, we show that CHI3L1 is increased in the serum of tumor bearing mice. We found that CHI3L1 levels are increased at both the “pre-metastatic” and “metastatic stage” and that tumor cells, splenic, alveolar and interstitial macrophages; and myeloid derived population produce CHI3L1. Furthermore, we demonstrated that CHI3L1 has an inhibitory role on the expression of interferon-gamma (IFN γ) by T cells, while enhancing the production of pro-inflammatory mediators by macrophages such as Cchemokine ligand 2 (CCL2/MCP-1), Chemokine CX motif ligand 2 (CXCL2/IL-8) and matrix metalloproteinase-9 (MMP-9), all of which promote tumor growth and metastasis. We demonstrated that in vivo treatment of tumor-bearing mice with chitin microparticles, a TH1 adjuvant and a substrate for CHI3L1, promoted immune effector functions with increased production of IFN-γ but decreased CCL2/MCP-1, CXCL2/IL-8 and MMP-9 expression by splenic and pulmonary macrophages. Significantly, in vivo administration of chitin microparticles decreased tumor growth and pulmonary metastasis in mammary tumor bearing mice. These results suggest that CHI3L1 may play a role in tumor progression. Inflammation plays a pivotal role during tumor progression and metastasis by promoting the production of pro-inflammatory molecules such as CHI3L1. However, little is known about how CHI3L1 expression can affect secondary sites to enhance metastasis. In these studies, we demonstrated that CHI3L1 alters the cellular composition and inflammatory mediators that aid in the establishment of a metastatic niche for the support of infiltrating tumor cells leading to accelerated tumor progression. Since previous studies showed that CHI3L1 modulates inflammation, we determined the role of CHI3L1 in the context of pre-existing inflammation and metastasis. We found that CHI3L1 deficient mice with preexisting inflammation had decreased pro-inflammatory mediators, and significant reduction in tumor volume and metastasis compared to wild type controls. Preexisting inflammation and CHI3L1 may be driving the establishment of a premetastatic milieu in the lungs and aiding in the establishment of metastasis. Understanding the role of CHI3L1 in inflammation during tumor progression could result in the design of targeted therapies for breast cancer patients. / Includes bibliography. / Dissertation (Ph.D.)--Florida Atlantic University, 2015. / FAU Electronic Theses and Dissertations Collection

Carbohydrate recognition is one of the most sophisticated recognition processes in biological systems, mediating many important aspects of cell-cell recognition, such as inflammation, cell differentiation, and metastasis. Consequently, lectin-glycan interactions have been intensively studied in order to mimic their actions for potential bioanalytical and biomedical applications. Galectins, a class of ß-galactoside-specific animal lectins, have been strongly implicated in inflammation and cancer. Galectin-3 is involved in carbohydrate-mediated metastatic cell heterotypic and homotypic adhesion via interaction with Thomsen-Friedenreich (TF) antigen on cancer-associated MUC1. However, the precise mechanism by which galectin-3 recognizes TF antigen is poorly understood. Our thermodynamic studies have shown that the presentation of the carbohydrate ligand by MUC1-based peptide scaffolds can have a major impact on recognition, and may facilitate the design of more potent and specific galectin-3 inhibitors that can be used as novel chemical tools in dissecting the precise role of galectin-3 in cancer and inflammatory diseases. Another lectin, odorranalectin (OL), has been recently identified from Odorrana grahami skin secretions as the smallest cyclic peptide lectin, has a particular selectivity for L-fucose and very low toxicity and immunogenicity, rendering OL an excellent candidate for drug delivery to targeted sites, such as: (1) tumor-associated fucosylated antigens implicated in the pathogenesis of several cancers, for overcoming the nonspecificity of most anticancer agents; (2) the olfactory epithelium of nasal mucosa for enhanced delivery of peptide-based drugs to the brain. / Includes bibliography. / Dissertation (Ph.D.)--Florida Atlantic University, 2015.

Restrictive cardiomyopathy (RCM) is a cardiac muscle disorder characterized by increased ventricular stiffness and diastolic dysfunction. Patients with RCM often present severe cardiac problems which usually lead to heart failure and sudden death. No effective treatment is available for RCM which makes the finding of novel efficient therapies an urgent necessity. Great progress in molecular biology techniques and advances in transgenic animal development provide great opportunities for the study of RCM and other cardiovascular diseases encountered in clinical patients.... Our laboratory is among the first to generate transgenic mouse models of RCM based on cardiac troponin I (cTnI) missense mutations. In this study, transgenic mice that suffer from RCM have been generated to understand the factors behind the diastolic dysfunction associated with that myocardial disease.... The information obtained from this study allows a better understanding of the role of troponin in RCM and the factors behind the physiopathology of the disease. It will also offer a therapeutic strategy taking into account the physiological characteristic of RCM. / by Pierre-Ives Jean-Charles. / Thesis (Ph.D.)--Florida Atlantic University, 2012. / Includes bibliography. / Mode of access: World Wide Web. / System requirements: Adobe Reader.

Restrictive cardiomyopathy (RCM) is represented in part by left ventricular stiffness and diastolic dysfunction. Missense mutations of the cardiac troponin I (cTnI) gene cause idiopathic RCM. These mutations are located in the C-terminus of cTnI and affect cardiac relaxation. Transgenic mouse models presenting the pathology observed in clinical patients with RCM have been generated previously and express the mutant cTnI in their hearts. RCM-linked mutations increase cardiac myofilament Ca2+ sensitivity and promote diastolic dysfunction in the heart. Previous studies using double transgenic mice (cTnI/R193H/ND) showed that ventricular relaxation is enhanced in the cTnI/R193H transgenic mice. In this study, another double transgenic mouse model, (cTnI/R193H/ND/KO), provides an avenue to investigate its rescuing effects on RCMlinked mutations in the cTnI /R193H/KO mouse. Use of molecular biological techniques, transgenic animal developments and murine echocardiography in this study has culminated into a greater understanding of RCM and diastolic dysfunction. / Includes bibliography. / Thesis (M.S.)--Florida Atlantic University, 2014. / FAU Electronic Theses and Dissertations Collection

Transcription and translation of proteins are required for the consolidation of episodic memory. Arc, an effector immediate early gene, has been linked to synaptic plasticity following learning and memory. It is well established that the rodent hippocampus is essential for processing spatial memory, but its role in processing object memory is a point of contention. Using immunohistochemical techniques, hippocampal sections were stained for arc proteins in the CA1 region of the dorsal hippocampus in mice following two variations of the novel object recognition (NOR) task. Results suggest mice that acquired strong object memory showed significant hippocampal activation. In mice that acquired weak object memory, hippocampal activation was not significantly different from controls. Arc expression was also examined in other hippocampal sub-regions, as well as in the perirhinal cortex. These results suggest that the mice must acquire a threshold amount of object information before the hippocampal CA1 region is engaged. / Includes bibliography. / Thesis (M.A.)--Florida Atlantic University, 2015 / FAU Electronic Theses and Dissertations Collection

by Stefanie P.C. Czerwinski. / Thesis (M.S.)--Florida Atlantic University, 2013. / Includes bibliography. / Mode of access: World Wide Web. / System requirements: Adobe Reader. / Sjèogren's Syndrome (SS) is a chronic, inflammatory autoimmune disease affecting mostly the exocrine cells of lacrimal and salivary glands, leading to diminished secretory function and resulting in keratoconjunctivitis sicca (dry eye disease) and/or stomatitis sicca (dry mouth disease). Despite several decades of studies focusing on autoimmune diseases and dry eye diseases, the exact etiology and mechanisms of SS remain unknown. Besides the fact that SS is often unreported, unrecognized and untreated, today's therapies rely exclusively on treating the symptoms after disease progression; there exists neither prevention therapy nor cure for SS. In addition, SS has been diagnosed predominantly in post-menopausal women with the female to male ratio reaching 9:1, suggesting a role of ovarian sex hormones in the pathogenesis of SS. However, not all postmenopausal women develop SS, indicating the contribution of other factors such as a genetic background to the onset of SS. In the present study, ovariectomized (OVX) NOD.B10.H2b mice provide a model of menopause with a genetic predisposition to SS, as compared to non-predisposed C57BL/10 mice. Both strands of mice were either sham operated, OVX, OVX and treated with 17(Sb (Bestradiol (E2), or OVX and treated with dihydrotestosterone (DHT). Lacrimal glands were collected 3, 7, 21, and 30 days after surgery and processed for RNA analysis by rt-qPCR and protein assays by ELISA to evaluate cytokine expression and concentrations of IL- 1\U+fffd\, TNF-a, IFN-(Sd(B, IL-10, and IL-4 on a timeline. Overall, our results showed a significant increase in IL-1\U+fffd\ TNF-a, IL-10, and IL-4 expression and levels in the lacrimal glands of OVX NOD.B10.H2b mice as compared to sham operated animals, and treatment with E2 or DHT at time of OVX prevented the increase in cytokine expression and levels.