Nanoencapsulated capsaicin changes migration behavior and morphology of madin darby canine kidney cell monolayers

We have developed a drug delivery nanosystem based on chitosan and capsaicin. Both substances have a wide range of biological activities. We investigated the nanosystem’s influence on migration and morphology of Madin Darby canine kidney (MDCK-C7) epithelial cells in comparison to the capsaicin-free nanoformulation, free capsaicin, and control cells. For minimally-invasive quantification of cell migration, we applied label-free digital holographic microscopy (DHM) and single-cell tracking. Moreover, quantitative DHM phase images were used as novel stain-free assay to quantify the temporal course of global cellular morphology changes in confluent cell layers. Cytoskeleton alterations and tight junction protein redistributions were complementary analyzed by fluorescence microscopy. Calcium influx measurements were conducted to characterize the influence of the nanoformulations and capsaicin on ion channel activities. We found that both, capsaicin-loaded and unloaded chitosan nanocapsules, and also free capsaicin, have a significant impact on directed cell migration and cellular motility. Increase of velocity and directionality of cell migration correlates with changes in the cell layer surface roughness, tight junction integrity and cytoskeleton alterations. Calcium influx into cells occurred only after nanoformulation treatment but not upon addition of free capsaicin. Our results pave the way for further studies on the biological significance of these findings and potential biomedical applications, e.g. as drug and gene carriers.

[1]  Per Artursson,et al.  Chitosans as Absorption Enhancers for Poorly Absorbable Drugs. 1: Influence of Molecular Weight and Degree of Acetylation on Drug Transport Across Human Intestinal Epithelial (Caco-2) Cells , 1996, Pharmaceutical Research.

[2]  H. Isoda,et al.  Cofilin mediates tight-junction opening by redistributing actin and tight-junction proteins. , 2008, Biochemical and biophysical research communications.

[3]  H. Junginger,et al.  Chitosans for enhanced delivery of therapeutic peptides across intestinal epithelia: in vitro evaluation in Caco-2 cell monolayers , 1997 .

[4]  T. Yen,et al.  Opening of epithelial tight junctions and enhancement of paracellular permeation by chitosan: microscopic, ultrastructural, and computed-tomographic observations. , 2012, Molecular pharmaceutics.

[5]  M. de la Fuente,et al.  Novel hyaluronan-based nanocarriers for transmucosal delivery of macromolecules. , 2008, Macromolecular bioscience.

[6]  Michael S. Goldberg,et al.  Nanostructured materials for applications in drug delivery and tissue engineering , 2007, Journal of biomaterials science. Polymer edition.

[7]  V. Gupta,et al.  Naga chilli: a potential source of capsaicinoids with broad-spectrum ethnopharmacological applications. , 2010, Journal of ethnopharmacology.

[8]  M. J. Santander-Ortega,et al.  Chitosan-based nanocapsules: physical characterization, stability in biological media and capsaicin encapsulation , 2012, Colloid and Polymer Science.

[9]  F. Goycoolea,et al.  In Vitro and Sensory Evaluation of Capsaicin-Loaded Nanoformulations , 2015, PloS one.

[10]  M. Alonso,et al.  Transmucosal macromolecular drug delivery. , 2005, Journal of controlled release : official journal of the Controlled Release Society.

[11]  J. Nah,et al.  Influence of molecular weight on oral absorption of water soluble chitosans. , 2005, Journal of controlled release : official journal of the Controlled Release Society.

[12]  H. Isoda,et al.  Capsaicin induces cofilin dephosphorylation in human intestinal cells: the triggering role of cofilin in tight-junction signaling. , 2007, Biochemical and biophysical research communications.

[13]  D. Ingber,et al.  Ultra-rapid activation of TRPV4 ion channels by mechanical forces applied to cell surface beta1 integrins. , 2010, Integrative biology : quantitative biosciences from nano to macro.

[14]  Figen Korel,et al.  Ground red peppers: capsaicinoids content, Scoville scores, and discrimination by an electronic nose. , 2002, Journal of agricultural and food chemistry.

[15]  B. Kemper,et al.  Facing glycosphingolipid–Shiga toxin interaction: dire straits for endothelial cells of the human vasculature , 2012, Cellular and Molecular Life Sciences.

[16]  A. Schwab,et al.  Role of ion channels and transporters in cell migration. , 2012, Physiological reviews.

[17]  H. Isoda,et al.  Capsaicin-enhanced Ribosomal Protein P2 Expression in Human Intestinal Caco-2 Cells , 2005, Cytotechnology.

[18]  C. Gorzelanny,et al.  Chitosan encapsulation modulates the effect of capsaicin on the tight junctions of MDCK cells , 2015, Scientific Reports.

[19]  B. Isakson,et al.  The connexin 43/ZO-1 complex regulates cerebral endothelial F-actin architecture and migration. , 2015, American journal of physiology. Cell physiology.

[20]  Á. González-Fernández,et al.  A Polymer/Oil Based Nanovaccine as a Single-Dose Immunization Approach , 2013, PloS one.

[21]  Keiji Numata,et al.  Biopolymer-Based Nanoparticles for Drug/Gene Delivery and Tissue Engineering , 2013, International journal of molecular sciences.

[22]  B. Bussolati,et al.  TRPV4 mediates tumor-derived endothelial cell migration via arachidonic acid-activated actin remodeling , 2012, Oncogene.

[23]  P. Friedl,et al.  Collective cell migration: guidance principles and hierarchies. , 2015, Trends in cell biology.

[24]  B. Nilius,et al.  A novel function of capsaicin-sensitive TRPV1 channels: involvement in cell migration. , 2007, Cell calcium.

[25]  M. Alonso,et al.  Chitosan nanocapsules: a new carrier for nasal peptide delivery , 2006 .

[26]  T. Kissel,et al.  Controlled pulmonary drug and gene delivery using polymeric nano-carriers. , 2012, Journal of controlled release : official journal of the Controlled Release Society.

[27]  M. Thanou,et al.  Biodegradation, biodistribution and toxicity of chitosan. , 2010, Advanced drug delivery reviews.

[28]  H. Isoda,et al.  Capsaicin induced the upregulation of transcriptional and translational expression of glycolytic enzymes related to energy metabolism in human intestinal epithelial cells. , 2009, Journal of agricultural and food chemistry.

[29]  Desirée Teijeiro-Osorio,et al.  New generation of hybrid poly/oligosaccharide nanoparticles as carriers for the nasal delivery of macromolecules. , 2009, Biomacromolecules.

[30]  E. Cuche,et al.  Digital holographic microscopy: a noninvasive contrast imaging technique allowing quantitative visualization of living cells with subwavelength axial accuracy. , 2005, Optics letters.

[31]  Myung K. Kim.  Principles and techniques of digital holographic microscopy , 2010 .

[32]  Á. González-Fernández,et al.  Co-delivery of viral proteins and a TLR7 agonist from polysaccharide nanocapsules: a needle-free vaccination strategy. , 2013, Journal of controlled release : official journal of the Controlled Release Society.

[33]  M. Tominaga,et al.  Effects of capsaicin on human intestinal cell line Caco-2 , 2001, Cytotechnology.

[34]  F. Zernike.  How I discovered phase contrast. , 1955, Science.

[35]  M. Schrader,et al.  Activation of Transient Receptor Potential Vanilloid Type-1 Channel Prevents Adipogenesis and Obesity , 2007, Circulation Research.

[36]  H. Isoda,et al.  Analysis of the mechanism of the tight-junctional permeability increase by capsaicin treatment on the intestinal Caco-2 cells , 2002, Cytotechnology.

[37]  Daniel Carl,et al.  Investigation of living pancreas tumor cells by digital holographic microscopy. , 2006, Journal of biomedical optics.

[38]  M. Alonso,et al.  Chitosan nanocapsules as carriers for oral peptide delivery: effect of chitosan molecular weight and type of salt on the in vitro behaviour and in vivo effectiveness. , 2006, Journal of nanoscience and nanotechnology.

[39]  Daniel Carl,et al.  Parameter-optimized digital holographic microscope for high-resolution living-cell analysis. , 2004, Applied optics.

[40]  Y. Zambito,et al.  Effect of different chitosan derivatives on in vitro scratch wound assay: a comparative study. , 2015, International journal of biological macromolecules.

[41]  Joseph M. DeSimone,et al.  Strategies in the design of nanoparticles for therapeutic applications , 2010, Nature Reviews Drug Discovery.

[42]  R. Duncan.  The dawning era of polymer therapeutics , 2003, Nature Reviews Drug Discovery.

[43]  M. Alonso,et al.  Chitosan-alginate blended nanoparticles as carriers for the transmucosal delivery of macromolecules. , 2009, Biomacromolecules.

[44]  A. Hensel,et al.  Structure of chitosan determines its interactions with mucin. , 2014, Biomacromolecules.

[45]  Hsing-Wen Sung,et al.  Mechanism and consequence of chitosan-mediated reversible epithelial tight junction opening. , 2011, Biomaterials.

[46]  Driton Vllasaliu,et al.  Tight junction modulation by chitosan nanoparticles: comparison with chitosan solution. , 2010, International journal of pharmaceutics.

[47]  M. Hatta,et al.  Integrin α6β4 and TRPV1 channel coordinately regulate directional keratinocyte migration. , 2015, Biochemical and biophysical research communications.

[48]  Eleonore Fröhlich,et al.  The role of surface charge in cellular uptake and cytotoxicity of medical nanoparticles , 2012, International journal of nanomedicine.

[49]  Lihong Liu,et al.  Modern methods for delivery of drugs across the blood-brain barrier. , 2012, Advanced drug delivery reviews.

[50]  M. Alonso,et al.  Efficacy and Mechanism of Action of Chitosan Nanocapsules for Oral Peptide Delivery , 2006, Pharmaceutical Research.

[51]  P. Artursson,et al.  Chitosans as Absorption Enhancers for Poorly Absorbable Drugs 2: Mechanism of Absorption Enhancement , 1997, Pharmaceutical Research.

[52]  Björn Kemper,et al.  Virulence from vesicles: Novel mechanisms of host cell injury by Escherichia coli O104:H4 outbreak strain , 2015, Scientific Reports.

[53]  M. V. Lozano,et al.  Highly efficient system to deliver taxanes into tumor cells: docetaxel-loaded chitosan oligomer colloidal carriers. , 2008, Biomacromolecules.

[54]  M. Alonso,et al.  Evaluation of cationic polymer-coated nanocapsules as ocular drug carriers , 1997 .

[55]  G. Bally,et al.  Digital Holographic Microscopy , 2007, PhotonicsViews.

[56]  D. Julius,et al.  The capsaicin receptor: a heat-activated ion channel in the pain pathway , 1997, Nature.

[57]  Lisbeth Illum,et al.  Effect of Chitosan on the Permeability of Monolayers of Intestinal Epithelial Cells (Caco-2) , 1994, Pharmaceutical Research.

[58]  H. Isoda,et al.  The Reversible Increase in Tight Junction Permeability Induced by Capsaicin Is Mediated via Cofilin-Actin Cytoskeletal Dynamics and Decreased Level of Occludin , 2013, PloS one.

[59]  M. Alonso,et al.  Chitosan nanoparticles: a new vehicle for the improvement of the delivery of drugs to the ocular surface. Application to cyclosporin A. , 2001, International journal of pharmaceutics.

[60]  Gaurav Kumar Jain,et al.  Metallic nanoparticles: technology overview & drug delivery applications in oncology , 2010, Expert opinion on drug delivery.

[61]  V. Dodane,et al.  Effect of chitosan on epithelial permeability and structure. , 1999, International journal of pharmaceutics.

[62]  B. He,et al.  Carboxymethylated chitosan stimulates proliferation of Schwann cells in vitro via the activation of the ERK and Akt signaling pathways. , 2011, European journal of pharmacology.

[63]  C. Remuñán-López,et al.  Enhancement of Nasal Absorption of Insulin Using Chitosan Nanoparticles , 1999, Pharmaceutical Research.