The facts of the matter: What is a hormone?

Authors: Gerard Karsenty ^aff001
Authors place of work: Department of Genetics and Development, Columbia University Irving Medical Center, New York, New York, United States of America ^aff001
Published in the journal: The facts of the matter: What is a hormone?. PLoS Genet 16(6): e32767. doi:10.1371/journal.pgen.1008938
Category: Formal Comment
doi: https://doi.org/10.1371/journal.pgen.1008938

By choice, scientists value facts over beliefs. This is why science is constantly moving forward; it embraces novelty, provided it is fortified by evidence and all sorts of controls, checks and balances, as a sign of and a tool for progress.

A goal common to all individuals that devote their life to science is to advance knowledge. They may have other goals but this particular one has to be common to all scientists. This goal also creates a burden and not such an easy one to bear—the burden of restraint. Scientists live under the public light: they publish, i.e., share their findings with the scientific community at large and sometimes with the general public. What comes with that are huge responsibilities, the one to be right of course, but more importantly the responsibility to be cautious, to stay within the data and to not drag them beyond what they actually show. Otherwise the pursuit of knowledge is, unwillingly but definitely, tainted. Results have to be controlled over and over, and of course, just as importantly, their presentation has to be neutral as opposed to clever. This serves as a necessary preamble to a series of facts.

The word hormone means “that which sets in motion”, and was first used in 1902 before the emergence of molecular biology, yet its carefully worded definition has essentially remained unchanged. This definition is at the heart of this piece and our work on the endocrinology of bone over the past 13 years.

To be clear, a hormone is defined both experimentally and physiologically by what it does when present in abundance in animals or in humans because of an environmental stimulus, physiological perturbation, tumor, genetic abnormality, or when injected in an otherwise normal human being or animal of any species. As long as a molecule has not been shown to affect a particular physiological function when injected in a normal animal, it may be many things, it may have many names but a hormone it may not be. I cite here a recent and vivid example of what I write. Mice lacking the GLP1 receptor have a mild metabolic phenotype at best, yet GLP1 plays a critical role in physiology and its analogs are best-selling drugs for the treatment of diabetes [1]. Likewise, the blossoming field of FGF21 biology is based on gain of function experiments. What I write is not meant to disparage mouse genetics since it is part of what my lab has used to address many questions in the field of bone biology and I have learnt to appreciate its value. It is rather meant to remind myself and everyone else that biology is an assembly of disciplines and approaches. Scientists are not and should not be children in a toy store. The newest tool does not, and is not meant to, replace the older one, rather it enriches the tool kit. Even more to the point, one experimental approach with very small N, based on erroneous protocols, using mixed genetic backgrounds, does not suffice to invalidate, even if carefully worded, the work done by many others using multiple and state-of-the-art experimental approaches.

I write the preceding to introduce another fact: osteocalcin was shown to have properties of a hormone in mice, rats, monkeys and subsequently humans through gain-of-function experiments [2–52]. Those experiments occupy more than half of the original paper [50]. It was the analysis of mice generated for another purpose, but that happened to have too much osteocalcin that set this field in motion. These mice, the Esp-/- mice (Esp is a gene preferentially expressed in osteoblasts), were not only hyper-osteocalcinemic they were also hyperinsulinemic, hypertestosteronemic, agitated, and had the ability to run much longer than their wild type littermates [50,53]. Granted, this could have had many causes, but one of them that deserved to be tested was whether it was explained in part by their increased circulating osteocalcin levels. Thereafter it was only by 1) realizing that the phenotypes of Esp-/- and Osteocalcin-/- mice were mirror images of each other 2) performing genetic complementation experiments and 3) injecting osteocalcin in wild type mice or treating cells with osteocalcin that the notion that osteocalcin was a hormone was advanced [6,50,53–55].

I stand by these results unequivocally. I stand by them because they were so unexpected by us and everybody else that reviewers, rightly so, asked for control after control, each of which made the results more robust. The results were more robust because of the large number of mice (N over 50 by now in both sexes for every experiment). They were also more robust because they used cross-validating assays, every genetic control, different genetic backgrounds, both sexes, and various ages. They were more robust, and more credible, because for each experiment we used gold-standard techniques exactly as described by experts in fields other than our own. In fact, in most cases these experiments were conducted not by us, but by labs with whom we had shared the Ocn-/- or Esp-/- or other relevant mutant mice and who were world experts in the particular assays being conducted. To name just a very small portion, euglycemic clamps were done in the laboratory of Jason Kim, islet perifusion in the lab of Klaus Kaestner, indirect calorimetry in the lab of Jeffrey Pessin, conditional fear conditioning in the lab of Rene Hen, electron micrographs of testes in the one of Louis Hermo, whole cell electrophysiology in the lab of Xiao-Bing Gao, and in vivo nerve fiber recordings in the lab of Kamal Rahmouni. These are, by all criteria, renowned experts in their respective fields and there are many more that cannot be listed here. Together they brought histology, electron microscopy, electrophysiology, indirect calorimetry and metabolomics evidence to the picture. I repeat, by our own cautious design, each of these scientists worked independently with full latitude to verify or contradict our results and it is thanks to their work that our results are so robust. Thirteen years later, even if it was painful at the time, I take this opportunity to thank each and every reviewer for having made this work what it is.

Since the initial publication proposing that osteocalcin is a hormone there have been hundreds of publications confirming that osteocalcin behaves as a hormone that signals through two specific receptors that were notably identified by two competing groups and that elicits particular physiological responses in mice and humans. These studies, through different in vivo and in vitro assays, all show that osteocalcin, and by that I mean bioactive (undercarboxylated) and not total osteocalcin, has the properties of a hormone. These publications come from investigators across many fields; investigators I know, investigators I do not know, and I have never interacted with and even, investigators that were fierce competitors in making discoveries about the endocrine functions of osteocalcin. Some of these publications are from prominent investigators or a Nobel laureate, others from less recognized but equally hard-working, truth-seeking scientists who were not scared off or were perhaps attracted by novelty. Where the papers were published is irrelevant because, in my view, science is not driven by impact factor but by the rigorous, controlled, verified, pursuit of knowledge, novel knowledge. What is rewarding as an endocrinologist is to see that a large portion of this body of work was done as it should be, through gain-of-function experiments, and part of it in human cells or patients [9,39,51,52,56–61]. These publications come from over 30 groups and counting, across 5 continents, they were reviewed at journals by colleagues who were not necessarily benevolent, revised, re-reviewed, and finally published. As such they earned their credibility and deserve the respect that goes to any scientific publication. Not more respect of course, but certainly (why would it be?) not less. A few of them are listed below [2–49,52]. Taken together, over the past 13 years, these papers form a body of work, the very beginnings of a field. This near unanimity (science is a human activity after all and there is never complete unanimity) despite using different hands, models and assays is what more than anything else supports the idea that osteocalcin is a hormone. Let’s be absolutely clear, even if there were 1000 of these publications, the sheer number them would not be enough to make them right. But to prove them wrong the very same experiments from body weight measurements, to electron microscopy, and electrophysiology, etc. have to be performed in the same conditions and achieve different results, otherwise we are not speaking of scientific facts.

In all fairness, and I’m well aware of it, for every 20 publications or so showing that osteocalcin is a hormone there was always one failing to do so. Often there are technical or biological reasons for the discrepancy and the reason has often taught us new things about this new hormone whose nature we are only beginning to uncover. For example, one of them uses an outbred CRISPR generated, but not sequenced, rat model with N of 3 to 5 per group per experiment. Others may have measured total (as opposed to undercarboxylated) osteocalcin in humans, which makes it hard to rigorously determine the endocrinological relevance because these levels do not necessarily correlate with levels of bioactive osteocalcin. Those discrepancies and others have often served as reminders that this new hormone will likely be fertile territory for discovery for many years to come.

Incidentally, none of them, until now, came with an editorial [62] declaring that the book is closed and osteocalcin is not a hormone for the rest of time and insinuating (for most readers I know) scientific incompetence or superficiality on my part. I personally do not know of a scientific book that is ever closed. But PLOS Genetics just published 2 papers that contradict each other on many points but claim that 2 mouse models lacking osteocalcin are more or less wild type. They are accompanied with a vivid perspective, cleverly written that unquestionably raises doubt on the way I conduct science.

If a hormone is defined by what it does when in excess of baseline in normal animals, then unlike what the Perspective claims from its first to its last paragraph, neither of these papers addresses the question of whether osteocalcin is a hormone. They did not inject wild–type animals with any of the many preparations of osteocalcin shown by many other studies to work [2–49]. They simply tested whether the mice they generated and analyzed, through protocols different from those used by mainstream investigators, do not have a phenotype that we described in mice we generated and analyzed through different protocols. As such they join the small but respectable contingent of people who do not believe that osteocalcin is a hormone and do not believe bone is an endocrine organ. There is nothing wrong with that view a priori. It becomes wrong when these two papers are presented as the final truth. Although I will not address in detail whether or not there are experimental flaws in these studies, I will note that in the published reviews one of the reviewers commented that the genetic backgrounds, standard deviations and the number of animals analyzed per experiment were all questionable. The reviewer also mentioned that the conclusions were “too sweeping” and should be corrected.

Besides overt qualitative and quantitative differences in the ways the assays were performed the reasons for the discrepancies between their results and those of 30 labs is for now unknown and I am fully aware of it. But such discrepancies are the way novel discoveries, especially those effecting human health, often occur. The answer to this question can only emerge by exchanging mice, performing controls, analyzing them extensively, using the complete panel of appropriate endocrine tests and verifying comprehensively the outcome and specificity of the genetic methodologies used to inactivate Osteocalcin.

I need to state here that the Ocn-/- mice we generated have been and are available on demand and are already found all over the world. Moreover, these mutant mice have also been deposited at, and are available from, the Jackson Laboratories (https://www.jax.org/strain/034070). In contrast, and despite several solemn pronouncements in their papers to journalists and in social media for about a year, mice from the other parties have not been deposited at the Jackson Laboratories at the time I write this article. This is why I have contacted Dr. Williams who acknowledged that indeed his mouse model has not been submitted to the Jackson Laboratories. We are working diligently with him to exchange mice.

In addition to this exchange of reagents that I hope will occur for the sake of the entire community there is another question that scientists and journalists alike have asked me. If the papers published in PLOS Genetics have not addressed the definition of a hormone, why would their conclusions and a Perspective written by one of their reviewers, who is an osteoporosis expert, attack, with ardor, the work of the many labs who have addressed this question in mice, rats, monkeys and humans? I am not the one who can answer this question.

Science does not work through inference. Novelty comes through hard work, through the use of the entire experimental gamut, control after control, literally I would say through blood, sweat (a lot of it) and tears (some). Before telling hundreds of different scientists from different fields that they are all wrong, have always been, that their results are serving a dogma proposed by a slow-witted scientist that burdens federal funds one must conduct thoughtful, exhaustive and robust experimentation. I have been asked, often by the same people, after each of my contributions to the field: “Runx2 does not control bone formation”, “central neuronal control of bone mass is a fantasy”, “serotonin does not regulate bone mass”, “bone mass is not coordinated with energy metabolism hormones and reproduction” and now “osteocalcin is not a hormone”. None of these questions or their accompanying papers stood the test of time. Science is not politics and there is no room in science for what has the appearance of veiled character assassination. Before saying that a single novel truth based on negative and incomplete data has emerged overnight, one needs to do more than simply say it, one needs to walk the walk and demonstrate it. This was not done. Do these results discredit, unwillingly or not, previous work? In the short-term, with some people, maybe; but scientific life is much longer than that.

Zdroje

1. Knudsen LB. Inventing Liraglutide, a Glucagon-Like Peptide-1 Analogue, for the Treatment of Diabetes and Obesity. ACS Pharmacol Transl Sci. 2019;2(6):468–84. doi: 10.1021/acsptsci.9b00048 32259078

2. Al Rifai O, Chow J, Lacombe J, Julien C, Faubert D, Susan-Resiga D, et al. Proprotein convertase furin regulates osteocalcin and bone endocrine function. J Clin Invest. 2017;127(11):4104–17. doi: 10.1172/JCI93437 28972540

3. Boraschi-Diaz I, Tauer JT, El-Rifai O, Guillemette D, Lefebvre G, Rauch F, et al. Metabolic phenotype in the mouse model of osteogenesis imperfecta. J Endocrinol. 2017;234(3):279–89. doi: 10.1530/JOE-17-0335 28716975

4. Brennan-Speranza TC, Henneicke H, Gasparini SJ, Blankenstein KI, Heinevetter U, Cogger VC, et al. Osteoblasts mediate the adverse effects of glucocorticoids on fuel metabolism. J Clin Invest. 2012;122(11):4172–89. doi: 10.1172/JCI63377 23093779

5. Du J, Zhang M, Lu J, Zhang X, Xiong Q, Xu Y, et al. Osteocalcin improves nonalcoholic fatty liver disease in mice through activation of Nrf2 and inhibition of JNK. Endocrine. 2016;53(3):701–9. doi: 10.1007/s12020-016-0926-5 26994931

6. Fulzele K, Riddle RC, DiGirolamo DJ, Cao X, Wan C, Chen D, et al. Insulin receptor signaling in osteoblasts regulates postnatal bone acquisition and body composition. Cell. 2010;142(2):309–19. doi: 10.1016/j.cell.2010.06.002 20655471

7. Glatigny M, Moriceau S, Rivagorda M, Ramos-Brossier M, Nascimbeni AC, Lante F, et al. Autophagy Is Required for Memory Formation and Reverses Age-Related Memory Decline. Curr Biol. 2019;29(3):435–48 e8. doi: 10.1016/j.cub.2018.12.021 30661803

8. Guedes JAC, Esteves JV, Morais MR, Zorn TM, Furuya DT. Osteocalcin improves insulin resistance and inflammation in obese mice: Participation of white adipose tissue and bone. Bone. 2018;115:68–82. doi: 10.1016/j.bone.2017.11.020 29183784

9. Guo Q, Li H, Xu L, Wu S, Sun H, Zhou B. Undercarboxylated osteocalcin reverts insulin resistance induced by endoplasmic reticulum stress in human umbilical vein endothelial cells. Sci Rep. 2017;7(1):46. doi: 10.1038/s41598-017-00163-2 28246389

10. Guo XZ, Shan C, Hou YF, Zhu G, Tao B, Sun LH, et al. Osteocalcin Ameliorates Motor Dysfunction in a 6-Hydroxydopamine-Induced Parkinson's Disease Rat Model Through AKT/GSK3beta Signaling. Front Mol Neurosci. 2018;11:343. doi: 10.3389/fnmol.2018.00343 30319352

11. Gupte AA, Sabek OM, Fraga D, Minze LJ, Nishimoto SK, Liu JZ, et al. Osteocalcin protects against nonalcoholic steatohepatitis in a mouse model of metabolic syndrome. Endocrinology. 2014;155(12):4697–705. doi: 10.1210/en.2014-1430 25279794

12. Hayashi Y, Kawakubo-Yasukochi T, Mizokami A, Hazekawa M, Yakura T, Naito M, et al. Uncarboxylated Osteocalcin Induces Antitumor Immunity against Mouse Melanoma Cell Growth. J Cancer. 2017;8(13):2478–86. doi: 10.7150/jca.18648 28900485

13. Hayashi Y, Kawakubo-Yasukochi T, Mizokami A, Takeuchi H, Nakamura S, Hirata M. Differential Roles of Carboxylated and Uncarboxylated Osteocalcin in Prostate Cancer Growth. J Cancer. 2016;7(12):1605–9. doi: 10.7150/jca.15523 27698897

14. Huesa C, Zhu D, Glover JD, Ferron M, Karsenty G, Milne EM, et al. Deficiency of the bone mineralization inhibitor NPP1 protects mice against obesity and diabetes. Dis Model Mech. 2014;7(12):1341–50. doi: 10.1242/dmm.017905 25368121

15. Kawakubo-Yasukochi T, Kondo A, Mizokami A, Hayashi Y, Chishaki S, Nakamura S, et al. Maternal oral administration of osteocalcin protects offspring from metabolic impairment in adulthood. Obesity (Silver Spring). 2016;24(4):895–907.

16. Kode A, Mosialou I, Silva BC, Joshi S, Ferron M, Rached MT, et al. FoxO1 protein cooperates with ATF4 protein in osteoblasts to control glucose homeostasis. J Biol Chem. 2012;287(12):8757–68. doi: 10.1074/jbc.M111.282897 22298775

17. Kosmidis S, Polyzos A, Harvey L, Youssef M, Denny CA, Dranovsky A, et al. RbAp48 Protein Is a Critical Component of GPR158/OCN Signaling and Ameliorates Age-Related Memory Loss. Cell Rep. 2018;25(4):959–73 e6. doi: 10.1016/j.celrep.2018.09.077 30355501

18. Lacombe J, Karsenty G, Ferron M. In vivo analysis of the contribution of bone resorption to the control of glucose metabolism in mice. Mol Metab. 2013;2(4):498–504. doi: 10.1016/j.molmet.2013.08.004 24327965

19. Levinger I, Lin X, Zhang X, Brennan-Speranza TC, Volpato B, Hayes A, et al. The effects of muscle contraction and recombinant osteocalcin on insulin sensitivity ex vivo. Osteoporos Int. 2016;27(2):653–63. doi: 10.1007/s00198-015-3273-0 26259649

20. Li H, Zhou B, Xu L, Liu J, Zang W, Wu S, et al. The reciprocal interaction between autophagic dysfunction and ER stress in adipose insulin resistance. Cell Cycle. 2014;13(4):565–79. doi: 10.4161/cc.27406 24309597

21. Lin X, Parker L, McLennan E, Hayes A, McConell G, Brennan-Speranza TC, et al. Undercarboxylated Osteocalcin Improves Insulin-Stimulated Glucose Uptake in Muscles of Corticosterone-Treated Mice. J Bone Miner Res. 2019;34(8):1517–30. doi: 10.1002/jbmr.3731 30908701

22. Lin X, Parker L, McLennan E, Zhang X, Hayes A, McConell G, et al. Recombinant Uncarboxylated Osteocalcin Per Se Enhances Mouse Skeletal Muscle Glucose Uptake in both Extensor Digitorum Longus and Soleus Muscles. Front Endocrinol (Lausanne). 2017;8:330.

23. Lin X, Parker L, McLennan E, Zhang X, Hayes A, McConell G, et al. Uncarboxylated Osteocalcin Enhances Glucose Uptake Ex Vivo in Insulin-Stimulated Mouse Oxidative But Not Glycolytic Muscle. Calcif Tissue Int. 2018;103(2):198–205. doi: 10.1007/s00223-018-0400-x 29427234

24. Matsumoto Y, La Rose J, Lim M, Adissu HA, Law N, Mao X, et al. Ubiquitin ligase RNF146 coordinates bone dynamics and energy metabolism. J Clin Invest. 2017;127(7):2612–25. doi: 10.1172/JCI92233 28581440

25. Mizokami A, Mukai S, Gao J, Kawakubo-Yasukochi T, Otani T, Takeuchi H, et al. GLP-1 signaling is required for improvement of glucose tolerance by osteocalcin. J Endocrinol. 2020;244(2):285–96. doi: 10.1530/JOE-19-0288 31693486

26. Mizokami A, Wang D, Tanaka M, Gao J, Takeuchi H, Matsui T, et al. An extract from pork bones containing osteocalcin improves glucose metabolism in mice by oral administration. Biosci Biotechnol Biochem. 2016;80(11):2176–83. doi: 10.1080/09168451.2016.1214530 27460506

27. Mizokami A, Yasutake Y, Gao J, Matsuda M, Takahashi I, Takeuchi H, et al. Osteocalcin induces release of glucagon-like peptide-1 and thereby stimulates insulin secretion in mice. PLoS One. 2013;8(2):e57375. doi: 10.1371/journal.pone.0057375 23437377

28. Mizokami A, Yasutake Y, Higashi S, Kawakubo-Yasukochi T, Chishaki S, Takahashi I, et al. Oral administration of osteocalcin improves glucose utilization by stimulating glucagon-like peptide-1 secretion. Bone. 2014;69:68–79. doi: 10.1016/j.bone.2014.09.006 25230237

29. Namai F, Shigemori S, Sudo K, Sato T, Yamamoto Y, Nigar S, et al. Recombinant Mouse Osteocalcin Secreted by Lactococcus lactis Promotes Glucagon-Like Peptide-1 Induction in STC-1 Cells. Curr Microbiol. 2018;75(1):92–8. doi: 10.1007/s00284-017-1354-3 28905106

30. Otani T, Matsuda M, Mizokami A, Kitagawa N, Takeuchi H, Jimi E, et al. Osteocalcin triggers Fas/FasL-mediated necroptosis in adipocytes via activation of p300. Cell Death Dis. 2018;9(12):1194. doi: 10.1038/s41419-018-1257-7 30546087

31. Otani T, Mizokami A, Hayashi Y, Gao J, Mori Y, Nakamura S, et al. Signaling pathway for adiponectin expression in adipocytes by osteocalcin. Cell Signal. 2015;27(3):532–44. doi: 10.1016/j.cellsig.2014.12.018 25562427

32. Pi M, Kapoor K, Ye R, Nishimoto SK, Smith JC, Baudry J, et al. Evidence for Osteocalcin Binding and Activation of GPRC6A in beta-Cells. Endocrinology. 2016;157(5):1866–80. doi: 10.1210/en.2015-2010 27007074

33. Pi M, Wu Y, Quarles LD. GPRC6A mediates responses to osteocalcin in beta-cells in vitro and pancreas in vivo. J Bone Miner Res. 2011;26(7):1680–3. doi: 10.1002/jbmr.390 21425331

34. Pi M, Xu F, Ye R, Nishimoto SK, Williams RW, Lu L, et al. Role of GPRC6A in Regulating Hepatic Energy Metabolism in Mice. Sci Rep. 2020;10(1):7216. doi: 10.1038/s41598-020-64384-8 32350388

35. Qaradakhi T, Gadanec LK, Tacey AB, Hare DL, Buxton BF, Apostolopoulos V, et al. The Effect of Recombinant Undercarboxylated Osteocalcin on Endothelial Dysfunction. Calcif Tissue Int. 2019;105(5):546–56. doi: 10.1007/s00223-019-00600-6 31485687

36. Rached MT, Kode A, Silva BC, Jung DY, Gray S, Ong H, et al. FoxO1 expression in osteoblasts regulates glucose homeostasis through regulation of osteocalcin in mice. J Clin Invest. 2010;120(1):357–68. doi: 10.1172/JCI39901 20038793

37. Rashdan NA, Sim AM, Cui L, Phadwal K, Roberts FL, Carter R, et al. Osteocalcin Regulates Arterial Calcification Via Altered Wnt Signaling and Glucose Metabolism. J Bone Miner Res. 2020;35(2):357–67. doi: 10.1002/jbmr.3888 31596966

38. Riddle RC, Frey JL, Tomlinson RE, Ferron M, Li Y, DiGirolamo DJ, et al. Tsc2 is a molecular checkpoint controlling osteoblast development and glucose homeostasis. Mol Cell Biol. 2014;34(10):1850–62. doi: 10.1128/MCB.00075-14 24591652

39. Sabek OM, Nishimoto SK, Fraga D, Tejpal N, Ricordi C, Gaber AO. Osteocalcin Effect on Human beta-Cells Mass and Function. Endocrinology. 2015;156(9):3137–46. doi: 10.1210/EN.2015-1143 26151356

40. Solhjoo S, Akbari M, Toolee H, Mortezaee K, Mohammadipour M, Nematollahi-Mahani SN, et al. Roles for osteocalcin in proliferation and differentiation of spermatogonial cells cocultured with somatic cells. J Cell Biochem. 2019;120(4):4924–34. doi: 10.1002/jcb.27767 30302795

41. Tsuka S, Aonuma F, Higashi S, Ohsumi T, Nagano K, Mizokami A, et al. Promotion of insulin-induced glucose uptake in C2C12 myotubes by osteocalcin. Biochem Biophys Res Commun. 2015;459(3):437–42. doi: 10.1016/j.bbrc.2015.02.123 25735975

42. Wei J, Hanna T, Suda N, Karsenty G, Ducy P. Osteocalcin promotes beta-cell proliferation during development and adulthood through Gprc6a. Diabetes. 2014;63(3):1021–31. doi: 10.2337/db13-0887 24009262

43. Yasutake Y, Mizokami A, Kawakubo-Yasukochi T, Chishaki S, Takahashi I, Takeuchi H, et al. Long-term oral administration of osteocalcin induces insulin resistance in male mice fed a high-fat, high-sucrose diet. Am J Physiol Endocrinol Metab. 2016;310(8):E662–E75. doi: 10.1152/ajpendo.00334.2015 26884384

44. Ye R, Pi M, Cox JV, Nishimoto SK, Quarles LD. CRISPR/Cas9 targeting of GPRC6A suppresses prostate cancer tumorigenesis in a human xenograft model. J Exp Clin Cancer Res. 2017;36(1):90. doi: 10.1186/s13046-017-0561-x 28659174

45. Yi M, Wu Y, Long J, Liu F, Liu Z, Zhang YH, et al. Exosomes secreted from osteocalcin-overexpressing endothelial progenitor cells promote endothelial cell angiogenesis. Am J Physiol Cell Physiol. 2019;317(5):C932–C41. doi: 10.1152/ajpcell.00534.2018 31411920

46. Yoshikawa Y, Kode A, Xu L, Mosialou I, Silva BC, Ferron M, et al. Genetic evidence points to an osteocalcin-independent influence of osteoblasts on energy metabolism. J Bone Miner Res. 2011;26(9):2012–25. doi: 10.1002/jbmr.417 21557308

47. Zhou B, Li H, Liu J, Xu L, Zang W, Wu S, et al. Intermittent injections of osteocalcin reverse autophagic dysfunction and endoplasmic reticulum stress resulting from diet-induced obesity in the vascular tissue via the NFkappaB-p65-dependent mechanism. Cell Cycle. 2013;12(12):1901–13. doi: 10.4161/cc.24929 23708521

48. Zhou B, Li H, Xu L, Zang W, Wu S, Sun H. Osteocalcin reverses endoplasmic reticulum stress and improves impaired insulin sensitivity secondary to diet-induced obesity through nuclear factor-kappaB signaling pathway. Endocrinology. 2013;154(3):1055–68. doi: 10.1210/en.2012-2144 23407450

49. De Toni L, De Filippis V, Tescari S, Ferigo M, Ferlin A, Scattolini V, et al. Uncarboxylated osteocalcin stimulates 25-hydroxy vitamin D production in Leydig cell line through a GPRC6a-dependent pathway. Endocrinology. 2014;155(11):4266–74. doi: 10.1210/en.2014-1283 25093461

50. Lee NK, Sowa H, Hinoi E, Ferron M, Ahn JD, Confavreux C, et al. Endocrine regulation of energy metabolism by the skeleton. Cell. 2007;130(3):456–69. doi: 10.1016/j.cell.2007.05.047 17693256

51. Chowdhury S, Schulz LC, Palmisano B, Singh P, Berger JM, Yadav VK, et al. Muscle derived interleukin-6 increases exercise capacity by signaling in osteoblasts. J Clin Invest. 2020.

52. Millar SA, Anderson SI, O'Sullivan S E. Human vascular cell responses to the circulating bone hormone osteocalcin. J Cell Physiol. 2019;234(11):21039–48. doi: 10.1002/jcp.28707 31026070

53. Oury F, Sumara G, Sumara O, Ferron M, Chang H, Smith CE, et al. Endocrine regulation of male fertility by the skeleton. Cell. 2011;144(5):796–809. doi: 10.1016/j.cell.2011.02.004 21333348

54. Oury F, Khrimian L, Denny CA, Gardin A, Chamouni A, Goeden N, et al. Maternal and offspring pools of osteocalcin influence brain development and functions. Cell. 2013;155(1):228–41. doi: 10.1016/j.cell.2013.08.042 24074871

55. Ferron M, McKee MD, Levine RL, Ducy P, Karsenty G. Intermittent injections of osteocalcin improve glucose metabolism and prevent type 2 diabetes in mice. Bone. 2012;50(2):568–75. doi: 10.1016/j.bone.2011.04.017 21550430

56. Di Nisio A, Rocca MS, Fadini GP, De Toni L, Marcuzzo G, Marescotti MC, et al. The rs2274911 polymorphism in GPRC6A gene is associated with insulin resistance in normal weight and obese subjects. Clin Endocrinol (Oxf). 2017;86(2):185–91.

57. De Toni L, Di Nisio A, Speltra E, Rocca MS, Ghezzi M, Zuccarello D, et al. Polymorphism rs2274911 of GPRC6A as a Novel Risk Factor for Testis Failure. J Clin Endocrinol Metab. 2016;101(3):953–61. doi: 10.1210/jc.2015-3967 26735260

58. Korostishevsky M, Malkin I, Trofimov S, Pei Y, Deng HW, Livshits G. Significant association between body composition phenotypes and the osteocalcin genomic region in normative human population. Bone. 2012;51(4):688–94. doi: 10.1016/j.bone.2012.07.010 22842327

59. Das SK, Sharma NK, Elbein SC. Analysis of osteocalcin as a candidate gene for type 2 diabetes (T2D) and intermediate traits in Caucasians and African Americans. Dis Markers. 2010;28(5):281–6. doi: 10.3233/DMA-2010-0701 20592451

60. Oury F, Ferron M, Huizhen W, Confavreux C, Xu L, Lacombe J, et al. Osteocalcin regulates murine and human fertility through a pancreas-bone-testis axis. J Clin Invest. 2013;123(6):2421–33. doi: 10.1172/JCI65952 23728177

61. Confavreux CB, Borel O, Lee F, Vaz G, Guyard M, Fadat C, et al. Osteoid osteoma is an osteocalcinoma affecting glucose metabolism. Osteoporos Int. 2012;23(5):1645–50. doi: 10.1007/s00198-011-1684-0 21681611

62. Manolagas SC. Osteocalcin promotes bone mineralization but is not a hormone. PLoS Genet. 2020;16(6):e1008714.

Článek Duplication and divergence of the retrovirus restriction gene Fv1 in Mus caroli allows protection from multiple retroviruses

Článek Super-resolution imaging of RAD51 and DMC1 in DNA repair foci reveals dynamic distribution patterns in meiotic prophase

Článek Steroid hormones regulate genome-wide epigenetic programming and gene transcription in human endometrial cells with marked aberrancies in endometriosis

Článek Regulation of olfactory-based sex behaviors in the silkworm by genes in the sex-determination cascade

Článek Osteocalcin promotes bone mineralization but is not a hormone

Článek A conserved, N-terminal tyrosine signal directs Ras for inhibition by Rabex-5

Článek Integrins regulate epithelial cell shape by controlling the architecture and mechanical properties of basal actomyosin networks

Článek Cancer-associated mutations in the iron-sulfur domain of FANCJ affect G-quadruplex metabolism

Článek NRF2 loss recapitulates heritable impacts of paternal cigarette smoke exposure

Článek Pax6 organizes the anterior eye segment by guiding two distinct neural crest waves

Článek Transcriptomic stratification of late-onset Alzheimer's cases reveals novel genetic modifiers of disease pathology