Dissecting the mechanism of cytokine release induced by T-cell engagers highlights the contribution of neutrophils.

T cell engagers represent a novel promising class of cancer-immunotherapies redirecting T cells to tumor cells and have some promising outcomes in the clinic. These molecules can be associated with a mode-of-action related risk of cytokine release syndrome (CRS) in patients. CRS is characterized by the rapid release of pro-inflammatory cytokines such as TNF-α, IFN-γ, IL-6 and IL-1ß and immune cell activation eliciting clinical symptoms of fever, hypoxia and hypotension. In this work, we investigated the biological mechanisms triggering and amplifying cytokine release after treatment with T cell bispecific antibodies (TCBs) employing an in vitro co-culture assay of human PBMCs or total leukocytes (PBMCs + neutrophils) and corresponding target antigen-expressing cells with four different TCBs. We identified T cells as the triggers of the TCB-mediated cytokine cascade and monocytes and neutrophils as downstream amplifier cells. Furthermore, we assessed the chronology of events by neutralization of T-cell derived cytokines. For the first time, we demonstrate the contribution of neutrophils to TCB-mediated cytokine release and confirm these findings by single-cell RNA sequencing of human whole blood incubated with a B-cell depleting TCB. This work could contribute to the construction of mechanistic models of cytokine release and definition of more specific molecular and cellular biomarkers of CRS in the context of treatment with T-cell engagers. In addition, it provides insight for the elaboration of prophylactic mitigation strategies that can reduce the occurrence of CRS and increase the therapeutic index of TCBs.

Supplementation with Bifidobacterium longum Bar33 and Lactobacillus helveticus Bar13 mixture improves immunity in elderly humans (over 75 years) and aged mice.

OBJECTIVE: Aging induces several physiologic and immune changes. The usefulness of probiotics in ameliorating age-related disorders remains largely unexplored. The aim of this study was to evaluate the effectiveness of a Bifidobacterium longum Bar33 and Lactobacillus helveticus Bar13 mixture in improving the physiologic status and immunity of older adults (over 75 years). Furthermore, the possible role of such mixture in ameliorating gut immunity in aged mice was investigated. METHODS: A randomized, double-blind, placebo-controlled trial was conducted with 98 adults (84.6 ± 7.8 y), supplemented for 30 d with a biscuit containing a probiotic mixture of B. longum Bar33 and L. helveticus Bar13 (1:1), or no probiotics, as placebo. Blood was collected for analysis of biochemical parameters, lymphocyte subpopulations, natural killer activity, and cytokine release. Aged Balb/c mice received the same probiotic mixture or placebo daily for 28 d, then blood and intestinal lymphocyte subpopulations were analyzed. RESULTS: The probiotic mixture ameliorated immune response in older adults by increasing naive, activated memory, regulatory T cells, B cells, and natural killer activity and decreasing memory T cells compared with placebo (P < 0.05). The biochemical parameters did not change after probiotic supplementation. In the gut of old mice, the two probiotics modulated cells crucial for gut immune homeostasis by increasing regulatory T (Treg and Tr1) and decreasing γδ T cells compared with control mice (P < 0.05). In addition, B cells increased in the gut and blood of probiotic-treated mice. CONCLUSION: Results from the present study data indicated that B. longum Bar33 and L. helveticus Bar13 improve immune function at intestinal and peripheral sites in aging.

A Consensus Proposal for Nutritional Indicators to Assess the Sustainability of a Healthy Diet: The Mediterranean Diet as a Case Study.

BACKGROUND: There is increasing evidence of the multiple effects of diets on public health nutrition, society, and environment. Sustainability and food security are closely interrelated. The traditional Mediterranean Diet (MD) is recognized as a healthier dietary pattern with a lower environmental impact. As a case study, the MD may guide innovative inter-sectorial efforts to counteract the degradation of ecosystems, loss of biodiversity, and homogeneity of diets due to globalization through the improvement of sustainable healthy dietary patterns. This consensus position paper defines a suite of the most appropriate nutrition and health indicators for assessing the sustainability of diets based on the MD. METHODS: In 2011, an informal International Working Group from different national and international institutions was convened. Through online and face-to-face brainstorming meetings over 4 years, a set of nutrition and health indicators for sustainability was identified and refined. RESULTS: Thirteen nutrition indicators of sustainability relating were identified in five areas. Biochemical characteristics of food (A1. Vegetable/animal protein consumption ratios; A2. Average dietary energy adequacy; A3. Dietary Energy Density Score; A4. Nutrient density of diet), Food Quality (A5. Fruit and vegetable consumption/intakes; A6. Dietary Diversity Score), Environment (A7. Food biodiversity composition and consumption; A8. Rate of Local/regional foods and seasonality; A9. Rate of eco-friendly food production and/or consumption), Lifestyle (A10. Physical activity/physical inactivity prevalence; A11. Adherence to the Mediterranean dietary pattern), Clinical Aspects (A12. Diet-related morbidity/mortality statistics; A13. Nutritional Anthropometry). A standardized set of information was provided for each indicator: definition, methodology, background, data sources, limitations of the indicator, and references. CONCLUSION: The selection and analysis of these indicators has been performed (where possible) with specific reference to the MD. Sustainability of food systems is an urgent priority for governments and international organizations to address the serious socioeconomic and environmental implications of short-sighted and short-term practices for agricultural land and rural communities. These proposed nutrition indicators will be a useful methodological framework for designing health, education, and agricultural policies in order, not only to conserve the traditional diets of the Mediterranean area as a common cultural heritage and lifestyle but also to enhance the sustainability of diets in general.

Investigations of the effects of the antimalarial drug dihydroartemisinin (DHA) using the Frog Embryo Teratogenesis Assay-Xenopus (FETAX).

Artemisinin derivatives are effective and safe drugs for treating malaria, but they are not recommended during the first trimester of pregnancy because of resorptions and abnormalities observed in animal reproduction studies. Previous studies in rats showed that artemisinin embryotoxicity derives from the depletion of primitive red blood cells (RBCs) over a narrow critical time window (gestation Days 9-14). In order to further investigate the susceptibility of primitive RBCs to artemisinins and to establish whether this susceptibility is species-specific or inherent to the compound, we studied dihydroartemisinin (DHA), both a drug in its own right and the main metabolite of current artemisinin derivatives in use, in the Frog Embryo Teratogenesis Assay-Xenopus (FETAX). This model readily allows investigation and monitoring of primitive and definitive RBCs. Effects on frog larvae exposed to DHA for 48 h during early embryonic development, starting from 24 h post fertilization, were similar to those on rat embryos in terms of reduction in the number of primitive RBCs (clonally produced within the ventral blood island). In contrast, RBCs of older larvae (stage 47, produced at the definitive sites of hematopoiesis) were affected minimally and subsequently recovered. Compared to rat embryos, the frog larvae had no areas of necrosis but they shared similar heart defects. The mitochondrion appeared to be the main subcellular target, similar to observations in Plasmodium. These results implicate artemisinin-induced embryotoxicity through perturbation of metabolically active RBCs; whereas this mode of action does not appear to be species-specific, the stages of susceptibility varied between different species. The window of susceptibility and duration of exposure must be considered to evaluate the clinical relevance of these findings.

In vivo and in vitro investigations of the effects of the antimalarial drug dihydroartemisinin (DHA) on rat embryos.

Artemisinin derivatives are clinically effective and safe antimalarials, but are not recommended during the first trimester of pregnancy because of the resorptions and abnormalities seen in animal reproduction studies. Understanding how, when and what toxicity occurs is crucial to any assessment of clinical relevance. Previously, DHA has been shown in the rat whole embryo culture (WEC) to primarily affect primitive red blood cells (RBCs) causing subsequent tissue damage and dysmorphogenesis. To verify the primary target of DHA in vivo and to detect consequences induced by early damage on embryo development, pregnant female rats were orally treated on gestation days (GD) 9.5 and 10.5 with 7.5 or 15 mg/kg/day DHA and caesarean sectioned on GD11.5, 12.5, 13.5, 15 and 20. A parallel in vitro WEC study evaluated the role of oxidative damage and examined blood islands and primitive RBCs. In accordance with the WEC results, primitive RBCs from yolk sac hematopoiesis were the target of DHA in vivo. The resulting anemia led to cell damage, which depending on its degree, was either diffuse or focal. Embryonic response to acute anemia varied from complete recovery to malformation and death, depending on the extent of cell death. Malformations occurred only in litters with embryonic deaths. DHA induced low glutathione levels in RBCs, indicating that oxidative stress may be involved in artemisinin toxicity; effects were extremely rapid, with altered RBCs seen as early as GD10. In establishing the relevance of these findings to humans, one should consider differences in the development of rodents and humans. While yolk sac hematopoiesis occurs similarly in the two species, early placentation and extent of exposure differ. In particular, early hematopoiesis takes only 7 days in rats (during which RBCs expand in a clonal fashion) compared with 6 weeks in humans; thus the susceptible period in relation to the duration of exposure to an artemisinin-based treatment may be substantially different.