Advertisement

Metabolomics

, Volume 11, Issue 5, pp 1287–1301 | Cite as

Non-targeted metabolomics of Brg1/Brm double-mutant cardiomyocytes reveals a novel role for SWI/SNF complexes in metabolic homeostasis

  • Ranjan Banerjee
  • Scott J. Bultman
  • Darcy Holley
  • Carolyn Hillhouse
  • James R. Bain
  • Christopher B. Newgard
  • Michael J. Muehlbauer
  • Monte S. Willis
Original Article

Abstract

Mammalian SWI/SNF chromatin-remodeling complexes utilize either BRG1 or Brm as alternative catalytic subunits to alter the position of nucleosomes and regulate gene expression. Genetic studies have demonstrated that SWI/SNF complexes are required during cardiac development and also protect against cardiovascular disease and cancer. However, Brm constitutive null mutants do not exhibit a cardiomyocyte phenotype and inducible Brg1 conditional mutations in cardiomyocyte do not demonstrate differences until stressed with transverse aortic constriction, where they exhibit a reduction in cardiac hypertrophy. We recently demonstrated the overlapping functions of Brm and Brg1 in vascular endothelial cells and sought here to test if this overlapping function occurred in cardiomyocytes. Brg1/Brm double mutants died within 21 days of severe cardiac dysfunction associated with glycogen accumulation and mitochondrial defects based on histological and ultrastructural analyses. To determine the underlying defects, we performed nontargeted metabolomics analysis of cardiac tissue by GC/MS from a line of Brg1/Brm double-mutant mice, which lack both Brg1 and Brm in cardiomyocytes in an inducible manner, and two groups of controls. Metabolites contributing most significantly to the differences between Brg1/Brm double-mutant and control-group hearts were then determined using the variable importance in projection analysis. Increased cardiac linoleic acid and oleic acid suggest alterations in fatty acid utilization or intake are perturbed in Brg1/Brm double mutants. Conversely, decreased glucose-6-phosphate, fructose-6-phosphate, and myoinositol suggest that glycolysis and glycogen formation are impaired. These novel metabolomics findings provide insight into SWI/SNF-regulated metabolic pathways and will guide mechanistic studies evaluating the role of SWI/SNF complexes in homeostasis and cardiovascular disease prevention.

Keywords

SWI/SNF complex BRG1 BRM Cardiomyocyte Metabolomics Fatty acid Glucose 

Abbreviation

Ang II

Angiotensin II

BAFs

BRG1- or BRM-associated factors

Brg1

Brahma-related gene 1

Brm

Brahma

Flx

LoxP-flanked DNA polymerase gene

F-6-P

Fructose-6-phosphate

G-6-P

Glucose-6-phosphate

α-MHC-Cre-ERT

Cre recombinase fused to a mutated ligand-binding domain of human estrogen receptor

RXRα

Retinoid X receptor alpha

PPAR

Peroxisome proliferator activated receptor

PGC-1α

PPAR-gamma coactivator 1-alpha

VECs

Vascular endothelial cells

VIP

Variable importance in projection

Notes

Acknowledgments

The authors would like to thank Kumar Pandya for providing the αMHC-Cre-ERT transgenic mice and advice on adding tamoxifen to the rodent chow. The authors would like to acknowledge Janice Weaver (University of North Carolina Animal Histopathology Laboratory) and Victoria Madden (University of North Carolina Microscopy Services Laboratory) for assistance in preparing histological specimens and performing the TEM, respectively. Finally, the authors would also like to thank Tim Koves for his guidance and valuable discussion and suggestions for harvesting and preparing heart samples for metabolomics analysis. This work was supported by the National Institutes of Health (R01HL104129 to M.W. and RO1 CA125237 to S.J.B.), a Jefferson-Pilot Corporation Fellowship (to M.W.), and the Fondation Leducq (to M.W.).

Conflict of interest

The authors declare that they have no conflict of interest.

Compliance with Ethical Standards

All applicable international, national, and/or institutional guidelines for the care and use of animals were followed.

Supplementary material

11306_2015_786_MOESM1_ESM.pptx (3.7 mb)
Supplementary material 1 (PPTX 3782 kb)
11306_2015_786_MOESM2_ESM.xlsx (161 kb)
Supplementary material 2 (XLSX 160 kb)

References

  1. Bain, J. R., Stevens, R. D., Wenner, B. R., Ilkayeva, O., Muoio, D. M., & Newgard, C. B. (2009). Metabolomics applied to diabetes research: Moving from information to knowledge. Diabetes, 58, 2429–2443. doi: 10.2337/db09-0580.PubMedCentralCrossRefPubMedGoogle Scholar
  2. Bevilacqua, A., Willis, M. S., & Bultman, S. J. (2013). SWI/SNF chromatin-remodeling complexes in cardiovascular development and disease. Cardiovascular Pathology,. doi: 10.1016/j.carpath.2013.09.003.PubMedCentralPubMedGoogle Scholar
  3. Bultman, S., et al. (2000). A Brg1 null mutation in the mouse reveals functional differences among mammalian SWI/SNF complexes. Molecular Cell, 6, 1287–1295.CrossRefPubMedGoogle Scholar
  4. Carley, A. N., Taegtmeyer, H., & Lewandowski, E. D. (2014). Matrix revisited: Mechanisms linking energy substrate metabolism to the function of the heart. Circulation Research, 114, 717–729. doi: 10.1161/CIRCRESAHA.114.301863.PubMedCentralCrossRefPubMedGoogle Scholar
  5. Chang, C. P., & Bruneau, B. G. (2012). Epigenetics and cardiovascular development. Annual Review of Physiology, 74, 41–68. doi: 10.1146/annurev-physiol-020911-153242.CrossRefPubMedGoogle Scholar
  6. Curtis, C. D., Davis, R. B., Ingram, K. G., & Griffin, C. T. (2012). Chromatin-remodeling complex specificity and embryonic vascular development. Cellular and Molecular Life Sciences,. doi: 10.1007/s00018-012-1023-4.PubMedCentralPubMedGoogle Scholar
  7. Debril, M. B., Gelman, L., Fayard, E., Annicotte, J. S., Rocchi, S., & Auwerx, J. (2004). Transcription factors and nuclear receptors interact with the SWI/SNF complex through the BAF60c subunit. Journal of Biological Chemistry, 279, 16677–16686. doi: 10.1074/jbc.M312288200.CrossRefPubMedGoogle Scholar
  8. Doenst, T., Nguyen, T. D., & Abel, E. D. (2013). Cardiac metabolism in heart failure: Implications beyond ATP production. Circulation Research, 113, 709–724. doi: 10.1161/CIRCRESAHA.113.300376.PubMedCentralCrossRefPubMedGoogle Scholar
  9. Drogan, D., et al. (2014). Untargeted metabolic profiling identifies altered serum metabolites of type 2 diabetes mellitus in a prospective, nested case-control study. Clinical Chemistry,. doi: 10.1373/clinchem.2014.228965.PubMedGoogle Scholar
  10. Fiehn, O., et al. (2008). Quality control for plant metabolomics: Reporting MSI-compliant studies. Plant Journal, 53, 691–704. doi: 10.1111/j.1365-313X.2007.03387.x.CrossRefPubMedGoogle Scholar
  11. Files, D. C., et al. (2012). A critical role for muscle ring finger-1 in acute lung injury-associated skeletal muscle wasting. American Journal of Respiratory and Critical Care Medicine, 185, 825–834. doi: 10.1164/rccm.201106-1150OC.PubMedCentralCrossRefPubMedGoogle Scholar
  12. Floegel, A., et al. (2013). Identification of serum metabolites associated with risk of type 2 diabetes using a targeted metabolomic approach. Diabetes, 62, 639–648. doi: 10.2337/db12-0495.PubMedCentralCrossRefPubMedGoogle Scholar
  13. Frazier, D. M., et al. (2006). The tandem mass spectrometry newborn screening experience in North Carolina: 1997-2005. Journal of Inherited Metabolic Disease, 29, 76–85. doi: 10.1007/s10545-006-0228-9.CrossRefPubMedGoogle Scholar
  14. Garcia, M., et al. (2009). Phosphofructo-1-kinase deficiency leads to a severe cardiac and hematological disorder in addition to skeletal muscle glycogenosis. PLOS Genetics, 5, e1000615. doi: 10.1371/journal.pgen.1000615.PubMedCentralCrossRefPubMedGoogle Scholar
  15. Gatfield, D., et al. (2009). Integration of microRNA miR-122 in hepatic circadian gene expression. Genes and Development, 23, 1313–1326. doi: 10.1101/gad.1781009.PubMedCentralCrossRefPubMedGoogle Scholar
  16. Gresh, L., et al. (2005). The SWI/SNF chromatin-remodeling complex subunit SNF5 is essential for hepatocyte differentiation. EMBO Journal, 24, 3313–3324. doi: 10.1038/sj.emboj.7600802.PubMedCentralCrossRefPubMedGoogle Scholar
  17. Griffin, C. T., Brennan, J., & Magnuson, T. (2008). The chromatin-remodeling enzyme BRG1 plays an essential role in primitive erythropoiesis and vascular development. Development, 135, 493–500. doi: 10.1242/dev.010090.PubMedCentralCrossRefPubMedGoogle Scholar
  18. Griffin, C. T., Curtis, C. D., Davis, R. B., Muthukumar, V., & Magnuson, T. (2011). The chromatin-remodeling enzyme BRG1 modulates vascular Wnt signaling at two levels. Proceedings of the National Academy of Sciences USA, 108, 2282–2287. doi: 10.1073/pnas.1013751108.CrossRefGoogle Scholar
  19. Halket, J. M., Przyborowska, A., Stein, S. E., Mallard, W. G., Down, S., & Chalmers, R. A. (1999). Deconvolution gas chromatography/mass spectrometry of urinary organic acids–potential for pattern recognition and automated identification of metabolic disorders. Rapid Communications in Mass Spectrometry, 13, 279–284. doi: 10.1002/(SICI)1097-0231(19990228).CrossRefPubMedGoogle Scholar
  20. Han, P., Hang, C. T., Yang, J., & Chang, C. P. (2011). Chromatin remodeling in cardiovascular development and physiology. Circulation Research, 108, 378–396. doi: 10.1161/CIRCRESAHA.110.224287.PubMedCentralCrossRefPubMedGoogle Scholar
  21. Hang, C. T., et al. (2010). Chromatin regulation by Brg1 underlies heart muscle development and disease. Nature, 466, 62–67. doi: 10.1038/nature09130.PubMedCentralCrossRefPubMedGoogle Scholar
  22. He, Q., & Han, X. (2014). Cardiolipin remodeling in diabetic heart. Chemistry and Physics of Lipids, 179, 75–81. doi: 10.1016/j.chemphyslip.2013.10.007.CrossRefPubMedGoogle Scholar
  23. Kadoch, C., et al. (2013). Proteomic and bioinformatic analysis of mammalian SWI/SNF complexes identifies extensive roles in human malignancy. Nature Genetics, 45, 592–601. doi: 10.1038/ng.2628.PubMedCentralCrossRefPubMedGoogle Scholar
  24. Kind, T., et al. (2009). FiehnLib: Mass spectral and retention index libraries for metabolomics based on quadrupole and time-of-flight gas chromatography/mass spectrometry. Analytical Chemistry, 81, 10038–10048. doi: 10.1021/ac9019522.PubMedCentralCrossRefPubMedGoogle Scholar
  25. Koitabashi, N., et al. (2009). Avoidance of transient cardiomyopathy in cardiomyocyte-targeted tamoxifen-induced MerCreMer gene deletion models. Circulation Research, 105, 12–15. doi: 10.1161/CIRCRESAHA.109.198416.PubMedCentralCrossRefPubMedGoogle Scholar
  26. Kolwicz, S. C, Jr, Purohit, S., & Tian, R. (2013). Cardiac metabolism and its interactions with contraction, growth, and survival of cardiomyocytes. Circulation Research, 113, 603–616. doi: 10.1161/CIRCRESAHA.113.302095.CrossRefPubMedGoogle Scholar
  27. Kopka, J., Schauer, N., Krueger, S., Birkemeyer, C., Usadel, B., Bergmueller, E., Doermann, P., Weckwerth, W., Gibon, Y., Stitt, M., Willmitzer, L., Fernie, A.R., Steinhauser, D. (2005). GMD@CSB.DB: The Golm Metabolome Database. Bioinformatics, 21(8), 1635–1638. doi: 10.1093/bioinformatics/bti236.CrossRefPubMedGoogle Scholar
  28. Kosho, T., Kuniba, H., Tanikawa, Y., Hashimoto, Y., & Sakurai, H. (2013a). Natural history and parental experience of children with trisomy 18 based on a questionnaire given to a Japanese trisomy 18 parental support group. American Journal of Medical Genetics A, 161A, 1531–1542. doi: 10.1002/ajmg.a.35990.CrossRefGoogle Scholar
  29. Kosho, T., et al. (2013b). Clinical correlations of mutations affecting six components of the SWI/SNF complex: Detailed description of 21 patients and a review of the literature. American Journal of Medical Genetics A, 161, 1221–1237. doi: 10.1002/ajmg.a.35933.CrossRefGoogle Scholar
  30. Lei, I., Gao, X., Sham, M. H., & Wang, Z. (2012). SWI/SNF protein component BAF250a regulates cardiac progenitor cell differentiation by modulating chromatin accessibility during second heart field development. Journal of Biological Chemistry, 287, 24255–24262. doi: 10.1074/jbc.M112.365080.PubMedCentralCrossRefPubMedGoogle Scholar
  31. Lemon, B., Inouye, C., King, D. S., & Tjian, R. (2001). Selectivity of chromatin-remodelling cofactors for ligand-activated transcription. Nature, 414, 924–928. doi: 10.1038/414924a.CrossRefPubMedGoogle Scholar
  32. Li, S., et al. (2008). Genome-wide coactivation analysis of PGC-1alpha identifies BAF60a as a regulator of hepatic lipid metabolism. Cell Metabolism, 8, 105–117. doi: 10.1016/j.cmet.2008.06.013.PubMedCentralCrossRefPubMedGoogle Scholar
  33. Lickert, H., et al. (2004). Baf60c is essential for function of BAF chromatin remodelling complexes in heart development. Nature, 432, 107–112. doi: 10.1038/nature03071.CrossRefPubMedGoogle Scholar
  34. Mallard, W. G., & Reed, J. (1997). Automated mass spectral deconvolution and identification system: AMDIS user guide. Gaithersburg: National Institute of Standards and Technology, US Department of Commerce iv. 58.Google Scholar
  35. McDonald, T. S., Tan, K. N., Hodson, M. P., & Borges, K. (2014). Alterations of hippocampal glucose metabolism by even versus uneven medium chain triglycerides. Journal of Cerebral Blood Flow and Metabolism, 34, 153–160. doi: 10.1038/jcbfm.2013.184.PubMedCentralCrossRefPubMedGoogle Scholar
  36. Mervaala, E., et al. (2010). Metabolomics in angiotensin II-induced cardiac hypertrophy. Hypertension, 55, 508–515. doi: 10.1161/HYPERTENSIONAHA.109.145490.CrossRefPubMedGoogle Scholar
  37. Mulligan, C. M., et al. (2012). Dietary linoleate preserves cardiolipin and attenuates mitochondrial dysfunction in the failing rat heart. Cardiovascular Research, 94, 460–468. doi: 10.1093/cvr/cvs118.PubMedCentralCrossRefPubMedGoogle Scholar
  38. Nakajima, H., Raben, N., Hamaguchi, T., & Yamasaki, T. (2002). Phosphofructokinase deficiency; past, present and future. Current Molecular Medicine, 2, 197–212.CrossRefPubMedGoogle Scholar
  39. Nelson, T. J., Balza, R, Jr, Xiao, Q., & Misra, R. P. (2005). SRF-dependent gene expression in isolated cardiomyocytes: Regulation of genes involved in cardiac hypertrophy. Journal of Molecular and Cellular Cardiology, 39, 479–489. doi: 10.1016/j.yjmcc.2005.05.004.CrossRefPubMedGoogle Scholar
  40. Neubauer, S. (2007). The failing heart–an engine out of fuel. New England Journal of Medicine, 356, 1140–1151. doi: 10.1056/NEJMra063052.CrossRefPubMedGoogle Scholar
  41. Oakley, R. H., et al. (2013). Essential role of stress hormone signaling in cardiomyocytes for the prevention of heart disease. Proceedings of the National Academy of Sciences USA, 110, 17035–17040. doi: 10.1073/pnas.1302546110.CrossRefGoogle Scholar
  42. Raben, N., & Sherman, J. B. (1995). Mutations in muscle phosphofructokinase gene. Human Mutation, 6, 1–6. doi: 10.1002/humu.1380060102.CrossRefPubMedGoogle Scholar
  43. Reyes, J. C., Barra, J., Muchardt, C., Camus, A., Babinet, C., & Yaniv, M. (1998). Altered control of cellular proliferation in the absence of mammalian brahma (SNF2alpha). EMBO Journal, 17, 6979–6991. doi: 10.1093/emboj/17.23.6979.PubMedCentralCrossRefPubMedGoogle Scholar
  44. Roessner, U., Wagner, C., Kopka, J., Trethewey, R. N., & Willmitzer, L. (2000). Technical advance: Simultaneous analysis of metabolites in potato tuber by gas chromatography-mass spectrometry. Plant J, 23, 131–142.CrossRefPubMedGoogle Scholar
  45. Ronan, J. L., Wu, W., & Crabtree, G. R. (2013). From neural development to cognition: Unexpected roles for chromatin. Nature Reviews Genetics, 14, 347–359. doi: 10.1038/nrg3413.PubMedCentralCrossRefPubMedGoogle Scholar
  46. Salma, N., Xiao, H., Mueller, E., & Imbalzano, A. N. (2004). Temporal recruitment of transcription factors and SWI/SNF chromatin-remodeling enzymes during adipogenic induction of the peroxisome proliferator-activated receptor gamma nuclear hormone receptor. Molecular and Cellular Biology, 24, 4651–4663. doi: 10.1128/MCB.24.11.4651-4663.2004.PubMedCentralCrossRefPubMedGoogle Scholar
  47. Santen, G. W., et al. (2012). Mutations in SWI/SNF chromatin remodeling complex gene ARID1B cause Coffin-Siris syndrome. Nature Genetics, 44, 379–380. doi: 10.1038/ng.2217.CrossRefPubMedGoogle Scholar
  48. Sohal, D. S., et al. (2001). Temporally regulated and tissue-specific gene manipulations in the adult and embryonic heart using a tamoxifen-inducible Cre protein. Circulation Research, 89, 20–25.CrossRefPubMedGoogle Scholar
  49. Sparagna, G. C., & Lesnefsky, E. J. (2009). Cardiolipin remodeling in the heart. Journal of Cardiovascular Pharmacology, 53, 290–301. doi: 10.1097/FJC.0b013e31819b5461.CrossRefPubMedGoogle Scholar
  50. Stankunas, K., et al. (2008). Endocardial Brg1 represses ADAMTS1 to maintain the microenvironment for myocardial morphogenesis. Developmental Cell, 14, 298–311. doi: 10.1016/j.devcel.2007.11.018.PubMedCentralCrossRefPubMedGoogle Scholar
  51. Stein, S. E. (1999). An integrated method for spectrum extraction and compound identification from GC/MS data. Journal of the American Society for Mass Spectrometry, 10(8), 770–781.CrossRefGoogle Scholar
  52. Styczynski, M. P., Moxley, J. F., Tong, L. V., Walther, J. L., Jensen, K. L., & Stephanopoulos, G. N. (2007). Systematic identification of conserved metabolites in GC/MS data for metabolomics and biomarker discovery. Analytical Chemistry, 79, 966–973. doi: 10.1021/ac0614846.CrossRefPubMedGoogle Scholar
  53. Sumi-Ichinose, C., Ichinose, H., Metzger, D., & Chambon, P. (1997). SNF2beta-BRG1 is essential for the viability of F9 murine embryonal carcinoma cells. Molecular and Cellular Biology, 17, 5976–5986.PubMedCentralPubMedGoogle Scholar
  54. Takeuchi, J. K., & Bruneau, B. G. (2009). Directed transdifferentiation of mouse mesoderm to heart tissue by defined factors. Nature, 459, 708–711. doi: 10.1038/nature08039.PubMedCentralCrossRefPubMedGoogle Scholar
  55. Takeuchi, J. K., et al. (2011). Chromatin remodelling complex dosage modulates transcription factor function in heart development. Nature Communications, 2, 187. doi: 10.1038/ncomms1187.PubMedCentralCrossRefPubMedGoogle Scholar
  56. Trotter, K. W., & Archer, T. K. (2007). Nuclear receptors and chromatin remodeling machinery. Molecular and Cellular Endocrinology, 265–266, 162–167. doi: 10.1016/j.mce.2006.12.015.PubMedCentralCrossRefPubMedGoogle Scholar
  57. Tsurusaki, Y., et al. (2012). Mutations affecting components of the SWI/SNF complex cause Coffin-Siris syndrome. Nature Genetics, 44, 376–378. doi: 10.1038/ng.2219.CrossRefPubMedGoogle Scholar
  58. Van Houdt, J. K., et al. (2012). Heterozygous missense mutations in SMARCA2 cause Nicolaides-Baraitser syndrome. Nature Genetics, 44(445–9), S1. doi: 10.1038/ng.1105.Google Scholar
  59. Viswakarma, N., et al. (2010). Coactivators in PPAR-regulated gene expression. PPAR Research,. doi: 10.1155/2010/250126.Google Scholar
  60. Wang, W., Xue, Y., Zhou, S., Kuo, A., Cairns, B. R., & Crabtree, G. R. (1996). Diversity and specialization of mammalian SWI/SNF complexes. Genes and Development, 10, 2117–2130.CrossRefPubMedGoogle Scholar
  61. Wang, Z., et al. (2004). Polybromo protein BAF180 functions in mammalian cardiac chamber maturation. Genes and Development, 18, 3106–3116. doi: 10.1101/gad.1238104.PubMedCentralCrossRefPubMedGoogle Scholar
  62. Willis, M. S., et al. (2009a). Muscle ring finger 1 mediates cardiac atrophy in vivo. American Journal of Physiology Heart and Circulatory Physiology, 296, H997–H1006. doi: 10.1152/ajpheart.00660.2008.PubMedCentralCrossRefPubMedGoogle Scholar
  63. Willis, M. S., et al. (2009b). Cardiac muscle ring finger-1 increases susceptibility to heart failure in vivo. Circulation Research, 105, 80–88. doi: 10.1161/CIRCRESAHA.109.194928.PubMedCentralCrossRefPubMedGoogle Scholar
  64. Willis, M. S., et al. (2012). Functional redundancy of SWI/SNF catalytic subunits in maintaining vascular endothelial cells in the adult heart. Circulation Research, 111, e111–e122. doi: 10.1161/CIRCRESAHA.112.265587.PubMedCentralCrossRefPubMedGoogle Scholar
  65. Wu, J. I., Lessard, J., & Crabtree, G. R. (2009). Understanding the words of chromatin regulation. Cell, 136, 200–206. doi: 10.1016/j.cell.2009.01.009.PubMedCentralCrossRefPubMedGoogle Scholar
  66. Xia, J., Mandal, R., Sinelnikov, I. V., Broadhurst, D., & Wishart, D. S. (2012). MetaboAnalyst 2.0–a comprehensive server for metabolomic data analysis. Nucleic Acids Research, 40, W127–W133. doi: 10.1093/nar/gks374.PubMedCentralCrossRefPubMedGoogle Scholar
  67. Xia, J., Psychogios, N., Young, N., & Wishart, D. S. (2009). MetaboAnalyst: A web server for metabolomic data analysis and interpretation. Nucleic Acids Research, 37, W652–W660. doi: 10.1093/nar/gkp356.PubMedCentralCrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Ranjan Banerjee
    • 1
  • Scott J. Bultman
    • 2
  • Darcy Holley
    • 2
  • Carolyn Hillhouse
    • 3
  • James R. Bain
    • 4
    • 5
  • Christopher B. Newgard
    • 4
    • 5
  • Michael J. Muehlbauer
    • 4
  • Monte S. Willis
    • 3
    • 6
  1. 1.University of North Carolina School of MedicineChapel HillUSA
  2. 2.Department of GeneticsUniversity of North CarolinaChapel HillUSA
  3. 3.Department of Pathology & Laboratory MedicineUniversity of North CarolinaChapel HillUSA
  4. 4.Sarah W. Stedman Nutrition and Metabolism Center, Duke Molecular Physiology InstituteDuke University Medical CenterDurhamUSA
  5. 5.Division of Endocrinology, Metabolism, and Nutrition, Department of MedicineDuke University Medical CenterDurhamUSA
  6. 6.McAllister Heart InstituteUniversity of North CarolinaChapel HillUSA

Personalised recommendations