Pharmaceutical Research

, 36:50 | Cite as

Evaluation of the Immunomodulatory Effects of All-Trans Retinoic Acid Solid Lipid Nanoparticles and Human Mesenchymal Stem Cells in an A549 Epithelial Cell Line Model

  • Christina M. Payne
  • Liam P. Burke
  • Brenton Cavanagh
  • Daniel O’Toole
  • Sally-Ann Cryan
  • Helena M. KellyEmail author
Research Paper



To investigate two potential strategies aimed at targeting the inflammatory pathogenesis of COPD: a small molecule, all trans retinoic acid (atRA) and human mesenchymal stem cells (hMSCs).


atRA was formulated into solid lipid nanoparticles (SLNs) via the emulsification-ultrasonication method, and these SLNs were characterised physicochemically. Assessment of the immunomodulatory effects of atRA-SLNs on A549 cells in vitro was determined using ELISA. hMSCs were suspended in a previously developed methylcellulose, collagen and beta-glycerophosphate hydrogel prior to investigating their immunomodulatory effects in vitro.


SLNs provided significant encapsulation of atRA and also sustained its release over 72 h. A549 cells were viable following the addition of atRA SLNs and showed a reduction in IL-6 and IL-8 levels. A549 cells also remained viable following addition of the hMSC/hydrogel formulation – however, this formulation resulted in increased levels of IL-6 and IL-8, indicating a potentially pro-inflammatory effect.


Both atRA SLNs and hMSCs show potential for modulating the environment in inflammatory disease, though through different mechanisms and leading to different outcomes – despite both being explored as strategies for use in inflammatory disease. atRA shows promise by acting in a directly anti-inflammatory manner, whereas further research into the exact mechanisms and behaviours of hMSCs in inflammatory diseases is required.


all trans retinoic acid chronic obstructive pulmonary disease human mesenchymal stem cells immunomodulatory solid lipid nanoparticles 



Analysis of Variance


All trans Retinoic Acid


Alveolar type II cells


Bronchoalveolar lavage




Cell counting kit 8


Chronic Obstructive Pulmonary Disease


Enzyme linked immunosorbent assay


High dose


Human mesenchymal stem cells




Low dose




Standard deviation


Standard error of mean


Solid lipid nanoparticle


Transmission electron microscopy


Tumour necrosis factor


Nanoparticle average size


Zeta potential



  1. 1.
    Hind M, Maden M. Is a regenerative approach viable for the treatment of COPD? Br J Pharmacol. 2011;163(1):106–15.PubMedPubMedCentralCrossRefGoogle Scholar
  2. 2.
    O’Donnell R, Breen D, Wilson S, Djukanovic R. Inflammatory cells in the airways in COPD. Thorax. 2006;61(5):448–54 Available from: Scholar
  3. 3.
    MacNee, W. ABC of chronic obstructive pulmonary disease: Pathology, pathogenesis, and pathophysiology. Br Med J. 2006 [cited 2017 Jul 7];332. Available from:
  4. 4.
    Chronic obstructive pulmonary disease: Management of chronic obstructive pulmonary disease in adults in primary and secondary care [Internet]. 2010 [cited 2015 Jun 24]. Available from:
  5. 5.
    Kam RKT, Deng Y, Chen Y, Zhao H. Retinoic acid synthesis and functions in early embryonic development. Cell Biosci. 2012;2(1):11 Available from: Scholar
  6. 6.
    Massaro GD, Massaro D. Postnatal treatment with retinoic acid increases the number of pulmonary alveoli in rats. Am J Physiol Lung Cell Mol Physiol. 1996;270(2):L305–10 Available from: Scholar
  7. 7.
    Massaro GD, Massaro D, Chan WY, Clerch LB, Ghyselinck N, Chambon P, et al. Retinoic acid receptor-beta: an endogenous inhibitor of the perinatal formation of pulmonary alveoli. Physiol Genomics. 2000;4(1):51–7 Available from: Scholar
  8. 8.
    Nozaki Y, Yamagata T, Sugiyama M, Ikoma S, Kinoshita K, Funauchi M. Anti-inflammatory effect of all-trans-retinoic acid in inflammatory arthritis. Clin Immunol. 2006;119(3):272–9.PubMedCrossRefGoogle Scholar
  9. 9.
    Wolf JE. Potential anti-inflammatory effects of topical retinoids and retinoid analogues. Adv Ther. 2002;19(3):109–18 Available from: Scholar
  10. 10.
    Balato A, Schiattarella M, Lembo S, Mattii M, Prevete N, Balato N, et al. Interleukin-1 family members are enhanced in psoriasis and suppressed by vitamin D and retinoic acid. Arch Dermatol Res. 2013;305(3):255–62 Available from: Scholar
  11. 11.
    March TH, Cossey PY, Esparza DC, Dix KJ, McDonald JD, Bowen LE. Inhalation administration of all-trans-retinoic acid for treatment of elastase-induced pulmonary emphysema in Fischer 344 rats. Exp Lung Res. 2004;30(5):383–404.PubMedCrossRefGoogle Scholar
  12. 12.
    Mao JT, Goldin JG, Dermand J, Ibrahim G, Brown MS, Emerick A, et al. A pilot study of all- trans -Retinoic acid for the treatment of human emphysema. Crit Care Med. 2002;165:718–23.CrossRefGoogle Scholar
  13. 13.
    Roth MD, Connett JE, D’Armiento JM, Foronjy RF, Friedman PJ, Goldin JG, et al. Feasibility of retinoids for the treatment of emphysema study. Chest. 2006;130(5):1334–45.PubMedCrossRefGoogle Scholar
  14. 14.
    Kubo H. Concise review: clinical prospects for treating chronic obstructive pulmonary disease with regenerative approaches. Stem Cells Transl Med. 2012;1(8):627–31 Available from: Scholar
  15. 15.
    Krause DS, Theise ND, Collector MI, Henegariu O, Hwang S, Gardner R, et al. Multi-Organ, Multi-Lineage Engraftment by a Single Bone Marrow-Derived Stem Cell. Cell. 2001;105(3):369–77 Available from: Scholar
  16. 16.
    Kotton DN, Ma BY, Cardoso WV, Sanderson EA, Summer RS, Williams MC, et al. Bone marrow-derived cells as progenitors of lung alveolar epithelium. Development. 2001;128(24):5181–8 Available from: Scholar
  17. 17.
    Gupta N, Su X, Popov B, Lee JW, Serikov V, Matthay MA. Intrapulmonary delivery of bone marrow-derived mesenchymal stem cells improves survival and attenuates endotoxin-induced acute lung injury in mice. J Immunol. 2007;179(3):1855–63 Available from: Scholar
  18. 18.
    Yuhgetsu H, Ohno Y, Funaguchi N, Asai T, Sawada M, Takemura G, et al. Beneficial effects of autologous bone marrow mononuclear cell transplantation against elastase-induced emphysema in rabbits. Exp Lung Res. 2006;32:413–26 Available from: Scholar
  19. 19.
    Shigemura N, Okumura M, Mizuno S, Imanishi Y, Matsuyama A, Shiono H, et al. Lung Tissue Engineering Technique with Adipose Stromal Cells Improves Surgical Outcome for Pulmonary Emphysema. Am J Respir Crit Care Med. 2006;174(11):1199–205 Available from: Scholar
  20. 20.
    Shigemura N, Okumura M, Mizuno S, Imanishi Y, Nakamura T, Sawa Y. Autologous transplantation of adipose tissue-derived stromal cells ameliorates pulmonary emphysema. Am J Transplant. 2006;6(11):2592–600 Available from: Scholar
  21. 21.
    Weiss DJ, Casaburi R, Flannery R, LeRoux-Williams M, Tashkin DP. A Placebo-Controlled, Randomized Trial of Mesenchymal Stem Cells in COPD. Chest. 2013;143(6):1590 Available from: Scholar
  22. 22.
    Jenning V, Gohla SH. Encapsulation of retinoids in solid lipid nanoparticles (SLN). J Microencapsul. 2001;18(2):149–58 Available from: Scholar
  23. 23.
    Crowe DL, Kim R, RAS C. Retinoic acid differentially regulates cancer cell proliferation via dose-dependent modulation of the mitogen-activated protein kinase pathway. Mol cancer Res. 2003;1(7):532–40 Available from: Scholar
  24. 24.
    Müller RH, Maassen S, Weyhers H, Mehnert W. Phagocytic uptake and cytotoxicity of solid lipid nanoparticles (SLN) sterically stabilized with poloxamine 908 and poloxamer 407. J Drug Target. 1996;4(3):161–70 Available from: Scholar
  25. 25.
    Payne C, Dolan EB, O’Sullivan J, Cryan S-A, Kelly HM. A methylcellulose and collagen based temperature responsive hydrogel promotes encapsulated stem cell viability and proliferation in vitro. Drug Deliv Transl Res [Internet]. 2016;7(1):1–15 Available from: Scholar
  26. 26.
    Das S, Ng WK, Kanaujia P, Kim S, Tan RBH. Formulation design, preparation and physicochemical characterizations of solid lipid nanoparticles containing a hydrophobic drug: effects of process variables. Colloids surfaces B biointerfaces. 2011;88(1):483–489. Available from
  27. 27.
    Cirpanli Y, Unlü N, Caliş S. Hincal a A. Formulation and in-vitro characterization of retinoic acid loaded poly (lactic-co-glycolic acid) microspheres. J Microencapsul. 2005;22(8):877–89.PubMedCrossRefGoogle Scholar
  28. 28.
    Almouazen E, Bourgeois S, Boussaïd A, Valot P, Malleval C, Fessi H, et al. Development of a nanoparticle-based system for the delivery of retinoic acid into macrophages. Int J Pharm. 2012;430(1–2):207–15.PubMedCrossRefGoogle Scholar
  29. 29.
    O’Gorman MT, Jatoi NA, Lane SJ, Mahon BP. IL-1b and TNF-a induce increased expression of CCL28 by airway epithelial cells via an NFkB-dependent pathway. Cell Immunol. 2005;238:87–96 Available from: Scholar
  30. 30.
    Westesen K, Bunjes H, Koch MH. Physicochemical characterization of lipid nanoparticles and evaluation of their drug loading capacity and sustained release potential. J Control Release. 1997;48(2–3):223–36 Available from: Scholar
  31. 31.
    Mitragotri S, Burke PA, Langer R. Overcoming the challenges in administering biopharmaceuticals: formulation and delivery strategies. Nat Rev Drug Discov. 2014;13(9):655–72 Available from: Scholar
  32. 32.
    Liu J, Gong T, Fu H, Wang C, Wang X, Chen Q, et al. Solid lipid nanoparticles for pulmonary delivery of insulin. Int J Pharm. 2008;356(1–2):333–44.PubMedCrossRefGoogle Scholar
  33. 33.
    Weber S, Zimmer A. Pardeike J. Solid lipid nanoparticles (SLN) and nanostructured lipid carriers (NLC) for pulmonary application: A review of the state of the art. Eur J pharm biopharm. 2014;86(1):7–22. Available from:
  34. 34.
    Makled S, Nafee N, Boraie N. Nebulized solid lipid nanoparticles for the potential treatment of pulmonary hypertension via targeted delivery of phosphodiesterase-5-inhibitor. Int J pharm. 2017;517(1–2):312–321. Available from:
  35. 35.
    Hu L, Tang X, Cui F. Solid lipid nanoparticles (SLNs) to improve oral bioavailability of poorly soluble drugs. J Pharm Pharmacol. 2004;56(12):1527–35.PubMedCrossRefGoogle Scholar
  36. 36.
    Lee C-M, Jeong H-J, Park J-W, Kim J, Lee K-Y. Temperature-induced release of all-trans-retinoic acid loaded in solid lipid nanoparticles for topical delivery. Macromol Res. 2008;16(8):682–5 Available from: Scholar
  37. 37.
    Aburahma MH, Badr-Eldin SM. Compritol 888 ATO: a multifunctional lipid excipient in drug delivery systems and nanopharmaceuticals. Expert Opin Drug Deliv. 2014;11(12):1865–1883. Available from:
  38. 38.
    Cai S, Yang Q, Bagby TR. Lymphatic drug delivery using engineered liposomes and solid lipid nanoparticles. Adv Drug Deliv Rev. 2011;63(10):901–8 Available from: Scholar
  39. 39.
    Müller RH, Radtke M, Wissing SA. Solid lipid nanoparticles (SLN) and nanostructured lipid carriers (NLC) in cosmetic and dermatological preparations. Adv Drug Deliv Rev. 2002;54:S131–55 Available from: Scholar
  40. 40.
    Vivek K, Reddy H, RSR M. Investigations of the effect of the lipid matrix on drug entrapment, in vitro release, and physical stability of olanzapine-loaded solid lipid nanoparticles. AAPS PharmSciTech. 2007;8(4):16–24 Available from: Scholar
  41. 41.
    Larsson M, Hill A, Duffy J. Suspension Stability: Why Particle Size, Zeta Potential and rheology are Important. Annu Trans Nord Rheol Soc. 2012;20:209–14 Available from: Scholar
  42. 42.
    Foster KA, Oster CG, Mayer MM, Avery ML, Audus KL. Characterization of the A549 Cell Line as a Type II Pulmonary Epithelial Cell Model for Drug Metabolism. Exp Cell Res. 1998;243(2):359–66 Available from: Scholar
  43. 43.
    International Organisation for Standardisation. ISO 10993-5: Biological evaluation of medical devices. Part 5: Tests for in vitro cytotoxicity [Internet]. 2009 [cited 2017 Jul 5]. p. 34. Available from:
  44. 44.
    Crestani B, Cornillet P, Dehoux M, Rolland C, Guenounou M, Aubier M. Alveolar type II epithelial cells produce interleukin-6 in vitro and in vivo. Regulation by alveolar macrophage secretory products. J Clin Invest. 1994;94(2):731–40 Available from: Scholar
  45. 45.
    Chung KF. Cytokines in chronic obstructive pulmonary disease. Eur Respir J Suppl. 2001;34:50s–9s Available from: Scholar
  46. 46.
    Kips JC, Tavernier J, Pauwels RA. Tumor Necrosis Factor Causes Bronchial Hyperresponsiveness in Rats. Am Rev Respir Dis. 1992;145(2_pt_1):332–6 Available from: Scholar
  47. 47.
    Keatings VM, Collins PD, Scott DM, Barnes PJ. Differences in interleukin-8 and tumor necrosis factor-alpha in induced sputum from patients with chronic obstructive pulmonary disease or asthma. Am J Respir Crit Care Med. 1996;153(2):530–4 Available from: Scholar
  48. 48.
    Tschopp J, Burns K, Clatworthy J, Martin L, Martinon F, Plumpton C, et al. TOLLIP, a new component of the IL-1RI pathway, links IRAK to the IL-1 receptor. Nat Cell Biol. 2000;2(6):346–51 Available from: Scholar
  49. 49.
    Chen G, Goeddel DV. TNF-R1 Signaling: A Beautiful Pathway. Science. 2002;296(5573):1634–5 Available from: Scholar
  50. 50.
    Cooke EL, Uings IJ, Xia CL, Woo P, Ray KP. Functional analysis of the interleukin-1-receptor-associated kinase (IRAK-1) in interleukin-1 beta-stimulated nuclear factor kappa B (NF-kappa B) pathway activation: IRAK-1 associates with the NF-kappa B essential modulator (NEMO) upon receptor stimulatio. Biochem J. 2001;359(Pt 2):403–10 Available from: Scholar
  51. 51.
    Hageman GJ, Larik I, Pennings H-J, Haenen GRMM, Wouters EFM, Aalt B. Systemic poly(ADP-ribose) polymerase-1 activation, chronic inflammation, and oxidative stress in COPD patients. Free Radic Biol Med. 2003;35(2):140–8 Available from: Scholar
  52. 52.
    Eid AA, Ionescu AA, Nixon LS, Lewis-Jenkins V, Matthews SB, Griffiths TL, et al. Inflammatory Response and Body Composition in Chronic Obstructive Pulmonary Disease. Am J Respir Crit Care Med. 2001;164(8):1414–8 Available from: Scholar
  53. 53.
    Bhowmik A, Seemungal TA, Sapsford RJ, Wedzicha JA. Relation of sputum inflammatory markers to symptoms and lung function changes in COPD exacerbations. Thorax. 2000;55(2):114–20 Available from: Scholar
  54. 54.
    Wedzicha JA, Seemungal TAR, MacCallum PK, Paul EA, Donaldson GC, Bhowmik A, et al. Acute Exacerbations of Chronic Obstructive Pulmonary Disease Are Accompanied by Elevations of Plasma Fibrinogen and Serum IL-6 Levels. Thromb Haemost. 2000;84(2):210–5 Available from: Scholar
  55. 55.
    Pesci A, Balbi B, Majori M, Cacciani G, Bertacco S, Alciato P, et al. Inflammatory cells and mediators in bronchial lavage of patients with chronic obstructive pulmonary disease. Eur Respir J. 1998;12(2) Available from:
  56. 56.
    Criner GJ, Pinto-Plata V, Strange C, Dransfield M, Gotfried M, Leeds W, et al. Biologic lung volume reduction in advanced upper lobe emphysema: phase 2 results. Am J Respir Crit Care Med. 2009;179(9):791–8 Available from: Scholar
  57. 57.
    Dey A, Wong ET, Cheok CF, Tergaonkar V, Lane DP. R-Roscovitine simultaneously targets both the p53 and NF-κB pathways and causes potentiation of apoptosis: implications in cancer therapy. Cell Death Differ. 2008;15(2):263–73 Available from: Scholar
  58. 58.
    Kyurkchiev D, Bochev I, Ivanova-Todorova E, Mourdjeva M, Oreshkova T, Belemezova K, et al. Secretion of immunoregulatory cytokines by mesenchymal stem cells. World J Stem Cells. 2014;6(5):552–70 Available from: Scholar
  59. 59.
    Bernardo ME, Fibbe WE. Mesenchymal Stromal Cells: Sensors and Switchers of Inflammation. Cell Stem Cell. 2013;13(4):392–402 Available from: Scholar
  60. 60.
    Scheller J, Chalaris A, Schmidt-Arras D, Rose-John S. The pro- and anti-inflammatory properties of the cytokine interleukin-6. Biochim Biophys Acta - Mol Cell Res. 2011;1813(5):878–88 Available from: Scholar
  61. 61.
    McLoughlin RM, Jenkins BJ, Grail D, Williams AS, Fielding CA, Parker CR, et al. IL-6 trans-signaling via STAT3 directs T cell infiltration in acute inflammation. Proc Natl Acad Sci U S A. 2005;102(27):9589–94 Available from: Scholar
  62. 62.
    Ehrhardt C, Kim K-J, Lehr C-M. Isolation and Culture of Human Alveolar Epithelial Cells. In: Human Cell Culture Protocols [Internet]. New Jersey: Humana Press; 2005 [cited 2017 Sep 10]. p. 207–16. Available from:
  63. 63.
    Cryan SA, Sivadas N, Garcia-Contreras L. In vivo animal models for drug delivery across the lung mucosal barrier. Adv Drug Deliv Rev. 2007;59(11):1133–51.PubMedCrossRefGoogle Scholar
  64. 64.
    Roche ET, Hastings CL, Lewin SA, Shvartsman DE, Brudno Y, Vasilyev NV, et al. Comparison of biomaterial delivery vehicles for improving acute retention of stem cells in the infarcted heart. Biomaterials. 2014;35(25):6850–8 Available from: Scholar
  65. 65.
    Groneberg DA, Chung KF. Models of chronic obstructive pulmonary disease. Respir Res. 2004;5(1):18 Available from: Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.School of PharmacyRoyal College of Surgeons in IrelandDublin 2Ireland
  2. 2.Tissue Engineering Research Group, Department of AnatomyRoyal College of Surgeons in IrelandDublin 2Ireland
  3. 3.Discipline of Bacteriology, School of MedicineNational University of Ireland GalwayGalwayIreland
  4. 4.Cellular and Molecular Imaging CoreRoyal College of Surgeons in IrelandDublin 2Ireland
  5. 5.Discipline of Anaesthesia, School of MedicineNational University of Ireland GalwayGalwayIreland
  6. 6.Centre for Research in Medical Devices (CÚRAM)National University of Ireland GalwayGalwayIreland
  7. 7.Trinity Centre for BioengineeringTrinity College DublinDublin 2Ireland

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