Progress 11/01/07 to 09/30/12
Outputs
OUTPUTS: Major activity of the current project is focusing on the identification of new factors involved in lipid metabolism and adipocyte development in food animals. Through this project, we have identified several genes involved in the breakdown of triglyceride in most of food animals. The results from the studies on developmental, hormonal and nutritional regulations of expression of these genes provide basic information that can be used to potentially develop new nutritional and hormonal interventions to regulate fat accretion in food animals. We also developed and injected lentiviral vectors to deliver these target genes into quail embryos, and successfully generated several lines of transgenic quail that exclusively express a target gene in adipose tissue. This is the first transgenic model in the avian species that showed a targeted expression of the transgene in adipose tissue. The avian transgenesis technologies developed from this hatch project will lead to future studies focusing on the effect of other novel target genes in adipose development and lipid metabolism in vivo using transgenic quail. The outcomes from this hatch project will increase our understanding of the roles of novel genes, gene products, and their metabolites on the growth and development of avian adipose tissue. The scientific knowledge gained will be used to develop more effective strategies to regulate fat accretion by providing important selection markers for superior lines of leaner chickens with greater feed efficiency and for providing new targets to drug companies for designing new pharmaceutical interventions for lean chicken production. Because the fundamental biology of adipocyte development and lipid metabolism is very similar across vertebrates, the results of the hatch project will lead to strategies to control fat accretion, not only in poultry, but in other food animals and humans as well. Through the project, 9 graduate students, 3 postdocs, 2 visiting scholars and 2 technicians were trained to learn new assay skills and tasks. The results from the project were presented as a Keynote Lecture in the annual meeting (2012) for the Korean Society of Animal Science and Technology and as an invited talk in the international obesity meeting (2011). Other outreach activities include initiation of two new collaborative research funds with two groups of research partners at Seoul National University for the avian transgenesis and the Rural Development Administration in the Korea Department of Agriculture and Fisheries for cloning of transgenic pigs for xenotransplantation. PARTICIPANTS: Principal investigator, Dr. Kichoon Lee led the project by managing the budget, designing experiments, interpreting data, and preparing data and manuscript for journal publication. Visiting Scholars (Jongteak Yoon, Shin-Ae Oh), Postdoc researchers (Sangsu Shin, Yan Song), graduate students (Jeffery Deiuliis, Jonghyun Shin, Julie Serr, Sarah Donley, Xiang Li, Jinsoo Ahn, Shujin Yang, and Chen Zhang), and laboratory technicians (Yeunsu Suh and Ba-Reum Kim) performed the experiments reported in 2007-2012 and prepared manuscripts for submission to journals. TARGET AUDIENCES: Target audiences will be researchers in the area of animal science, endocrinology, developmental biology, and obesity and diabetes. New information in this project will provide a scientific knowledge for education and a possible application for animal production and pharmaceutical industry. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
1)Generation of transgenic quail expressing a target gene in adipose tissue: Previously, our group produced transgenic quail expressing GFP in various tissues under the Rous sarcoma virus (RSV) promoter. Our recent transgenic quail containing a transgene under the promoter of an adipose-specific gene, fatty acid binding protein 4, have an exclusive expression of the transgene in adipose tissues. This is the first transgenic model in the avian species that showed a targeted expression of the transgene in adipose tissue. The outcome of these studies is that using the avian transgenic technology, other novel genes can be overexpressed in adipose tissue of transgenic quail to understand their functions in adipose tissue growth and development. 2)Loss of Fat with Increased ATGL-Mediated Lipolysis in Adipose Tissue during Laying Stages in Quail: We investigated regulation of key genes involved in lipid metabolism in the adipose to relate lipolytic capacities with physiological changes at the pre-laying, onset of laying, and actively laying stages of quail. We demonstrated for that for the first time in any species the genes involved in lipolysis are upregulated and ATGL protein is phosphorylated during actively laying stages to release NEFA. Our data suggest that the laying birds undergo active lipolysis in the adipocyte with reduction of fat mass, and increase VLDL secretion from the liver in order to secure a lipid supply for yolk maturation. The outcomes of these studies were published in Lipids. 3)Developmental, hormonal, and nutritional regulations of genes involved in lipolysis in food animals: We cloned three genes (ATGL, G0S2 and CGI-58) involved in lipolysis for four different breeds of cattle and three breeds of pigs. The sequences of these genes were compared to relate variations of gene sequences with adiposities of the different breeds. We identified novel alternative splicing isoforms of CGI-58 in cattle and pigs. In addition, developmental, hormonal, and nutritional regulations (fasting/refeeding and food restriction) of expression of these genes were also investigated. The results were published in high impact journals (JAS and Lipids). 4)Identification of adipose-specific novel genes by comparative analysis of microarray data: In addition to extensively analyzing these data sets to identify adipose-specific genes in the mouse and human during last year, we recently performed microarray analysis, using the Affymetrix chicken GeneChips containing 28,000 transcripts (80% of the total transcripts) and identified new sets of adipose-specific genes in chickens. The selected adipose-specific genes for the chicken were compared to the data for the mouse and human. The selected candidate genes were also exclusively expressed in adipose tissues of human and mice, demonstrating conservation of tissue-specific expression of genes across species of animals. We also identified several novel genes that are tissue specific genes for various organs and tissues. Our strategies and results from the comparative analysis of microarray databases were prepared for submission to a high impact journal (PLos ONE).
Publications
- Ahn J, Oh S, Suh Y, Moeller SJ, Lee K. (2013)Porcine G0/G1 Switch Gene 2 (G0S2) expression is regulated during adipogenesis and short-term in vivo nutritional interventions. Lipids. DOI : 10.1007/s11745-013-3756-8.
- Yang S, Suh Y, Choi YM, Shin S, Han JY, Lee K. (2013)Loss of Fat with Increased Adipose Triglyceride Lipase-Mediated Lipolysis in Adipose Tissue during Laying Stages in Quail. Lipids. 48(1):13-21.
- Yasmeen R, Reichert B, Deiuliis J, Yang F, Lynch A, Meyers J, Sharlach M, Shin S, Volz KS, Green KB, Lee K, Alder H, Duester G, Zechner R, Rajagopalan S, Ziouzenkova O. (2013)Autocrine Function of Aldehyde Dehydrogenase 1 as a Determinant of Diet- and Sex-Specific Differences in Visceral Adiposity. Diabetes. 62(1):124-136.
- Li X, Suh Y, Kim BR, Moeller SJ, Lee K. (2012)Alternative splicing and developmental and hormonal regulation of porcine comparative gene identification-58 (CGI-58) mRNA. J Anim Sci. 90(12):4346-4354.
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Progress 01/01/11 to 12/31/11
Outputs
OUTPUTS: Major activity of the current project is focusing on the identification of new factors involved in lipid metabolism and adipocyte development in food animals. Collaborative research is being conducted to understand their roles in adipose development using a primary cell culture system and regulation of gene expression in response to hormonal and nutritional changes in vivo. These results were presented at scientific meetings and reported in peer review journals (Journal of Animal Science and Lipids). Some of these new findings are actively used as teaching materials to provide a current understanding of lipid metabolism and adipocyte development. We also developed and injected lentiviral vectors to deliver these target genes into quail embryos, and successfully generated several lines of transgenic quail. In 2012, using these transgenic quail we will continue to study gene function in adipose and muscle development. PARTICIPANTS: Principal investigator, Dr. Kichoon Lee led the project by managing budget, designing experiments, interpreting data, and preparing data and manuscript for journal publication. Postdoc researchers (Sangsu Shin), graduate students (Julie Serr, Sarah Donley, Xiang Li, Jinsoo Ahn, Shujin Yang, and Chen Zhang), and a laboratory technician (Yeunsu Suh and Ba-Reum Kim) performed the experiments reported in 2011 and prepared manuscript for submission to Journals. Through the project, graduate students, postdoc and technicians were trained to learn new assay skills and tasks. TARGET AUDIENCES: Target audiences will be researchers in the area of animal science, endocrinology, developmental biology, and obesity and diabetes. New information in this project will provide a scientific knowledge for education and a possible application for animal production and pharmaceutical area. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
Research outcomes for 2011 are summarized below. 1) The production of poultry with high growth-rate and low-fat by using the avian transgenesis technologies: We successfully generated several recombinant lentivirus containing ATGL or DLK1 genes that regulate adipose mass. Using these viral vectors, we generated several lines of transgenic quail to study to; 1) determine the effect of overexpression of the avian adipose triglyceride lipase (ATGL) gene in adipose tissue on differentiation of poultry adipocytes, alteration of lipid metabolism, and accretion of adipose tissue, 2) determine whether the overexpression of DLK1 in adipose tissue would inhibit adipocyte development and decrease fat mass in transgenic poultry, and 3) determine the effect of overexpression of ATGL or DLK1 in adipose on the growth-rate and feed efficiency of poultry. We believe high impact discoveries from this project will yield valuable data for publications in high impact journals. 2) Hormonal and nutritional regulation of genes involved in lipolysis of avian adipose tissue: We found that amounts of ATGL protein could not perfectly reflect lipolytic activities, suggesting other regulatory mechanisms are involved in these processes. We demonstrated for the first time that ATGL activity is regulated by modulation of other proteins, an activator and inhibitor of ATGL protein in chicken adipose tissue. The genes encoding activator and inhibitor proteins (CGI-58 and G0S2 genes) were successfully cloned for chickens, turkeys and quail and reported in the Genbank. The hormonal developmental and nutritional regulation of these genes in adipose tissue was performed to relate the function of these genes in the lipolysis of the avian species. The outcomes of these studies were published in three papers in JAS and Lipids. 3) Identification of variation in sequences of genes involved in lipolysis among different breeds of cattle and pigs: Recent studies revealed that mutations of mouse and human genes (ATGL, G0S2 and CGI-58) involved in lipolysis result in increased fat accumulation in adipose tissue and muscle. We cloned three genes for four different breeds of cattle and three breeds of pigs. The sequences of these genes were compared to relate variations of gene sequences with adiposities of the different breeds. We identified novel alternative splicing isoforms of G0S2 and CGI-58 in cattle and pigs. In addition, developmental and hormonal regulation of expression of these genes was also investigated. The results of this project are being prepared for submission of manuscripts to high impact journals. 4) Identification of adipose-specific novel genes by comparative analysis of microarray data: We have initiated a novel strategy to identify adipose-specific genes in this year. Several novel genes were identified recently by comparative analysis of microarray data bases obtained from the Genbank. We are focusing on the function of these genes in the development of adipose tissue and obesity in humans. Two manuscripts are being prepared for submission to high impact journals.
Publications
- Relling AE, Lee K, Loerch SC, Reynolds CK. 2011. Effects of Glucose, Propionate and Splanchnic Hormones on Neuropeptide mRNA concentrations in the Ovine Hypothalamus. Journal of Animal Physiology and Animal Nutrition. 2011 Jul 7. doi: 10.1111/j.1439-0396.2011.01192.x.
- Serr J, Suh Y, and Lee K. 2011. Cloning of comparative gene identification-58 gene in avian species and investigation of its developmental and nutritional regulation in chicken adipose tissue. Journal of Animal Science. Vol. 89, no. 11. : 3490-3500.
- J. Serr, Y. Suh, S.A. Oh, S. Shin, MS. Kim, J.D. Latshaw and K. Lee. 2011. Acute up-regulation of adipose triglyceride lipase and release of non-esterified fatty acids by dexamethasone in chicken adipose tissue. Lipids. Vol. 46. (September): 813-820.
- Jang HJ, Choi JW, Kim YM, Shin SS, Lee K, Han JY. 2011. Reactivation of Transgene Expression by Alleviating CpG Methylation of the Rous sarcoma virus Promoter in Transgenic Quail Cells. Molecular Biotechnology. Vol. 3, no. 49. (November): 222-228.
- W. Huang, H. C. Hines, K. M. Irvin, K. Lee, and M. E. Davis. 2011. Response to divergent selection for insulin-like growth factor-I concentration and correlated responses in growth traits in Angus cattle. Journal of Animal Science. Vol. 89. (December): 3924-3934.
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Progress 01/01/10 to 12/31/10
Outputs
OUTPUTS: The major activity of this project is focusing on the identification of new factors involved in lipid metabolism and adipocyte development in food animals. Our research is being conducted to understand their roles in adipose development using transgenic quail models that are being generated in my laboratory. We also cloned additional genes that are involved in the processes of fat breakdown in adipocytes. In addition, regulation of gene expression in response to hormonal and nutritional changes in vitro and in vivo was studied. These results were presented at scientific meetings and reported in peer review journals (Journal of Animal Science, Poultry Science, and Lipids), and used as preliminary data that led to two international and federal grants (a total of $600,000) in 2010. Some of these new findings are actively used as teaching materials to provide a current understanding of lipid metabolism and adipocyte development. We successfully generated several recombinant lentivirus containing transgenes. These viral vectors were micro-injected into about 400 quail embryos and about 60 embryos were hatched. These potential chimeras were mated with wild-type quail for the production of progenies that are being screened by PCR for germ line transmission of the transgenes. We will continue to work on this project in 2011 to study gene function in adipose and muscle development. PARTICIPANTS: Principal investigator, Dr. Kichoon Lee, led the project by managing the budget, designing experiments, interpreting data, and preparing the data and manuscript for journal publication. Post-doctoral researchers; Dr. Sangsu Shin and Shin-Ae Oh, graduate research associates; Julie Serr, Sarah Donley and Xiang Li, and a research assistant, Yeunsu Suh, performed the experiments reported in 2010 and prepared the manuscript for submission to the various scientific journals. Through the project, graduate students, postdocs and technicians were trained to learn new assay skills and tasks. TARGET AUDIENCES: Target audiences are researchers in the area of animal science, endocrinology, developmental biology, obesity, and diabetes. New information in this project will provide a scientific knowledge for education and a possible application for animal production and pharmaceutical areas. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
Improving muscle growth and decreasing fat accretion in the poultry species is essential in maximizing poultry production. We identified and characterized several genes for enhancing muscle growth and reducing fat mass in poultry species to achieve this goal. In addition, a series of collaborative studies were conducted to provide new genetic and nutritional approaches with positive impacts on poultry production. A multi-disciplinary research team was composed of scientists from the Department of Animal Sciences, and the Department of Human Nutrition at OSU, and internationally well-known scientists in avian transgenesis at Seoul National University in Korea. Our team put together three proposals to determine the role of several new genes in adipose and muscle growth and development in poultry. The two proposals received support ($600,000) from the Global Research Network Program, and USDA-AFRI. Cell culture systems for avian muscle cells and adipocytes, and transgenic birds are being constructed to understand the mechanisms by which these genes increase muscle mass and decrease fat accretion. Under the current economic conditions, the value of a gain in breast yield and feed conversion to the poultry industry are huge. A single percentage increase in breast yield translates to about $36 million per year. In addition, a reduction in fat accretion will also lead to a significant reduction of the production cost. A 0.1 increase in feed conversion is valued at $315 million per year in the poultry industry. The knowledge gained from the current studies will lead to the development of new breeding strategies for selection of superior lines of avian species with greater muscle growth and lesser fat accretion. Additionally, the development of new genetic, nutritional, and hormonal approaches to modulate the expression of the target genes in the muscle and adipose tissues of the poultry species is expected to have positive impacts on poultry production.
Publications
- 1.Oh S, Suh Y, Pang MG, K Lee. 2010. Cloning of avian G(0)/G(1) switch gene 2 (G0S2) genes and developmental and nutritional regulation of G0S2 in chicken adipose tissue. J. Anim Sci. 10.2527/jas.2010-333.
- 2.Deiuliis J, Shin J, Murphy E, Kronberg SL, Eastridge ML, Suh Y, Yoon JT, K Lee. 2010. Bovine adipose triglyceride lipase is not altered and adipocyte fatty acid-binding protein is increased by dietary flaxseed. Lipids. 45:963-973.
- 3.Deiuliis JA, Liu LF, Belury MA, Rim JS, Shin S, and K Lee. 2010. Beta(3)-adrenergic signaling acutely down regulates adipose triglyceride lipase in brown adipocytes. Lipids. 45:479-489.
- 4.Shin S, JY Han, and K Lee. 2010. Cloning of Avian delta-like 1 homolog (Dlk1) gene: The biallelic expression of Dlk1 in avian species. Poult Sci. 89(5):948-955.
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Progress 01/01/09 to 12/31/09
Outputs
OUTPUTS: The major activity of this project is focusing on the identification of new factors involved in lipid metabolism and adipocyte development in food animals. Collaborative research is being conducted to understand their roles in adipose development using primary cell culture system and regulation of gene expression in response to hormonal and nutritional changes in vivo. These results were presented at scientific meetings and reported in peer review journals (Journal of Animal Science, Poultry Science, and Endocrinology). Some of these new findings are actively used as teaching materials to provide a current understanding of lipid metabolism and adipocyte development. We also developed an adenoviral vector system to deliver a transgene into poultry cells for studying gene function in adipose and muscle development. PARTICIPANTS: Principal investigator, Dr. Kichoon Lee led the project by managing the budget, designing experiments, interpreting data, and preparing the data and manuscript for journal publication. Post-doctoral researcher, Dr. Sangsu Shin, graduate research associates, Julie Serr and Jonghyun Shin, and a research assistant,Yeunsu Suh, performed the experiments reported in 2009 and prepared the manuscript for submission to the various scientific journals. Through the project, graduate students, postdocs and technicians were trained to learn new assay skills and tasks. TARGET AUDIENCES: Target audiences are researchers in the area of animal science, endocrinology, developmental biology, obesity and diabetes. New information in this project will provide a scientific knowledge for education and a possible application for animal production and pharmaceutical areas. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
Reduction of subcutaneous fat and enhanced intramuscular fat will be beneficial to livestock producers and the consumer. Our research focuses on discovery of novel factors and understanding their roles in adipose tissue development. Net accretion of fat is determined by a balance between fat synthesis and breakdown. We have identified a new lipase called adipose triglyceride lipase, an enzyme that breaks down triglyceride into fatty acids, for the first time in the pig, chicken, turkey and quail (Poultry Science, Lipids). We reported the developmental, nutritional, and hormonal regulation of the ATGL gene in adipose tissue (Poultry Science). To investigate the expression of candidate genes for marbling in food animals, we developed the cell culture system to convert muscle primary cells into fat cells. Expression of two candidate genes, adipocyte- and muscle-fatty acid binding protein (A-FABP and M-FABP) increased during these processes, suggesting a positive association of A-FABP and M-FABP expression with marbling (Journal of Animal Science). Given the role of these genes in trafficking of fatty acids inside of cells, these genes will be positive selection markers for food animals with high marbling. We have employed DNA microarray and bioinformatics technologies for the rapid genetic analysis of about 12,000 genes to discover novel genes involved in adipocyte development. With the further development of a series of experimental methods, we selected 15 candidate genes as a high priority group to intensively study the role of these candidate genes in adipocyte development and lipid metabolism. The results from the studies on one of these genes, called Interferon-stimulated gene 12 b1 (ISG12B1), were published in Endocrinology. Adenovirus has been used in vivo and in vitro as a vector to carry a foreign gene for gene transfer. However, the feasibility of the adenoviral vector system has not been evaluated in poultry. We found the recombinant adenovirus could successfully deliver the transgene into the various chicken cell types (Journal of Animal Science). Using the adenoviral vector system, target genes can be introduced into poultry species in vitro and in vivo to assess its role in growth and development.
Publications
- Shin J, SG Velleman, JD Latshaw, MP Wick, Y Suh, and K Lee. The ontogeny of Delta-Like Protein 1 mRNA expression during muscle development and regeneration: Comparison of broiler and leghorn chickens. Poult Sci. 2009. 88 (7):1427-1437.
- Liu LF, A Purushotham, AA Wendel, K Koba, J Deiuliis, K Lee, and MA Belury. Regulation of adipose triglyceride lipase by rosiglitazone. Diabetes Obes. Metab. 2009.11(2):131-142.
- Lee K, J Shin, JD Latshaw, Y Suh, and J Serr. Cloning of adipose triglyceride lipase complementary deoxyribonucleic acid in poultry and expression of adipose triglyceride lipase during development of adipose in chickens. Poult. Sci. 2009. 88(3):620-630.
- Serr J, Y Suh, and K Lee. Regulation of adipose triglyceride lipase by fasting and re-feeding in avian species. Poult Sci. 2009. 88(12):2585-2591.
- Shin J, B Li, ME Davis, Y Suh, and K Lee. Comparative analysis of fatty acid binding protein 4 promoters: Conservation of PPAR binding sites. J. Anim. Sci. 2009. 87(12): 3923-3934.
- Shin J, DR Bae, JD Latshaw, MP Wick, JM Reddish, and K Lee. Technical note: A gene delivery system in the embryonic cells of avian species using a human adenoviral vector. J. Anim. Sci. 2009. 87(9):2791-2795.
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Progress 01/01/08 to 12/31/08
Outputs
OUTPUTS: Obesity caused by excessive fat accretion in adipose tissue is a major health issue for the human population. Microarray technologies have been successfully used to discover new genes and their pathways. We have screened 12,000 genes using a gene-chip microarray in order to identify those genes that are highly expressed in adipocytes when compared to preadipocytes. Twenty genes were selected based on >10-fold induction and possible relevance in adipose tissue biology. These genes are further selected for temporal expression during adipose development in vivo and in vitro, nutritional regulation, and tissue distribution of gene expression. The final three candidate genes were dominantly expressed in adipose tissue, and their expression is dramatically increased during adipocyte differentiation. In addition, they are nutritionally regulated, suggesting an important role in adipocyte development. The function of a candidate gene, called interferon stimulated gene B1 (ISG12B1) was evaluated by overexpression of this gene using well-established adenoviral gene expression systems in our lab. In addition, using bioinformatics and genetic tools we assessed the localization of ISG12b1 protein in cellular compartments. The function of this protein in cellular compartment was also investigated. The other two genes are extensively investigated to find their role in fat accretion in animals. PARTICIPANTS: Principal investigator, Dr. Kichoon Lee lead the project by managing budget, designing experiments, interpreting data, and preparing data and manuscript for journal publication. Postdoc researcher, Dr. Bing Li performed most of the experiments reported in 2008 and prepared manuscript for submission to Journal. In addition, a graduate student, Jonghyun Shin assisted experiments and collection of data. Through the project, graduate students and postdocs were trained to learn new assay skills and tasks. TARGET AUDIENCES: Target audiences will be researchers in the area of animal science, endocrinology, developmental biology, and obesity and diabetes. New information in this project will provide a scientific knowledge for education and a possible application for animal production and pharmaceutical area. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
Obesity caused by excessive fat accretion in adipose tissue or the whole body is a major health issue in a significant portion of the US population. The increase in the prevalence of obesity over the last few decades has raised concerns about associated health risks. Major health risks associated with obesity number more than 55 including type 2 diabetes (T2D). Understanding the underlying genetic, nutritional, and hormonal mechanisms that regulate adipocyte glucose and lipid metabolism and development of insulin resistance will provide new strategies for the advancement of obesity and diabetes therapies. As central organelles for glucose and lipid metabolism, mitochondria have gained recent attention as targets for diabetes therapy and dysregulation of mitochondrial function in adipose and muscle has been implicated in the etiology of T2D. Our studies revealed 1) abundant expression of the ISG12b1 gene in adipose tissue; 2) ISG12b1 protein is clearly localized in the mitochondria; 3) adenovirus-mediated expression of the ISG12b1 gene inhibited mitochondrial biogenesis and differentiation of 3T3-L1 preadipocytes into adipocytes; and 4) anti-diabetic drugs, PPAR agonists, that activate mitochondria biogenesis in adipocytes and muscle cells decreased ISG12b1 gene expression. Our experimental evidence provides new insight for an inhibitory role of ISG12b1 in mitochondrial biogenesis and adipocyte lipid accumulation. The scientific knowledge gained from the studies will be used to develop new strategies for the treatment of obesity and the improvement of livestock carcass quality by modulating fat content. For the first year of a total five year project, the results gained from 2008 promise to lead more discoveries of new genes through this project.
Publications
- Li B, Shin J, Lee K.Interferon alpha stimulated gene 12 inhibits adipocyte differentiation by inhibiting mitochondrial biogenesis and function. Endocrinology. 2009. 150(3):1217-24.
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