{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,18]],"date-time":"2026-08-18T04:47:00Z","timestamp":1787028420117,"version":"build-2736575974"},"reference-count":56,"publisher":"MDPI AG","issue":"22","license":[{"start":{"date-parts":[[2019,11,7]],"date-time":"2019-11-07T00:00:00Z","timestamp":1573084800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>(1) Background: Central congenital hypothyroidism (CCH) is a rare endocrine disorder that can be caused by mutations in the \u03b2-subunit of thyrotropin (TSHB). The TSHB mutation C105Vfs114X leads to isolated thyroid-stimulating-hormone-(TSH)-deficiency and results in a severe phenotype. The aim of this study was to gain more insight into the underlying molecular mechanism and the functional effects of this mutation based on two assumptions: a) the three-dimensional (3D) structure of TSH should be modified with the C105V substitution, and\/or b) whether the C-terminal modifications lead to signaling differences. (2) Methods: wild-type (WT) and different mutants of hTSH were generated in human embryonic kidney 293 cells (HEK293 cells) and TSH preparations were used to stimulate thyrotropin receptor (TSHR) stably transfected into follicular thyroid cancer cells (FTC133-TSHR cells) and transiently transfected into HEK293 cells. Functional characterization was performed by determination of Gs, mitogen activated protein kinase (MAPK) and Gq\/11 activation. (3) Results: The patient mutation C105Vfs114X and further designed TSH mutants diminished cyclic adenosine monophosphate (cAMP) signaling activity. Surprisingly, MAPK signaling for all mutants was comparable to WT, while none of the mutants induced PLC activation. (4) Conclusion: We characterized the patient mutation C105Vfs114X concerning different signaling pathways. We identified a strong decrease of cAMP signaling induction and speculate that this could, in combination with diverse signaling regarding the other pathways, accounting for the patient\u2019s severe phenotype.<\/jats:p>","DOI":"10.3390\/ijms20225564","type":"journal-article","created":{"date-parts":[[2019,11,7]],"date-time":"2019-11-07T11:17:25Z","timestamp":1573125445000},"page":"5564","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":5,"title":["The Pathogenic TSH \u03b2-Subunit Variant C105Vfs114X Causes a Modified Signaling Profile at TSHR"],"prefix":"10.3390","volume":"20","author":[{"given":"Laura","family":"Kalveram","sequence":"first","affiliation":[{"name":"Institute of Experimental Pediatric Endocrinology, Charit\u00e9-Universit\u00e4tsmedizin Berlin, corporate member of Freie Universit\u00e4t Berlin, Humboldt-Universit\u00e4t zu Berlin, and Berlin Institute of Health, 13353 Berlin, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Gunnar","family":"Kleinau","sequence":"additional","affiliation":[{"name":"Institute of Medical Physics and Biophysics, Group Protein X-ray Crystallography and Signal Transduction, Charit\u00e9 \u2013 Universit\u00e4tsmedizin Berlin, corporate member of Freie Universit\u00e4t Berlin, Humboldt-Universit\u00e4t zu Berlin, and Berlin Institute of Health, 10117 Berlin, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8892-1290","authenticated-orcid":false,"given":"Kamila","family":"Szyma\u0144ska","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Molecular Biology, Medical University of Lublin, 20-093 Lublin, Poland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5028-2075","authenticated-orcid":false,"given":"Patrick","family":"Scheerer","sequence":"additional","affiliation":[{"name":"Institute of Medical Physics and Biophysics, Group Protein X-ray Crystallography and Signal Transduction, Charit\u00e9 \u2013 Universit\u00e4tsmedizin Berlin, corporate member of Freie Universit\u00e4t Berlin, Humboldt-Universit\u00e4t zu Berlin, and Berlin Institute of Health, 10117 Berlin, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9794-802X","authenticated-orcid":false,"given":"Adolfo","family":"Rivero-M\u00fcller","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Molecular Biology, Medical University of Lublin, 20-093 Lublin, Poland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Annette","family":"Gr\u00fcters-Kieslich","sequence":"additional","affiliation":[{"name":"Institute of Experimental Pediatric Endocrinology, Charit\u00e9-Universit\u00e4tsmedizin Berlin, corporate member of Freie Universit\u00e4t Berlin, Humboldt-Universit\u00e4t zu Berlin, and Berlin Institute of Health, 13353 Berlin, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Heike","family":"Biebermann","sequence":"additional","affiliation":[{"name":"Institute of Experimental Pediatric Endocrinology, Charit\u00e9-Universit\u00e4tsmedizin Berlin, corporate member of Freie Universit\u00e4t Berlin, Humboldt-Universit\u00e4t zu Berlin, and Berlin Institute of Health, 13353 Berlin, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2019,11,7]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"959","DOI":"10.1016\/S0022-3476(86)80276-1","article-title":"Detection of congenital hypopituitary hypothyroidism: Ten-year experience in the Northwest Regional Screening Program","volume":"109","author":"Hanna","year":"1986","journal-title":"J. Pediatrics"},{"key":"ref_2","first-page":"24","article-title":"Neonatal thyroid disorders","volume":"59","author":"Gruters","year":"2003","journal-title":"Horm. Res."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"15","DOI":"10.3928\/0090-4481-19920101-06","article-title":"Congenital hypothyroidism","volume":"21","author":"Gruters","year":"1992","journal-title":"Pediatr. Ann."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"33","DOI":"10.1111\/j.1651-2227.1999.tb14401.x","article-title":"Combined pituitary hormone deficiency: Role of Pit-1 and Prop-1","volume":"88","author":"Pfaffle","year":"1999","journal-title":"Acta Paediatr. Suppl."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"5779","DOI":"10.1210\/jc.2003-032124","article-title":"A familial form of congenital hypopituitarism due to a PROP1 mutation in a large kindred: Phenotypic and in vitro functional studies","volume":"89","author":"Reynaud","year":"2004","journal-title":"J. Clin. Endocrinol. Metab."},{"key":"ref_6","first-page":"50","article-title":"Clinical and biochemical phenotype of familial anterior hypopituitarism from mutation of the PROP1 gene","volume":"84","author":"Rosenbloom","year":"1999","journal-title":"J. Clin. Endocrinol. Metab."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"379","DOI":"10.1159\/000350013","article-title":"Isolated central hypothyroidism in young siblings as a manifestation of PROP1 deficiency: Clinical impact of whole exome sequencing","volume":"79","author":"Wassner","year":"2013","journal-title":"Horm. Res. Paediatr."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"147","DOI":"10.1016\/B978-0-12-381296-4.00009-9","article-title":"Constitutively active thyrotropin and thyrotropin-releasing hormone receptors and their inverse agonists","volume":"485","author":"Neumann","year":"2010","journal-title":"Methods Enzymol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"345","DOI":"10.1210\/jc.2011-1591","article-title":"Central hypothyroidism in a patient with pituitary autoimmunity: Evidence for TSH-independent thyroid hormone synthesis","volume":"97","author":"Barbesino","year":"2012","journal-title":"J. Clin. Endocrinol. Metab."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"3068","DOI":"10.1210\/jc.2012-1616","article-title":"Clinical review: Central hypothyroidism: Pathogenic, diagnostic, and therapeutic challenges","volume":"97","author":"Persani","year":"2012","journal-title":"J. Clin. Endocrinol. Metab."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"1250","DOI":"10.1172\/JCI118540","article-title":"A circulating, biologically inactive thyrotropin caused by a mutation in the beta subunit gene","volume":"97","author":"Herodotou","year":"1996","journal-title":"J. Clin. Investig."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"355","DOI":"10.1515\/JPEM.2004.17.3.355","article-title":"Novel TSHbeta subunit gene mutation causing congenital central hypothyroidism in a newborn male","volume":"17","author":"Morales","year":"2004","journal-title":"J. Pediatric Endocrinol. Metab."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"E98","DOI":"10.1210\/jc.2010-0223","article-title":"Two novel mutations of the TSH-beta subunit gene underlying congenital central hypothyroidism undetectable in neonatal TSH screening","volume":"95","author":"Baquedano","year":"2010","journal-title":"J. Clin. Endocrinol. Metab."},{"key":"ref_14","first-page":"1762","article-title":"Congenital central hypothyroidism due to a homozygous mutation in the thyrotropin beta-subunit gene follows an autosomal recessive inheritance","volume":"83","author":"Doeker","year":"1998","journal-title":"J. Clin. Endocrinol. Metab."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"387","DOI":"10.1089\/thy.2000.10.387","article-title":"Congenital central isolated hypothyroidism caused by a homozygous mutation in the TSH-beta subunit gene","volume":"10","author":"Heinrichs","year":"2000","journal-title":"Thyroid Off. J. Am. Thyroid Assoc."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"4811","DOI":"10.1210\/jc.2002-020297","article-title":"Congenital central hypothyroidism due to homozygous thyrotropin beta 313 Delta T mutation is caused by a Founder effect","volume":"87","author":"Brumm","year":"2002","journal-title":"J. Clin. Endocrinol. Metab."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"669","DOI":"10.1515\/JPEM.2004.17.4.669","article-title":"Hypothyroidism in siblings due to a homozygous mutation of the TSH-beta subunit gene","volume":"17","author":"Felner","year":"2004","journal-title":"J. Pediatric Endocrinol. Metab."},{"key":"ref_18","first-page":"41","article-title":"The C105fs114X is the prevalent thyrotropin beta-subunit gene mutation in Argentinean patients with congenital central hypothyroidism","volume":"61","author":"Domene","year":"2004","journal-title":"Horm. Res."},{"key":"ref_19","first-page":"149","article-title":"Compound heterozygous and homozygous mutations of the TSHbeta gene as a cause of congenital central hypothyroidism in Europe","volume":"62","author":"Karges","year":"2004","journal-title":"Horm. Res."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"227","DOI":"10.1055\/s-2006-924232","article-title":"Initially elevated TSH and congenital central hypothyroidism due to a homozygous mutation of the TSH beta subunit gene: Case report and review of the literature","volume":"114","author":"Partsch","year":"2006","journal-title":"Exp. Clin. Endocrinol. Diabetes"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"170","DOI":"10.1203\/00006450-199908000-00007","article-title":"Severe congenital hypothyroidism due to a homozygous mutation of the betaTSH gene","volume":"46","author":"Biebermann","year":"1999","journal-title":"Pediatric Res."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"639","DOI":"10.1089\/thy.2009.0462","article-title":"New cases of isolated congenital central hypothyroidism due to homozygous thyrotropin beta gene mutations: A pitfall to neonatal screening","volume":"20","author":"Ramos","year":"2010","journal-title":"Thyroid Off. J. Am. Thyroid Assoc."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"410","DOI":"10.1111\/cen.13149","article-title":"Molecular spectrum of TSHbeta subunit gene defects in central hypothyroidism in the UK and Ireland","volume":"86","author":"Nicholas","year":"2017","journal-title":"Clin. Endocrinol."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"1141","DOI":"10.1089\/105072502321085252","article-title":"Congenital isolated central hypothyroidism caused by a \u201chot spot\u201d mutation in the thyrotropin-beta gene","volume":"12","author":"McDermott","year":"2002","journal-title":"Thyroid Off. J. Am. Thyroid Assoc."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"553","DOI":"10.1089\/105072503322238818","article-title":"Congenital secondary hypothyroidism due to a mutation C105Vfs114X thyrotropin-beta mutation: Genetic study of five unrelated families from Switzerland and Argentina","volume":"13","author":"Deladoey","year":"2003","journal-title":"Thyroid Off. J. Am. Thyroid Assoc."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"954","DOI":"10.1210\/me.2016-1065","article-title":"Minireview: Insights Into the Structural and Molecular Consequences of the TSH-beta Mutation C105Vfs114X","volume":"30","author":"Kleinau","year":"2016","journal-title":"Mol. Endocrinol."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"87","DOI":"10.1136\/adc.67.1.87","article-title":"Congenital hypothyroidism detected by neonatal screening: Relationship between biochemical severity and early clinical features","volume":"67","author":"Grant","year":"1992","journal-title":"Arch Dis. Child"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"473","DOI":"10.1152\/physrev.00031.2001","article-title":"Thyroid-stimulating hormone and thyroid-stimulating hormone receptor structure-function relationships","volume":"82","author":"Szkudlinski","year":"2002","journal-title":"Physiol. Rev."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"S8","DOI":"10.1186\/1756-6614-4-S1-S8","article-title":"Molecular description of non-autoimmune hyperthyroidism at a neonate caused by a new thyrotropin receptor germline mutation","volume":"4","author":"Biebermann","year":"2011","journal-title":"Thyroid Res."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"2399","DOI":"10.1172\/JCI30380","article-title":"Thyrocyte-specific Gq\/G11 deficiency impairs thyroid function and prevents goiter development","volume":"117","author":"Kero","year":"2007","journal-title":"J. Clin. Investig."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"13733","DOI":"10.1016\/S0021-9258(17)36705-4","article-title":"The human thyrotropin receptor activates G-proteins Gs and Gq\/11","volume":"269","author":"Allgeier","year":"1994","journal-title":"J. Biol. Chem."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"116","DOI":"10.1073\/pnas.93.1.116","article-title":"The human thyrotropin receptor: A heptahelical receptor capable of stimulating members of all four G protein families","volume":"93","author":"Laugwitz","year":"1996","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"R1","DOI":"10.1016\/0303-7207(90)90209-Q","article-title":"Thyrotropin activates both the cyclic AMP and the PIP2 cascades in CHO cells expressing the human cDNA of TSH receptor","volume":"74","author":"Raspe","year":"1990","journal-title":"Mol. Cell. Endocrinol."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"197","DOI":"10.1530\/JOE-16-0384","article-title":"A vital region for human glycoprotein hormone trafficking revealed by an LHB mutation","volume":"231","author":"Potorac","year":"2016","journal-title":"J. Endocrinol."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"691","DOI":"10.1210\/er.2012-1072","article-title":"Novel Insights on Thyroid Stimulating Hormone Receptor Signal Transduction","volume":"34","author":"Kleinau","year":"2013","journal-title":"Endocr. Rev."},{"key":"ref_36","doi-asserted-by":"crossref","unstructured":"Krause, G., Kreuchwig, A., and Kleinau, G. (2012). Extended and structurally supported insights into extracellular hormone binding, signal transduction and organization of the thyrotropin receptor. PLoS ONE, 7.","DOI":"10.1371\/journal.pone.0052920"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"12491","DOI":"10.1073\/pnas.1206643109","article-title":"Structure of follicle-stimulating hormone in complex with the entire ectodomain of its receptor","volume":"109","author":"Jiang","year":"2012","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_38","first-page":"473","article-title":"Potent thyrotropic activity of human chorionic gonadotropin variants in terms of 125I incorporation and de novo synthesized thyroid hormone release in human thyroid follicles","volume":"80","author":"Yamazaki","year":"1995","journal-title":"J. Clin. Endocrinol. Metab."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"1257","DOI":"10.1038\/nbt1096-1257","article-title":"Engineering human glycoprotein hormone superactive analogues","volume":"14","author":"Szkudlinski","year":"1996","journal-title":"Nat. Biotechnol."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"18048","DOI":"10.1074\/jbc.M800449200","article-title":"Extended hormone binding site of the human thyroid stimulating hormone receptor: Distinctive acidic residues in the hinge region are involved in bovine thyroid stimulating hormone binding and receptor activation","volume":"283","author":"Mueller","year":"2008","journal-title":"J. Biol. Chem."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"1131","DOI":"10.1006\/bbrc.1993.1334","article-title":"Expression of the human TSH receptor in a human thyroid carcinoma cell line that lacks an endogenous TSH receptor: Growth inhibition by cAMP","volume":"191","author":"Derwahl","year":"1993","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"3605","DOI":"10.1210\/jc.2010-0112","article-title":"A new phenotype of nongoitrous and nonautoimmune hyperthyroidism caused by a heterozygous thyrotropin receptor mutation in transmembrane helix 6","volume":"95","author":"Winkler","year":"2010","journal-title":"J. Clin. Endocrinol. Metab."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"6904","DOI":"10.1016\/S0021-9258(20)89587-8","article-title":"Delineation via site-directed mutagenesis of the carboxyl-terminal region of human choriogonadotropin beta required for subunit assembly and biological activity","volume":"266","author":"Chen","year":"1991","journal-title":"J. Biol. Chem."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"49","DOI":"10.1016\/S1570-9639(02)00501-0","article-title":"Disulfide bonds Cys(9)-Cys(57), Cys(34)-Cys(88) and Cys(38)-Cys(90) of the beta-subunit of human chorionic gonadotropin are crucial for heterodimer formation with the alpha-subunit: Experimental evidence for the conclusions from the crystal structure of hCG","volume":"1645","author":"Mishra","year":"2003","journal-title":"Biochim. Biophys. Acta"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"940","DOI":"10.1038\/s41598-017-01078-8","article-title":"Human Luteinizing Hormone and Chorionic Gonadotropin Display Biased Agonism at the LH and LH\/CG Receptors","volume":"7","author":"Riccetti","year":"2017","journal-title":"Sci. Rep."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"62","DOI":"10.1016\/j.regpep.2008.03.002","article-title":"Biochemical roles of the oligosaccharide chains in thyrostimulin, a heterodimeric hormone of glycoprotein hormone subunits alpha 2 (GPA2) and beta 5 (GPB5)","volume":"148","author":"Okajima","year":"2008","journal-title":"Regul. Pept."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"263","DOI":"10.1530\/rep.1.00663","article-title":"An update of the pathophysiology of human gonadotrophin subunit and receptor gene mutations and polymorphisms","volume":"130","author":"Themmen","year":"2005","journal-title":"Reproduction"},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"290","DOI":"10.1080\/09513590.2018.1529159","article-title":"FSH beta-subunit mutations in two sisters: The first report from the Indian sub-continent and review of previous cases","volume":"35","author":"Misgar","year":"2019","journal-title":"Gynecol. Endocrinol."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"379","DOI":"10.1093\/molehr\/gas018","article-title":"Structural and functional analysis of rare missense mutations in human chorionic gonadotrophin beta-subunit","volume":"18","author":"Nagirnaja","year":"2012","journal-title":"Mol. Hum. Reprod."},{"key":"ref_50","first-page":"4468","article-title":"New autosomal recessive mutation of the TSH-beta subunit gene causing central isolated hypothyroidism","volume":"86","author":"Vuissoz","year":"2001","journal-title":"J. Clin. Endocrinol. Metab."},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"1972","DOI":"10.1172\/JCI26031","article-title":"Thyrotropin receptor-associated diseases: From adenomata to Graves disease","volume":"115","author":"Davies","year":"2005","journal-title":"J. Clin. Investig."},{"key":"ref_52","first-page":"673","article-title":"The thyrotropin (TSH) receptor: Interaction with TSH and autoantibodies","volume":"19","author":"Rapoport","year":"1998","journal-title":"Endocr. Rev."},{"key":"ref_53","first-page":"988","article-title":"Familial hypothyroidism caused by a nonsense mutation in the thyroid-stimulating hormone beta-subunit gene","volume":"46","author":"Feltquate","year":"1990","journal-title":"Am. J. Hum. Genet."},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"19121","DOI":"10.1038\/srep19121","article-title":"REPLACR-mutagenesis, a one-step method for site-directed mutagenesis by recombineering","volume":"6","author":"Trehan","year":"2016","journal-title":"Sci. Rep."},{"key":"ref_55","first-page":"1898","article-title":"Constitutive activation of the Gs alpha protein-adenylate cyclase pathway may not be sufficient to generate toxic thyroid adenomas","volume":"81","author":"Derwahl","year":"1996","journal-title":"J. Clin. Endocrinol. Metab."},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"2227","DOI":"10.1007\/s00018-017-2728-1","article-title":"Evidence of G-protein-coupled receptor and substrate transporter heteromerization at a single molecule level","volume":"75","author":"Fischer","year":"2018","journal-title":"Cell Mol Life Sci"}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/20\/22\/5564\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T13:32:41Z","timestamp":1760189561000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/20\/22\/5564"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2019,11,7]]},"references-count":56,"journal-issue":{"issue":"22","published-online":{"date-parts":[[2019,11]]}},"alternative-id":["ijms20225564"],"URL":"https:\/\/doi.org\/10.3390\/ijms20225564","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2019,11,7]]}}}