Advertisement

Two-photon microscopy analysis of leukocyte trafficking and motility

Abstract

During the last several years, live tissue imaging, in particular using two-photon laser microscopy, has advanced our understanding of leukocyte trafficking mechanisms. Studies using this technique are revealing distinct molecular requirements for leukocyte migration in different tissue environments. Also emerging from the studies are the ingenious infrastructures for leukocyte trafficking, which are produced by stromal cells. This review summarizes the recent imaging studies that provided novel mechanistic insights into in vivo leukocyte migration essential for immunosurveillance.

This is a preview of subscription content, log in to check access.

We’re sorry, something doesn't seem to be working properly.

Please try refreshing the page. If that doesn't work, please contact support so we can address the problem.

Fig. 1

References

  1. 1.

    Cyster JG (2005) Chemokines, sphingosine-1-phosphate, and cell migration in secondary lymphoid organs. Annu Rev Immunol 23:127–159

  2. 2.

    Luster AD, Alon R, von Andrian UH (2005) Immune cell migration in inflammation: present and future therapeutic targets. Nat Immunol 6:1182–1190

  3. 3.

    Koizumi K, Hojo S, Akashi T, Yasumoto K, Saiki I (2007) Chemokine receptors in cancer metastasis and cancer cell-derived chemokines in host immune response. Cancer Sci 98:1652–1658

  4. 4.

    Brinkmann V (2007) Sphingosine 1-phosphate receptors in health and disease: mechanistic insights from gene deletion studies and reverse pharmacology. Pharmacol Ther 115:84–105

  5. 5.

    Germain RN, Miller MJ, Dustin ML, Nussenzweig MC (2006) Dynamic imaging of the immune system: progress, pitfalls and promise. Nat Rev Immunol 6:497–507

  6. 6.

    Cahalan MD, Parker I (2008) Choreography of cell motility and interaction dynamics imaged by two-photon microscopy in lymphoid organs. Annu Rev Immunol 26:585–626

  7. 7.

    Miller MJ, Wei SH, Parker I, Cahalan MD (2002) Two-photon imaging of lymphocyte motility and antigen response in intact lymph node. Science 296:1869–1873

  8. 8.

    Miller MJ, Wei SH, Cahalan MD, Parker I (2003) Autonomous T cell trafficking examined in vivo with intravital two-photon microscopy. Proc Natl Acad Sci USA 100:2604–2609

  9. 9.

    Phan TG, Grigorova I, Okada T, Cyster JG (2007) Subcapsular encounter and complement-dependent transport of immune complexes by lymph node B cells. Nat Immunol 8:992–1000

  10. 10.

    Phan TG, Green JA, Gray EE, Xu Y, Cyster JG (2009) Immune complex relay by subcapsular sinus macrophages and noncognate B cells drives antibody affinity maturation. Nat Immunol 10:786–793

  11. 11.

    Mempel TR, Henrickson SE, Von Andrian UH (2004) T-cell priming by dendritic cells in lymph nodes occurs in three distinct phases. Nature 427:154–159

  12. 12.

    Bajenoff M, Egen JG, Koo LY, Laugier JP, Brau F, Glaichenhaus N, Germain RN (2006) Stromal cell networks regulate lymphocyte entry, migration, and territoriality in lymph nodes. Immunity 25:989–1001

  13. 13.

    Bajenoff M, Glaichenhaus N, Germain RN (2008) Fibroblastic reticular cells guide T lymphocyte entry into and migration within the splenic T cell zone. J Immunol 181:3947–3954

  14. 14.

    Wilson EH, Harris TH, Mrass P, John B, Tait ED, Wu GF, Pepper M, Wherry EJ, Dzierzinski F, Roos D, Haydon PG, Laufer TM, Weninger W, Hunter CA (2009) Behavior of parasite-specific effector CD8+ T cells in the brain and visualization of a kinesis-associated system of reticular fibers. Immunity 30:300–311

  15. 15.

    Lindquist RL, Shakhar G, Dudziak D, Wardemann H, Eisenreich T, Dustin ML, Nussenzweig MC (2004) Visualizing dendritic cell networks in vivo. Nat Immunol 5:1243–1250

  16. 16.

    Garrod KR, Wei SH, Parker I, Cahalan MD (2007) Natural killer cells actively patrol peripheral lymph nodes forming stable conjugates to eliminate MHC-mismatched targets. Proc Natl Acad Sci USA 104:12081–12086

  17. 17.

    Beuneu H, Deguine J, Breart B, Mandelboim O, Di Santo JP, Bousso P (2009) Dynamic behavior of NK cells during activation in lymph nodes. Blood 114:3227–3234

  18. 18.

    Mueller SN, Germain RN (2009) Stromal cell contributions to the homeostasis and functionality of the immune system. Nat Rev Immunol 9:618–629

  19. 19.

    Kaldjian EP, Gretz JE, Anderson AO, Shi Y, Shaw S (2001) Spatial and molecular organization of lymph node T cell cortex: a labyrinthine cavity bounded by an epithelium-like monolayer of fibroblastic reticular cells anchored to basement membrane-like extracellular matrix. Int Immunol 13:1243–1253

  20. 20.

    Matheu MP, Beeton C, Garcia A, Chi V, Rangaraju S, Safrina O, Monaghan K, Uemura MI, Li D, Pal S, de la Maza LM, Monuki E, Flugel A, Pennington MW, Parker I, Chandy KG, Cahalan MD (2008) Imaging of effector memory T cells during a delayed-type hypersensitivity reaction and suppression by Kv1.3 channel block. Immunity 29:602–614

  21. 21.

    Mrass P, Takano H, Ng LG, Daxini S, Lasaro MO, Iparraguirre A, Cavanagh LL, von Andrian UH, Ertl HC, Haydon PG, Weninger W (2006) Random migration precedes stable target cell interactions of tumor-infiltrating T cells. J Exp Med 203:2749–2761

  22. 22.

    Friedl P, Weigelin B (2008) Interstitial leukocyte migration and immune function. Nat Immunol 9:960–969

  23. 23.

    Wolf K, Muller R, Borgmann S, Brocker EB, Friedl P (2003) Amoeboid shape change and contact guidance: T-lymphocyte crawling through fibrillar collagen is independent of matrix remodeling by MMPs and other proteases. Blood 102:3262–3269

  24. 24.

    Bartholomaus I, Kawakami N, Odoardi F, Schlager C, Miljkovic D, Ellwart JW, Klinkert WE, Flugel-Koch C, Issekutz TB, Wekerle H, Flugel A (2009) Effector T cell interactions with meningeal vascular structures in nascent autoimmune CNS lesions. Nature 462:94–98

  25. 25.

    Park EJ, Peixoto A, Imai Y, Goodarzi A, Cheng G, Carman CV, von Andrian UH, Shimaoka M (2009) Distinct roles for LFA-1 affinity regulation during T cell adhesion, diapedesis, and interstitial migration in lymph nodes. Blood 115(8):1572–1581

  26. 26.

    Grigorova IL, Schwab SR, Phan TG, Pham TH, Okada T, Cyster JG (2009) Cortical sinus probing, S1P1-dependent entry and flow-based capture of egressing T cells. Nat Immunol 10:58–65

  27. 27.

    Bousso P, Robey E (2003) Dynamics of CD8+ T cell priming by dendritic cells in intact lymph nodes. Nat Immunol 4:579–585

  28. 28.

    Miller MJ, Safrina O, Parker I, Cahalan MD (2004) Imaging the single cell dynamics of CD4+ T cell activation by dendritic cells in lymph nodes. J Exp Med 200:847–856

  29. 29.

    Okada T, Miller MJ, Parker I, Krummel MF, Neighbors M, Hartley SB, O’Garra A, Cahalan MD, Cyster JG (2005) Antigen-engaged B cells undergo chemotaxis toward the T zone and form motile conjugates with helper T cells. PLoS Biol 3:e150

  30. 30.

    Allen CD, Okada T, Cyster JG (2007) Germinal-Center Organization and Cellular Dynamics. Immunity 27:190–202

  31. 31.

    Qi H, Cannons JL, Klauschen F, Schwartzberg PL, Germain RN (2008) SAP-controlled T-B cell interactions underlie germinal centre formation. Nature 455:764–769

  32. 32.

    Lammermann T, Sixt M (2009) Mechanical modes of ‘amoeboid’ cell migration. Curr Opin Cell Biol 21:636–644

  33. 33.

    Link A, Vogt TK, Favre S, Britschgi MR, Acha-Orbea H, Hinz B, Cyster JG, Luther SA (2007) Fibroblastic reticular cells in lymph nodes regulate the homeostasis of naive T cells. Nat Immunol 8:1255–1265

  34. 34.

    Okada T, Ngo VN, Ekland EH, Forster R, Lipp M, Littman DR, Cyster JG (2002) Chemokine requirements for B cell entry to lymph nodes and Peyer’s patches. J Exp Med 196:65–75

  35. 35.

    Ebisuno Y, Tanaka T, Kanemitsu N, Kanda H, Yamaguchi K, Kaisho T, Akira S, Miyasaka M (2003) Cutting edge: the B cell chemokine CXC chemokine ligand 13/B lymphocyte chemoattractant is expressed in the high endothelial venules of lymph nodes and Peyer’s patches and affects B cell trafficking across high endothelial venules. J Immunol 171:1642–1646

  36. 36.

    Sigmundsdottir H, Butcher EC (2008) Environmental cues, dendritic cells and the programming of tissue-selective lymphocyte trafficking. Nat Immunol 9:981–987

  37. 37.

    Allen CD, Ansel KM, Low C, Lesley R, Tamamura H, Fujii N, Cyster JG (2004) Germinal center dark and light zone organization is mediated by CXCR4 and CXCR5. Nat Immunol 5:943–952

  38. 38.

    Woolf E, Grigorova I, Sagiv A, Grabovsky V, Feigelson SW, Shulman Z, Hartmann T, Sixt M, Cyster JG, Alon R (2007) Lymph node chemokines promote sustained T lymphocyte motility without triggering stable integrin adhesiveness in the absence of shear forces. Nat Immunol 8:1076–1085

  39. 39.

    Parish CR (2006) The role of heparan sulphate in inflammation. Nat Rev Immunol 6:633–643

  40. 40.

    Alon R, Ley K (2008) Cells on the run: shear-regulated integrin activation in leukocyte rolling and arrest on endothelial cells. Curr Opin Cell Biol 20:525–532

  41. 41.

    Giannone G, Dubin-Thaler BJ, Rossier O, Cai Y, Chaga O, Jiang G, Beaver W, Dobereiner HG, Freund Y, Borisy G, Sheetz MP (2007) Lamellipodial actin mechanically links myosin activity with adhesion-site formation. Cell 128:561–575

  42. 42.

    Wolf K, Friedl P (2009) Mapping proteolytic cancer cell-extracellular matrix interfaces. Clin Exp Metastasis 26:289–298

  43. 43.

    Friedl P, Entschladen F, Conrad C, Niggemann B, Zanker KS (1998) CD4+ T lymphocytes migrating in three-dimensional collagen lattices lack focal adhesions and utilize beta1 integrin-independent strategies for polarization, interaction with collagen fibers and locomotion. Eur J Immunol 28:2331–2343

  44. 44.

    Kinashi T (2005) Intracellular signalling controlling integrin activation in lymphocytes. Nat Rev Immunol 5:546–559

  45. 45.

    Ebisuno Y, Katagiri K, Katakai T, Ueda Y, Nemoto T, Inada H, Nabekura J, Okada T, Kannagi R, Tanaka T, Miyasaka M, Hogg N, Kinashi T (2009) Rap1 controls lymphocyte adhesion cascades and interstitial migration within lymph nodes in RAPL-dependent and -independent manners. Blood 115(4):804–814

  46. 46.

    Lammermann T, Bader BL, Monkley SJ, Worbs T, Wedlich-Soldner R, Hirsch K, Keller M, Forster R, Critchley DR, Fassler R, Sixt M (2008) Rapid leukocyte migration by integrin-independent flowing and squeezing. Nature 453:51–55

  47. 47.

    Graham DB, Zinselmeyer BH, Mascarenhas F, Delgado R, Miller MJ, Swat W (2009) ITAM signaling by Vav family Rho guanine nucleotide exchange factors regulates interstitial transit rates of neutrophils in vivo. PLoS ONE 4:e4652

  48. 48.

    Mrass P, Kinjyo I, Ng LG, Reiner SL, Pure E, Weninger W (2008) CD44 mediates successful interstitial navigation by killer T cells and enables efficient antitumor immunity. Immunity 29:971–985

  49. 49.

    Boissonnas A, Fetler L, Zeelenberg IS, Hugues S, Amigorena S (2007) In vivo imaging of cytotoxic T cell infiltration and elimination of a solid tumor. J Exp Med 204:345–356

  50. 50.

    Breart B, Lemaitre F, Celli S, Bousso P (2008) Two-photon imaging of intratumoral CD8+ T cell cytotoxic activity during adoptive T cell therapy in mice. J Clin Invest 118:1390–1397

  51. 51.

    Han SB, Moratz C, Huang NN, Kelsall B, Cho H, Shi CS, Schwartz O, Kehrl JH (2005) Rgs1 and Gnai2 regulate the entrance of B lymphocytes into lymph nodes and B cell motility within lymph node follicles. Immunity 22:343–354

  52. 52.

    Okada T, Cyster JG (2007) CC chemokine receptor 7 contributes to Gi-dependent T cell motility in the lymph node. J Immunol 178:2973–2978

  53. 53.

    Asperti-Boursin F, Real E, Bismuth G, Trautmann A, Donnadieu E (2007) CCR7 ligands control basal T cell motility within lymph node slices in a phosphoinositide 3-kinase-independent manner. J Exp Med 204:1167–1179

  54. 54.

    Huang JH, Cardenas-Navia LI, Caldwell CC, Plumb TJ, Radu CG, Rocha PN, Wilder T, Bromberg JS, Cronstein BN, Sitkovsky M, Dewhirst MW, Dustin ML (2007) Requirements for T lymphocyte migration in explanted lymph nodes. J Immunol 178:7747–7755

  55. 55.

    Worbs T, Mempel TR, Bolter J, von Andrian UH, Forster R (2007) CCR7 ligands stimulate the intranodal motility of T lymphocytes in vivo. J Exp Med 204:489–495

  56. 56.

    Halin C, Scimone ML, Bonasio R, Gauguet JM, Mempel TR, Quackenbush E, Proia RL, Mandala S, von Andrian UH (2005) The S1P-analog FTY720 differentially modulates T-cell homing via HEV: T-cell-expressed S1P1 amplifies integrin activation in peripheral lymph nodes but not in Peyer patches. Blood 106:1314–1322

  57. 57.

    Suzuki K, Grigorova I, Phan TG, Kelly LM, Cyster JG (2009) Visualizing B cell capture of cognate antigen from follicular dendritic cells. J Exp Med 206:1485–1493

  58. 58.

    Schwickert TA, Lindquist RL, Shakhar G, Livshits G, Skokos D, Kosco-Vilbois MH, Dustin ML, Nussenzweig MC (2007) In vivo imaging of germinal centres reveals a dynamic open structure. Nature 446:83–87

  59. 59.

    Hauser AE, Junt T, Mempel TR, Sneddon MW, Kleinstein SH, Henrickson SE, von Andrian UH, Shlomchik MJ, Haberman AM (2007) Definition of germinal-center B cell migration in vivo reveals predominant intrazonal circulation patterns. Immunity 26:655–667

  60. 60.

    Okada T, Cyster JG (2006) B cell migration and interactions in the early phase of antibody responses. Curr Opin Immunol 18:278–285

  61. 61.

    Hwang IY, Park C, Harrison K, Kehrl JH (2009) TLR4 signaling augments B lymphocyte migration and overcomes the restriction that limits access to germinal center dark zones. J Exp Med 206:2641–2657

  62. 62.

    Davalos D, Grutzendler J, Yang G, Kim JV, Zuo Y, Jung S, Littman DR, Dustin ML, Gan WB (2005) ATP mediates rapid microglial response to local brain injury in vivo. Nat Neurosci 8:752–758

  63. 63.

    Haynes SE, Hollopeter G, Yang G, Kurpius D, Dailey ME, Gan WB, Julius D (2006) The P2Y12 receptor regulates microglial activation by extracellular nucleotides. Nat Neurosci 9:1512–1519

  64. 64.

    Ishii M, Egen JG, Klauschen F, Meier-Schellersheim M, Saeki Y, Vacher J, Proia RL, Germain RN (2009) Sphingosine-1-phosphate mobilizes osteoclast precursors and regulates bone homeostasis. Nature 458:524–528

  65. 65.

    Benvenuti F, Hugues S, Walmsley M, Ruf S, Fetler L, Popoff M, Tybulewicz VL, Amigorena S (2004) Requirement of Rac1 and Rac2 expression by mature dendritic cells for T cell priming. Science 305:1150–1153

  66. 66.

    Lammermann T, Renkawitz J, Wu X, Hirsch K, Brakebusch C, Sixt M (2009) Cdc42-dependent leading edge coordination is essential for interstitial dendritic cell migration. Blood 113:5703–5710

  67. 67.

    Nombela-Arrieta C, Mempel TR, Soriano SF, Mazo I, Wymann MP, Hirsch E, Martinez AC, Fukui Y, von Andrian UH, Stein JV (2007) A central role for DOCK2 during interstitial lymphocyte motility and sphingosine-1-phosphate-mediated egress. J Exp Med 204:497–510

  68. 68.

    Nishikimi A, Fukuhara H, Su W, Hongu T, Takasuga S, Mihara H, Cao Q, Sanematsu F, Kanai M, Hasegawa H, Tanaka Y, Shibasaki M, Kanaho Y, Sasaki T, Frohman MA, Fukui Y (2009) Sequential regulation of DOCK2 dynamics by two phospholipids during neutrophil chemotaxis. Science 324:384–387

  69. 69.

    Kunisaki Y, Nishikimi A, Tanaka Y, Takii R, Noda M, Inayoshi A, Watanabe K, Sanematsu F, Sasazuki T, Sasaki T, Fukui Y (2006) DOCK2 is a Rac activator that regulates motility and polarity during neutrophil chemotaxis. J Cell Biol 174:647–652

  70. 70.

    Gotoh K, Tanaka Y, Nishikimi A, Inayoshi A, Enjoji M, Takayanagi R, Sasazuki T, Fukui Y (2008) Differential requirement for DOCK2 in migration of plasmacytoid dendritic cells versus myeloid dendritic cells. Blood 111:2973–2976

  71. 71.

    Werr J, Johansson J, Eriksson EE, Hedqvist P, Ruoslahti E, Lindbom L (2000) Integrin alpha(2)beta(1) (VLA-2) is a principal receptor used by neutrophils for locomotion in extravascular tissue. Blood 95:1804–1809

  72. 72.

    Matheu MP, Deane JA, Parker I, Fruman DA, Cahalan MD (2007) Class IA phosphoinositide 3-kinase modulates basal lymphocyte motility in the lymph node. J Immunol 179:2261–2269

  73. 73.

    Foger N, Rangell L, Danilenko DM, Chan AC (2006) Requirement for coronin 1 in T lymphocyte trafficking and cellular homeostasis. Science 313:839–842

  74. 74.

    Shiow LR, Roadcap DW, Paris K, Watson SR, Grigorova IL, Lebet T, An J, Xu Y, Jenne CN, Foger N, Sorensen RU, Goodnow CC, Bear JE, Puck JM, Cyster JG (2008) The actin regulator coronin 1A is mutant in a thymic egress-deficient mouse strain and in a patient with severe combined immunodeficiency. Nat Immunol 9:1307–1315

  75. 75.

    Shiow LR, Paris K, Akana MC, Cyster JG, Sorensen RU, Puck JM (2009) Severe combined immunodeficiency (SCID) and attention deficit hyperactivity disorder (ADHD) associated with a Coronin-1A mutation and a chromosome 16p11.2 deletion. Clin Immunol 131:24–30

  76. 76.

    Combaluzier B, Pieters J (2009) Chemotaxis and phagocytosis in neutrophils is independent of coronin 1. J Immunol 182:2745–2752

  77. 77.

    Sakata D, Taniguchi H, Yasuda S, Adachi-Morishima A, Hamazaki Y, Nakayama R, Miki T, Minato N, Narumiya S (2007) Impaired T lymphocyte trafficking in mice deficient in an actin-nucleating protein, mDia1. J Exp Med 204:2031–2038

  78. 78.

    Bardi G, Niggli V, Loetscher P (2003) Rho kinase is required for CCR7-mediated polarization and chemotaxis of T lymphocytes. FEBS Lett 542:79–83

  79. 79.

    Jacobelli J, Chmura SA, Buxton DB, Davis MM, Krummel MF (2004) A single class II myosin modulates T cell motility and stopping, but not synapse formation. Nat Immunol 5:531–538

  80. 80.

    Jacobelli J, Bennett FC, Pandurangi P, Tooley AJ, Krummel MF (2009) Myosin-IIA and ICAM-1 regulate the interchange between two distinct modes of T cell migration. J Immunol 182:2041–2050

  81. 81.

    Narumiya S, Tanji M, Ishizaki T (2009) Rho signaling, ROCK and mDia1, in transformation, metastasis and invasion. Cancer Metastasis Rev 28:65–76

  82. 82.

    Morley SC, Wang C, Lo WL, Lio CW, Zinselmeyer BH, Miller MJ, Brown EJ, Allen PM (2010) The actin-bundling protein L-plastin dissociates CCR7 proximal signaling from CCR7-induced motility. J Immunol 184:3628–3638

  83. 83.

    Katagiri K, Imamura M, Kinashi T (2006) Spatiotemporal regulation of the kinase Mst1 by binding protein RAPL is critical for lymphocyte polarity and adhesion. Nat Immunol 7:919–928

  84. 84.

    Katagiri K, Katakai T, Ebisuno Y, Ueda Y, Okada T, Kinashi T (2009) Mst1 controls lymphocyte trafficking and interstitial motility within lymph nodes. EMBO J 28:1319–1331

  85. 85.

    Kinashi T, Katagiri K (2005) Regulation of immune cell adhesion and migration by regulator of adhesion and cell polarization enriched in lymphoid tissues. Immunology 116:164–171

  86. 86.

    Jeon TJ, Lee DJ, Merlot S, Weeks G, Firtel RA (2007) Rap1 controls cell adhesion and cell motility through the regulation of myosin II. J Cell Biol 176:1021–1033

  87. 87.

    Reif K, Ekland EH, Ohl L, Nakano H, Lipp M, Forster R, Cyster JG (2002) Balanced responsiveness to chemoattractants from adjacent zones determines B-cell position. Nature 416:94–99

  88. 88.

    Ueno T, Saito F, Gray DH, Kuse S, Hieshima K, Nakano H, Kakiuchi T, Lipp M, Boyd RL, Takahama Y (2004) CCR7 signals are essential for cortex-medulla migration of developing thymocytes. J Exp Med 200:493–505

  89. 89.

    Witt CM, Raychaudhuri S, Schaefer B, Chakraborty AK, Robey EA (2005) Directed migration of positively selected thymocytes visualized in real time. PLoS Biol 3:e160

  90. 90.

    Ehrlich LI, Oh DY, Weissman IL, Lewis RS (2009) Differential contribution of chemotaxis and substrate restriction to segregation of immature and mature thymocytes. Immunity 31:986–998

  91. 91.

    Kurobe H, Liu C, Ueno T, Saito F, Ohigashi I, Seach N, Arakaki R, Hayashi Y, Kitagawa T, Lipp M, Boyd RL, Takahama Y (2006) CCR7-dependent cortex-to-medulla migration of positively selected thymocytes is essential for establishing central tolerance. Immunity 24:165–177

  92. 92.

    Castellino F, Huang AY, Altan-Bonnet G, Stoll S, Scheinecker C, Germain RN (2006) Chemokines enhance immunity by guiding naive CD8+ T cells to sites of CD4+ T cell-dendritic cell interaction. Nature 440:890–895

  93. 93.

    Beuneu H, Garcia Z, Bousso P (2006) Cutting edge: cognate CD4 help promotes recruitment of antigen-specific CD8 T cells around dendritic cells. J Immunol 177:1406–1410

  94. 94.

    Castellino F, Germain RN (2007) Chemokine-guided CD4+ T cell help enhances generation of IL-6RalphahighIL-7Ralpha high prememory CD8+ T cells. J Immunol 178:778–787

  95. 95.

    Schwab SR, Cyster JG (2007) Finding a way out: lymphocyte egress from lymphoid organs. Nat Immunol 8:1295–1301

  96. 96.

    Wei SH, Rosen H, Matheu MP, Sanna MG, Wang SK, Jo E, Wong CH, Parker I, Cahalan MD (2005) Sphingosine 1-phosphate type 1 receptor agonism inhibits transendothelial migration of medullary T cells to lymphatic sinuses. Nat Immunol 6:1228–1235

  97. 97.

    Sinha RK, Park C, Hwang IY, Davis MD, Kehrl JH (2009) B lymphocytes exit lymph nodes through cortical lymphatic sinusoids by a mechanism independent of sphingosine-1-phosphate-mediated chemotaxis. Immunity 30:434–446

  98. 98.

    Matloubian M, Lo CG, Cinamon G, Lesneski MJ, Xu Y, Brinkmann V, Allende ML, Proia RL, Cyster JG (2004) Lymphocyte egress from thymus and peripheral lymphoid organs is dependent on S1P receptor 1. Nature 427:355–360

  99. 99.

    Pflicke H, Sixt M (2009) Preformed portals facilitate dendritic cell entry into afferent lymphatic vessels. J Exp Med 206:2925–2935

  100. 100.

    Ohl L, Mohaupt M, Czeloth N, Hintzen G, Kiafard Z, Zwirner J, Blankenstein T, Henning G, Forster R (2004) CCR7 governs skin dendritic cell migration under inflammatory and steady-state conditions. Immunity 21:279–288

  101. 101.

    Cinamon G, Shinder V, Alon R (2001) Shear forces promote lymphocyte migration across vascular endothelium bearing apical chemokines. Nat Immunol 2:515–522

  102. 102.

    Shulman Z, Shinder V, Klein E, Grabovsky V, Yeger O, Geron E, Montresor A, Bolomini-Vittori M, Feigelson SW, Kirchhausen T, Laudanna C, Shakhar G, Alon R (2009) Lymphocyte crawling and transendothelial migration require chemokine triggering of high-affinity LFA-1 integrin. Immunity 30:384–396

  103. 103.

    Pereira JP, An J, Xu Y, Huang Y, Cyster JG (2009) Cannabinoid receptor 2 mediates the retention of immature B cells in bone marrow sinusoids. Nat Immunol 10:403–411

  104. 104.

    Sapoznikov A, Pewzner-Jung Y, Kalchenko V, Krauthgamer R, Shachar I, Jung S (2008) Perivascular clusters of dendritic cells provide critical survival signals to B cells in bone marrow niches. Nat Immunol 9:388–395

  105. 105.

    Cariappa A, Mazo IB, Chase C, Shi HN, Liu H, Li Q, Rose H, Leung H, Cherayil BJ, Russell P, von Andrian U, Pillai S (2005) Perisinusoidal B cells in the bone marrow participate in T-independent responses to blood-borne microbes. Immunity 23:397–407

  106. 106.

    Allen CD, Cyster JG (2008) Follicular dendritic cell networks of primary follicles and germinal centers: phenotype and function. Semin Immunol 20:14–25

  107. 107.

    Le Borgne M, Ladi E, Dzhagalov I, Herzmark P, Liao YF, Chakraborty AK, Robey EA (2009) The impact of negative selection on thymocyte migration in the medulla. Nat Immunol 10:823–830

  108. 108.

    Lin A, Loughman JA, Zinselmeyer BH, Miller MJ, Caparon MG (2009) Streptolysin S inhibits neutrophil recruitment during the early stages of Streptococcus pyogenes infection. Infect Immun 77:5190–5201

  109. 109.

    Allen CD, Okada T, Tang HL, Cyster JG (2007) Imaging of germinal center selection events during affinity maturation. Science 315:528–531

Download references

Acknowledgment

I thank Jason Cyster for providing Hy10 mice and CFP mice, Yoshikazu Ando for helping figure preparation, and the reviewers for valuable comments. I also acknowledge Masahiro Kitano, Michiyuki Matsudsa, Yasuo Mori, Tomohiro Kurosaki, and Takashi Saito for their help in setting up the two-photon microscope. This work was supported by the Japan Society for the Promotion of Science, the Ministry of Education, Culture, Sports, Science and Technology of Japan, the Uehara Memorial Foundation, the Sumitomo Foundation, and the Mochida Memorial Foundation for Medical and Pharmaceutical Research,

Open Access

This article is distributed under the terms of the Creative Commons Attribution Noncommercial License which permits any noncommercial use, distribution, and reproduction in any medium, provided the original author(s) and source are credited.

Author information

Correspondence to Takaharu Okada.

Additional information

This article is published as part of the Special Issue on Immunoimaging of Immune System Function.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Migration of the B cell-T cell conjugate in the stromal cell network in the lymph node. The movie shows the 45-min time lapse images recorded every 30 s. Cognate antigen-specific B cells (green) and helper T cells (red) are stably conjugated. In cyan are primarily irradiation-resistant stromal cells. The mouse lymph node was imaged ex vivo by TPLM 36 h after immunization. Elapsed time is shown as hours/minutes/seconds. The image volume is 100 μm (x) × 100 μm (y) × 60 μm (z). The scale bar shows 30 μm on the nearest x-y plane from the viewpoint (MPG 4.71 MB)

Video 1

Migration of the B cell-T cell conjugate in the stromal cell network in the lymph node. The movie shows the 45-min time lapse images recorded every 30 s. Cognate antigen-specific B cells (green) and helper T cells (red) are stably conjugated. In cyan are primarily irradiation-resistant stromal cells. The mouse lymph node was imaged ex vivo by TPLM 36 h after immunization. Elapsed time is shown as hours/minutes/seconds. The image volume is 100 μm (x) × 100 μm (y) × 60 μm (z). The scale bar shows 30 μm on the nearest x-y plane from the viewpoint (MPG 4.71 MB)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Okada, T. Two-photon microscopy analysis of leukocyte trafficking and motility. Semin Immunopathol 32, 215–225 (2010). https://doi.org/10.1007/s00281-010-0210-3

Download citation

Keywords

  • Live imaging
  • Leukocyte migration
  • Chemokine
  • Integrin
  • Stromal cells