published_or_final_version / Anatomy / Doctoral / Doctor of Philosophy

published_or_final_version / abstract / Medical Sciences / Master / Master of Medical Sciences

published_or_final_version / Obstetrics and Gynaecology / Doctoral / Doctor of Philosophy

Master of Science / Department of Anatomy and Physiology / Mark Weiss / Artificial insemination and cryopreservation of sperm are important components of any transgenic animal facility because they allow for the reduction in animal colony size and the safe storage of germplasm from valuable strains. In addition, they allow long-term storage of these strains and easy transportation of the genetic material to other research facilities internationally. Thus far, only one laboratory has created live rat pups after sperm cryopreservation and intrauterine insemination. Another laboratory made advances in cryopreservation media that improved sperm motility post-thawing, but no pups resulted from this work. In my study, these two cryopreservation media were utilized to perform intrauterine inseminations with both fresh samples of rat sperm as well as samples that were cryopreserved in liquid nitrogen to replicate and extend these studies. Pharmacoejaculation was tested as a means to obtain spermatozoa without euthanizing the male to collect the epididymis, but results were inconsistent and the samples were not useful for intrauterine inseminations or cryopreservation. Epidiymal sperm was then collected into the various media and frozen in liquid nitrogen. In my hands, the frozen/thawed rat sperm achieved motility of less than 1%. Next, the impact of altering the freezing rate on sperm motility was evaluated. Epididymal sperm was collected and processed using a modified protocol and were then frozen at 2, 4 or 6 cm above the level of liquid nitrogen. Four to six days after freezing, samples were thawed and post-thaw sperm motility was evaluated. Sperm motility was measured prior to freezing as well as after-thawing. The sperm motility was correlated with LIVE/DEAD® staining. Sperm motility did not differ between the groups as a result of the freezing rate (Friedman test p=0.23). The published techniques are not robust and require further development to improve the motility of rat sperm after cryopreservation and achieve pregnancy via intrauterine insemination.

It is now more than 35 years since the first world test-tube baby, Louise Brown, was born (1978) in England and it is estimated that since then more than 4 000 000 of children were produced by in vitro fertilization (IVF) worldwide. The initial success of IVF was less than 20% in best clinics, but now it reaches about 40%. This is a consequence of introduction of new methods, standardization and exploitation of new manipulation and culture media, as well as the incorporation of research results. Nevertheless, the most important still remains the skill and experience of IVF clinics and IVF laboratories staff, especially their ability to critically evaluate the quality of biological material and to decide which cure and treatment are the best one. At least, some biological material (immature and low quality oocytes) can be used for training and also for some experiments aiming to explain some questions, which are not yet fully understood (for example aneuploidies in human oocytes and embryos). In addition, this training can facilitate the introduction of new progressive approaches and may also improve indirectly the quality of infertility treatments. The first part of thesis is focused on the quality evaluation of oocytes collected by aspiration from follicles of stimulated patients. For labeling...

Os parasitas nematóides afetam homens e animais, causando graves prejuízos à saúde pública e consideráveis perdas econômicas. A disponibilidade de anti-helmínticos de amplo espectro de ação tem auxiliado na redução de um grande número de perdas em decorrência das infestações parasitárias. Foram objetivos deste trabalho avaliar os efeitos do tratamento com moxidectina, na forma de longa-ação (LA), em sua dose terapêutica em touros, sobre a consistência e perímetro testicular, as características físicas (motilidade e vigor) e morfológicas do sêmen e o comportamento sexual. Foram utilizados 12 touros com idade de 48 ± 10 meses. Os animais foram alocados em 6 blocos com dois tratamentos. Os animais foram divididos nos grupos: controle (n=6), no qual cada animal recebeu 5 mL de solução fisiológica, via subcutânea, na orelha esquerda e grupo tratado (n=6), no qual cada animal recebeu 5 mL de moxidectina a 10% via subcutânea, na orelha esquerda. Os animais foram submetidos a exames andrológicos semanais, por um período de até 60 dias após o tratamento, sendo avaliadas as características testiculares (consistência e perímetro) e características seminais (motilidade, vigor e morfologia). Foram realizados testes de libido a cada 15 dias, num total de cinco testes. Os resultados obtidos indicam que não houve diferença significativa (P>0,05) para a consistência testicular, perímetro testicular, motilidade progressiva, vigor espermático, morfologia espermática e libido entre os touros dos grupos controle e tratado. Esses resultados indicam que o tratamento com moxidectina a 10% não influencia negativamente as características reprodutivas de touros. / The helminths affect human beings and animals, causing serious damage to public health and considerable economic losses. The availability of anthelminthic of a broad-spectrum has helped reducing a large number of losses due to parasitic infestation. The objectives of this work were to evaluate the effect of the treatment with moxidectin, in Long Action (LA) formulation, in therapeutic doses in bulls, on the testicular consistency and perimeter, physical characteristics (motility and strength) and morphological of the semen and sexual behaviour. Twelve bulls, age 48 ± 10 months used. These animals were put in six, 06 blocks with two treatments. The animals were divided in groups: Control group (n=6) each animal received 5 ml of physiological solution subcutaneously in the left ear, and Treated group (n=6) each animal received 5 ml of moxidectin at 10% subcutaneously in the right ear. The animals were submitted to andrological examinations weekly, within a period of 60 days after the treatment. The testicular characteristics (consistency and perimeter) and seminal characteristic (motility, strength and morphology) were evaluated. Libido tests were done every fortnight, in five tests all together. The results indicated that there were not significant differences. (P>0.05) for the testicular consistency, perimeter, progressive motility, spermatic strength, morphology and libido among bulls of the control and treated groups. These results indicated that the treatment with moxidectin at 10% does not influence negatively on the reproductive characteristics of the bulls.

Thesis (M.A.)--Boston University / PLEASE NOTE: Boston University Libraries did not receive an Authorization To Manage form for this thesis or dissertation. It is therefore not openly accessible, though it may be available by request. If you are the author or principal advisor of this work and would like to request open access for it, please contact us at open-help@bu.edu. Thank you. / The incorporation of C14 label into the lipids of the spermatozoa of man and fish (the alewife, Alosa pseudoharengus,) was studied. Washed spermatozoa of man and the alewife were incubated in vitro under aerobic conditions with glucose-C14 (r.1.). The sperm cells were extracted with chloroform-methanol, and the crude lipid extracts were purified to remove radioactive contaminants. The lipid extracts were fractionated into the major classes of lipids by column chromatography and further separated by thin-layer chromatography. Glycerides, phosphatides and cholesterol were isolated and identified and their radioactivity was determined. C14 label was found exclusively in the lycerol portion of the glycerolipids of the spermatozoa of both man and the alewife. The rates of incorporation of glucose-C14 into the total lipids were 54.4 ummoles/1010 human spermatozoa/hour and 1.8 ummoles/10^10 alewife spermatozoa/hour. In the sperm lipids of both species the glycerides were the most actively labeled components with the diglycerides having the highest specific activities and the triglycerides being the most abundant components. Whereas in human spermatozoa the glyceride content was 20% of the total lipids, in the alewife spermatozoa the fraction constituted only 4% Cholesterol, 14 - 15% of the total sperm lipids in both species, was not labeled. The polyglycerophosphatide fraction was the most abundant and the most actively C14-labeled component in the phosphatides of alewife spermatozoa. Most of the radioactivity incorporated in the phosphatides of human spermatozoa was found in the lecithin and cephalin fractions. Lecithin was the phosphatide of highest specific activity. By means of infrared spectroscopy, evidence of the presence of plasmalogen was found in the phosphatides of human, but not of alewife spermatozoa. Unlike mammalian spermatozoa, fish spermatozoa have poor glycolytic capacity. Nevertheless, the present work shows that fish spermatozoa can utilize glucose for the synthesis of glyceride glycerol at adequate rates. The demonstration of the biosynthesis of lipids from glucose by the spermatozoa of man and fish suggests that sperm may utilize carbohydrate not only for the maintenance of motility through the energy generated by glycolysis, but also for the replenishment of lipid reserves. / 2031-01-01

Os parasitas nematóides afetam homens e animais, causando graves prejuízos à saúde pública e consideráveis perdas econômicas. A disponibilidade de anti-helmínticos de amplo espectro de ação tem auxiliado na redução de um grande número de perdas em decorrência das infestações parasitárias. Foram objetivos deste trabalho avaliar os efeitos do tratamento com moxidectina, na forma de longa-ação (LA), em sua dose terapêutica em touros, sobre a consistência e perímetro testicular, as características físicas (motilidade e vigor) e morfológicas do sêmen e o comportamento sexual. Foram utilizados 12 touros com idade de 48 ± 10 meses. Os animais foram alocados em 6 blocos com dois tratamentos. Os animais foram divididos nos grupos: controle (n=6), no qual cada animal recebeu 5 mL de solução fisiológica, via subcutânea, na orelha esquerda e grupo tratado (n=6), no qual cada animal recebeu 5 mL de moxidectina a 10% via subcutânea, na orelha esquerda. Os animais foram submetidos a exames andrológicos semanais, por um período de até 60 dias após o tratamento, sendo avaliadas as características testiculares (consistência e perímetro) e características seminais (motilidade, vigor e morfologia). Foram realizados testes de libido a cada 15 dias, num total de cinco testes. Os resultados obtidos indicam que não houve diferença significativa (P>0,05) para a consistência testicular, perímetro testicular, motilidade progressiva, vigor espermático, morfologia espermática e libido entre os touros dos grupos controle e tratado. Esses resultados indicam que o tratamento com moxidectina a 10% não influencia negativamente as características reprodutivas de touros. / The helminths affect human beings and animals, causing serious damage to public health and considerable economic losses. The availability of anthelminthic of a broad-spectrum has helped reducing a large number of losses due to parasitic infestation. The objectives of this work were to evaluate the effect of the treatment with moxidectin, in Long Action (LA) formulation, in therapeutic doses in bulls, on the testicular consistency and perimeter, physical characteristics (motility and strength) and morphological of the semen and sexual behaviour. Twelve bulls, age 48 ± 10 months used. These animals were put in six, 06 blocks with two treatments. The animals were divided in groups: Control group (n=6) each animal received 5 ml of physiological solution subcutaneously in the left ear, and Treated group (n=6) each animal received 5 ml of moxidectin at 10% subcutaneously in the right ear. The animals were submitted to andrological examinations weekly, within a period of 60 days after the treatment. The testicular characteristics (consistency and perimeter) and seminal characteristic (motility, strength and morphology) were evaluated. Libido tests were done every fortnight, in five tests all together. The results indicated that there were not significant differences. (P>0.05) for the testicular consistency, perimeter, progressive motility, spermatic strength, morphology and libido among bulls of the control and treated groups. These results indicated that the treatment with moxidectin at 10% does not influence negatively on the reproductive characteristics of the bulls.

Abstract not available

Typewritten ms. Includes bibliographical references