京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第21336号 / 医博第4394号 / 新制||医||1031(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 横出 正之, 教授 岩井 一宏, 教授 戸口田 淳也 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

Thermogenic brown adipose tissue generates heat via mitochondrial uncoupling protein-1 (UCP-1), increases whole-body energy expenditure and may protects against obesity and metabolic disorders. White adipocytes store excess energy in the form of triglycerides. UCP-1 positive adipocytes develop within white adipose tissue (beige or brite adipocytes) in response to cold exposure or β3 adrenergic agonists. It was known that beige adipocytes arise from a distinct lineage compared with brown adipocytes, but the developmental origin of the beige adipocytes is still unclear. Signaling pathways that control beige adipocyte determination and formation are essentially unknown. Here, we identified a novel signaling pathway that regulates the lineage specification of beige adipocytes. Bone morphogenetic protein 7 (BMP7), a known brown adipogenesis inducer, suppresses Rho-GTPase kinase (ROCK) and depolymerizes F-actin (filamentous actin) into G-actin (globular actin) in mesenchymal stem cells. G-actin regulates myocardin-related transcription factor A (MRTFA) that co-transactivates serum response factor (SRF) and promotes smooth muscle cell differentiation in various organs. Subcutaneous white adipose tissue from MRTFA-/- mice had enhanced accumulation of UCP-1+ adipocytes and elevated levels of brown-selective proteins. Compared with wild type (WT) controls, MRTFA-/- mice exhibited improved metabolic profiles and were protected from diet-induced obesity and insulin resistance, suggesting that the beige adipocytes are physiologically functional. Compared to WT mice, stromal vascular cells from MRTFA-/- mice expressed higher levels of distinct beige progenitor markers and reduced levels of smooth muscle markers. Our studies demonstrate a novel ROCK-actin-MRTFA/SRF pathway that contributes to the development of beige adipocytes.

Brown and beige fat are specialized adipose tissues found in almost all mammals that can increase energy expenditure and produce heat. Cold exposure and b3-adrenergic stimulation has been extensively shown to activate brown adipose tissue (BAT) in rodents, which promotes uncoupled respiration of glucose and lipid substrates via uncoupling protein 1 (UCP1). Prolonged stimulation can induce white adipose browning, which leads to the emergence of thermogenic cells within white fat depots, called beige adipocytes. The beige adipocyte possesses a unique molecular signature, yet shares several characteristics of brown adipocytes, including high mitochondrial content. When activated, beige fat can be induced to initiate a thermogenic transcriptional program similar to that of BAT. Recent human studies have identified brown and/or beige fat in the supraclavicular region using various radiation imaging modalities. This remarkable discovery has reinvigorated scientific interest in adipose browning and brown/beige fat activation as possible therapeutic targets for obesity. Like in rodents, several groups have previously tested the potential impact of cold exposure and b3-adrenergic agonism on BAT-mediated thermogenesis in humans. However, even though these approaches were shown to significantly increase energy expenditure and promote weight loss in obese individuals, they are not ideal clinical interventions. Cold exposure is uncomfortable and requires prolonged treatment, while b3-adrenergic agonists may lead to many adverse effects like cardiovascular problems. This thesis will evaluate the therapeutic potential and clinical relevance of alternative anti-obesity approaches that target adipose browning and beige adipose activation.

Adipose tissue is an organ that plays an important role in energy storage, nutritional balance and thermogenesis. White and brown adipose tissues have distinct cell morphology and metabolic functions. White adipose tissue (WAT) with unilocular lipid droplets serves as a major site of energy storage, while brown adipose tissue (BAT) with multilocular lipid droplets plays an important role in thermogenesis via a mitochondrial protein, uncoupling protein 1 (UCP1). These cells are derived from mesenchymal stem cells (MSCs). Newly discovered beige adipocytes are derived from the same MSC precursors as WAT but resemble BAT due to expression of UCP1. Due to side effects of drugs for treating obesity, activation of UCP1 positive beige adipocytes in WAT has become a new therapeutic target. The interaction of extracellular matrix (ECM) with integrin was found to regulate cell specification of mesenchymal stem cells (MSCs) via intracellular signaling. However, the role of individual ECM proteins in beige adipogenesis in WAT remains unknown. Therefore, we established a system for culturing stromal vascular fraction (SVF) cells from inguinal WAT on ECM protein coated plates and differentiating the cells into either white or beige adipocytes. We found that cells cultured on type I collagen had more round cell morphology and higher mRNA expression of thermogenic genes, UCP1 and type II iodothyronine deiodinase (DIO2),which was further enhanced in myocardin-related transcription factor A (MRTFA) knockout SVF cells. MRTFA has been reported to regulate beige adipogenesis in BMP-ROCK signaling pathway. Based on our data, we found that type I collagen-integrin signaling regulates beige adipogenesis by controlling the activity of MRTFA in MSCs. Our study has provided an insight into developing therapeutic drugs to enhance beige adipocytes formation in WAT for reducing obesity in the future.

京都大学 / 0048 / 新制・論文博士 / 博士(農学) / 乙第13055号 / 論農博第2839号 / 新制||農||1045(附属図書館) / 学位論文||H28||N5012(農学部図書室) / 33145 / (主査)教授 河田 照雄, 教授 保川 清, 教授 橋本 渉 / 学位規則第4条第2項該当 / Doctor of Agricultural Science / Kyoto University / DFAM

During embryonic development, adipocytes emerge from microvasculature. Lineage-­‐tracing studies in mice have shown that adipocyte progenitors reside in the adipose tissue capillaries. However, the direct evidence of an association between adipocyte progenitors and vasculature in humans is lacking. A specific class of adipocytes (brown and beige/brite) expresses the uncoupling protein 1 (UCP1), which consumes glucose and fatty acids to generate heat. The abundance of UCP1- containing adipocytes correlates with a lean metabolically healthy phenotype in human. However, a causal relationship between the presence of these cells and metabolic benefits in human is not clear. In this thesis, I report human adipocyte progenitors proliferate in response to pro-angiogenic factors in association with adipose capillary networks in-vitro. The capillary-derived adipocytes transform from being UCP1-negative to positive upon adenylate cyclase activation, a defining feature of the brite/beige phenotype. Activated cells have denser, round mitochondria with UCP1 protein, and display uncoupled respiration. When implanted into NOD-scid IL2rgnull (NSG) mice, the adipocytes can form a vascularized fat pad that induces vascularization and becomes integrated into mouse circulatory system. In normal or high fat diet-fed NSG mice, activated brite/beige adipocytes enhance systemic glucose tolerance and improved hepatic steatosis, thus providing evidence for their potential therapeutic use. The adipocytes also express neuroendocrine and secretory factors such as Interleukin-33, proprotein convertase PCSK1 and proenkephalin PENK, which are correlated with human obesity. Finally, analyses on single-cell clones of capillary-sprout cells reveal the existence of diverse adipogenic progenitor populations. Further characterization of the clones will define the identifying features of the diverse adipocyte progenitor types that exist in human adipose tissue.