Biological insights from multi-omic analysis of 31 genomic risk loci for adult hearing difficulty


Autoři: Gurmannat Kalra aff001;  Beatrice Milon aff003;  Alex M. Casella aff001;  Brian R. Herb aff001;  Elizabeth Humphries aff001;  Yang Song aff001;  Kevin P. Rose aff001;  Ronna Hertzano aff001;  Seth A. Ament aff001;  Kevin P. Rose aff002
Působiště autorů: Institute for Genome Sciences, University of Maryland School of Medicine, Baltimore, MD, United States of America aff001;  Program in Molecular Medicine, University of Maryland School of Medicine, Baltimore, MD, United States of America aff002;  Department of Otorhinolaryngology-Head & Neck Surgery, University of Maryland School of Medicine, Baltimore, MD, United States of America aff003;  Physician Scientist Training Program, University of Maryland School of Medicine, Baltimore, MD, United States of America aff004;  Program in Molecular Epidemiology, University of Maryland School of Medicine, Baltimore, MD, United States of America aff005;  Department of Anatomy and Neurobiology, University of Maryland School of Medicine, Baltimore, MD, United States of America aff006;  Department of Psychiatry, University of Maryland School of Medicine, Baltimore, MD, United States of America aff007
Vyšlo v časopise: Biological insights from multi-omic analysis of 31 genomic risk loci for adult hearing difficulty. PLoS Genet 16(9): e32767. doi:10.1371/journal.pgen.1009025
Kategorie: Research Article
doi: 10.1371/journal.pgen.1009025

Souhrn

Age-related hearing impairment (ARHI), one of the most common medical conditions, is strongly heritable, yet its genetic causes remain largely unknown. We conducted a meta-analysis of GWAS summary statistics from multiple hearing-related traits in the UK Biobank (n = up to 330,759) and identified 31 genome-wide significant risk loci for self-reported hearing difficulty (p < 5x10-8), of which eight have not been reported previously in the peer-reviewed literature. We investigated the regulatory and cell specific expression for these loci by generating mRNA-seq, ATAC-seq, and single-cell RNA-seq from cells in the mouse cochlea. Risk-associated genes were most strongly enriched for expression in cochlear epithelial cells, as well as for genes related to sensory perception and known Mendelian deafness genes, supporting their relevance to auditory function. Regions of the human genome homologous to open chromatin in epithelial cells from the mouse were strongly enriched for heritable risk for hearing difficulty, even after adjusting for baseline effects of evolutionary conservation and cell-type non-specific regulatory regions. Epigenomic and statistical fine-mapping most strongly supported 50 putative risk genes. Of these, 39 were expressed robustly in mouse cochlea and 16 were enriched specifically in sensory hair cells. These results reveal new risk loci and risk genes for hearing difficulty and suggest an important role for altered gene regulation in the cochlear sensory epithelium.

Klíčová slova:

Cochlea – Deafness – Gene regulation – Genetic loci – Genome-wide association studies – Chromatin – Medical risk factors – Single nucleotide polymorphisms – Mammalian genomics


Zdroje

1. Yamasoba T, Lin FR, Someya S, Kashio A, Sakamoto T, Kondo K. Current concepts in age-related hearing loss: epidemiology and mechanistic pathways. Hear Res. 2013;303: 30–38. doi: 10.1016/j.heares.2013.01.021 23422312

2. Momi SK, Wolber LE, Fabiane SM, MacGregor AJ, Williams FMK. Genetic and Environmental Factors in Age-Related Hearing Impairment. Twin Res Hum Genet. 2015;18: 383–392. doi: 10.1017/thg.2015.35 26081266

3. Van Camp G, Smith R. Hereditary Hearing Loss Homepage. 2019. https://hereditaryhearingloss.org/

4. Hoffmann TJ, Keats BJ, Yoshikawa N, Schaefer C, Risch N, Lustig LR. A Large Genome-Wide Association Study of Age-Related Hearing Impairment Using Electronic Health Records. PLoS Genet. 2016;12: e1006371. doi: 10.1371/journal.pgen.1006371 27764096

5. Vuckovic D, Dawson S, Scheffer DI, Rantanen T, Morgan A, Di Stazio M, et al. Genome-wide association analysis on normal hearing function identifies PCDH20 and SLC28A3 as candidates for hearing function and loss. Hum Mol Genet. 2015;24: 5655–64. doi: 10.1093/hmg/ddv279 26188009

6. Van Laer L, Huyghe JR, Hannula S, Van Eyken E, Stephan DA, Maki-Torkko E, et al. A genome-wide association study for age-related hearing impairment in the Saami. Eur J Hum Genet. 2010;18: 685–693. doi: 10.1038/ejhg.2009.234 20068591

7. Girotto G, Pirastu N, Sorice R, Biino G, Campbell H, d’Adamo AP, et al. Hearing function and thresholds: a genome-wide association study in European isolated populations identifies new loci and pathways. J Med Genet. 2011;48: 369–374. doi: 10.1136/jmg.2010.088310 21493956

8. Friedman RA, Van Laer L, Huentelman MJ, Sheth SS, Van Eyken E, Corneveaux JJ, et al. GRM7 variants confer susceptibility to age-related hearing impairment. Hum Mol Genet. 2009;18: 785–796. doi: 10.1093/hmg/ddn402 19047183

9. Wells HRR, Freidin MB, Zainul Abidin FN, Payton A, Dawes P, Munro KJ, et al. GWAS Identifies 44 Independent Associated Genomic Loci for Self-Reported Adult Hearing Difficulty in UK Biobank. Am J Hum Genet. 2019;105: 788–802. doi: 10.1016/j.ajhg.2019.09.008 31564434

10. Fromer M, Roussos P, Sieberts SK, Johnson JS, Kavanagh DH, Perumal TM, et al. Gene expression elucidates functional impact of polygenic risk for schizophrenia. Nat Neurosci. 2016;19: 1442–1453. doi: 10.1038/nn.4399 27668389

11. Gusev A, Lee SH, Trynka G, Finucane H, Vilhjálmsson BJ, Xu H, et al. Partitioning heritability of regulatory and cell-type-specific variants across 11 common diseases. Am J Hum Genet. 2014. doi: 10.1016/j.ajhg.2014.10.004 25439723

12. Claussnitzer M, Dankel SN, Kim K-H, Quon G, Meuleman W, Haugen C, et al. FTO Obesity Variant Circuitry and Adipocyte Browning in Humans. N Engl J Med. 2015; 150819140043007. doi: 10.1056/NEJMoa1502214 26287746

13. Maurano MT, Humbert R, Rynes E, Thurman RE, Haugen E, Wang H, et al. Systematic localization of common disease-associated variation in regulatory DNA. Science. 2012;337: 1190–5. doi: 10.1126/science.1222794 22955828

14. Sudlow C, Gallacher J, Allen N, Beral V, Burton P, Danesh J, et al. UK Biobank: An Open Access Resource for Identifying the Causes of a Wide Range of Complex Diseases of Middle and Old Age. PLoS Med. 2015. doi: 10.1371/journal.pmed.1001779 25826379

15. Bulik-Sullivan BK, Loh P-R, Finucane HK, Ripke S, Yang J, Patterson N, et al. LD Score regression distinguishes confounding from polygenicity in genome-wide association studies. Nat Genet. 2015;47: 291–295. doi: 10.1038/ng.3211 25642630

16. Zheng J, Erzurumluoglu AM, Elsworth BL, Kemp JP, Howe L, Haycock PC, et al. LD Hub: a centralized database and web interface to perform LD score regression that maximizes the potential of summary level GWAS data for SNP heritability and genetic correlation analysis. Bioinformatics. 2016/09/22. 2017;33: 272–279. doi: 10.1093/bioinformatics/btw613 27663502

17. Rutherford BR, Brewster K, Golub JS, Kim AH, Roose SP. Sensation and Psychiatry: Linking Age-Related Hearing Loss to Late-Life Depression and Cognitive Decline. Am J Psychiatry. 2018;175: 215–224. doi: 10.1176/appi.ajp.2017.17040423 29202654

18. Dhanda N, Taheri S. A narrative review of obesity and hearing loss. Int J Obes (Lond). 2017;41: 1066–1073. doi: 10.1038/ijo.2017.32 28163314

19. Turley P, Walters RK, Maghzian O, Okbay A, Lee JJ, Fontana MA, et al. Multi-trait analysis of genome-wide association summary statistics using MTAG. Nat Genet. 2018;50: 229–237. doi: 10.1038/s41588-017-0009-4 29292387

20. Verweij N, van de Vegte YJ, van der Harst P. Genetic study links components of the autonomous nervous system to heart-rate profile during exercise. Nat Commun. 2018;9: 898. doi: 10.1038/s41467-018-03395-6 29497042

21. Huffman JE. Examining the current standards for genetic discovery and replication in the era of mega-biobanks. Nat Commun. 2018;9: 5054. doi: 10.1038/s41467-018-07348-x 30498205

22. Fransen E, Bonneux S, Corneveaux JJ, Schrauwen I, Di Berardino F, White CH, et al. Genome-wide association analysis demonstrates the highly polygenic character of age-related hearing impairment. Eur J Hum Genet. 2015;23: 110–115. doi: 10.1038/ejhg.2014.56 24939585

23. Euesden J, Lewis CM, O’Reilly PF. PRSice: Polygenic Risk Score software. Bioinformatics. 2015;31: 1466–8. doi: 10.1093/bioinformatics/btu848 25550326

24. de Leeuw CA, Mooij JM, Heskes T, Posthuma D. MAGMA: generalized gene-set analysis of GWAS data. PLoS Comput Biol. 2015;11: e1004219. doi: 10.1371/journal.pcbi.1004219 25885710

25. Mchugh RK, Friedman RA. Genetics of hearing loss: Allelism and modifier genes produce a phenotypic continuum. Anat Rec Part A Discov Mol Cell Evol Biol. 2006;288A: 370–381. doi: 10.1002/ar.a.20297 16550584

26. Scheffer DI, Shen J, Corey DP, Chen Z-Y. Gene Expression by Mouse Inner Ear Hair Cells during Development. J Neurosci. 2015;35: 6366–6380. doi: 10.1523/JNEUROSCI.5126-14.2015 25904789

27. Li Y, Liu H, Giffen KP, Chen L, Beisel KW, He DZZ. Transcriptomes of cochlear inner and outer hair cells from adult mice. Sci data. 2018;5: 180199. doi: 10.1038/sdata.2018.199 30277483

28. Liu H, Chen L, Giffen KP, Stringham ST, Li Y, Judge PD, et al. Cell-Specific Transcriptome Analysis Shows That Adult Pillar and Deiters’ Cells Express Genes Encoding Machinery for Specializations of Cochlear Hair Cells. Front Mol Neurosci. 2018;11: 356. doi: 10.3389/fnmol.2018.00356 30327589

29. Aguet F, Ardlie KG, Cummings BB, Gelfand ET, Getz G, Hadley K, et al. Genetic effects on gene expression across human tissues. Nature. 2017;550: 204–213. doi: 10.1038/nature24277 29022597

30. de la Torre-Ubieta L, Stein JL, Won H, Opland CK, Liang D, Lu D, et al. The Dynamic Landscape of Open Chromatin during Human Cortical Neurogenesis. Cell. 2018;172: 289–304.e18. doi: 10.1016/j.cell.2017.12.014 29307494

31. Huang K-L, Marcora E, Pimenova AA, Di Narzo AF, Kapoor M, Jin SC, et al. A common haplotype lowers PU.1 expression in myeloid cells and delays onset of Alzheimer’s disease. Nat Neurosci. 2017;20: 1052–1061. doi: 10.1038/nn.4587 28628103

32. Friedman LM, Dror AA, Avraham KB. Mouse models to study inner ear development and hereditary hearing loss. Int J Dev Biol. 2007;51: 609–631. doi: 10.1387/ijdb.072365lf 17891721

33. Elkon R, Milon B, Morrison L, Shah M, Vijayakumar S, Racherla M, et al. RFX transcription factors are essential for hearing in mice. Nat Commun. 2015;6: 8549. doi: 10.1038/ncomms9549 26469318

34. Hertzano R, Elkon R, Kurima K, Morrisson A, Chan S-L, Sallin M, et al. Cell type-specific transcriptome analysis reveals a major role for Zeb1 and miR-200b in mouse inner ear morphogenesis. PLoS Genet. 2011;7: e1002309. doi: 10.1371/journal.pgen.1002309 21980309

35. Yue F, Cheng Y, Breschi A, Vierstra J, Wu W, Ryba T, et al. A comparative encyclopedia of DNA elements in the mouse genome. Nature. 2014;515: 355–64. doi: 10.1038/nature13992 25409824

36. Roccio M, Perny M, Ealy M, Widmer HR, Heller S, Senn P. Molecular characterization and prospective isolation of human fetal cochlear hair cell progenitors. Nat Commun. 2018;9: 4027. doi: 10.1038/s41467-018-06334-7 30279445

37. Visel A, Minovitsky S, Dubchak I, Pennacchio LA. VISTA Enhancer Browser—a database of tissue-specific human enhancers. Nucleic Acids Res. 2007;35: D88–D92. doi: 10.1093/nar/gkl822 17130149

38. Hume CR, Bratt DL, Oesterle EC. Expression of LHX3 and SOX2 during mouse inner ear development. Gene Expr Patterns. 2007;7: 798–807. doi: 10.1016/j.modgep.2007.05.002 17604700

39. Ahn KJ, Passero FJ, Crenshaw EB 3rd. Otic mesenchyme expression of Cre recombinase directed by the inner ear enhancer of the Brn4/Pou3f4 gene. Genesis. 2009;47: 137–141. doi: 10.1002/dvg.20454 19217071

40. Hinrichs AS, Karolchik D, Baertsch R, Barber GP, Bejerano G, Clawson H, et al. The UCSC Genome Browser Database: update 2006. Nucleic Acids Res. 2006;34: D590–8. doi: 10.1093/nar/gkj144 16381938

41. Kundaje A, Meuleman W, Ernst J, Bilenky M, Yen A, Heravi-Moussavi A, et al. Integrative analysis of 111 reference human epigenomes. Nature. 2015;518: 317–30. doi: 10.1038/nature14248 25693563

42. Finucane HK, Bulik-Sullivan B, Gusev A, Trynka G, Reshef Y, Loh P-R, et al. Partitioning heritability by functional annotation using genome-wide association summary statistics. Nat Genet. 2015;47: 1228–1235. doi: 10.1038/ng.3404 26414678

43. Consortium T 1000 GP. An integrated map of genetic variation from 1,092 human genomes. Nature. 2012;491: 56–65. doi: 10.1038/nature11632 23128226

44. Schmitt AD, Hu M, Jung I, Xu Z, Qiu Y, Tan CL, et al. A Compendium of Chromatin Contact Maps Reveals Spatially Active Regions in the Human Genome. Cell Rep. 2016;17: 2042–2059. doi: 10.1016/j.celrep.2016.10.061 27851967

45. Schrauwen I, Chakchouk I, Liaqat K, Jan A, Nasir A, Hussain S, et al. A variant in LMX1A causes autosomal recessive severe-to-profound hearing impairment. Hum Genet. 2018;137: 471–478. doi: 10.1007/s00439-018-1899-7 29971487

46. Wesdorp M, de Koning Gans PAM, Schraders M, Oostrik J, Huynen MA, Venselaar H, et al. Heterozygous missense variants of LMX1A lead to nonsyndromic hearing impairment and vestibular dysfunction. Hum Genet. 2018. doi: 10.1007/s00439-018-1880-5 29754270

47. Borck G, Ur Rehman A, Lee K, Pogoda H-M, Kakar N, von Ameln S, et al. Loss-of-function mutations of ILDR1 cause autosomal-recessive hearing impairment DFNB42. Am J Hum Genet. 2011;88: 127–137. doi: 10.1016/j.ajhg.2010.12.011 21255762

48. Zhang Y, Zhang X, Long R, Yu L. A novel deletion mutation of the SOX2 gene in a child of Chinese origin with congenital bilateral anophthalmia and sensorineural hearing loss. J Genet. 2018;97: 1007–1011. 30262714

49. Wayne S, Robertson NG, DeClau F, Chen N, Verhoeven K, Prasad S, et al. Mutations in the transcriptional activator EYA4 cause late-onset deafness at the DFNA10 locus. Hum Mol Genet. 2001;10: 195–200. doi: 10.1093/hmg/10.3.195 11159937

50. Schonberger J, Wang L, Shin JT, Kim S Do, Depreux FFS, Zhu H, et al. Mutation in the transcriptional coactivator EYA4 causes dilated cardiomyopathy and sensorineural hearing loss. Nat Genet. 2005;37: 418–422. doi: 10.1038/ng1527 15735644

51. Daoud H, Zhang D, McMurray F, Yu A, Luco SM, Vanstone J, et al. Identification of a pathogenic FTO mutation by next-generation sequencing in a newborn with growth retardation and developmental delay. J Med Genet. 2016;53: 200–207. doi: 10.1136/jmedgenet-2015-103399 26378117

52. Pollak A, Lechowicz U, Murcia Pienkowski VA, Stawinski P, Kosinska J, Skarzynski H, et al. Whole exome sequencing identifies TRIOBP pathogenic variants as a cause of post-lingual bilateral moderate-to-severe sensorineural hearing loss. BMC Med Genet. 2017;18: 142. doi: 10.1186/s12881-017-0499-z 29197352

53. Shahin H, Walsh T, Sobe T, Abu Sa’ed J, Abu Rayan A, Lynch ED, et al. Mutations in a novel isoform of TRIOBP that encodes a filamentous-actin binding protein are responsible for DFNB28 recessive nonsyndromic hearing loss. Am J Hum Genet. 2006;78: 144–152. doi: 10.1086/499495 16385458

54. Manji SSM, Williams LH, Miller KA, Ooms LM, Bahlo M, Mitchell CA, et al. A mutation in synaptojanin 2 causes progressive hearing loss in the ENU-mutagenised mouse strain mozart. PLoS One. 2011. doi: 10.1371/journal.pone.0017607 21423608

55. Ohlemiller KK, Rybak Rice ME, Lett JM, Gagnon PM. Absence of strial melanin coincides with age-associated marginal cell loss and endocochlear potential decline. Hear Res. 2009. doi: 10.1016/j.heares.2008.12.005 19141317

56. Eguchi M, Kariya S, Okano M, Higaki T, Makihara S, Fujiwara T, et al. Lipopolysaccharide induces proinflammatory cytokines and chemokines in experimental otitis media through the prostaglandin D2 receptor (DP)-dependent pathway. Clin Exp Immunol. 2011;163: 260–269. doi: 10.1111/j.1365-2249.2010.04292.x 21166666

57. Wu J, Han W, Chen X, Guo W, Liu K, Wang R, et al. Matrix metalloproteinase-2 and -9 contribute to functional integrity and noise-induced damage to the blood-labyrinth-barrier. Mol Med Rep. 2017;16: 1731–1738. doi: 10.3892/mmr.2017.6784 28627704

58. Mahuzier A, Gaude H-M, Grampa V, Anselme I, Silbermann F, Leroux-Berger M, et al. Dishevelled stabilization by the ciliopathy protein Rpgrip1l is essential for planar cell polarity. J Cell Biol. 2012;198: 927–940. doi: 10.1083/jcb.201111009 22927466

59. Rocha-Sanchez SM, Scheetz LR, Contreras M, Weston MD, Korte M, McGee J, et al. Mature mice lacking Rbl2/p130 gene have supernumerary inner ear hair cells and supporting cells. J Neurosci. 2011;31: 8883–8893. doi: 10.1523/JNEUROSCI.5821-10.2011 21677172

60. Bowl MR, Simon MM, Ingham NJ, Greenaway S, Santos L, Cater H, et al. A large scale hearing loss screen reveals an extensive unexplored genetic landscape for auditory dysfunction. Nat Commun. 2017;8: 886. doi: 10.1038/s41467-017-00595-4 29026089

61. Butler A, Hoffman P, Smibert P, Papalexi E, Satija R. Integrating single-cell transcriptomic data across different conditions, technologies, and species. Nat Biotechnol. 2018;36: 411. Available: doi: 10.1038/nbt.4096 29608179

62. Shen J, Scheffer DI, Kwan KY, Corey DP. SHIELD: an integrative gene expression database for inner ear research. Database (Oxford). 2015;2015: bav071. doi: 10.1093/database/bav071 26209310

63. Chessum L, Matern MS, Kelly MC, Johnson SL, Ogawa Y, Milon B, et al. Helios is a key transcriptional regulator of outer hair cell maturation. Nature. 2018;563: 696–700. doi: 10.1038/s41586-018-0728-4 30464345

64. Burns JC, Kelly MC, Hoa M, Morell RJ, Kelley MW. Single-cell RNA-Seq resolves cellular complexity in sensory organs from the neonatal inner ear. Nat Commun. 2015;6: 8557. doi: 10.1038/ncomms9557 26469390

65. Boyle EA, Li YI, Pritchard JK. An Expanded View of Complex Traits: From Polygenic to Omnigenic. Cell. 2017;169: 1177–1186. doi: 10.1016/j.cell.2017.05.038 28622505

66. Ohlemiller KK, Hughes RM, Lett JM, Ogilvie JM, Speck JD, Wright JS, et al. Progression of cochlear and retinal degeneration in the tubby (rd5) mouse. Audiol Neurootol. 1997;2: 175–85. doi: 10.1159/000259242 9390831

67. Schuknecht HF, Watanuki K, Takahashi T, Belal AA, Kimura RS, Jones DD, et al. Atrophy of the stria vascularis, a common cause for hearing loss. Laryngoscope. 1974;84: 1777–821. doi: 10.1288/00005537-197410000-00012 4138750

68. Puligilla C, Kelley MW. Dual role for Sox2 in specification of sensory competence and regulation of Atoh1 function. Dev Neurobiol. 2017;77: 3–13. doi: 10.1002/dneu.22401 27203669

69. Koo SK, Hill JK, Hwang CH, Lin ZS, Millen KJ, Wu DK. Lmx1a maintains proper neurogenic, sensory, and non-sensory domains in the mammalian inner ear. Dev Biol. 2009;333: 14–25. doi: 10.1016/j.ydbio.2009.06.016 19540218

70. Bok J, Raft S, Kong K-A, Koo SK, Dräger UC, Wu DK. Transient retinoic acid signaling confers anterior-posterior polarity to the inner ear. Proc Natl Acad Sci U S A. 2011;108: 161–6. doi: 10.1073/pnas.1010547108 21173260

71. Buenrostro JD, Wu B, Chang HY, Greenleaf WJ. ATAC-seq: A Method for Assaying Chromatin Accessibility Genome-Wide. Curr Protoc Mol Biol. 2015;109: 21.29.1–9. doi: 10.1002/0471142727.mb2129s109 25559105

72. Watanabe K, Taskesen E, van Bochoven A, Posthuma D. Functional mapping and annotation of genetic associations with FUMA. Nat Commun. 2017;8: 1826. doi: 10.1038/s41467-017-01261-5 29184056

73. Boyle AP, Guinney J, Crawford GE, Furey TS. F-Seq: a feature density estimator for high-throughput sequence tags. Bioinformatics. 2008;24: 2537–2538. doi: 10.1093/bioinformatics/btn480 18784119

74. Wang K, Li M, Hakonarson H. ANNOVAR: functional annotation of genetic variants from high-throughput sequencing data. Nucleic Acids Res. 2010;38: e164–e164. doi: 10.1093/nar/gkq603 20601685

75. Rentzsch P, Witten D, Cooper GM, Shendure J, Kircher M. CADD: predicting the deleteriousness of variants throughout the human genome. Nucleic Acids Res. 2019;47: D886–D894. doi: 10.1093/nar/gky1016 30371827

76. Ay F, Bailey TL, Noble WS. Statistical confidence estimation for Hi-C data reveals regulatory chromatin contacts. Genome Res. 2014;24: 999–1011. doi: 10.1101/gr.160374.113 24501021

77. Kircher M, Witten DM, Jain P, O’Roak BJ, Cooper GM, Shendure J. A general framework for estimating the relative pathogenicity of human genetic variants. Nat Genet. 2014;46: 310–5. doi: 10.1038/ng.2892 24487276


Článek vyšel v časopise

PLOS Genetics


2020 Číslo 9

Nejčtenější v tomto čísle

Tomuto tématu se dále věnují…


Kurzy

Zvyšte si kvalifikaci online z pohodlí domova

Imunitní trombocytopenie (ITP) u dospělých pacientů
nový kurz
Autoři: prof. MUDr. Tomáš Kozák, Ph.D., MBA

Pěnová skleroterapie
Autoři: MUDr. Marek Šlais

White paper - jak vidíme optimální péči o zubní náhrady
Autoři: MUDr. Jindřich Charvát, CSc.

Hemofilie - série kurzů

Faktory ovlivňující léčbu levotyroxinem
Autoři:

Všechny kurzy
Kurzy Doporučená témata Časopisy
Přihlášení
Zapomenuté heslo

Nemáte účet?  Registrujte se

Zapomenuté heslo

Zadejte e-mailovou adresu se kterou jste vytvářel(a) účet, budou Vám na ni zaslány informace k nastavení nového hesla.

Přihlášení

Nemáte účet?  Registrujte se

VIRTUÁLNÍ ČEKÁRNA ČR Jste praktický lékař nebo pediatr? Zapojte se! Jste praktik nebo pediatr? Zapojte se!

×