Adult adipose derived mesenchymal stromal cells (ASCs) have been characterized in various species. Many factors may affect ASC fate and ASCs from different species may have different response to these factors. The first study was to identify the differences of the canine ASCs isolated from subcutaneous and infrapatellar adipose tissues, and evaluate the impact of cryopreservation on the cells. Based on paired comparisons of fresh and cryopreserved ASCs, cryopreserved ASCs had lower CD29 and CD44 protein expression and lower proliferation rates. The cryopreserved ASCs had relative lower mitochondria in the cytoplasm compared to the fresh ASCs regardless of tissue sources. The second study was to apply human ASCs for bone regeneration. The spinner flask bioreactor system was employed to load human ASCs onto three commercial scaffolds and the cell-scaffold constructs were cultured in stromal, osteogenic, or osteogenic for 48 hours followed by stromal medium for up to 28 days. The distinct scaffold upregulated different osteogenic signaling pathways, suggesting distinct osteogenic cell signaling pathways were selectively upregulated by scaffold composition. The third study was designed to quantify in vivo equine multipotent stromal cell (MSC) osteogenesis on synthetic polymer scaffolds with distinct mineral combinations 9 weeks after implantation in a murine model. Addition of mineral to polymer scaffolds enhanced equine MSC osteogenesis over polymer alone, and contributions by both exo- and endogenous MSCs were confirmed. The fourth study was designed to evaluate the effects of collagenase digestion and cryopreservation on equine ASCs. Higher collagenase concentration yielded more nucleated cells, and the percentages of MHCII-, CD44+, CD105+ cells in freshly isolated and cryopreserved cells were similar. The embryonic gene expression was enhanced and the essential gene expression decreased after cryopreservation. The fifth study was to demonstrate the endodermal transdifferentiation capability in feline ASCs. xvi Feline-specific pancreatic β cell induction medium was developed in the study, and islet-like cell clusters that secrete insulin in response to glucose stimulation were created. Overall, the investigations in this dissertation provide critical information for canine, feline, equine and human MSC-based tissue engineering therapies and may contribute to better efficiency and efficacy of cell

The purpose of this study was to measure the competency of problem solving skills of fourth-year veterinary students. The study identified two primary objectives, (a) define clinical competency for fourth-year veterinary medical students, and (b) construct an instrument to assess the student’s level of clinical competency. A faculty advisory panel identified three themes in the development of a definition of clinical competency, (a) competency was situational, (b) competency was described by ability, and (c) the definition of fourth-year student competence was descriptive within the context of primary patient care. The description of competency contributed to the establishment of parameters applied to the rubric. Student self-assessments were taken twice; initially at the beginning of the clinical rotation, and again at the end of the rotation as a post-clinical assessment. The faculty instructor provided a comparison post-clinical assessment to serve as an authentication of the reliability of the instrument. Overall, students qualified themselves to be “Fully Competent Students” at the beginning of the clinical rotation, and also at the end of the rotation. However, in the disaggregated quantified scores, the student self-assessment peaked at the highest competence level, “Among The Most Competent Students,” for five descriptors of values and beliefs listed under Responsibility, Professionalism, and Interpersonal Skills. The faculty comparison assessment was in agreement with the student’s qualified description as “Fully Competent Students” but did not agree with the higher quantified scores posted for values and beliefs. The descriptive statistics of the data reflect that the mean increased between the pre-clinical (M = 8.43, SD = 1.36) and the post-clinical (M = 9.10, SD = 1.32) assessments. The comparison mean for the faculty assessment (M = 9.01, SD = 1.52) was slightly lower than the student post-assessment. The assessment provided evidence supporting a confidence statement that the instrument has the sensitivity to detect changes in skills, and is consistent with research validated measures of problem-solving skills. Faculty authentication provided evidence of intra-rater reliability, while statistical analysis provided evidence that a relationship appears to be present between live-patient animal instruction and the increase in problem-solving competence of fourth-year veterinary students.

Assisted reproductive technologies developed in domestic cattle serve as a starting point in similar studies on nondomestic bovids. The common eland is a useful model species for studies on rare tragelaphine antelopes. In Chapter 3 of the present study, effects of components/attributes of protein-free embryo culture media on the in vitro development of in vitro-derived bovine embryos were evaluated. A 2 x 2 factorial study comparing effects of groups of amino acids (20aa or 11aa) in two base media (modified KSOM or BM-3) demonstrated that amino acids and base medium affected embryonic development. A subsequent 7 x 2 factorial experiment to evaluate effects of osmotic pressure and supplement type in BM-3-20aa showed that embryonic development was largely affected by supplements and identified glucose (0.2 mM) as a crucial supplement. In Chapter 4, the use of behavioral training and handling of elands in a hydraulic chute to perform transvaginal ultrasound-guided oocyte retrieval without inducing general anesthesia were evaluated. Nine of 10 females associated specific sound cues with food treats. Females varied in their response interval to audio cues and to training for voluntary entry into the chute. Handling elands for oocyte retrieval required sedation and increased blood glucose levels. In Chapter 5, type of estrous synchronization or ovarian stimulation protocol did not affect ovarian response. Animals, but not month of the year, affected ovarian response. In 37 oocyte retrieval procedures using seven females, an average of 12.8 follicles yielded 9.8 oocytes, of which up to 73% matured to metaphase II. In vitro fertilization, intracytoplasmic sperm injection and nuclear transfer resulted in embryonic development. In conclusion, the bovine embryo culture study suggests that the beneficial effects of amino acids are influenced by the base medium and glucose plays more important roles in non-ATP producing pathways. Behavioral training and handling of sedated females in a hydraulic chute is a reliable method for collecting eland oocytes, which can undergo in vitro maturation and some in vitro embryonic development.

This thesis focuses on the study of heparanase which is an enzyme involved as a molecular determinant of cancer metastasis. The purpose of this study was to first quantitate heparanase mRNA levels in both normal and tumorigenic samples from the same tissue specimen. Laser capture microdissection was used in the isolation and extraction of melanoma cell populations from normal tissue. There was a 29 fold upregulation of heparanase expression, detected by real-time PCR, in metastatic melanoma of the lung in comparison to normal lung tissue in mice. Immunohistochemistry (IHC) showed stronger staining in human metastatic melanoma when compared to primary melanoma tumors. IHC also showed a propensity for darker heparanase staining around blood vessels and vascular regions. These results further emphasized the importance of heparanase in invasive and angiogenic mechanisms in melanoma. Once heparanase was determined to be upregulated in melanoma tissue in vivo both at the mRNA and protein level, the next part of this thesis was directed to the development of an orthotopic brain slice model. A novel model that would provide a relatively efficient way to study the biological relevance and mechanisms involved in the invasive process of brain metastatic melanoma and the role that heparanase plays in this invasive process. We showed that this model could be used to determine invasion into brain tissue at both qualitative and quantitative levels. We showed that HPSE-1 augmented invasion of brain metastatic melanoma cells into brain tissue. We also showed that melanoma cells show a time dependent expression of heparanase while invading into brain tissue. Thus, we showed that heparanase is involved in cancer metastasis development and could have important implications in the development of potential drugs aimed to combat cancer metastasis.

Los escarabajos coprófagos (Coleoptera: Scarabaeoidea) son un grupo de insectos muy importante en términos de diversidad y biomasa en los pastos de la cuenca del Mediterráneo. A pesar de su importancia ecológica, los escarabajos coprófagos están amenazados por varios factores, entre los cuales se destacan los cambios en el manejo de la ganadería tradicional. El objetivo general de esta tesis fue evaluar los efectos del abandono de la ganadería, su extensificación y uso histórico de productos médico veterinarios sobre las comunidades de escarabajos coprófagos usando medidas de biodiversidad y cuantificando la capacidad de remoción de estiércol en un paisaje sub-montano de la provincia de Pesaro-Urbino (Italia). En este trabajo se colectaron un total de 156 936 individuos, pertenecientes a 58 especies de escarabajos coprófagos. Los resultados de este trabajo sugieren que el uso histórico de productos médico veterinarios tiene un impacto negativo y ubiquitario sobre todas las especies de la comunidad de estudio, llevando a un empobrecimiento de los ensambles y a una pérdida de abundancia y biomasa. No obstante se ha registrado el mantenimiento de la estructura de la comunidad. Estos resultados son corroborados igualmente cuando nos enfocamos en la diversidad funcional. En efecto, el uso histórico de productos médico veterinarios lleva a una pérdida en el número de roles funcionales dentro de la comunidad, sin embargo las abundancias se encuentran repartidas equitativamente dentro de cada nicho funcional. Todos estos efectos sobre la comunidad de escarabajos coprófagos debido al uso de productos médico veterinarios, al final quedan reflejados en una pérdida del 70% en la capacidad de remoción de estiércol, comparados con áreas donde estos productos no son aplicados. La extensificación de la ganadería está representada por áreas que tienen una intensidad de pastoreo muy baja, las cuales pueden ser consideradas como la primera etapa hacia el abandono completo. En este sentido, los resultados de este trabajo demuestran como una simple reducción de la carga ganadera lleva a efectos negativos sobre las comunidades de escarabajos coprófagos, registrando una disminución en la diversidad alpha y en el número de especies indicadoras. Esta disminución en la cantidad de recurso trófico tiene implicaciones sobre la composición de la comunidad, favoreciendo especies oportunistas cuya biología le permite evitar la competencia, como por ejemplo; especies cleptoparásitas o especies cuyas larvas tienen un comportamiento saprófago. Este “efecto filtro” (filter effect) debido a la disminución de la cantidad trófica, está confirmado por los resultados de diversidad funcional, en los cuales se registró una disminución en el número de nichos funcionales y una fuerte redundancia por la presencia de nichos funcionales muy similares entre ellos. Los efectos sobre la comunidad de escarabajos coprófagos en los sitios con baja carga ganadera reflejan finalmente una pérdida en la capacidad de enterramiento del estiércol del 30-40% respecto a las comunidades en áreas con una mayor carga ganadera. Los resultados de este trabajo muestran que en las áreas con abandono total de la ganadería se observó una disminución en el número de especies, en la abundancia y biomasa total, en el número de especies indicadoras, en la abundancia de las especies de gran tamaño y en las especies que no tienen un comportamiento de nidificación durante el periodo reproductivo. Sin embargo, el mantenimiento de una comunidad bien estructurada, subraya la importancia de dos factores locales: el excremento de la fauna silvestre y la mayor complejidad del hábitat debido a un proceso de invasión de árboles en el pasto. Este último factor es probablemente responsable de la presencia de especies indicadoras típicas de hábitats más cerrados como los bosques o matorrales. En cuanto a los resultados de diversidad funcional, estos confirman la interpretación del “efecto filtro” debido a la disminución en la cantidad de recurso trófico. En efecto, estos resultados muestran una unicidad funcional significativa en esta comunidad como consecuencia del fuerte cambio composicional que el abandono de la ganadería ha implicado, a pesar de una disminución en el número de nichos funcionales y en el mantenimiento de una buena estructura funcional. Así mismo, se registró una disminución en la capacidad de enterramiento del estiércol entre 27% y 47% respecto a las áreas con baja carga ganadera y con carga moderada por todos estos cambios cuali y cuantitativos en la comunidad de escarabajos coprófagos de los sitios abandonados. Se propone la ganadería ecológica como forma correcta de gestión de los pastos, la cual permitiría mantener un nivel de carga moderado, así como el control en uso de compuestos médico veterinarios.

This item contains the agenda, minutes, and attachments for the Faculty Senate meeting on this date. There may be additional materials from the meeting available at the Faculty Center.