京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第19175号 / 医博第4017号 / 新制||医||1010(附属図書館) / 32167 / 京都大学大学院医学研究科医学専攻 / (主査)教授 渡邊 直樹, 教授 松原 和夫, 教授 横出 正之 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

京都大学 / 0048 / 新制・課程博士 / 博士(人間・環境学) / 甲第19903号 / 人博第789号 / 新制||人||190(附属図書館) / 28||人博||789(吉田南総合図書館) / 32980 / 京都大学大学院人間・環境学研究科相関環境学専攻 / (主査)准教授 三浦 智行, 教授 川本 卓男, 教授 宮下 英明 / 学位規則第4条第1項該当 / Doctor of Human and Environmental Studies / Kyoto University / DFAM

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第20971号 / 医博第4317号 / 新制||医||1026(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 河本 宏, 教授 濵﨑 洋子, 教授 松田 秀一 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

Canine Oral Papillomavirus (COPV) induces warts on the oral mucosa in domestic dogs and other canids. The canine oral papillomatosis (COP) is a well established animal model of mucosal papillomatosis. While the regression of a current infection is mediated by cellular immunity, humoral immunity does prevent reinfection. Question of the present study was, if unspecific stimulation of the immune system with the inducer of paramunity Baypamune® during the period of growth would have an influence on the course of canine oral papillomatosis. Clinical criteria for this purpose were period of growth, period of regression and the size and morphology of Papillomas at the end of growth. Additionally ALAT and ASAT were quantified in order to rule out deterioration of liver cells. PCV and leucocytes where monitored and a differentiation was performed. In a second part of the study, L1-antibodies and IFNg, TNF-a as well as IL-18 were determined by the Institute of Virology of the Faculty of Veterinary Medicine of the University of Leipzig. At the end of the study, biopsies of mucosa were taken for detection of viral genome. In a pre-study, three Labrador Retrievers at 14 weeks of age were challenged with 15 μl of virus suspension (40 μg of COPV-L1 Protein/ml) per site by scarification of oral mucosa. Papillomas developed at all challenged sites. These were removed during the period of growth and handed over to the Institute of Virology of the Faculty of Summary 70 Veterinary Medicine of the University of Leipzig for preparation of the suspension used in the main study. In the main study, 13 Labrador Retrievers at 14 weeks of age were challenged with 10 μl of virus suspension (40 μg of COPV-L1 Protein/ml) per site by scarification of three areas of the left oral mucosa. All animals developed at least at one site warts, totally 88 % of the challenged sites showed papillomas. 44 Days after infection, 12 dogs were divided into two groups which did not differ in time of incubation and size of the papillomas. Over a period of four weeks, one group received a total of five dosis of Baypamune®, the other group five dosis of placebo. There was no statistically significant difference between the groups regarding the period of growth, period of regression, size and morphology of the papillomas. The administration of Baypamune® during the period of growth had no effect on the clinical course of COP in this trial. However, time of application of the substances should be considered critically, since papillomas of five animals had been in regression before the first application of Baypamune® and placebo had taken place. No deterioration of liver cells could be ascertained. There was no difference between the groups regarding packed cell volume, white blood count and differentiation throughout the study. Parameters of unspecific immunity determined by the Institute of Virology, IFN-γ, TNF-a und IL-18, delivered no evaluable results. However, four animals of the placebogroup showed Papilloma-DNA in the Biopsies taken, but none of the animals, that received Baypamune®, being this a possible indicator for stimulation of cellular immunity. Anti-L1-Antibodies rose earlier in the Baypamune®group than in the placebogroup. In conclusion therapeutic efficacy of Baypamune® in dogs with present COP could not be shown in this trial, making future investigation necessary.

The 5-HT agonist drug mCPP contributed to a 5-HT hypothesis of obsessive-compulsive disorder (OCD), but the effects of the drug in human and animal studies have been inconsistent. The objective of this thesis was to shed light on the behavioural and neurobiological effects of mCPP using the quinpirole sensitization rat model of OCD and in a reciprocal manner, to use the drug to further reveal behavioural and neurobiological components of the animal model. The utility in using the quinpirole model is that the process of analysis by experimentation can be employed to observe effects of the drug on three separate behavioural components identified to underlie the model compulsive behaviour: vigor, focus and satiety. Four original studies were designed to address this objective, and the findings yielded novel contributions to the literature. We suggest that mCPP attenuates compulsive checking by attenuating the exacerbated vigor and satiety characteristic of compulsive behavior, but this effect may not have been captured in previous clinical studies because OCD was measured as a unitary phenomenon across different symptom subtypes. We also reveal that separate systems underlie the development and performance of compulsive behaviour in the animal model, and mCPP reduces its performance but not its development. Hence, the animal model findings suggest that mCPP can attenuate performance of OCD behavior but the drug does not reverse the pathology of OCD or arrest the pathogenesis of OCD. Neurobiologically, we hypothesize that the underlying mechanism mediating the response to mCPP is mediated downstream of the nucleus accumbens core (NAc), at the substantia nigra pars reticulata, based on the finding that the effects of mCPP on vigor and satiety are present in NAc lesioned animals. Finally, although findings of this thesis indicate that 5-HT2A/C receptors do not mediate the response to mCPP, an oppositional role for DA and 5-HT on the model of compulsive behaviour is proposed, consistent with a security motivation theory of OCD. Overall, this thesis shed new light on the effects of mCPP on OCD, and reveals novel neurobiological and behavioural correlates of the quinpirole model. / Thesis / Doctor of Philosophy (PhD)

<p>  </p> <p>Posttraumatic stress disorder involves a constellation of neural and behavioral alterations in response to trauma exposure. Aside from symptoms involved in posttraumatic stress disorder diagnosis, patients frequently present with working memory impairments. Working memory training has been established as an effective intervention to reduce posttraumatic stress symptoms. Working memory is associated with posttraumatic stress disorder in that it is commonly impaired in patients and that training can reduce the severity of posttraumatic stress symptoms. Taken together, these points suggest the possibility of a shared mechanism between working memory and posttraumatic stress disorder but working memory has not been studied thoroughly in rodent models of posttraumatic stress disorder. The present study utilizes footshock trauma to induce a posttraumatic stress state in rats and evaluates the effect of trauma and trauma-paired cues on working memory performance. Results demonstrate the emergence of chronic deficits in working memory among traumatized animals three weeks post-trauma. Presentation of trauma-paired cues caused further decrement in working memory performance. Regression analysis indicates that the degree of working memory impairment in response to a trauma-paired cue can be significantly predicted by behavioral phenotypes typic of diagnostic symptoms for posttraumatic stress disorder. This study enhances existing animal models by replicating the clinical observations of working memory deficits associated with posttraumatic stress disorder. This will pave the way for future work to probe underlying mechanistic dysregulation of working memory following trauma exposure and for future development of novel treatment strategies. </p>

Tryptophan depletion is a non-pharmacological and non-invasive method extensively used to investigate the role of serotonin (5-hydroxytryptamine in humans and animals. The method is based on reducing the availability of the essential amino acid tryptophan, the dietary serotonin precursor. As a precursor of serotonin, L-Tryptophan has a key role in the regulation of many physiological processes and, inter alia, in the pathology and pathophysiology of neuropsychiatric disorders and diseases. Despite the fact, that the method of tryptophan depletion has been applied in many experimental studies, the exact mechanism, by which tryptophan depletion inducted neurophysiological effects, remain unclear. Also, the protentional use of this method together with other drug coadministration has not been explored in detail yet. In this thesis, the most possible mechanisms of tryptophan depletion are discussed. Biochemical and behavioural effects of low dose of dizocilpine (0.1 mg/kg and 0.15 mg/kg) in animal model of tryptophan depletion are investigated as well. And finally, effects of administration of allopregnanolone and tacrine in model of tryptophan depletion with coadministration of MK-801 are studied. The results show that acute tryptophan depletion with prior starvation, not chronic depletion, caused...

The increasing incidence of disc degeneration and its correlation with lower back pain is an alarming trend in modern society. The research of intervertebral disc degeneration and low back pain would greatly benefit from additional methods to study its etiology and possible treatment methods. A large animal model that maintains the biological and mechanical environment that is most similar to the human lumbar spine could provide substantial improvements in understanding and resolving the problem of intervertebral disc related low back pain.This dissertation presents my doctoral work of investigating the potential for the camelid cervical spine to serve as a suitable animal model for advancing biomechanical research of low back pain and intervertebral disc degeneration in the human lumbar spine. Specifically, this work identifies the cellular, morphological and biomechanical characteristics of the camelid cervical spine and intervertebral disc as compared to the human lumbar spine. My results demonstrate that there are remarkable similarities in all aspects. Many of the similarities with respect to the cellular environment of the intervertebral disc are a consequence of the camelid status as a large mammal. Additional testing of the cellular makeup of the camelid intervertebral disc cells revealed that many human qRT-PCR primers associated with disc degeneration are suitable for use in alpacas without modification. From a biomechanics standpoint, the camelid cervical spine also has a vertically oriented spinal posture and is unsupported near the end in an open kinetic chain, providing a mechanical parallel with the human lumbar spine. The camelid cervical intervertebral disc size is closer to the human lumbar intervertebral disc than all other currently used animal models available for comparison in the literature. Average flexibility (range of motion) of a camelid spinal motion segment showed similarities in all modes of loading. Based on magnetic resonance imaging and radiologic grading of the intervertebral disc, almost 90% of elderly camelids exhibited advanced degeneration (Pfirrmann grade 3 or higher) in their cervical spine, and about half of aged camelids have developed severe degeneration (Pfirrmann grade 4 or higher) in at least one or more of their cervical segments, most commonly within the two lowest cervical segments (e.g. c6c7 and/or c7t1). Thus, while there remain differences, the remarkable similarities between the camelid and human spine strengthen the case for using camelids as a model for human disc degeneration, normal and pathological biomechanics and fluid transport, and potentially as a pre-clinical model for investigating the efficacy of novel spinal devices.

Objective: Atherosclerosis, the main pathology underlying cardiovascular diseases is accelerated in diabetic patients. Genetic mouse models require breeding efforts which are time-consuming and costly. Our aim was to establish a new nongenetic model of inducible metabolic risk factors that mimics hyperlipidemia, hyperglycemia, or both and allows the detection of phenotypic differences dependent on the metabolic stressor(s). Methods and Results: Wild-typemice were injected with gain-of-function PCSK9D377Y (proprotein convertase subtilisin/kexin type 9) mutant adeno-associated viral particles (AAV) and streptozotocin and fed either a high-fat diet (HFD) for 12 or 20 weeks or a high-cholesterol/high-fat diet (Paigen diet, PD) for 8 weeks. To evaluate atherosclerosis, two different vascular sites (aortic sinus and the truncus of the brachiocephalic artery) were examined in the mice. Combined hyperlipidemic and hyperglycemic (HGHCi) mice fed a HFD or PD displayed characteristic features of aggravated atherosclerosis when compared to hyperlipidemia (HCi HFD or PD) mice alone. Atherosclerotic plaques of HGHCi HFD animals were larger, showed a less stable phenotype (measured by the increased necrotic core area, reduced fibrous cap thickness, and less a-SMA-positive area) and had more inflammation (increased plasma IL-1b level, aortic pro-inflammatory gene expression, and MOMA-2-positive cells in the BCA) after 20 weeks of HFD. Differences between the HGHCi and HCi HFD models were confirmed using RNA-seq analysis of aortic tissue, revealing that significantly more genes were dysregulated in mice with combined hyperlipidemia and hyperglycemia than in the hyperlipidemia-only group. The HGHCi-associated genes were related to pathways regulating inflammation (increased Cd68, iNos, and Tnfa expression) and extracellular matrix degradation (Adamts4 and Mmp14). When comparing HFD with PD, the PD aggravated atherosclerosis to a greater extent in mice and showed plaque formation after 8 weeks. Hyperlipidemic and hyperglycemicmice fed a PD (HGHCi PD) showed less collagen (Sirius red) and increased inflammation (CD68-positive cells) within aortic plaques than hyperlipidemic mice (HCi PD). HGHCi-PD mice represent a directly inducible hyperglycemic atherosclerosis model compared with HFD-fed mice, in which atherosclerosis is severe by 8 weeks. Conclusion: We established a nongenetically inducible mouse model allowing comparative analyses of atherosclerosis in HCi and HGHCi conditions and its modification by diet, allowing analyses of multiple metabolic hits in mice.

With chronic post surgical pain affecting up to one third of patients undergoing surgeries and the price of treatment being astoundingly high there has been a transition in research to investigate and identify risk factors. Through identification of risk factors new preventative measures can be taken to ensure better surgical outcomes. The role that psychosocial factors can play in the development of chronic post surgical pain has long been recognized yet its mechanisms are still unknown. We aim to investigate how environment can play a direct role in pain perception and sensitivity. We used a Chronic Mild Stress (CMS) paradigm to induce depression in 10 adult male mice, we used 10 control mice who were left in standard opti cages, and 10 enriched mice who were placed in large enrichment cages. CMS mice were exposed to a series of stressors and all mice underwent spared nerve injury surgery. During spared nerve injury the common peroneal and tibial branches of the sciatic nerve were severed while the sural branch was left intact. Overall well-being and pain threshold of mice were tested via Von Frey, Hot Plate, Heat Place Preference, Dynamic Weight Bearing, Hole Board, and Social Interaction. It was found that CMS mice experienced thermal hyperalgesia yet normal thermal threshold sensation. CMS mice also spent less time interacting with novel mice in social interaction, and less amount of time exploring the center of the hole board arena than control or enriched mice. While Von Frey results did not change over the course of the experiment, dynamic weight bearing results indicated spared nerve injury surgery was successful and produced chronic pain. Results indicate that environment plays a role in thermal pain perception and CMS affected overall well being of mice as CMS mice exhibited more timid and anxious behavior.