In Vitro and In Vivo Analysis of Extracellular Vesicle-Mediated Metastasis Using a Bright, Red-Shifted Bioluminescent Reporter Protein.

Cancer cells produce heterogeneous extracellular vesicles (EVs) as mediators of intercellular communication. This study focuses on a novel method to image EV subtypes and their biodistribution in vivo. A red-shifted bioluminescence resonance energy transfer (BRET) EV reporter is developed, called PalmReNL, which allows for highly sensitive EV tracking in vitro and in vivo. PalmReNL enables the authors to study the common surface molecules across EV subtypes that determine EV organotropism and their functional differences in cancer progression. Regardless of injection routes, whether retro-orbital or intraperitoneal, PalmReNL positive EVs, isolated from murine mammary carcinoma cells, localized to the lungs. The early appearance of metastatic foci in the lungs of mammary tumor-bearing mice following multiple intraperitoneal injections of the medium and large EV (m/lEV)-enriched fraction derived from mammary carcinoma cells is demonstrated. In addition, the results presented here show that tumor cell-derived m/lEVs act on distant tissues through upregulating LC3 expression within the lung.

Immunoadjuvant therapy in sepsis: novel strategies for immunosuppressive sepsis coming down the pike.

Recent efforts have focused on immunoadjuvant therapies for sepsis. The host inflammatory response consequent to initial exposure to pathogens is often followed by anti-inflammatory forces, resulting in increased morbidity and mortality in such critically ill patients. In the subacute stage of sepsis, apoptosis (type I programmed cell death) and subsequently autophagy (type II programmed cell death) have been attracting recent research interest. Although many patients may die during the initial cytokine storm, those who survive this phase might acquire defining characteristics of profound immunosuppression, including failure to clear the primary infection, development of secondary opportunistic infections, and reactivation of latent viruses. Both types of cell death are currently thought to be associated with this subacute immunosuppressive phase of sepsis, and acceleration of autophagy might alleviate immunosuppression through regulation of apoptosis of key immune effector cells. Programmed cell death 1 (PD-1) and its corresponding ligand play a major pathological role in immunosuppression not only in cancer but also in sepsis. Positive costimulatory pathways in T cells, such as CD28 signaling, permit the effector T cell to expand, persist, and effectively clear antigen. However, PD-1 is a negative costimulatory pathway on T cells that broadly enhances immunosuppressive signals across the innate and adaptive immune system. To counter this immunosuppression in sepsis, checkpoint blockade has garnered attention in an area of clinical research. In this review, we introduce some approaches of immunotherapy using anti-PD-1 antibody in infectious diseases and share our future perspectives.

Restoration of T Cell function in multi-drug resistant bacterial sepsis after interleukin-7, anti-PD-L1, and OX-40 administration.

BACKGROUND: Multidrug resistant (MDR) bacterial pathogens are a serious problem of increasing importance facing the medical community. MDR bacteria typically infect the most immunologically vulnerable: patients in intensive care units, patients with extensive comorbidities, oncology patients, hemodialysis patients, and other immune suppressed individuals are likely to fall victim to these pathogens. One promising novel approach to treatment of MDR bacteria is immuno-adjuvant therapy to boost patient immunity. Success with this strategy would have the major benefit of providing protection against a number of MDR pathogens. OBJECTIVES: This study had two main objectives. First, immunophenotyping of peripheral blood mononuclear cells from patients with sepsis associated with MDR bacteria was performed to examine for findings indicative of immunosuppression. Second, the ability of three immuno-adjuvants with distinct mechanisms of action to reverse CD4 and CD8 T cell dysfunction, a pathophysiological hallmark of sepsis, was evaluated. RESULTS: Septic patients with MDR bacteria had increased expression of the inhibitory receptor PD-1 and its ligand PD-L1 and decreased monocyte HLA-DR expression compared to non-septic patients. All three immuno-adjuvants, IL-7, anti-PD-L1, and OX-40L, increased T cell production of IFN-γ in a subset of septic patients with MDR bacteria: IL-7 was most efficacious. There was a strong trend toward increased mortality in patients whose T cells failed to increase IFN-γ production in response to the three treatments. CONCLUSION: Immuno-adjuvant therapy reversed T cell dysfunction, a key pathophysiological mechanism in septic patients with MDR bacteria.