Progress 10/01/18 to 09/30/19
Outputs
Progress Report Objectives (from AD-416): 1: Identify, using ex vivo model of leptin signaling cascades in hypothalamus, cellular signaling networks that modify leptin-signal transducer and activator of transcription 3 (STAT3) signaling and potentially contribute to leptin resistance. Determine signaling networks that regulate expression of suppressor of cytokine signaling (SOCS3). 1A: Employing ex vivo model, determine if STAT3 signaling acts as signaling hub for distinct signaling pathways mediating cellular leptin resistance. 1B: Identify cellular signaling pathway(s) that regulate suppressor of SOCS3 gene expression by using a SOCS3-Luciferase mouse model. 2: Determine role of a cellular leptin signaling modifier in high fat diet-induced leptin resistance and subsequent alterations in energy and glucose homeostasis, and adiposity. 2A: Determine if SOCS3 commonly mediates ER (endoplasmic reticulum) stress, inflammatory signals, PPARy'agonists, and hyperleptinemia induced- leptin resistance in vivo. 2B: Determine whether conditional deletion of brain SOCS3 is sufficient to reverse diet-induced leptin resistance and obesity. 3: Study mechanism of circadian dysfunction-induced leptin resistance and role of leptin resistance in obesity development by defining mechanism of reciprocal interactions of central circadian clock with leptin-controlled arcuate neurons in hypothalamus, role of hypothalamic central circadian clock in controlling leptin expression in fat tissue, and role of leptin resistance in development of metabolic syndromes and diet-induced obesity. 3A: Study mechanism of reciprocal interactions between central circadian clock and hypothalamic arcuate nucleus in maintaining homeostasis of leptin signaling. 3B: Study role of circadian dysfunction of sympathetic nervous system (SNS) signaling in development of leptin resistance and diet-induced obesity. Approach (from AD-416): Obesity is associated with several co-morbidities, including type 2 diabetes mellitus, several types of cancer, and cardiovascular disease. Reduction in body weight has a beneficial impact on a number of metabolic and cardiovascular risk factors. Thus, developing effective strategies to fight obesity has the potential to reduce the incidence of a myriad of diseases. Leptin is a potent appetite suppressant which negatively regulates energy balance via activation of leptin receptors, particularly those within the central nervous system. Shortly after leptin was identified, it was established that obesity is commonly associated with increased circulating leptin levels which promote the development of leptin resistance. Despite its key role in the development of obesity and obesity-related metabolic disorders, the mechanisms inducing leptin resistance are not well-understood. Recent studies suggest that disruption of normal circadian rhythms induces leptin resistance, thus promoting obesity. Leptin is also known to significantly impact function of the cardiovascular and immune systems, which may have direct consequences for the development of metabolic syndrome and related co- morbidities. Using a combination of unique investigative tools, including novel genetically-modified mouse models, tissue explant and embryo culture techniques, this research will address multiple aspects of the role leptin plays in the development of obesity. This includes the study of molecular mechanisms which contribute to the development of leptin resistance, and mechanisms by which circadian rhythms regulate leptin expression in adipose tissues and signaling within the hypothalamic arcuate nucleus. A significant body of data also suggests an important role for leptin signaling in the regulation of vascular and hematopoietic cells, which may contribute to altered metabolism and function of these cell types in obesity. We will therefore characterize the functional role of leptin in early hemato-vascular cell development, and study its impact on normal cardiovascular and immune cell function during adult homeostasis. For Objective 1, this year we completed all of the necessary studies for generation, validation, and assessment of the genetically engineered reporter mice that can monitor inflammation, and screening of signaling pathways that potentially affect inflammation in Objective 1. We collected and analyzed the results demonstrating that a novel signaling molecule called glycogen synthase kinase 3B (GSK3B) can lead to the inflammatory-like condition in dose- and time-dependent manners. Over the life of Objective 1, the following was accomplished: (1) we generated and validated SOCS-3-Luciferase mouse model, and (2) using the reporter mouse model, we identified a previously unrecognized link between GSK3B and neural inflammation. For Objective 2, this year we used genetically engineered mouse models to define the role of a key signaling molecule called SOCS-3 in controlling body weight and glucose balance, in the brain through leptin, a critical hormone maintaining normal body weight. Following up on our efforts to establish and validate the forebrain- specific SOCS3 knockout mice in the last year, we have been continuing to expand and characterize the mice. We would expect to complete the metabolic study by the end of this year. Over the life of Objective 2, we found that SOCS3 is necessary to mediate endoplasmic reticulum stress- induced leptin resistance. The molecular analysis of the reciprocal interaction between the suprachiasmatic nucleus clock and arcuate nucleus proposed in Objective 3A was delayed in Year 4 due to a critical vacancy. In 2019, we resumed this study and expect to make significant progress and/or complete these studies in the next year. We completed all studies proposed in Objective 3B. Accomplishments 01 Discovery of a molecular pathway in the brain that impacts obesity. The brain is a critical site for the control of body weight and an understanding of how the brain reacts to excess nutrition and mediates obesity remains incomplete. To gain a better understanding of the underlying neural mechanisms of obesity, researchers in Houston, Texas, have developed a new laboratory tool which will help identify a critical molecular pathway that potentially promotes obesity. Using cellular imaging techniques enabled the scientists to visualize gene expression of a molecule that promotes obesity and identify a specific neural pathway involved in the cellular process that may drive obesity. Neural levels of the gene were successfully altered by manipulating the activity of the signaling pathway. These findings will provide a clue regarding the mechanisms that determine body weight and may lead to a novel approach to control body weight by manipulating this particular pathway. 02 The role of sympathetic circadian dysfunction in obesity and cancer. Obesity is closely associated with hypertension in humans, which, in addition to uncontrolled fat gain, is also characterized by over- activation of the sympathetic nervous system and increases the risk of chronic kidney and cardiovascular diseases. The sympathetic nervous system is important in the body's fight or flight response and helps to maintain homeostasis in the body. Researchers in Houston, Texas, found that a disruption in the circadian rhythm (our internal biological awake/sleep clock) elevates sympathetic activity. Blocking circadian disruption induced sympathetic over-activation by treatment with a drug beta-blocker) can reduce diet-induced obesity and completely inhibit obesity-related liver cancer. Beta-blockers have been safely used in treating various hypertension and cardiovascular disorders in humans since 1960s. These findings suggest that beta-blockers also have important therapeutic potentials in the prevention and treatment of obesity and obesity-related liver cancer in humans.
Impacts
(N/A)
Publications
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Progress 10/01/17 to 09/30/18
Outputs
Progress Report Objectives (from AD-416): 1: Identify, using ex vivo model of leptin signaling cascades in hypothalamus, cellular signaling networks that modify leptin-signal transducer and activator of transcription 3 (STAT3) signaling and potentially contribute to leptin resistance. Determine signaling networks that regulate expression of suppressor of cytokine signaling (SOCS3). 1A: Employing ex vivo model, determine if STAT3 signaling acts as signaling hub for distinct signaling pathways mediating cellular leptin resistance. 1B: Identify cellular signaling pathway(s) that regulate suppressor of SOCS3 gene expression by using a SOCS3-Luciferase mouse model. 2: Determine role of a cellular leptin signaling modifier in high fat diet-induced leptin resistance and subsequent alterations in energy and glucose homeostasis, and adiposity. 2A: Determine if SOCS3 commonly mediates ER (endoplasmic reticulum) stress, inflammatory signals, PPARy'agonists, and hyperleptinemia induced- leptin resistance in vivo. 2B: Determine whether conditional deletion of brain SOCS3 is sufficient to reverse diet-induced leptin resistance and obesity. 3: Study mechanism of circadian dysfunction-induced leptin resistance and role of leptin resistance in obesity development by defining mechanism of reciprocal interactions of central circadian clock with leptin-controlled arcuate neurons in hypothalamus, role of hypothalamic central circadian clock in controlling leptin expression in fat tissue, and role of leptin resistance in development of metabolic syndromes and diet-induced obesity. 3A: Study mechanism of reciprocal interactions between central circadian clock and hypothalamic arcuate nucleus in maintaining homeostasis of leptin signaling. 3B: Study role of circadian dysfunction of sympathetic nervous system (SNS) signaling in development of leptin resistance and diet-induced obesity. Approach (from AD-416): Obesity is associated with several co-morbidities, including type 2 diabetes mellitus, several types of cancer, and cardiovascular disease. Reduction in body weight has a beneficial impact on a number of metabolic and cardiovascular risk factors. Thus, developing effective strategies to fight obesity has the potential to reduce the incidence of a myriad of diseases. Leptin is a potent appetite suppressant which negatively regulates energy balance via activation of leptin receptors, particularly those within the central nervous system. Shortly after leptin was identified, it was established that obesity is commonly associated with increased circulating leptin levels which promote the development of leptin resistance. Despite its key role in the development of obesity and obesity-related metabolic disorders, the mechanisms inducing leptin resistance are not well-understood. Recent studies suggest that disruption of normal circadian rhythms induces leptin resistance, thus promoting obesity. Leptin is also known to significantly impact function of the cardiovascular and immune systems, which may have direct consequences for the development of metabolic syndrome and related co- morbidities. Using a combination of unique investigative tools, including novel genetically-modified mouse models, tissue explant and embryo culture techniques, this research will address multiple aspects of the role leptin plays in the development of obesity. This includes the study of molecular mechanisms which contribute to the development of leptin resistance, and mechanisms by which circadian rhythms regulate leptin expression in adipose tissues and signaling within the hypothalamic arcuate nucleus. A significant body of data also suggests an important role for leptin signaling in the regulation of vascular and hematopoietic cells, which may contribute to altered metabolism and function of these cell types in obesity. We will therefore characterize the functional role of leptin in early hemato-vascular cell development, and study its impact on normal cardiovascular and immune cell function during adult homeostasis. Following the completion of generation and functional validation of the genetically engineered reporter mice that can monitor inflammation last year, we started to screen a signaling pathway that potentially affects inflammation. Using brain slices of the reporter mice, we found a novel signaling molecule called glycogen synthase kinase 3B (GSK3B) leads to the inflammatory-like condition. We further confirmed the effect by performing dose- and time-course studies. These results uncover a previously unrecognized link between GSK3B and neural inflammation. In Objective 2, we used genetically engineered mouse models to define the role of a key signaling molecule called SOCS-3 in controlling body weight and glucose balance, in the brain through leptin, a critical hormone maintaining normal body weight. To overcome the problem with the generation of SOCS-3 knockout mice we had last year, we have been generating a new group of the knockout mouse model. We have produced the experimental animals. We confirmed the deletion of the gene and have started the experiment by feeding the knockout mice with a high-fat diet. We would expect to complete the metabolic study by the end of this year. In Objective 3A and 3B, the experiments proposed objectives 3A and 3B both require a large number of animals of several different strains, diverse approaches, and heavy workloads. All mouse models and experimental conditions are established in the Fu lab. However, the experiments in Objective 3A were unable to progress as planned due to critical vacancy (lack of the key personnel to carry on the proposed studies) and fund limitation. In Objective 3B, we have completed all phenotype and major mechanistic studies proposed in Objective 3B. These have led to two major publications. We are in the last stage to complete all mechanistic studies proposed in Objective 3B, and expect a major publication for these studies the next 12 months of time.
Impacts
(N/A)
Publications
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Progress 10/01/16 to 09/30/17
Outputs
Progress Report Objectives (from AD-416): 1: Identify, using ex vivo model of leptin signaling cascades in hypothalamus, cellular signaling networks that modify leptin-signal transducer and activator of transcription 3 (STAT3) signaling and potentially contribute to leptin resistance. Determine signaling networks that regulate expression of suppressor of cytokine signaling (SOCS3). 1A: Employing ex vivo model, determine if STAT3 signaling acts as signaling hub for distinct signaling pathways mediating cellular leptin resistance. 1B: Identify cellular signaling pathway(s) that regulate suppressor of SOCS3 gene expression by using a SOCS3-Luciferase mouse model. 2: Determine role of a cellular leptin signaling modifier in high fat diet-induced leptin resistance and subsequent alterations in energy and glucose homeostasis, and adiposity. 2A: Determine if SOCS3 commonly mediates ER (endoplasmic reticulum) stress, inflammatory signals, PPARy'agonists, and hyperleptinemia induced- leptin resistance in vivo. 2B: Determine whether conditional deletion of brain SOCS3 is sufficient to reverse diet-induced leptin resistance and obesity. 3: Study mechanism of circadian dysfunction-induced leptin resistance and role of leptin resistance in obesity development by defining mechanism of reciprocal interactions of central circadian clock with leptin-controlled arcuate neurons in hypothalamus, role of hypothalamic central circadian clock in controlling leptin expression in fat tissue, and role of leptin resistance in development of metabolic syndromes and diet-induced obesity. 3A: Study mechanism of reciprocal interactions between central circadian clock and hypothalamic arcuate nucleus in maintaining homeostasis of leptin signaling. 3B: Study role of circadian dysfunction of sympathetic nervous system (SNS) signaling in development of leptin resistance and diet-induced obesity. Approach (from AD-416): Obesity is associated with several co-morbidities, including type 2 diabetes mellitus, several types of cancer, and cardiovascular disease. Reduction in body weight has a beneficial impact on a number of metabolic and cardiovascular risk factors. Thus, developing effective strategies to fight obesity has the potential to reduce the incidence of a myriad of diseases. Leptin is a potent appetite suppressant which negatively regulates energy balance via activation of leptin receptors, particularly those within the central nervous system. Shortly after leptin was identified, it was established that obesity is commonly associated with increased circulating leptin levels which promote the development of leptin resistance. Despite its key role in the development of obesity and obesity-related metabolic disorders, the mechanisms inducing leptin resistance are not well-understood. Recent studies suggest that disruption of normal circadian rhythms induces leptin resistance, thus promoting obesity. Leptin is also known to significantly impact function of the cardiovascular and immune systems, which may have direct consequences for the development of metabolic syndrome and related co- morbidities. Using a combination of unique investigative tools, including novel genetically-modified mouse models, tissue explant and embryo culture techniques, this research will address multiple aspects of the role leptin plays in the development of obesity. This includes the study of molecular mechanisms which contribute to the development of leptin resistance, and mechanisms by which circadian rhythms regulate leptin expression in adipose tissues and signaling within the hypothalamic arcuate nucleus. A significant body of data also suggests an important role for leptin signaling in the regulation of vascular and hematopoietic cells, which may contribute to altered metabolism and function of these cell types in obesity. We will therefore characterize the functional role of leptin in early hemato-vascular cell development, and study its impact on normal cardiovascular and immune cell function during adult homeostasis. For Objective 1, we validated the genetically engineered reporter mice that can monitor inflammation. First, we showed that the reporter properly detects inflammation in a living animal. Second, we used brain slices from the mice to monitor changes in inflammation in the brain areas involved in body weight control. These results would support the development a novel screening system that enables us to discover novel chemicals or compounds that control the gene of the brain�s control of body weight. In Objective 2, we used genetically engineered mouse models to define the role of a key signaling molecule called SOCS-3 in controlling body weight and glucose balance. This occurs in the brain through leptin, a critical hormone maintaining normal body weight. During the course of the experiments, we found that the mice used, which are supposed to have the deleted gene, still show the gene expression. Thus, results generated using this mouse model cannot be explained by the deletion of the targeted gene. To further address this problem, we decided to utilize a newly generated knockout mouse model. To this end, we have purchased a new mouse type and are currently producing the mouse lines. We would expect that the mouse model would be generated by the end of this year. In Objective 3A, our milestone was partially met due to unexpected difficulties in experiments and shortage of staff. These issues led us to modify our research plan to generate different mouse models for our studies. Currently, we are in the process of generating the last mouse model for the second time, because the mouse model generated initially showed an abnormal phenotype. These unexpected difficulties in generating animal models caused a significant delay in experiments proposed in Objective 3A due to an increased work load that exceeded the capacity of our staff. In Objective 3B, we have generated all mouse models for experiments proposed and completed the first part of our study on the role of sympathetic dysfunction in obesity, liver metabolic disorders, and obesity-related cancers. The results from these studies have been published in the journal Cancer Cell in 2016, which demonstrated that circadian disruption of sympathetic nervous system leads to increased metabolic syndrome and liver tumor. Accomplishments 01 Cellular signaling pathway that mediates obesity. Brain mechanisms that link diet and obesity are not fully understood. Researchers in Houston, Texas recently identified a novel signaling pathway that actively responds to high-fat diet feeding and mediates leptin resistance, obesity and glucose imbalance. Consuming a high-fat diet results in changes in the brain that lead to a decreased sensitivity to leptin, the 'satiety hormone' that helps regulate body weight by inhibiting appetite. We examined the role of a signaling molecule called Rap1 in body weight control and glucose balance and found that mice lacking the Rap 1 gene did not gain body fat when fed a high-fat diet. This new mechanism involving Rap1 in the brain may represent a potential therapeutic target for treating human obesity in the future. 02 The role of circadian dysfunction in metabolic syndrome and cancer. Researchers are attempting to answer if chronic circadian disruption alone leads to obesity and cancer. Scientists in Houston, Texas conducted research that demonstrated that chronic circadian disruption alone is sufficient to induce metabolic syndrome, fatty liver disease, and cancer. Chronic circadian disruption is affecting the majority people in the U.S. now due to lifestyle changes. Our findings lead to a better understanding of how unhealthy lifestyle factors can increase the risk of obesity and cancer, and will lead to development of novel strategies for obesity and cancer prevention and treatment.
Impacts
(N/A)
Publications
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Progress 10/01/15 to 09/30/16
Outputs
Progress Report Objectives (from AD-416): 1: Identify, using ex vivo model of leptin signaling cascades in hypothalamus, cellular signaling networks that modify leptin-signal transducer and activator of transcription 3 (STAT3) signaling and potentially contribute to leptin resistance. Determine signaling networks that regulate expression of suppressor of cytokine signaling (SOCS3). 1A: Employing ex vivo model, determine if STAT3 signaling acts as signaling hub for distinct signaling pathways mediating cellular leptin resistance. 1B: Identify cellular signaling pathway(s) that regulate suppressor of SOCS3 gene expression by using a SOCS3-Luciferase mouse model. 2: Determine role of a cellular leptin signaling modifier in high fat diet-induced leptin resistance and subsequent alterations in energy and glucose homeostasis, and adiposity. 2A: Determine if SOCS3 commonly mediates ER (endoplasmic reticulum) stress, inflammatory signals, PPARy'agonists, and hyperleptinemia induced- leptin resistance in vivo. 2B: Determine whether conditional deletion of brain SOCS3 is sufficient to reverse diet-induced leptin resistance and obesity. 3: Study mechanism of circadian dysfunction-induced leptin resistance and role of leptin resistance in obesity development by defining mechanism of reciprocal interactions of central circadian clock with leptin-controlled arcuate neurons in hypothalamus, role of hypothalamic central circadian clock in controlling leptin expression in fat tissue, and role of leptin resistance in development of metabolic syndromes and diet-induced obesity. 3A: Study mechanism of reciprocal interactions between central circadian clock and hypothalamic arcuate nucleus in maintaining homeostasis of leptin signaling. 3B: Study role of circadian dysfunction of sympathetic nervous system (SNS) signaling in development of leptin resistance and diet-induced obesity. Approach (from AD-416): Obesity is associated with several co-morbidities, including type 2 diabetes mellitus, several types of cancer, and cardiovascular disease. Reduction in body weight has a beneficial impact on a number of metabolic and cardiovascular risk factors. Thus, developing effective strategies to fight obesity has the potential to reduce the incidence of a myriad of diseases. Leptin is a potent appetite suppressant which negatively regulates energy balance via activation of leptin receptors, particularly those within the central nervous system. Shortly after leptin was identified, it was established that obesity is commonly associated with increased circulating leptin levels which promote the development of leptin resistance. Despite its key role in the development of obesity and obesity-related metabolic disorders, the mechanisms inducing leptin resistance are not well-understood. Recent studies suggest that disruption of normal circadian rhythms induces leptin resistance, thus promoting obesity. Leptin is also known to significantly impact function of the cardiovascular and immune systems, which may have direct consequences for the development of metabolic syndrome and related co- morbidities. Using a combination of unique investigative tools, including novel genetically-modified mouse models, tissue explant and embryo culture techniques, this research will address multiple aspects of the role leptin plays in the development of obesity. This includes the study of molecular mechanisms which contribute to the development of leptin resistance, and mechanisms by which circadian rhythms regulate leptin expression in adipose tissues and signaling within the hypothalamic arcuate nucleus. A significant body of data also suggests an important role for leptin signaling in the regulation of vascular and hematopoietic cells, which may contribute to altered metabolism and function of these cell types in obesity. We will therefore characterize the functional role of leptin in early hemato-vascular cell development, and study its impact on normal cardiovascular and immune cell function during adult homeostasis. This year we have generated and established some of the mouse lines to be used in our research studies. Additionally, we established ex vivo models of leptin resistance induced by distinct conditions by treating organotypic brain slices with pharmacological agents that induce these conditions in Objective 1A. We have also started the initial assessment of the mice produced during FY2016 for Objectives 1B, 2A and 2B. We have tested the arcuate nucleus and central clock co-culture condition for Objective 3A, and completed analyzing the effects of high fat diet intake on body weight and body composition, physical activity and energy expenditure of the mutant mice as proposed in Objective 3B. While the milestone for Objective 2A is to prepare a mouse model during this period, due to an unexpected inefficiency in mouse breeding to produce an experimental cohort, we expect to obtain enough number of the mice needed to perform the assay by the end of 2016. In addition, there has been a delay in the preparation of a manuscript from Objectives 3A and 3B. This delay is due to the needs to change the mouse model expressing firefly luciferase reporter, which was fused with a circadian gene and used for detecting the activity of molecular clock in the brain of the original mouse model. However, we found that this mouse model displays a firefly luciferase level that was too low to be stably detected in mouse brain at all times over a 24 hour period. In addition, we also encountered significant difficulties in real-time recording the reporter activity over a consecutive 3 days as proposed in Objective 3A using a shared Real-Time Bioluminescence Monitoring System located at other labs. To address these problems, we generated two new reporter mouse models expressing higher basal levels of firefly reporter, and are currently seeking options of establishing an independent real- time reporter activity recording set-up for circadian research only. The studies in Objective 3B.1 were aimed to define the role of the sympathetic nervous system white fat innervation in body weight control via controlling the expression of leptin, a hormone that is exclusively produced in the white fat tissues and controls appetite. We have completed all experiments originally proposed in this objective. However, we found that mice used in this study, which are deficient in sympathetic signaling in all cells in the body, developed white fat in the brown fat tissue. As a result, these mice display a low energy expenditure rate and developed obesity regardless of leptin expression in the white fat. Thus, the results generated using these mouse model cannot be explained by serum levels of leptin. To address this problem, we decided to generate mouse models that lack sympathetic function only in the white fat. To this end, we have generated the founder mice that can be used for generating mice lacking sympathetic signaling in white fat in the future. In additional to the studies described above, our research activities in the last year have produced 4 manuscripts. The first manuscript demonstrates that chronic circadian disruption, which has reached the epidemic level in the U.S., induces metabolic syndrome including hyperinsulinemia, hyperglycemia, dyslipidemia and non- alcoholic fatty liver in mice, as well chronic liver inflammation. Together, these abnormalities lead to non-alcoholic steatohepatitis and liver fibrosis that promote liver tumor in mice. This pathophysiological pathway of liver tumor initiation found in our mouse model closely mimics that which is frequently observed for spontaneous liver tumor development in obese humans. Liver tumor was previously considered a rare type of cancer in the U.S. but there has been a rapid increase in recent years, coupled with the obesity pandemic. However, although fatty liver disease is predicted to be a leading cause of liver cancer in the 21st century, the lack of animal models that develop liver tumor following the same mechanism as obese humans leads to a poor understanding and a completely lack of early detection and prognosis makers for this disease. As a result, liver tumor is currently the fastest rising cause of cancer- related death. Our studies not only establish a useful animal model for studying the mechanisms of fatty liver disease and fatty liver-induced liver cancer, but also identified the key gene pathways that are potential therapeutic targets for liver tumor prevention and treatment in humans. Thus, these findings have important therapeutic implications. This study is currently in consideration for publication. The second manuscript resulted from a collaborative project, which demonstrates that chronic circadian disruption deregulates both innate and adaptive immune responses at the systemic level to promote fatty liver disease. The third manuscript is generated from a collaborative project, which defines the role of the gene encoding a transcriptional regulator, the small heterodimer partner (SHP), in controlling the time of food-intake over a 24 hour period. This study is currently in consideration for publication. Our fourth manuscript demonstrates a new molecular pathway that mediates neural leptin resistance, obesity and metabolic diseases under overnutrition. We have shown that a cellular signaling molecule (small GTPase Rap1) is expressed in hypothalamic areas that control whole-body metabolism and is activated in high-fat-diet (HFD)-induced obesity. Deletion of the molecule in the brain Rap1 protects mice from dietary obesity and its-associated metabolic disturbance in the periphery and from HFD-induced neuropathological changes in the hypothalamus. Furthermore, a small chemical inhibitor of the molecule normalizes hypothalamic pathological alterations, and reduces body weight in mice with dietary obesity. Based on these results, we concluded that neuronal Rap1 critically regulates leptin sensitivity and mediates HFD-induced obesity and hypothalamic pathology and may represent a potential therapeutic target for obesity treatment. Accomplishments 01 The role of circadian control of leptin expression and signaling. Research studies focusing on the pathophysiological mechanism of body weight control is difficult and new efficient tools and techniques are in need to be developed or obtained. Researchers at the Children's Nutrition Research Center in Houston, Texas have developed new mouse models for studying the interaction between the brain and the fat tissues which are essential for maintaining the homeostasis of body weight control. Our team also developed a new technique that can be used to monitor the dynamic interaction of the central circadian clock with the hypothalamus energy homeostasis center. These tools will aid our understanding of the physiological role of circadian expression and signaling of leptin and these new tools and approaches will be invaluable to our understanding the pathophysiological mechanisms of circadian dynamics in disease prevention and treatment.
Impacts
(N/A)
Publications
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Progress 10/01/14 to 09/30/15
Outputs
Progress Report Objectives (from AD-416): 1: Identify, using ex vivo model of leptin signaling cascades in hypothalamus, cellular signaling networks that modify leptin-signal transducer and activator of transcription 3 (STAT3) signaling and potentially contribute to leptin resistance. Determine signaling networks that regulate expression of suppressor of cytokine signaling (SOCS3). 1A: Employing ex vivo model, determine if STAT3 signaling acts as signaling hub for distinct signaling pathways mediating cellular leptin resistance. 1B: Identify cellular signaling pathway(s) that regulate suppressor of SOCS3 gene expression by using a SOCS3-Luciferase mouse model. 2: Determine role of a cellular leptin signaling modifier in high fat diet-induced leptin resistance and subsequent alterations in energy and glucose homeostasis, and adiposity. 2A: Determine if SOCS3 commonly mediates ER (endoplasmic reticulum) stress, inflammatory signals, PPARy'agonists, and hyperleptinemia induced- leptin resistance in vivo. 2B: Determine whether conditional deletion of brain SOCS3 is sufficient to reverse diet-induced leptin resistance and obesity. 3: Study mechanism of circadian dysfunction-induced leptin resistance and role of leptin resistance in obesity development by defining mechanism of reciprocal interactions of central circadian clock with leptin-controlled arcuate neurons in hypothalamus, role of hypothalamic central circadian clock in controlling leptin expression in fat tissue, and role of leptin resistance in development of metabolic syndromes and diet-induced obesity. 3A: Study mechanism of reciprocal interactions between central circadian clock and hypothalamic arcuate nucleus in maintaining homeostasis of leptin signaling. 3B: Study role of circadian dysfunction of sympathetic nervous system (SNS) signaling in development of leptin resistance and diet-induced obesity. Approach (from AD-416): Obesity is associated with several co-morbidities, including type 2 diabetes mellitus, several types of cancer, and cardiovascular disease. Reduction in body weight has a beneficial impact on a number of metabolic and cardiovascular risk factors. Thus, developing effective strategies to fight obesity has the potential to reduce the incidence of a myriad of diseases. Leptin is a potent appetite suppressant which negatively regulates energy balance via activation of leptin receptors, particularly those within the central nervous system. Shortly after leptin was identified, it was established that obesity is commonly associated with increased circulating leptin levels which promote the development of leptin resistance. Despite its key role in the development of obesity and obesity-related metabolic disorders, the mechanisms inducing leptin resistance are not well-understood. Recent studies suggest that disruption of normal circadian rhythms induces leptin resistance, thus promoting obesity. Leptin is also known to significantly impact function of the cardiovascular and immune systems, which may have direct consequences for the development of metabolic syndrome and related co- morbidities. Using a combination of unique investigative tools, including novel genetically-modified mouse models, tissue explant and embryo culture techniques, this research will address multiple aspects of the role leptin plays in the development of obesity. This includes the study of molecular mechanisms which contribute to the development of leptin resistance, and mechanisms by which circadian rhythms regulate leptin expression in adipose tissues and signaling within the hypothalamic arcuate nucleus. A significant body of data also suggests an important role for leptin signaling in the regulation of vascular and hematopoietic cells, which may contribute to altered metabolism and function of these cell types in obesity. We will therefore characterize the functional role of leptin in early hemato-vascular cell development, and study its impact on normal cardiovascular and immune cell function during adult homeostasis. We have recently initiated studies to investigate the molecular mechanisms of cellular leptin resistance (Objectives 1 and 2) and the role of the circadian clock in the prevention of obesity (Objectives 3 and 4). The current experiments are focused on generating mouse models and establishing essential techniques for the experiments proposed. Some of the mouse lines have already been established and we are currently validating the mouse models for the studies in Objectives 1 and 2. For Objectives 3 and 4, the milestone in the first 12 month is to prepare mouse models for proposed research. Due to an unexpected inefficiency in mouse breeding and a delay in generating new mouse models, we expect to obtain all mouse models needed in the next 6 months (at the end of year 2015). Furthermore, we established the ex vivo model system by assessing the effects of multiple reagents that are reported to induce cellular leptin resistance. The results clearly suggest that organotypic brain slices are an ideal system as the ex vivo model to recapitulate multiple forms of leptin resistance. In the following years, using our novel ex vivo system, we will perform further analysis to dissect signaling pathways mediating leptin resistance. Any animal research conducted have received IACUC approval. Any human research studies conducted have received IRB approval. Accomplishments 01 Circadian disruption increases the risk of obesity. Obesity continues to be a significant concern and is more prevalent in individuals that perform shift-work due a disruption in their circadian clock. Children's Nutrition Research Center scientists in Houston, TX reported that frequent disruption of the circadian homeostasis, which has reached epidemic levels in the U.S., can directly increase fat gain independent of all previously identified obesity risk factors, such as gene mutation, diet components and the amount of physical exercise. This discovery, will have a significant impact on obesity research and will support the development of more efficient weight watching programs for obesity prevention.
Impacts
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Publications
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