Ryan P. Larson

Mechanical injury polarizes skin dendritic cells to elicit a TH2 response by inducing cutaneous thymic stromal lymphopoietin expression

BACKGROUND Atopic dermatitis is characterized by scratching and by T(H)2-dominated immune response to cutaneously introduced antigens. Antigen application to skin mechanically injured by tape stripping results in T(H)2-dominated skin inflammation. OBJECTIVE To examine the effect of tape stripping on the capacity of skin dendritic cells (DCs) to polarize T cells toward a T(H)2 phenotype. METHODS CD11c(+) DCs were isolated from skin of BALB/c or C57BL/6 mice. Fluorescein isothiocyanate (FITC)(+) and FITC(-) DCs were isolated from draining lymph nodes (DLNs) 24 hours after painting the skin with FITC. DCs were assessed for their ability to induce cytokine secretion by ovalbumin-stimulated naive CD4(+) T cells from T cell receptor-ovalbumin transgenic DO11.10 mice. Cytokine mRNA levels were examined by quantitative PCR. RESULTS Dendritic cells isolated from the skin of wild-type, but not thymic stromal lymphopoietin (TSLP) receptor(-/-) or IL-10(-/-), mice 6 hours after tape stripping elicited significantly more IL-4 and IL-13 and significantly less IFN-γ production by CD4(+) cells than DCs isolated from unmanipulated skin, and expressed significantly more mRNA for the T(H)2 skewing molecules IL-10, Jagged1, and Jagged2, but significantly less mRNA for the T(H)1 skewing cytokine IL-12. CD11c(+)FITC(+) cells isolated from DLNs of shaved and tape stripped skin of wild-type, but not TSLP receptor(-/-) or IL-10(-/-), mice polarized T cells significantly more toward T(H)2 and expressed significantly more IL-10, Jagged1, and Jagged2 mRNA than CD11c(+)FITC(+) cells isolated from DLNs of shaved skin. Tape stripping significantly increased TSLP levels in the skin, and TSLP was shown to play an essential role in the T(H)2 polarization of skin DCs by tape stripping. CONCLUSIONS Tape stripping upregulates TSLP levels in the skin, which polarizes skin DCs to elicit a T(H)2 response via the induction of IL-10.

The transcription factor STAT5 is critical in dendritic cells for the development of TH2 but not TH1 responses.

Dendritic cells (DCs) are critical in immune responses, linking innate and adaptive immunity. We found here that DC-specific deletion of the transcription factor STAT5 was not critical for development but was required for T helper type 2 (TH2), but not TH1, allergic responses in both the skin and lungs. Loss of STAT5 in DCs led to the inability to respond to thymic stromal lymphopoietin (TSLP). STAT5 was required for TSLP-dependent DC activation, including upregulation of the expression of costimulatory molecules and chemokine production. Furthermore, TH2 responses in mice with DC-specific loss of STAT5 resembled those seen in mice deficient in the receptor for TSLP. Our results show that the TSLP-STAT5 axis in DCs is a critical component for the promotion of type 2 immunity at barrier surfaces.

Dibutyl Phthalate-Induced Thymic Stromal Lymphopoietin Is Required for Th2 Contact Hypersensitivity Responses

Thymic stromal lymphopoietin (TSLP) is an IL-7–related cytokine, produced by epithelial cells, that has been linked to atopic dermatitis and asthma; however, it remains unclear how TSLP shapes the adaptive immune response that causes these allergic disorders. In this study, we demonstrate a role for TSLP in a Th2 model of contact hypersensitivity in mice. TSLP is required for the development of Th2-type contact hypersensitivity induced by the hapten FITC in combination with the sensitizing agent dibutyl phthalate. TSLPR-deficient mice exhibited a dramatically reduced response, including markedly reduced local infiltration by eosinophils, Th2 cytokine production, and serum IgE levels, following FITC sensitization and challenge. The reduced response by TSLPR-deficient mice is likely due to decreased frequency and reduced T cell stimulatory function of skin-derived Ag-bearing FITC+CD11c+ dendritic cells in draining lymph nodes following FITC sensitization. These data suggest that skin-derived dendritic cells are direct or indirect targets of TSLP in the development of type 2 immune responses in the skin, where TSLP drives their maturation, accumulation in skin draining lymph nodes, and ability to induce proliferation of naive allergen-specific T cells.

ICOS and Bcl6-dependent pathways maintain a CD4 T cell population with memory-like properties during tuberculosis

Protective CD4 T cells specific for M. tuberculosis (Mtb) are maintained in the lungs during active Mtb infection. Similar to memory CD4 T cells, persistence of these Mtb-specific cells requires intrinsic expression of Bcl6 and ICOS.

Foxp3(+) regulatory T cells in tuberculosis.

The immune response to Mycobacterium tuberculosis (Mtb) must be tightly regulated to mount a sufficient response to limit bacterial growth and dissemination while avoiding excessive inflammation that could damage host tissues. A wide variety of cell types, cell surface molecules, and cytokines are likely to contribute to this regulation, but recent studies have revealed that a subset of CD4 T cells expressing the transcription factor Foxp3, called regulatory T (reg) cells, play a critical role [1-3]. Although the first reports of T reg cells in tuberculosis (TB) occurred only recently (i.e., 2006) [4, 5], we have already gained many insights into their activity during TB. While it is likely that T reg cells do play some beneficial roles by preventing inflammation-mediated damage to host tissues during TB, this aspect of their function has not been well studied to date. What is clear, however, is that during the initial T cell response to Mtb infection, Mtb induces the expansions of T reg cells that delay the onset of adaptive immunity, suggesting that Mtb has hijacked T reg cell-mediated immune suppression to allow it to replicate unabated in the lung until T cells finally arrive [6]. In this chapter, we will first provide an overview of the delayed T cell response to Mtb and a brief introduction to regulatory T cells. We will then review what is known about T reg cells from observations in human populations, discuss mechanistic insights revealed in the mouse model, and speculate about the relevance of this understanding for future efforts to prevent and treat TB.

MiR-155–regulated molecular network orchestrates cell fate in the innate and adaptive immune response to Mycobacterium tuberculosis

Significance The mechanism by which Mycobacterium tuberculosis (Mtb) modulates the host immune response is not fully understood. We have used a systems biology approach to generate a microRNA regulatory network composed of 77 microRNAs that are associated with Mtb–macrophage interactions. We have determined a unique and dual role for one of these regulators, miR-155, as a rheostat regulating the survival of both innate and adaptive immune cells. On the one hand, miR-155 maintains the survival of Mtb-infected macrophages, providing a niche favoring bacterial replication. On the other hand, miR-155 maintains the survival of Mtb-specific T cells, enabling an effective adaptive response. Our work underscores the value of systems-based prediction of pathogen-specific microRNA networks as a tool to define host–pathogen interactions. The regulation of host–pathogen interactions during Mycobacterium tuberculosis (Mtb) infection remains unresolved. MicroRNAs (miRNAs) are important regulators of the immune system, and so we used a systems biology approach to construct an miRNA regulatory network activated in macrophages during Mtb infection. Our network comprises 77 putative miRNAs that are associated with temporal gene expression signatures in macrophages early after Mtb infection. In this study, we demonstrate a dual role for one of these regulators, miR-155. On the one hand, miR-155 maintains the survival of Mtb-infected macrophages, thereby providing a niche favoring bacterial replication; on the other hand, miR-155 promotes the survival and function of Mtb-specific T cells, enabling an effective adaptive immune response. MiR-155–induced cell survival is mediated through the SH2 domain-containing inositol 5-phosphatase 1 (SHIP1)/protein kinase B (Akt) pathway. Thus, dual regulation of the same cell survival pathway in innate and adaptive immune cells leads to vastly different outcomes with respect to bacterial containment.

Thymic stromal lymphopoietin and the pathophysiology of atopic disease.

Thymic stromal lymphopoietin (TSLP) is an IL-7-related cytokine expressed predominantly by barrier epithelial cells. TSLP is a potent activator of several cell types, including myeloid-derived dendritic cells, monocytes/macrophages and mast cells. Recent studies have revealed an important role for TSLP in the initiation and progression of allergic inflammatory diseases. In this review, we will discuss the role of TSLP in atopic diseases, as well as its function in immune homeostasis.

Temporal Regulation of Ig Gene Diversification Revealed by Single-Cell Imaging

Rearranged Ig V regions undergo activation-induced cytidine deaminase (AID)-initiated diversification in sequence to produce either nontemplated or templated mutations, in the related pathways of somatic hypermutation and gene conversion. In chicken DT40 B cells, gene conversion normally predominates, producing mutations templated by adjacent pseudo-V regions, but impairment of gene conversion switches mutagenesis to a nontemplated pathway. We recently showed that the activator, E2A, functions in cis to promote diversification, and that G1 phase of cell cycle is the critical window for E2A action. By single-cell imaging of stable AID-yellow fluorescent protein transfectants, we now demonstrate that AID-yellow fluorescent protein can stably localize to the nucleus in G1 phase, but undergoes ubiquitin-dependent proteolysis later in cell cycle. By imaging of DT40 polymerized lactose operator-λR cells, in which polymerized lactose operator tags the rearranged λR gene, we show that both the repair polymerase Polη and the multifunctional factor MRE11/RAD50/NBS1 localize to λR, and that λR/Polη colocalizations occur predominately in G1 phase, when they reflect repair of AID-initiated damage. We find no evidence of induction of γ-H2AX, the phosphorylated variant histone that is a marker of double-strand breaks, and Ig gene conversion may therefore proceed by a pathway involving templated repair at DNA nicks rather than double-strand breaks. These results lead to a model in which Ig gene conversion initiates and is completed or nearly completed in G1 phase. AID deaminates ssDNA, and restriction of mutagenesis to G1 phase would contribute to protecting the genome from off-target attack by AID when DNA replication occurs in S phase.

IL-23 induces atopic dermatitis-like inflammation instead of psoriasis-like inflammation in CCR2-deficient mice.

Psoriasis is an immune-mediated chronic inflammatory skin disease, characterized by epidermal hyperplasia and infiltration of leukocytes into the dermis and epidermis. IL-23 is expressed in psoriatic skin, and IL-23 injected into the skin of mice produces IL-22-dependent dermal inflammation and acanthosis. The chemokine receptor CCR2 has been implicated in the pathogenesis of several inflammatory diseases, including psoriasis. CCR2-positive cells and the CCR2 ligand, CCL2 are abundant in psoriatic lesions. To examine the requirement of CCR2 in the development of IL-23-induced cutaneous inflammation, we injected the ears of wild-type (WT) and CCR2-deficient (CCR2−/−) mice with IL-23. CCR2−/− mice had increased ear swelling and epidermal thickening, which was correlated with increased cutaneous IL-4 levels and increased numbers of eosinophils within the skin. In addition, TSLP, a cytokine known to promote and amplify T helper cell type 2 (Th2) immune responses, was also increased within the inflamed skin of CCR2−/− mice. Our data suggest that increased levels of TSLP in CCR2−/− mice may contribute to the propensity of these mice to develop increased Th2-type immune responses.

Cutting edge: allergen-specific CD4 T cells respond indirectly to thymic stromal lymphopoietin to promote allergic responses in the skin.

Thymic stromal lymphopoietin (TSLP) is an epithelial-derived cytokine that has been implicated in the initiation of allergic responses. CD4 T cells and dendritic cells are able to respond to TSLP in vitro; however, there has not been a careful dissection of the spatiotemporal response to TSLP by CD4 T cells in vivo during an allergic response. Previous work has suggested a requirement for TSLP in amplifying Th2 responses during allergen challenge by direct action on CD4 T cells; however, these studies did not determine whether there is an effect of TSLP on CD4 T cells during allergen sensitization. In this study we demonstrate an indirect role for TSLP on CD4 T cells during sensitization and challenge phases of an allergic response. This indirect effect of TSLP on CD4 T cells is due in part to the presence of TSLP exclusively in the allergen-sensitized and -challenged skin, rather than the draining lymph nodes.