Engagement of TLR2 Reverses the Suppressor Function of Conjunctiva CD4⁺CD25⁺Regulatory T Cells and Promotes Herpes Simplex Virus Epitope-Specific CD4⁺CD25⁻Effector T Cell Responses

Abstract

PURPOSE. The authors recently reported that Foxp3(+)CD4(+) CD25(+(Bright)) "natural" regulatory T cells (nT(reg) cells) are abundant in rabbit conjunctiva and suppress herpes simplex virus (HSV)-1-specific CD4(+) and CD8(+) effector T cells (T(eff) cells). However, little is known about the overall regulatory mechanisms of these nT(reg) cells. The authors investigate the regulation of conjunctiva-resident nT(reg) cells through Toll-like receptors (TLRs) and their effect on ocular mucosal T(eff) cell immunity. METHODS. CD4(+)CD25(+) nT(reg) cells were purified from naive rabbit conjunctivas, and their TLR expression profile was determined. The effects of TLR engagement on nT(reg) cell-mediated suppression of CD4(+) T(eff) cells were determined in vitro and in vivo. RESULTS. The authors found that conjunctiva-resident nT(reg) cells express high levels of TLR2 and TLR9; exposure to the TLR2 ligand lipoteichoic acid (LTA) led to the increased activation and proliferation of nT(reg) cells, and the addition of autologous APCs further increased nT(reg) cell expansion; in contrast, the TLR9 ligand CpG(2007) inhibited the proliferation of nT(reg) cells, and the addition of autologous APCs had no effect on such inhibition; nT(reg) cells treated with LTA, but not with CpG(2007), expressed IFN-γ and IL-10 mRNA, but not TGF-β; consistent with in vitro data, rabbits immunized by topical ocular drops of HSV-gD peptides + TLR2 ligand (LTA) displayed enhanced CD4(+)CD25(-) T(eff) cell immune responses when compared with HSV-gD peptides + TLR9 ligand (CpG(2007)). CONCLUSIONS. Although conjunctiva-resident CD4(+)CD25(+) nT(reg) cells express high level of TLR2 and TLR9, their suppressive function is more significantly reversed after the administration of TLR2 ligand (LTA; P < 0.005) than of TLR9 ligand (CpG(200); P > 0.005). These findings will likely help optimize the topical ocular administration of immunotherapies.

Therapeutic Immunization with a Mixture of Herpes Simplex Virus 1 Glycoprotein D-Derived “Asymptomatic” Human CD8 + T-Cell Epitopes Decreases Spontaneous Ocular Shedding in Latently Infected HLA Transgenic Rabbits: Association with Low Frequency of Local PD-1 + TIM-3 + CD8 + Exhausted T Cells

“…We choose the Nε-palmitic acidlysine residue (a TLR-2 ligand) to better target the ASYMP CD4-CD8 peptide vaccine to the local ocular mucosal immune system, since, as shown in Fig. 2C and D, high levels of Toll-like receptor 2 (TLR-2), but not Toll-like receptor 4 (TLR-4), were expressed by rabbit cornea-and conjunctiva-derived CD11b/c ϩ antigen-presenting cells (APCs) (26). The final three TLR-2 ligand-peptide conjugate constructs (i.e., CD4-CD8 lipopeptide vaccines) were made using a chemoselective ligation method (25), which provides higher yield, purity, and solubility than the traditional method of lipopeptide synthesis.…”

Retinal Muller Glia Initiate Innate Response to Infectious Stimuli via Toll-Like Receptor Signaling

“…An increasing number of studies had shown that TLRs are expressed by a variety of tissues and cells in the eye [10], [26]. However, whether retinal Muller glia express various TLRs is not known.…”

Cutaneous RANK–RANKL Signaling Upregulates CD8-Mediated Antiviral Immunity during Herpes simplex Virus Infection by Preventing Virus-Induced Langerhans Cell Apoptosis

“…Upon cutaneous HSV-1 infection, K14-RANKL TG mice showed reduced disease severity as compared with WT controls, possibly suggesting that the infection might have impaired the RANKLmediated expansion of functional Treg. This hypothesis was supported by studies demonstrating that, during infection, Tregs frequently lose their suppressive activity in response to upregulated TLR signaling (Dasgupta et al, 2001;Peng et al, 2005;Zhang et al, 2010;Hackl et al, 2011). However, although we detected increased levels of various TLRs, such as TLR2, TLR7, and TLR9, in infected skin from TG mice (Klenner, unpublished observation), in vitro suppression assays revealed that Tregs from HSV-1-infected WT and TG mice inhibited the proliferation of effector T cells equally well, indicating that a potential loss of Treg function has a minor role in explaining the reduced disease severity in HSV-1-infected K14-RANKL TG mice.…”