Advertisement

Effect of intra-ovarian injection of mesenchymal stem cells in aged mares

  • Sicilia T. Grady
  • Ashlee E. Watts
  • James A. Thompson
  • M. Cecilia T. Penedo
  • Kranti Konganti
  • Katrin Hinrichs
Reproductive Physiology and Disease

Abstract

Purpose

This study aims to determine if intra-ovarian injection of bone marrow–derived mesenchymal stem cells (MSCs) improves or restores ovarian function in aged females.

Methods

Prospective randomized study of eight aged mares and six young mares receiving intra-ovarian injection of MSCs or vehicle. Main outcome measures were antral follicle count and serum anti-Müllerian hormone (AMH) (aged and young mares), and for aged mares, oocyte meiotic and developmental competence; gross and histological ovarian assessment; evaluation of presence of chimerism in recovered granulosa cells and in ovarian tissue samples; and gene expression in ovarian tissue as assessed by RNA sequencing.

Results

Injection of MSCs was not associated with significant changes in follicle number, oocyte recovery rate on follicle aspiration, oocyte maturation rate, or blastocyst rate after ICSI in aged mares, or in changes in follicle number in young mares. There were no significant changes in peripheral AMH concentrations, indicating a lack of effect on growing follicles. MSC donor DNA was not recovered in granulosa cells or in ovarian tissue, indicating lack of persistence of injected MSC. RNA sequencing revealed significant differences in gene expression between MSC- and vehicle-injected ovaries.

Conclusions

Intra-ovarian injection of bone marrow–derived MSCs altered gene expression but did not improve ovarian function in aged mares.

Keywords

Aging Anti-Müllerian hormone Equids Fertility Follicle-stimulating hormone Follicular development Oocyte Ovary Stem cells 

Notes

Acknowledgments

The authors thank Dr. Andrew Hillhouse for his help with RNA isolation; Dr. Gus Wright for his help with flow cytometry analyses; Dr. Young Ho Choi for performing ICSI on the recovered oocytes; Hsing Fann for her help with MSC culture, freezing, and thawing for intra-ovarian injections; and Angel del Valle for his help with mtDNA sequencing. The authors acknowledge Texas A&M Institute for Genome Sciences and Society (TIGSS) for providing computational resources for RNA-Seq data analysis and systems administration support for the TIGSS HPC Cluster.

Funding

Funded by the Clinical Equine ICSI Program at Texas A&M University, the Link Equine Research Fund at Texas A&M University, and a Postdoctoral Trainee Research Grant and a Graduate Student Core Facility Experiential Learning Program Grant from the College of Veterinary Medicine and Biomedical Sciences. S.T.G. was funded in part by a Texas A&M College of Veterinary Medicine & Biomedical Sciences Merit Scholars Fellowship.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

10815_2018_1371_MOESM1_ESM.docx (54 kb)
ESM 1 (DOCX 54 kb)

References

  1. 1.
    Itay S, Abramovici A, Nevo Z. Use of cultured embryonal chick epiphyseal chondrocytes as grafts for defects in chick articular-cartilage. Clin Orthop. 1987;220:284–303.Google Scholar
  2. 2.
    Wilke MM, Nydam DV, Nixon AJ. Enhanced early chondrogenesis in articular defects following arthroscopic mesenchymal stem cell implantation in an equine model. J Orthop Res. 2007;25:913–25.CrossRefGoogle Scholar
  3. 3.
    Lee RH, Seo MJ, Reger RL, Spees JL, Pulin AA, Olson SD, et al. Multipotent stromal cells from human marrow home to and promote repair of pancreatic islets and renal glomeruli in diabetic NOD/scid mice. Proc Natl Acad Sci U S A. 2006;103:17438–43.CrossRefGoogle Scholar
  4. 4.
    Phinney DG, Isakova I. Plasticity and therapeutic potential of mesenchymal stem cells in the nervous system. Curr Pharm Des. 2005;11:1255–65.CrossRefGoogle Scholar
  5. 5.
    Ortiz LA, Gambelli F, McBride C, Gaupp D, Baddoo M, Kaminski N, et al. Mesenchymal stem cell engraftment in lung is enhanced in response to bleomycin exposure and ameliorates its fibrotic effects. Proc Natl Acad Sci U S A. 2003;100:8407–11.CrossRefGoogle Scholar
  6. 6.
    Minguell JJ, Erices A. Mesenchymal stem cells and the treatment of cardiac disease. Exp Biol Med (Maywood). 2006;231:39–49.CrossRefGoogle Scholar
  7. 7.
    Salooja N, Szydlo RM, Socie G, Rio B, Chatterjee R, Ljungman P, et al. Pregnancy outcomes after peripheral blood or bone marrow transplantation: a retrospective survey. Lancet. 2001;358:271–6.CrossRefGoogle Scholar
  8. 8.
    Sanders JE, Buckner CD, Amos D, Levy W, Appelbaum FR, Doney K, et al. Ovarian function following marrow transplantation for aplastic anemia or leukemia. J Clin Oncol. 1988;6:813–8.CrossRefGoogle Scholar
  9. 9.
    Sanders JE, Hawley J, Levy W, Gooley T, Buckner CD, Deeg HJ, et al. Pregnancies following high-dose cyclophosphamide with or without high-dose busulfan or total-body irradiation and bone marrow transplantation. Blood. 1996;87:3045–52.PubMedGoogle Scholar
  10. 10.
    Takehara Y, Yabuuchi A, Ezoe K, Kuroda T, Yamadera R, Sano C, et al. The restorative effects of adipose-derived mesenchymal stem cells on damaged ovarian function. Lab Investig. 2013;93:181–93.CrossRefGoogle Scholar
  11. 11.
    Wang S, Yu L, Sun M, Mu S, Wang C, Wang D, et al. The therapeutic potential of umbilical cord mesenchymal stem cells in mice premature ovarian failure. Biomed Res Int. 2013;2013:690491.PubMedPubMedCentralGoogle Scholar
  12. 12.
    Lee H-J, Selesniemi K, Niikura Y, Niikura T, Klein R, Dombkowski DM, et al. Bone marrow transplantation generates immature oocytes and rescues long-term fertility in a preclinical mouse model of chemotherapy-induced premature ovarian failure. J Clin Oncol. 2007;25:3198–204.CrossRefGoogle Scholar
  13. 13.
    Fu X, He Y, Xie C, Liu W. Bone marrow mesenchymal stem cell transplantation improves ovarian function and structure in rats with chemotherapy-induced ovarian damage. Cytotherapy. 2008;10:353–63.CrossRefGoogle Scholar
  14. 14.
    Mohammed Ali AF, et al. Fertility treatment of aged women by laparoscopic intra ovarian injection of peripheral blood mononuclear cell (PBMNC) a new modality. Fertil Mag. 2013;52–5.Google Scholar
  15. 15.
    Herraiz S, Romeu M, Buigues A, Martinez S, Diaz-Garcia C, Gomez-Segui I, et al. Autologous stem cell ovarian transplantation to increase reproductive potential in patients who are poor responders. Fertil Steril. 2018;110:496–505 el.CrossRefGoogle Scholar
  16. 16.
    Li J, Mao Q, He J, She H, Zhang Z, Yin C. Human umbilical cord mesenchymal stem cells improve the reserve function of perimenopausal ovary via a paracrine mechanism. Stem Cell Res Ther. 2017;8:55.CrossRefGoogle Scholar
  17. 17.
    Carnevale EM, Ginther OJ. Defective oocytes as a cause of subfertility in old mares. Biol Reprod. 1995;Monograph 1:209–14.CrossRefGoogle Scholar
  18. 18.
    Fitzgerald C, Zimon AE, Jones EE. Aging and reproductive potential in women. Yale J Biol Med. 1998;71:367–81.PubMedPubMedCentralGoogle Scholar
  19. 19.
    Colleoni S, Barbacini S, Necchi D, Duchi R, Lazzari G, Galli C. Application of ovum pick-up, intracytoplasmic sperm injection and embryo culture in equine practice. Proc Am Assoc Equine Pract. 2007;53:554–9.Google Scholar
  20. 20.
    Jacobson CC, Choi YH, Hayden SS, Hinrichs K. Recovery of mare oocytes on a fixed biweekly schedule, and resulting blastocyst formation after intracytoplasmic sperm injection. Theriogenology. 2010;73:1116–26.CrossRefGoogle Scholar
  21. 21.
    Sellon DC. How to obtain a diagnostic bone marrow sample from the sternum of an adult horse. Proc Am Assoc Equine Pract. 2006;52:621–5.Google Scholar
  22. 22.
    Dominici M, Le Blanc K, Mueller I, Slaper-Cortenbach I, Marini F, Krause D, et al. Minimal criteria for defining multipotent mesenchymal stromal cells. The International Society for Cellular Therapy position statement. Cytotherapy. 2006;8:315–7.CrossRefGoogle Scholar
  23. 23.
    De Schauwer C, Piepers S, Van de Walle GR, Demeyere K, Hoogewijs MK, Govaere JL, et al. In search for cross-reactivity to immunophenotype equine mesenchymal stromal cells by multicolor flow cytometry. Cytometry A. 2012;81:312–23.CrossRefGoogle Scholar
  24. 24.
    Schnabel LV, Pezzanite LM, Antczak DF, Felippe MJ, Fortier LA. Equine bone marrow-derived mesenchymal stromal cells are heterogeneous in MHC class II expression and capable of inciting an immune response in vitro. Stem Cell Res Ther. 2014;5:13.CrossRefGoogle Scholar
  25. 25.
    Mitchell A, Rivas KA, Smith R, Watts AE. Cryopreservation of equine mesenchymal stem cells in 95% autologous serum and 5% DMSO does not alter post-thaw growth or morphology in vitro compared to fetal bovine serum or allogeneic serum at 20 or 95% and DMSO at 10 or 5%. Stem Cell Res Ther. 2015;6:1–12.CrossRefGoogle Scholar
  26. 26.
    Brück I, Raun K, Synnestvedt B, Greve T. Follicle aspiration in the mare using a transvaginal ultrasound-guided technique (short communication). Equine Vet J. 1992;24:58–9.CrossRefGoogle Scholar
  27. 27.
    Choi YH, Ross P, Velez IC, Macias-Garcia B, Riera FL, Hinrichs K. Cell lineage allocation in equine blastocysts produced in vitro under varying glucose concentrations. Reproduction. 2015;150:31–41.CrossRefGoogle Scholar
  28. 28.
    Choi YH, Love LB, Varner DD, Hinrichs K. Holding immature equine oocytes in the absence of meiotic inhibitors: effect on germinal vesicle chromatin and blastocyst development after intracytoplasmic sperm injection. Theriogenology. 2006;66:955–63.CrossRefGoogle Scholar
  29. 29.
    Rowland AL, Glass KG, Grady ST, Cummings KJ, Hinrichs K, Watts AE. Influence of caudal epidural analgesia on cortisol concentrations and pain-related behavioral responses in mares during and after ovariectomy via colpotomy. Vet Surg. 2018;47:715–21.CrossRefGoogle Scholar
  30. 30.
    Alves KA, Alves BG, Rocha CD, Visonna M, Mohallem RF, Gastal MO, et al. Number and density of equine preantral follicles in different ovarian histological section thicknesses. Theriogenology. 2015;83:1048–55.CrossRefGoogle Scholar
  31. 31.
    van de Goor LH, Panneman H, van Haeringen WA. A proposal for standardization in forensic equine DNA typing: allele nomenclature for 17 equine-specific STR loci. Anim Genet. 2010;41:122–7.CrossRefGoogle Scholar
  32. 32.
    Bowling AT, Del Valle A, Bowling M. A pedigree-based study of mitochondrial D-loop DNA sequence variation among Arabian horses. Anim Genet. 2000;31:1–7.CrossRefGoogle Scholar
  33. 33.
    Bolger AM, Lohse M, Usadel B. Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics. 2014;btu170.Google Scholar
  34. 34.
    Kim D, Langmead B, Salzberg SL. HISAT: a fast spliced aligner with low memory requirements. Nat Meth. 2015;12:357–60.CrossRefGoogle Scholar
  35. 35.
    Anders S, Pyl PT, Huber W. HTSeq—a Python framework to work with high-throughput sequencing data. Bioinformatics. 2014;btu638.Google Scholar
  36. 36.
    Love M, Anders S, Huber W. Differential analysis of count data—the DESeq2 package. Genome Biol. 2014;15:550.CrossRefGoogle Scholar
  37. 37.
    Zhang JL. Comparative investigation of three Bayesian p values. Comput Stat Data Anal. 2014;79:277–91.CrossRefGoogle Scholar
  38. 38.
    Hinrichs K, Schmidt AL. Meiotic competence in horse oocytes: interactions among chromatin configuration, follicle size, cumulus morphology, and season. Biol Reprod. 2000;62:1402–8.CrossRefGoogle Scholar
  39. 39.
    Velez I, Arnold C, Jacobson C, Norris J, Choi Y, Edwards J, et al. Effects of repeated transvaginal aspiration of immature follicles on mare health and ovarian status. Equine Vet J. 2012;44:78–83.CrossRefGoogle Scholar
  40. 40.
    Duchamp G, Bézard J, Palmer E. Oocyte yield and the consequences of puncture of all follicles larger than 8 millimetres in mares. Biol Reprod. 1995;Monograph 1:233–41.CrossRefGoogle Scholar
  41. 41.
    Abd-Allah SH, Shalaby SM, Pasha HF, El-Shal AS, Raafat N, Shabrawy SM, et al. Mechanistic action of mesenchymal stem cell injection in the treatment of chemically induced ovarian failure in rabbits. Cytotherapy. 2013;15:64–75.CrossRefGoogle Scholar
  42. 42.
    Ghadami M, El-Demerdash E, Zhang D, Salama SA, Binhazim AA, Archibong AE, et al. Bone marrow transplantation restores follicular maturation and steroid hormones production in a mouse model for primary ovarian failure. PLoS One. 2012;7:e32462.CrossRefGoogle Scholar
  43. 43.
    Rosen C, Shezen E, Aronovich A, Klionsky YZ, Yaakov Y, Assayag M, et al. Preconditioning allows engraftment of mouse and human embryonic lung cells, enabling lung repair in mice. Nat Med. 2015;21:869–79.CrossRefGoogle Scholar
  44. 44.
    Schnabel LV, Lynch ME, van der Meulen MC, Yeager AE, Kornatowski MA, Nixon AJ. Mesenchymal stem cells and insulin-like growth factor-I gene-enhanced mesenchymal stem cells improve structural aspects of healing in equine flexor digitorum superficialis tendons. J Orthop Res. 2009;27:1392–8.CrossRefGoogle Scholar
  45. 45.
    Iso Y, Spees JL, Serrano C, Bakondi B, Pochampally R, Song YH, et al. Multipotent human stromal cells improve cardiac function after myocardial infarction in mice without long-term engraftment. Biochem Biophys Res Commun. 2007;354:700–6.CrossRefGoogle Scholar
  46. 46.
    Spees JL, Olson SD, Whitney MJ, Prockop DJ. Mitochondrial transfer between cells can rescue aerobic respiration. Proc Natl Acad Sci U S A. 2006;103:1283–8.CrossRefGoogle Scholar
  47. 47.
    Claes A, Ball BA, Scoggin KE, Esteller-Vico A, Kalmar JJ, Conley AJ et al. The interrelationship between anti-Mullerian hormone, ovarian follicular populations and age in mares. Equine Vet J. 2014.Google Scholar
  48. 48.
    Behnam B, Modarressi MH, Conti V, Taylor KE, Puliti A, Wolfe J. Expression of Tsga10 sperm tail protein in embryogenesis and neural development: from cilium to cell division. Biochem Biophys Res Commun. 2006;344:1102–10.CrossRefGoogle Scholar
  49. 49.
    Mobasheri MB, Jahanzad I, Mohagheghi MA, Aarabi M, Farzan S, Modarressi MH. Expression of two testis-specific genes, TSGA10 and SYCP3, in different cancers regarding to their pathological features. Cancer Detect Prev. 2007;31:296–302.CrossRefGoogle Scholar
  50. 50.
    Barrett T, Wilhite SE, Ledoux P, Evangelista C, Kim IF, Tomashevsky M, et al. NCBI GEO: archive for functional genomics data sets—update. Nucleic Acids Res. 2013;41:D991–D5.CrossRefGoogle Scholar
  51. 51.
    Small CL, Shima JE, Uzumcu M, Skinner MK, Griswold MD. Profiling gene expression during the differentiation and development of the murine embryonic gonad. Biol Reprod. 2005;72:492–501.CrossRefGoogle Scholar
  52. 52.
    Kenigsberg S, Bentov Y, Chalifa-Caspi V, Potashnik G, Ofir R, Birk OS. Gene expression microarray profiles of cumulus cells in lean and overweight-obese polycystic ovary syndrome patients. Mol Hum Reprod. 2009;15:89–103.CrossRefGoogle Scholar
  53. 53.
    Miryounesi M, Nayernia K, Mobasheri MB, Dianatpour M, Oko R, Savad S, et al. Evaluation of in vitro spermatogenesis system effectiveness to study genes behavior: monitoring the expression of the testis specific 10 (Tsga10) gene as a model. Arch Iran Med. 2014;17:692–7.PubMedGoogle Scholar
  54. 54.
    Tanaka R, Ono T, Sato S, Nakada T, Koizumi F, Hasegawa K, et al. Over-expression of the testis-specific gene TSGA10 in cancers and its immunogenicity. Microbiol Immunol. 2004;48:339–45.CrossRefGoogle Scholar
  55. 55.
    Volodko N, Gordon M, Salla M, Ghazaleh HA, Baksh S. RASSF tumor suppressor gene family: biological functions and regulation. FEBS Lett. 2014;588:2671–84.CrossRefGoogle Scholar
  56. 56.
    Lee C-M, Yang P, Chen L-C, Chen C-C, Wu S-C, Cheng H-Y, et al. A novel role of RASSF9 in maintaining epidermal homeostasis. PLoS One. 2011;6:e17867.CrossRefGoogle Scholar
  57. 57.
    Mashaghi A, Marmalidou A, Tehrani M, Grace PM, Pothoulakis C, Dana R. Neuropeptide substance P and the immune response. Cell Mol Life Sci. 2016;73:4249–64.CrossRefGoogle Scholar
  58. 58.
    Ziche M, Morbidelli L, Pacini M, Geppetti P, Alessandri G, Maggi CA. Substance P stimulates neovascularization in vivo and proliferation of cultured endothelial cells. Microvasc Res. 1990;40:264–78.CrossRefGoogle Scholar
  59. 59.
    Ahmad M, Srinivasula SM, Wang L, Talanian RV, Litwack G, Fernandes-Alnemri T, et al. CRADD, a novel human apoptotic adaptor molecule for caspase-2, and FasL/tumor necrosis factor receptor-interacting protein RIP. Cancer Res. 1997;57:615–9.PubMedGoogle Scholar
  60. 60.
    Joswig AJ, Mitchell A, Cummings KJ, Levine GJ, Gregory CA, Smith R 3rd, et al. Repeated intra-articular injection of allogeneic mesenchymal stem cells causes an adverse response compared to autologous cells in the equine model. Stem Cell Res Ther. 2017;8:42.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Department of Veterinary Physiology and PharmacologyTexas A&M UniversityCollege StationUSA
  2. 2.Department of Large Animal Clinical SciencesTexas A&M UniversityCollege StationUSA
  3. 3.Veterinary Genetics LaboratoryUniversity of CaliforniaDavisUSA
  4. 4.Texas A&M Institute for Genome Sciences and SocietyTexas A&M UniversityCollege StationUSA

Personalised recommendations