Low-Dose Lung Radiation Therapy for COVID-19 Lung Disease: A Preclinical Efficacy Study in a Bleomycin Model of Pneumonitis.

PURPOSE: Low-dose whole lung radiation therapy (LDLR) has been proposed as a treatment for patients with acute respiratory distress syndrome associated with SARS-CoV-2 infection, and clinical trials are underway. There is an urgent need for preclinical evidence to justify this approach and inform dose, scheduling, and mechanisms of action. METHODS AND MATERIALS: Female C57BL/6 mice were treated with intranasal bleomycin sulfate (7.5 or 11.25 units/kg, day 0) and then exposed to whole lung radiation therapy (0.5, 1.0, or 1.5 Gy, or sham; day 3). Bodyweight was measured daily, and lung tissue was harvested for histology and flow cytometry on day 10. Computed tomography lung imaging was performed before radiation (day 3) and pre-endpoint (day 10). RESULTS: Bleomycin caused pneumonitis of variable severity, which correlated with weight loss. LDLR at 1.0 Gy was associated with a significant increase in the proportion of mice recovering to 98% of initial bodyweight, and a proportion of these mice exhibited less severe histopathologic lung changes. Mice experiencing moderate initial weight loss were more likely to respond to LDLR than those experiencing severe initial weight loss. In addition, LDLR (1.0 Gy) significantly reduced bleomycin-induced increases in interstitial macrophages, CD103+ dendritic cells (DCs), and neutrophil-DC hybrids. Overall, bleomycin-treated mice exhibited significantly higher percentages of nonaerated lung in left than right lungs, and LDLR (1.0 Gy) limited further reductions in aerated lung volume in right but not left lungs. LDLR at 0.5 and 1.5 Gy did not improve bodyweight, flow cytometric, or radiologic readouts of bleomycin-induced pneumonitis. CONCLUSIONS: Our data support the concept that LDLR can ameliorate acute inflammatory lung injury, identify 1.0 Gy as the most effective dose, and provide evidence that it is more effective in the context of moderate than severe pneumonitis. Mechanistically, LDLR at 1.0 Gy significantly suppressed bleomycin-induced accumulation of pulmonary interstitial macrophages, CD103+ DCs, and neutrophil-DC hybrids.

Stromal-immune cell crosstalk fundamentally alters the lung microenvironment following tissue insult.

Communication between stromal and immune cells is essential to maintain tissue homeostasis, mount an effective immune response and promote tissue repair. This 'crosstalk' occurs in both the steady state and following a variety of insults, for example, in response to local injury, at sites of infection or cancer. What do we mean by crosstalk between cells? Reciprocal activation and/or regulation occurs between immune and stromal cells, by direct cell contact and indirect mechanisms, including the release of soluble cytokines. Moving beyond cell-to-cell contact, this review investigates the complexity of 'cross-space' cellular communication. We highlight different examples of cellular communication by a variety of lung stromal and immune cells following tissue insults. This review examines how the 'geography of the lung microenvironment' is altered in various disease states; more specifically, we investigate how this influences lung epithelial cells and fibroblasts via their communication with immune cells and each other.

The Atypical Chemokine Receptor Ackr2 Constrains NK Cell Migratory Activity and Promotes Metastasis.

Chemokines have been shown to be essential players in a range of cancer contexts. In this study, we demonstrate that mice deficient in the atypical chemokine receptor Ackr2 display impaired development of metastasis in vivo in both cell line and spontaneous models. Further analysis reveals that this relates to increased expression of the chemokine receptor CCR2, specifically by KLRG1+ NK cells from the Ackr2-/- mice. This leads to increased recruitment of KLRG1+ NK cells to CCL2-expressing tumors and enhanced tumor killing. Together, these data indicate that Ackr2 limits the expression of CCR2 on NK cells and restricts their tumoricidal activity. Our data have important implications for our understanding of the roles for chemokines in the metastatic process and highlight Ackr2 and CCR2 as potentially manipulable therapeutic targets in metastasis.

Antigen-specific suppression of humoral immunity by anergic Ars/A1 B cells.

Autoreactive anergic B lymphocytes are considered to be dangerous because of their potential for activation and recruitment into autoimmune responses. However, they persist for days and constitute ∼5% of the B cell pool. We assessed their functional potential in the Ars/A1 transgene model, where anergic B cells express a dual-reactive Ag receptor that binds, in addition to a self-Ag, the hapten p-azophenylarsonate (Ars). When Ars/A1 B cells were transferred into adoptive recipients that were immunized with foreign proteins covalently conjugated with Ars, endogenous IgG immune responses to both were selectively and severely diminished, and the development of T helper cells was impaired. Approximately 95% inhibition of the anti-Ars response was attained with ∼4000 transferred Ars/A1 B cells through redundant mechanisms, one of which depended on their expression of MHC class II but not upon secretion of IL-10 or IgM. This Ag-specific suppressive activity implicates the autoreactive anergic B cell as an enforcer of immunological tolerance to self-Ags.

Vaccine adjuvants aluminum and monophosphoryl lipid A provide distinct signals to generate protective cytotoxic memory CD8 T cells.

Vaccines can greatly reduce the spread of and deaths from many infectious diseases. However, many infections have no successful vaccines. Better understanding of the generation of protective CD8 memory T cells by vaccination is essential for the rational design of new vaccines that aim to prime cellular immune responses. Here we demonstrate that the combination of two adjuvants that are currently licensed for use in humans can be used to prime long-lived memory CD8 T cells that protect mice from viral challenge. The universally used adjuvant, aluminum salts, primed long-lived memory CD8 T cells; however, effective cytotoxic T-cell differentiation occurred only in the presence of an additional adjuvant, monophosphoryl lipid A (MPL). MPL-induced IL-6 was required for cytotoxic differentiation. The IL-6 acted by inducing granzyme B production and reducing expression of inhibitory molecule PD1 on the surface of the primed CD8 T cells. CD8 memory T cells generated by antigen delivered with both aluminum salts and MPL provided significant protection from influenza A challenge. These adjuvants could be used in human vaccines to prime protective memory CD8 T cells.

Memory CD4 T cells that express CXCR5 provide accelerated help to B cells.

CD4 T cell help for B cells is critical for effective Ab responses. Although many of the molecules involved in helper functions of naive CD4 T cells have been characterized, much less is known about the helper capabilities of memory CD4 T cells, an important consideration for the design of vaccines that aim to prime protective memory CD4 T cells. In this study, we demonstrate that memory CD4 T cells enable B cells to expand more rapidly and class switch earlier than do primary responding CD4 T cells. This accelerated response does not require large numbers of memory cells, and similar numbers of primary responding cells provide less effective help than do memory cells. However, only memory CD4 T cells that express the B cell follicle homing molecule, CXCR5, are able to accelerate the response, suggesting that the rapidity of the Ab response depends on the ability of CD4 memory T cells to migrate quickly toward B cells.

CD4 memory T cells: what are they and what can they do?

Immunological memory provides the basis for successful vaccines. It is important to understand the properties of memory cells. There is much known about the phenotype and functions of memory CD8 T cells, less about memory B cells, while CD4 memory T cells have proved difficult to study. Differences in the types of memory CD4 cells studied and the difficulties of tracking the small number of cells have led to conflicting and unclear results. Here we discuss the different systems used to study CD4 memory cells and ask whether, and in what circumstances, memory CD4 cells could provide protection against infections.

Nedd4 augments the adaptive immune response by promoting ubiquitin-mediated degradation of Cbl-b in activated T cells.

Nedd4 and Itch are E3 ubiquitin ligases that ubiquitinate similar targets in vitro and thus are thought to function similarly. T cells lacking Itch show spontaneous activation and T helper type 2 polarization. To test whether loss of Nedd4 affects T cells in the same way, we generated Nedd4(+/+) and Nedd4(-/-) fetal liver chimeras. Nedd4(-/-) T cells developed normally but proliferated less, produced less interleukin 2 and provided inadequate help to B cells. Nedd4(-/-) T cells contained more of the E3 ubiquitin ligase Cbl-b, and Nedd4 was required for polyubiquitination of Cbl-b induced by CD28 costimulation. Our data demonstrate that Nedd4 promotes the conversion of naive T cells into activated T cells. We propose that Nedd4 and Itch ubiquitinate distinct target proteins in vivo.