The IL-10GFP (VeRT-X) mouse strain is not suitable for the detection of IL-10 production by granulocytes during lung inflammation

Abstract

The clear and unequivocal identification of immune effector functions is essential to understand immune responses. The cytokine IL-10 is a critical immune regulator and was shown, for example, to limit pathology during various lung diseases. However, the clear identification of IL-10-producing cells is challenging and, therefore, reporter mouse lines were developed to facilitate their detection. Several such reporter lines utilize GFP, including the IL-10GFP (VeRT-X) reporter strain studied here. In line with previous reports, we found that this IL-10GFP line faithfully reports on the IL-10 production of lymphoid cells. However, we show that the IL-10GFP reporter is not suitable to analyse IL-10 production of myeloid cells during inflammation. During inflammation, the autofluorescence of myeloid cells increased to an extent that entirely masked the IL-10-specific GFP-signal. Our data illustrate a general and important technical caveat using GFP-reporter lines for the analysis of myeloid cells and suggest that previous reports on effector functions of myeloid cells using such GFP-based reporters might require re-evaluation.

CD169+ macrophage intrinsic IL-10 production regulates immune homeostasis during sepsis

“…This discrepancy may be related to the inherent caveats of using cytokine reporter mice that may not report cytokine protein production with high fidelity. 32 The downstream targets of IL-10 are still unknown, although our data showing a reduction in inflammatory cytokine production such as IL-6 and TNF-α and an increase in regulatory cytokines such as G-CSF provide some clues. The scale of observed systemic changes in cytokine production in CD169-DTR mice is too large to be exclusively associated with the absence of CD169+ macrophages.…”

“…Thus, to determine if CD169+ macrophages are capable of producing IL-10 during homeostatic conditions and after the onset of septic shock, we utilized the IL-10 GFP mice (VeRT-X mouse model). The VeRT-X IL-10-GFP mouse to interrogate IL-10 production is a useful tool; however, since there are caveats associated with this model, 31 , 32 we only used these mice initially for the purposes of determining if CD169+ macrophages contribute to early IL-10 production following LPS treatment. First, we conducted flow cytometric analysis of IL-10 GFP splenocytes at 0 (naive), 1, and 3 h post LPS stimulation.…”

Staphylococcus aureus-induced immunosuppression mediated by IL-10 and IL-27 facilitates nasal colonisation

“…Here we describe for the first time that a similar mechanism is being used by the bacterium to survive within the nasal cavity during colonisation. Initial attempts to identify IL-10 + populations within the nasal tissue were carried out using flow cytometric methods however the identification of murine IL-10-producing cells by intracellular staining is challenging given that intracellular IL-10 is typically expressed at a relatively low intensity [ 35 , 36 ]. To circumvent this, we analysed IL-10 expression by individual cell subsets by RT-PCR.…”