Radiofrequency radiation reshapes tumor immune microenvironment into antitumor phenotype in pulmonary metastatic melanoma by inducing active transformation of tumor-infiltrating CD8+ T and NK cells.

Immunosuppression by the tumor microenvironment is a pivotal factor contributing to tumor progression and immunotherapy resistance. Priming the tumor immune microenvironment (TIME) has emerged as a promising strategy for improving the efficacy of cancer immunotherapy. In this study we investigated the effects of noninvasive radiofrequency radiation (RFR) exposure on tumor progression and TIME phenotype, as well as the antitumor potential of PD-1 blockage in a model of pulmonary metastatic melanoma (PMM). Mouse model of PMM was established by tail vein injection of B16F10 cells. From day 3 after injection, the mice were exposed to RFR at an average specific absorption rate of 9.7 W/kg for 1 h per day for 14 days. After RFR exposure, lung tissues were harvested and RNAs were extracted for transcriptome sequencing; PMM-infiltrating immune cells were isolated for single-cell RNA-seq analysis. We showed that RFR exposure significantly impeded PMM progression accompanied by remodeled TIME of PMM via altering the proportion and transcription profile of tumor-infiltrating immune cells. RFR exposure increased the activation and cytotoxicity signatures of tumor-infiltrating CD8+ T cells, particularly in the early activation subset with upregulated genes associated with T cell cytotoxicity. The PD-1 checkpoint pathway was upregulated by RFR exposure in CD8+ T cells. RFR exposure also augmented NK cell subsets with increased cytotoxic characteristics in PMM. RFR exposure enhanced the effector function of tumor-infiltrating CD8+ T cells and NK cells, evidenced by increased expression of cytotoxic molecules. RFR-induced inhibition of PMM growth was mediated by RFR-activated CD8+ T cells and NK cells. We conclude that noninvasive RFR exposure induces antitumor remodeling of the TIME, leading to inhibition of tumor progression, which provides a promising novel strategy for TIME priming and potential combination with cancer immunotherapy.

mTORC2 acts as a gatekeeper for mTORC1 deficiency-mediated impairments in ILC3 development.

Group 3 innate lymphoid cells (ILC3s) are mediators of intestinal immunity and barrier function. Recent studies have investigated the role of the mammalian target of rapamycin complex (mTOR) in ILC3s, whereas the mTORC1-related mechanisms and crosstalk between mTORC1 and mTORC2 involved in regulating ILC3 homeostasis remain unknown. In this study, we found that mTORC1 but not mTORC2 was critical in ILC3 development, IL-22 production, and ILC3-mediated intestinal homeostasis. Single-cell RNA sequencing revealed that mTORC1 deficiency led to disruption of ILC3 heterogeneity, showing an increase in differentiation into ILC1-like phenotypes. Mechanistically, mTORC1 deficiency decreased the expression of NFIL3, which is a critical transcription factor responsible for ILC3 development. The activities of both mTORC1 and mTORC2 were increased in wild-type ILC3s after activation by IL-23, whereas inhibition of mTORC1 by Raptor deletion or rapamycin treatment resulted in increased mTORC2 activity. Previous studies have demonstrated that S6K, the main downstream target of mTORC1, can directly phosphorylate Rictor to dampen mTORC2 activity. Our data found that inhibition of mTORC1 activity by rapamycin reduced Rictor phosphorylation in ILC3s. Reversing the increased mTORC2 activity via heterozygous or homozygous knockout of Rictor in Raptor-deleted ILC3s resulted in severe ILC3 loss and complete susceptibility to intestinal infection in mice with mTORC1 deficiency (100% mortality). Thus, mTORC1 acts as a rheostat of ILC3 heterogeneity, and mTORC2 protects ILC3s from severe loss of cells and immune activity against intestinal infection when mTORC1 activity is diminished.

Natural killer cell homing and trafficking in tissues and tumors: from biology to application.

Natural killer (NK) cells, a subgroup of innate lymphoid cells, act as the first line of defense against cancer. Although some evidence shows that NK cells can develop in secondary lymphoid tissues, NK cells develop mainly in the bone marrow (BM) and egress into the blood circulation when they mature. They then migrate to and settle down in peripheral tissues, though some special subsets home back into the BM or secondary lymphoid organs. Owing to its success in allogeneic adoptive transfer for cancer treatment and its "off-the-shelf" potential, NK cell-based immunotherapy is attracting increasing attention in the treatment of various cancers. However, insufficient infiltration of adoptively transferred NK cells limits clinical utility, especially for solid tumors. Expansion of NK cells or engineered chimeric antigen receptor (CAR) NK cells ex vivo prior to adoptive transfer by using various cytokines alters the profiles of chemokine receptors, which affects the infiltration of transferred NK cells into tumor tissue. Several factors control NK cell trafficking and homing, including cell-intrinsic factors (e.g., transcriptional factors), cell-extrinsic factors (e.g., integrins, selectins, chemokines and their corresponding receptors, signals induced by cytokines, sphingosine-1-phosphate (S1P), etc.), and the cellular microenvironment. Here, we summarize the profiles and mechanisms of NK cell homing and trafficking at steady state and during tumor development, aiming to improve NK cell-based cancer immunotherapy.

Prenatal inflammation exposure-programmed hypertension exhibits multi-generational inheritance via disrupting DNA methylome.

The multi-generation heredity trait of hypertension in human has been reported, but the molecular mechanisms underlying multi-generational inheritance of hypertension remain obscure. Recent evidence shows that prenatal inflammatory exposure (PIE) results in increased incidence of cardiovascular diseases, including hypertension. In this study we investigated whether and how PIE contributed to multi-generational inheritance of hypertension in rats. PIE was induced in pregnant rats by intraperitoneal injection of LPS or Poly (I:C) either once on gestational day 10.5 (transient stimulation, T) or three times on gestational day 8.5, 10.5, and 12.5 (persistent stimulation, P). Male offspring was chosen to study the paternal inheritance. We showed that PIE, irrespectively induced by LPS or Poly (I:C) stimulation during pregnancy, resulted in multi-generational inheritance of significantly increased blood pressure in rat descendants, and that prenatal LPS exposure led to vascular remodeling and vasoconstrictor dysfunction in both thoracic aorta and superior mesenteric artery of adult F2 offspring. Furthermore, we revealed that PIE resulted in global alteration of DNA methylome in thoracic aorta of F2 offspring. Specifically, PIE led to the DNA hypomethylation of G beta gamma (Gßγ) signaling genes in both the F1 sperm and the F2 thoracic aorta, and activation of PI3K/Akt signaling was implicated in the pathologic changes and dysregulated vascular tone of aortic tissue in F2 LPS-P offspring. Our data demonstrate that PIE reprogrammed DNA methylome of cells from the germline/mature gametes contributes to the development of hypertension in F2 PIE offspring. This study broadens the current knowledge regarding the multi-generation effect of the cumulative early life environmental factors on the development of hypertension.

[Prenatal exposure to lipopolysaccharide results in vascular renin-angiotensin system dysfunction in offspring rats].

OBJECTIVE: To explore the impact of prenatal exposure to lipopolysaccharide on renin-angiotensin system of offspring rats. METHODS: Six pregnant SD rats were randomly divided into 2 groups. The rats in the lipopolysaccharide (LPS) group were treated with LPS 0.79 mg/kg (i.p.) on the 8th, 10th and 12th day of gestation, and rats in the control group were treated with saline at the same time points. The blood pressure of offspring rats was measured by the tail cuff method. Protein expression of Angiotensin II (AngII) in thoracic aorta vessel was determined by immunohistochemistry. Protein expressions of AngII type 1 and type 2 receptor in thoracic aorta vessel were detected by Western blot. RESULTS: Blood pressure of 12-week-old offspring rats of LPS group was significantly higher than that of 12-week-old offspring rats of control group (P < 0.01). The protein expression of AngII and AngII type 1 receptor in thoracic aorta vessel were significantly higher while protein expression of AngII type 2 receptor was lower in 15-week-old offspring rats of LPS group than in control group, resulting in a significant increase in the ratio of AngII type 1 receptor/AngII type 2 receptor in the aorta at 15-week-old of offspring rats than in 15-week-old offspring rats of control group (P < 0.01). CONCLUSION: Prenatal lipopolysaccharide exposure results vascular renin-angiotensin system dysfunction, which may play an important role on the pathogenesis of hypertension development in offspring rats.