京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第18894号 / 医博第4005号 / 新制||医||1009(附属図書館) / 31845 / 京都大学大学院医学研究科医学専攻 / (主査)教授 山田 泰広, 教授 戸井 雅和, 教授 小川 誠司 / 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・論文博士 / 博士(医学) / 乙第13000号 / 論医博第2108号 / 新制||医||1016(附属図書館) / 32928 / (主査)教授 森田 智視, 教授 武藤 学, 教授 中山 健夫 / 学位規則第4条第2項該当 / Doctor of Medical Science / Kyoto University / DFAM

Background: Breastfeeding amongst HIV-infected and HIV-exposed mother-infant dyads is a wide-ranging and persistent field in which more investigation is needed. The literature widely recognizes the multifactorial and syndemic nature of HIV and infant feeding, specifically pertaining to maternal and other breastfeeding-associated risks. Findings differed regarding breastfeeding and general developmental outcomes amongst HIV-exposed and HIV-infected infants when compared with HIV-unexposed infants. Evidence, however, suggests slight neurodevelopmental differences in HIV-exposed infants when compared with HIV-unexposed infants, suggesting possible feeding differences. Recent literature also indicated a lack of knowledge among allied health care staff regarding evidence-based counselling content to be provided to mothers concerning single option feeding, breastfeeding outcomes and risks in HIV-affected mother-infant dyads in South Africa. Owing to these varied findings related to HIV-affected mother-infant dyads, synthesising of knowledge regarding HIV, infant breastfeeding outcomes and associated risk factors is warranted. Objective: To critically appraise recent literature regarding breastfeeding outcomes and associated risks in HIV-infected and HIV-exposed infants using the PRISMA-P statement guidelines. Method: Five electronic databases were systematically searched to obtain English publications from the last ten years pertaining to breastfeeding outcomes and associated risks of HIV-infected and HIV-exposed infants and children. Grey literature sources were also included. Data were extracted according to various data items and were synthesised using thematic synthesis. Results: Of the initial 7151 sources identified, 42 articles were deemed eligible for final inclusion. The final selection included 19 cohort studies and two expert committee reports, classified as grey literature. The remaining 21 studies compromised of case-control, cross-sectional, and randomized controlled trial studies. The following themes were identified from the review objectives: breastfeeding outcomes, breastfeeding risk factors, infant growth and developmental outcomes and barriers and facilitators to feeding decisions. Most studies focused on HIV-exposed infants’ growth and developmental outcomes. Exclusive breastfeeding was confirmed to have the best outcomes for all infants, regardless of their HIV status, which in turn supports national and international policies. The most prevalent factors that made it difficult for mothers to breastfeeding were maternal factors affecting decision-making for breastfeeding, followed by biological risk factors. Conclusion: Knowledge regarding breastfeeding outcomes in HIV-exposed and HIV-infected infants remains lacking and further research is necessary. This review emphasised that the majority of HIV-affected mother-infant dyads reside in sub-Saharan Africa, illustrating that health professionals, especially those in sub-Saharan Africa (SSA), have to look beyond their traditional assessment and management focuses to include the factors that can impact successful exclusive breastfeeding. Addressing both infants’ needs and maternal HIV-related needs and risks on macro, meso, and microsystem levels is necessary. / Dissertation (MA (Speech-Language Pathology))--University of Pretoria, 2021. / Speech-Language Pathology and Audiology / MA (Speech-Language Pathology) / Unrestricted

No description available.

Recent evidence has shown the important role of macrophages in both tumor development and progression. To investigate the role of macrophages we used a mouse model known as MaFIA (Macrophage Fas Induced Apoptosis) mice that allows for the selective deletion of macrophages. Mice were given melanoma cells at various stages of depletion. Tumor mass was measured and organs were processed for flow cytometry to measure melanoma cell migration. The results show that mice receiving depletion treatment have larger tumor sizes and weights than those mice retaining their macrophage population. We detect metastasis in both the lung and kidney in both macrophage depleted and non depleted mice. The more macrophages in an organ the larger the amount of melanoma positive cells are detected.

Thesis advisor: Eranthie Weerapana / Adeno-Associated Virus (AAV) has been developed into a powerful therapeutic tool - in the last ten years it has acted as a gene-delivery vehicle in several approved therapeutics and many more therapeutics on trial. Despite extensive research, gaps in our understanding of AAV’s infectious cycle still exist, and further development is needed for the creation of improved gene therapy vectors. Technology to incorporate Unnatural Amino Acids (UAAs) into the AAV capsid has recently been developed, and could aid in both furthering our understanding of AAV’s biology and in the therapeutic advancement of AAV. In this work, we demonstrate how the functionalization of the AAV capsid using UAA incorporation can advance our control over the AAV capsid and aid in probing and manipulating AAV biology. We describe our use UAA incorporation to place a bio-orthogonal reactive handle into AAV’s capsid followed by functionalization with a targeting moiety and demonstrate the unprecedented amount of control that UAA incorporation provides in the creation of a functional virus conjugate. We are able to control both the precise placement and the stoichiometry of the targeting moiety on the AAV capsid, providing a platform that, for the first time, can undergo rigorous optimization analogous to that which medicinal chemists put small molecules through. We also describe the creation of a new platform to site-specifically modify the AAV capsid using cysteine incorporation, a technique that retains the ability to site-specifically modify the capsid as UAA incorporation does, but does not require the excess machinery that UAA incorporation requires. Next we discuss the incorporation of a photocaging amino acid, NBK, into the AAV capsid. Using NBK, we were able to effectively block AAV’s primary binding interaction with Heparan Sulfate Proteoglycan (HSPG) and control the timing of AAV infection using light to chemically remove the photo-protecting group. While photocaging the HSPG interaction is only a proof of concept, it demonstrates the remarkable amount of control that UAA incorporation affords, and lends insight to what could be accomplished using the functionalities that can be placed on the AAV capsid with UAAs. / Thesis (PhD) — Boston College, 2020. / Submitted to: Boston College. Graduate School of Arts and Sciences. / Discipline: Chemistry.

No description available.

Apoptosis is essential for cellular homeostasis and is also a pathologic feature of various diseases. The balance between Bcl-2 family proteins determines whether a cell will live or die. Bax, a member of the BCL-2 family proteins, is a pro-apoptotic protein that exists in both a soluble, cytoplasmic form and a membrane-bound form. Upon apoptotic stimuli, Bax undergoes a conformational change and translocates to the mitochondria, initiating apoptotic events. However, little is known about whether Bax is involved in the regulation of mitochondrial function under non-apoptotic conditions, and how Bax binds to mitochondria to exert its activity. Here, we investigate the role of Bax in the regulation of mitochondrial function under non-apoptotic conditions and explore the molecular mechanisms for Bax binding mitochondria under apoptotic stimuli. Using Bax-containing and Bax-deficient (Bax⁻/⁻) HCT-116 cells, we examined Bax cellular localization and its effects on mitochondria bioenergetics, and also tested whether over-expression of full-length Bax in Bax⁻/⁻ cells would recover mitochondrial metabolic activity. To determine the effects of Bax localization upon mitochondrial function, we measured citrate synthase activity and ATP generation. We showed that Bax localized to the outer and inner mitochondrial membranes in non-apoptotic cells, enabling the activity of citrate synthase and the generation of ATP. Loss of Bax led to impairment of respiring mitochondria morphology and reduced oxidative capacity, all of which was restored by expression of full-length or C-terminal-deleted Bax. These findings indicate that under non-apoptotic conditions, the constitutive expression of Bax is necessary for mitochondrial bioenergetics. To determine the molecular mechanisms for Bax binding mitochondria under apoptotic stimuli, we previously performed in silico-mutagenesis and predicted that Lysines 189/190, in the C-terminal [alpha]9 helix, could regulate Bax binding to mitochondria. We demonstrated here that these lysines are the structural elements responsible for controlling how Bax interacts with the mitochondrial membrane. Expression of full-length Bax led to mitochondrial translocation and apoptosis, whereas deletion of the [alpha]9 helix resulted in cytosolic retention and dramatically reduced cell death. Mutation of the two lysine residues changed how Bax bound to mitochondrial membranes. We replicated the results achieved with full-length Bax by attaching the [alpha]9 helix of Bax to GFP or to a regulatory element, the degradation domain (DD), and induced apoptosis upon expression in cells. We demonstrated that the [alpha]9 helix alone promoted the mitochondrial translocation of Bax and increased apoptosis. These results indicate that the C-terminal [alpha]9 helix could be further studied for use in cancer therapies. Overall, we have demonstrated that the constitutive expression of the inactive form of Bax in non-apoptotic cells is necessary for mitochondrial bioenergetics, and have identified the C-terminal [alpha]9 helix of Bax as the effector domain of apoptotic function.

<p>Environmental exposure to lead (Pb) and cannabis use are two of the largest public health issues facing modern society in the United States and around the world. Exposure to Pb in early life has been unequivocally shown to have negative impacts on development, and recent research is mounting showing that it may also predispose individuals for risk of developing substance use disorders (SUD). At the same time, societal and legal attitudes towards cannabis (main psychoactive component delta-9-tetrahydrocannabinol) have been shifting, and many American states have legalized the recreational use of cannabis. It is also the 3^rdmost widely used drug of abuse in the US, and rates of cannabis use disorder are on the rise. This thesis sets out to establish whether there is a link between early life Pbexposure and later THC-related behavior in C57BL6/J mice, as has been demonstrated for other drugs of abuse. The first aim seeks to answer whether Pbexposure affects physiological THC sensitivity (as measured by the cannabinoid-induced tetrad). The secondaimseeks to answer whether Pbexposure affects edible THC self-administration and the development of THC tolerance (also measured by the tetrad).It was hypothesized that Pbexposure would decrease THC sensitivity (Aim 1), would enhance THC self-administration (Aim 2), enhance the development of THC tolerance (Aim 2), and finally that sex-dependent effects of Pb-exposure and THC would be observed (Aims 1 & 2). These hypotheses ended up not being supported, but Aim 1 produced findings indicating that THC sensitivity was increased by Pbexposure, but only in female mice. Future researchwill hopefully be able to fully explore the implications of these findings.</p>

Despite widespread availability of effective antiretroviral therapy (ART), people living with HIV (PWH) remain at risk of developing comorbidities including HIV-associated neurocognitive impairment (H-NCI). These individuals may then be at an increased risk for treatment non-adherence, which leads to poor quality of life and early mortality. Despite this risk, there is a paucity in trained professionals in low- and middle-income countries with appropriate knowledge and skills to identify H-NCI and make appropriate referrals for additional confirmatory testing or intervention, depending on the severity and context of the screening. General medical doctors, nurses and adherence counsellors provide most HIV related healthcare services at a primary healthcare level in South Africa. However, awareness of the clinical presentation of H-NCI, and their current screening practices among these cadres, is unclear. To address these knowledge gaps this thesis set out to explore the following aims (1) examine existing H-NCI knowledge and practices among healthcare workers delivering HIV services in South Africa, (2) develop an appropriate H-NCI training programme for primary healthcare workers, and (3) lastly, pilot the H-NCI training to determine whether H-NCI screening would be feasible at a primary healthcare level in South Africa. Methods To achieve these objectives, the study was divided into two phases. In phase one, a scoping review identified and summarised published studies addressing brain and/or behaviour training approaches, including H-NCI, targeting frontline HIV healthcare workers in Africa. An online survey was developed and administered to examine existing H-NCI knowledge and current practices among healthcare workers providing HIV services in South Africa. Focus group discussions and in-depth interviews were then conducted to explore knowledge gaps, previous H-NCI training and healthcare workers' perspectives of screening at a primary healthcare level. In phase two, an H-NCI training curriculum was developed and a work-integrated H-NCI training programme targeting primary healthcare workers was piloted. The pilot training assessed knowledge of H-NCI signs and symptoms, healthcare workers' attitude toward and comfort with H-NCI screening tools and healthcare workers ability to accurately administer an H-NCI screening tool. The assessments were repeated two months post-training to evaluate retention of knowledge and skills. Results The scoping review of the existing literature suggested that there were few brain and/ or behaviour training programs targeting healthcare workers providing HIV services in Africa. Of the ten studies identified in the scoping review, one study included H-NCI in the training curriculum. The online survey found that H-NCI knowledge was limited and screening practices virtually non-existent among healthcare workers providing HIV care in South Africa. Qualitative data gathered during the focus group discussions and the in-depth interviews provided greater insight on the existing knowledge and practices gaps as well as highlighting that healthcare workers had not received training on H-NCI. The results from the qualitative investigations showed that primary healthcare workers were in favour of receiving such training. Overall, knowledge of H-NCI improved among primary healthcare workers following the work-integrated H-NCI training programme. The training demonstrated that primary healthcare workers providing clinical services, such as medical doctors or professional nurses were able to administer an H-NCI screening tool. Although knowledge of the clinical presentation of H-NCI improved among adherence counsellors, these healthcare workers experienced challenges in administering the H-NCI screening tool. Conclusion As a body of work, the findings from this thesis suggest that healthcare professionals providing HIV services in South Africa have limited knowledge to identify H-NCI, and screening practices are uncommon. Although training revealed differences between cadres in administering screening tools, healthcare workers providing clinical care, including general medical doctors and professional nurses, may be able to provide H-NCI screening at routine annual visits. Although adherence counsellors are ideally situated in the clinic flow to provide targeted screening by flagging clinical presentation of H-NCI among PWH accessing care, this cadre will require additional training, mentorship and support to successfully administer H-NCI screening tools. However, the feasibility of H-NCI screening at a primary healthcare, timing and nature of any screening remains to be explored. This body of work is a step toward increasing the availability of skilled healthcare workers with appropriate knowledge and skills to screen and identify H-NCI in low- and middle-income countries. The work presented in this thesis provides a foundation for further development of the H-NCI training module and future investigations examining targeted screening strategies at a primary healthcare level, feasibility and access to existing interventions post-screening