• Th17 Reagents
  
  
• IC Flow Reagents
• Recombinant Cytokines
• ELISA Antibody Pairs
• ELISA Sets & Kits
• ELISPOT Sets
• In Vivo Cytokine Capture Assay (IVCCA)
• Cytokine & Chemokine Receptors

IL-17 Analysis by Intracellular Staining, ELISPOT, ELISA, and Bioassay

• IL-17 Data
• Phenotypic Profile of Th1, Th2 & Th-17 Cell Types
• Mouse Th-17 Cell Related Reagents
• Human Th-17 Cell Related Reagents

Th17 Lineage

IL-17-producing CD4+ T cells (Th-17 cells) have been identified as a unique subset of Th cells that develops along a pathway that is distinct from the Th1- and Th2-cell differentiation pathways. One proposed mechanism is IL-23-dependent, but independent of STAT-1, T-bet, STAT4, and STAT6. The hallmark effector molecules of Th1 and Th2 cells, e.g., IFN-γ and IL-4, have each been found to negatively regulate the generation of these Th-17 cells. By a second proposed mechanism, de novo differentiation of Th-17 cells in the absence of IL-23 has been demonstrated by treatment of naïve CD4 cells with TGF-β1 and IL-6, an effect heightened by IL-1β and TNF-α; IL-23, though absent during the Th-17 differentiation, was still required for survival and expansion of the Th-17 cells. In EAE experiments, adoptive transfer of IL-23-polarized IL-17 effector cells is required for disease progression. Moreover, inhibition of the (negative-regulatory) IFN-γ signal has been found to lead to enhanced development of pathogenic Th-17 effector cells and autoimmune disease exacerbation.

IL-17A Data

Human IL-17A ELISPOT: Left: Human PBMCs cultured for 24 hrs (no mitogen). Right: Human PBMCs activated with PMA/Ionomycin for 24 hrs.

IL-17 Biology

Interleukin-17 (IL-17) is a CD4+ T cell-derived cytokine that promotes inflammatory responses and is correlated with rheumatoid arthritis, asthma, multiple sclerosis, psoriasis, and transplant rejection. The cDNA encoding human IL-17 was isolated from a library of CD4+ T cells; the encoded protein exhibits 72 percent amino acid identity with HVS13 , an open reading frame from a T lymphotropic Herpesvirus saimiri, and 63 percent with mouse CTLA-8. Human IL-17 exists as glycosylated 20-30 kD homodimers. High levels of IL-17 homodimer are produced by activated peripheral blood CD4+ T-cells. IL-17 enhances expression of the intracellular adhesion molecule-1 (ICAM-1) in human fibroblasts. Human IL-17 also stimulates epithelial, endothelial, or fibroblastic cells to secrete IL-6, IL-8, G-CSF, and PGE2. In the presence of human IL-17, fibroblasts can sustain the proliferation of CD34+ hematopoietic progenitors and induce maturation into neutrophils. Mouse, rat, and human IL-17 can induce IL-6 secretion in mouse stromal cells, indicating that all homologs can recognize the mouse IL-17 receptor.

Development of IL-17-producing effector CD4+ T cells (Th-17)

Th17 Cells Express IL-17A, IL-17F, IL-22 & IL-17A/F Heterodimer

In addition to IL-17A, differentiated Th17 cells also produce IL-17F and IL-22 upon re-activation. Like IL-17A, IL-17F has been linked with inflammatory diseases. IL-17F and IL-17A expression has been observed in tissue samples from various autoimmune diseases, such as rheumatoid arthritis, multiple sclerosis, psoriasis, inflammatory bowel disease, and asthma. IL-17F treatment of airway epithelium, vein endothelial cells, and fibroblasts has been reported to induce expression of IL-6, IL-8, GROa, ENA-78, TGF-β, MCP-1, G-CSF, GM-CSF, and ICAM-1.

Of all the six IL-17 family members, IL-17F and IL-17A share the strongest homology (50% amino acid identity), and the two genes are located in the same chromosomal region. Recent studies have demonstrated coordinated regulation of IL-17A and IL-17F during Th17 differentiation. Expression of IL-17F and IL-17A has been detected in activated human peripheral blood lymphocytes, specifically by activated human CD4+ T cells.

Like IL-17A, IL-17F is a disulfide-linked homodimeric glycoprotein. The IL-17F homodimer includes a classical cysteine knot motif, which is found also in the TGF-β, bone morphogenetic protein, and nerve growth factor superfamilies. The presence of the cysteine knot motif suggested the possibility of a heterodimeric structure, as was reported for TGF-β and inhibin/activin. Recent reports by two research teams (Chen Dong et al.; Neil Wolfman et al.) confirm this hypothesis, namely that co-expression of IL-17F and IL-17A in HEK293 cells results in the formation of biologically active IL-17F/IL-17A heterodimers, in addition to the IL-17F homodimers and IL-17A homodimers. Moreover, activated human CD4+ T cells were found to produce the IL-17A/F heterodimer, along with the corresponding homodimers. In comparing the relative potency of IL-17A, IL-17F, and IL-17A/F, all three were found to induce GRO-a secretion; IL-17A was most potent, followed by IL-17A/F heterodimer, then IL-17F (100fold lower than IL-17A). In describing the relative abundance of the secreted proteins, activated human CD4+ T cells in culture were found to secret IL-17F homodimer at 10-fold higher levels than IL-17A homodimer, suggesting that the majority of the IL-17A protein expressed exists in the form of the IL-17A/F heterodimer. In the mouse, the IL-17A/F heterodimer (alone or in synergy with TNF-α) was found to regulate the expression of IL-6 and KC (mouse homolog of human GROa); this was found to be dependent on IL-17RA and TRAF6. This novel heterodimeric cytokine, IL-17A/F, is believed to regulate inflammatory responses.

IL-17F Data

IL-17F is Expressed by Th17 Cells upon re-activation (IL-17F vs. CD4): In vitro-differentiated TH17 cells were reactivated in the presence of monensin, then stained for CD4 followed by intracellular staining with Alexa Fluor® 647 anti-mouse IL-17F (clone eBio18F10) (left panel) or isotype control (right panel).			Th17 Cells co-express IL-17F & IL-17A upon re-activation (IL-17F vs. IL-17A): In vitro-differentiated Th17 cells were reactivated in the presence of monensin, then stained for CD4, fixed, permeabilized, and stained for Alexa Fluor® 647 anti-mouse IL-17F (clone eBio18F10) and PE anti-mouse IL-17A (clone eBio17B7) (left). Right panel depicts isotype control versus IL-17A.

Phenotypic Profile of Th1, Th2 & Th-17 Cell Types

References

• Chang SH, Dong C. A novel heterodimeric cytokine consisting of IL-17 and IL-17F regulates inflammatory responses. Cell Res. 2007 May;17(5):435-40. [Pubmed]
  
  
• Wright JF, Guo Y, Quazi A, Luxenberg DP, Bennett F, Ross JF, Qiu Y, Whitters MJ, Tomkinson KN, Dunussi-Joannopoulos K, Carreno BM, Collins M, Wolfman NM. Identification of an interleukin 17F/17A heterodimer in activated human CD4+ T cells. J Biol Chem. 2007 May 4;282(18):13447-55. Epub 2007 Mar 13. [Pubmed]
  
  
• Liang SC, Tan XY, Luxenberg DP, Karim R, Dunussi-Joannopoulos K, Collins M, Fouser LA. Interleukin (IL)-22 and IL-17 are coexpressed by Th17 cells and cooperatively enhance expression of antimicrobial peptides. J Exp Med. 2006 Oct 2;203(10):2271-9. Epub 2006 Sep 18. [Pubmed]
  
  
• Veldhoen M, Hocking RJ, Atkins CJ, Locksley RM, Stockinger B. TGFbeta in the context of an inflammatory cytokine milieu supports de novo differentiation of IL-17-producing T cells. Immunity. 2006 Feb;24(2):179-89. [PubMed]
  
  
• Harrington LE, Hatton RD, Mangan PR, Turner H, Murphy TL, Murphy KM, Weaver CT. Interleukin 17-producing CD4+ effector T cells develop via a lineage distinct from the T helper type 1 and 2 lineages. Nat Immunol. 2005 Nov;6(11):1123-32. [eBioTC11-18H10.1, ELISA, IC Staining, PubMed]
  
  
• Park H, Li Z, Yang XO, Chang SH, Nurieva R, Wang YH, Wang Y, Hood L, Zhu Z, Tian Q, Dong C. A distinct lineage of CD4 T cells regulates tissue inflammation by producing interleukin 17. Nat Immunol. 2005 Nov;6(11):1133-41. [eBioTC11-18H10.1, ELISA, IC Staining, PubMed]
  
  
• Nekrasova T, Shive C, Gao Y, Kawamura K, Guardia R, Landreth G, Forsthuber TG. ERK1-deficient mice show normal T cell effector function and are highly susceptible to experimental autoimmune encephalomyelitis. J Immunol. 2005 Aug 15;175(4):2374-80. [eBioTC11-18H10.1, ELISPOT, PubMed]
  
  
• Schubert D, Maier B, Morawietz L, Krenn V, Kamradt T. Immunization with glucose-6-phosphate isomerase induces T cell-dependent peripheral polyarthritis in genetically unaltered mice. J Immunol. 2004 Apr 1;172(7):4503-9. [eBioTC11-18H10.1, IC Staining, PubMed]
  
  
• Infante-Duarte C, Horton HF, Byrne MC, Kamradt T. Microbial lipopeptides induce the production of IL-17 in Th cells. J Immunol. 2000 Dec 1;165(11):6107-15. [eBioTC11-18H10.1, IC Staining, PubMed]
  
  
• Hofstetter HH, Ibrahim SM, Koczan D, Kruse N, Weishaupt A, Toyka KV, Gold R. Therapeutic efficacy of IL-17 neutralization in murine experimental autoimmune encephalomyelitis. Cell Immunol. 2005 Oct;237(2):123-30. [eBioTC11-18H10.1, ELISPOT, PubMed]

Mouse Th-17 Cell Related Reagents

Human Th-17 Cell Related Reagents