Single-cell analysis uncovers differential regulation of lung γδ T cell subsets by the co-inhibitory molecules, PD-1 and TIM-3

IL-17A-producing γδ T cells within the lung consist of both Vγ6+ tissue-resident cells and Vγ4+ circulating cells that play important roles in homeostasis, inflammation, infection, tumor progression and metastasis. How these γδ T cell subsets are regulated in the lung environment during homeostasis and cancer remains poorly understood. Using single-cell RNA sequencing and flow cytometry, we show that lung Vγ6+ cells express a repertoire of cell surface molecules distinctive from Vγ4+ cells, including PD-1 and ICOS. We found that PD-1 functions as a co-inhibitory molecule on Vγ6+ cells to reduce IL-17A production, whereas manipulation of ICOS signaling fails to affect IL-17A in Vγ6+ cells. In a mammary tumor model, ICOS and PD-1 expression on lung Vγ6+ cells remained stable. However, Vγ6+ and Vγ4+ cells within the lung pre-metastatic niche increased expression of IL-17A, IL-17F, amphiregulin (AREG) and TIM-3 in response to tumor-derived IL-1β and IL-23, where the upregulation of TIM-3 was specific to Vγ4+ cells. Inhibition of either PD-1 or TIM-3 in mammary tumor-bearing mice further increased IL-17A by Vγ6+ and Vγ4+ cells, indicating that both PD-1 and TIM-3 function as negative regulators of IL-17A-producing γδ T cell subsets. Together, these data demonstrate how lung γδ T cell subsets are differentially controlled by co-inhibitory molecules in steady-state and cancer.

Assessment of the WAP-Myc mouse mammary tumor model for spontaneous metastasis

Breast cancer is the most common form of cancer in women. Despite significant therapeutic advances in recent years, breast cancer also still causes the greatest number of cancer-related deaths in women, the vast majority of which (> 90%) are caused by metastases. However, very few mouse mammary cancer models exist that faithfully recapitulate the multistep metastatic process in human patients. Here we assessed the suitability of a syngrafting protocol for a Myc-driven mammary tumor model (WAP-Myc) to study autochthonous metastasis. A moderate but robust spontaneous lung metastasis rate of around 25% was attained. In addition, increased T cell infiltration was observed in metastatic tumors compared to donor and syngrafted primary tumors. Thus, the WAP-Myc syngrafting protocol is a suitable tool to study the mechanisms of metastasis in MYC-driven breast cancer.

Chemopreventive Effects of Propolis in the MNU-Induced Rat Mammary Tumor Model

Currently, one of the central problems in cancer management is the relapse of disease following conventional treatments, yet few therapeutic agents targeting resistance and tolerance exist. Propolis is known as a healing agent since ancient times. Therefore, over time, its curative properties have kept the interest of scientists, thus leading permanently to investigations of its other possible undiscovered effects. In this context, current experiments were performed to establish the chemopreventive potential of propolis extract (PE) (1.05 mg/kg BW/day) in N-methyl-N-nitrosourea- (MNU-) induced rat mammary tumors. MNU-inoculated/PE-treated rats had tumors of different physical attributes compared with control rats MNU-inoculated. The number of developed tumors (mean 49% versus 100%), incidence (mean 49% versus 100%), multiplicity (1.8 versus 3.7 (p<0.001)), tumor volume (mean 10 cm3 versus 16 cm3 (p<0.001)), and weight of the tumor mass (mean 7.42 g versus 9.00 g (p<0.05)) were noted. The numbers of grade I tumors recorded for MNU-inoculated rats were 24 (Group 1) and 7 (Group 2) for MNU-induced/PE-treated rats. In the serum of rats MNU-inoculated/PE-treated were found higher levels of antioxidative enzymes (SOD, CAT, and GPx) than in MNU-induced. Taken together, these data indicate that propolis could be a chemopreventive agent against MNU-induced mammary carcinogenesis.