CYRAD: a translational study assessing immune response to radiotherapy by photons or protons in postoperative head and neck cancer patients through circulating leukocyte subpopulations and cytokine levels.

BACKGROUND: Radiotherapy has both immunostimulant and immunosuppressive effects, particularly in radiation-induced lymphopenia. Proton therapy has demonstrated potential in mitigating this lymphopenia, yet the mechanisms by which different types of radiation affect the immune system function are not fully characterized. The Circulating Immunes Cells, Cytokines and Brain Radiotherapy (CYRAD) trial aims to compare the effects of postoperative X-ray and proton radiotherapy on circulating leukocyte subpopulations and cytokine levels in patients with head and neck (CNS and ear nose throat) cancer. METHODS: CYRAD is a prospective, non-randomized, single-center non interventional study assessing changes in the circulating leukocyte subpopulations and cytokine levels in head and neck cancer patients receiving X-ray or proton radiotherapy following tumor resection. Dosimetry parameters, including dose deposited to organs-at-risk such as the blood and cervical lymph nodes, are computed. Participants undergo 29 to 35 radiotherapy sessions over 40 to 50 days, followed by a 3-month follow-up. Blood samples are collected before starting radiotherapy (baseline), before the 11th (D15) and 30th sessions (D40), and three months after completing radiotherapy. The study will be conducted with 40 patients, in 2 groups of 20 patients per modality of radiotherapy (proton therapy and photon therapy). Statistical analyses will assess the absolute and relative relationship between variations (depletion, recovery) in immune cells, biomarkers, dosimetry parameters and early outcomes. DISCUSSION: Previous research has primarily focused on radiation-induced lymphopenia, paying less attention to the specific impacts of radiation on different lymphoid and myeloid cell types. Early studies indicate that X-ray and proton irradiation may lead to divergent outcomes in leukocyte subpopulations within the bloodstream. Based on these preliminary findings, this study aims to refine our understanding of how proton therapy can better preserve immune function in postoperative (macroscopic tumor-free) head and neck cancer patients, potentially improving treatment outcomes. PROTOCOL VERSION: Version 2.1 dated from January 18, 2023. TRIAL REGISTRATION: The CYRAD trial is registered from October 19, 2021, at the US National Library of Medicine, ClinicalTrials.gov ID NCT05082961.

Differential plasma cytokine variation following X-ray or proton brain irradiation using machine-learning approaches.

PURPOSE: X-ray and proton irradiation have been reported to induce distinct modifications in cytokine expression in vitro and in vivo, suggesting a dissimilar inflammatory response between X-rays and protons. We aimed to investigate the differences in cytokine profiles early following fractionated brain irradiation with X-rays or protons and their relationship with leukocyte subpopulations in rodents. MATERIALS AND METHODS: Our study utilized data from 80 tumor-free mice subjected to X-ray or proton brain irradiation in four fractions of 2.5Gy. Sixteen non-irradiated mice were used as the controls. Blood was collected 12h postirradiation to examine the profile of 13 cytokines. Correlation analysis, principal component analysis (PCA), and tree-based modeling were used to investigate the relationship between cytokine levels and leukocyte subpopulation variations following irradiation in the blood. RESULTS: Regardless of the irradiation type, brain irradiation resulted in a notable elevation in the plasma levels of IFN-γ and MCP-1. The use of either X-ray or proton beam had differential effect on plasma cytokine levels following brain irradiation. Specifically, X-ray irradiation was associated with significantly increased plasma levels of IFN-ß, IL-12p70, and IL-23, along with a decreased level of IL-1α, in comparison to proton irradiation. Correlation analysis revealed distinct cytokine regulatory patterns between X-ray and proton brain irradiation. PCA highlighted the association of MCP-1, IL-6, TNF-α, IL-17A, and IFN-γ with neutrophils, monocytes, and naïve T-cells following X-ray irradiation. TNF-α and IL-23 levels correlated with naïve CD4+-cells following proton irradiation. Tree-based models demonstrated that high TNF-α level resulted in an increase in naïve T-cells, neutrophils, and monocytes, whereas low IL-6 level was associated with decreases in these cell counts. CONCLUSION: Our findings revealed distinct inflammatory responses induced by X-ray irradiation in contrast to proton brain irradiation, as demonstrated by the differential regulation of cytokines in the bloodstream. Moreover, the study highlighted the association between specific cytokine levels and various leukocyte subpopulations. Further investigation is essential to accurately determine the impact of proton and X-ray brain irradiation on the inflammatory response and the efficacy of radiotherapy treatment.

Bayesian networks in modeling leucocyte interplay following brain irradiation: A comprehensive framework.

BACKGROUND AND OBJECTIVE: Understanding the intricate interactions among leucocyte subpopulations following radiotherapy is crucial for advancing cancer research and immunology. Recently, interest in recent radiotherapy modalities, such as protons, has increased. Herein, we present a framework utilizing Bayesian networks to uncover these complex relationships via an illustrative example of brain irradiation in rodents. METHODS: We utilized data from 96 healthy C57BL/6 adult mice subjected to either X-ray or proton brain irradiation. Leucocyte subpopulations in the blood collected 12 h after the final irradiated fraction were quantified. We employed Bayesian networks to detect causal interplay between physiological parameters, radiation variables and circulating leucocytes. The causal structure was learned via the use of the Bayesian information criterion as a scored criterion. Parameter estimation was performed to quantify the strength of the identified causal relationships. Cross-validation was used to validate our Bayesian network model's performance. RESULTS: In the X-ray model, we discovered previously undisclosed interactions between NK-cells and neutrophils, and between monocytes and T-CD4+ cells. The proton model revealed an interplay involving T-CD4+ cells and neutrophils. Both X-rays and protons led to heightened interactions between T-CD8+ cells and B cells, indicating their significant role in orchestrating immune responses. Additionally, the proton model displayed strengthened interactions between T-CD4+ and T-CD8+ cells, emphasizing a dynamic and coordinated immune response post-irradiation. Cross-validation results demonstrated the robustness of the Bayesian network model in explaining data uncertainty. CONCLUSION: The use of Bayesian networks as tools for causal structure discovery has revealed novel insights into the dynamics of immune responses to radiation exposure.

Revealing the effect of X-ray or proton brain irradiation on systemic inflammation and leukocyte subpopulation interplay in rodents.

The absolute lymphocyte count (ALC), lymphocyte-to-monocyte ratio (LMR), and neutrophil-to-lymphocyte ratio (NLR) offer convenient means to assess systemic inflammation post-cancer treatment, which influences treatment outcomes. Understanding these biomarker variations and leukocyte subpopulation interplay is crucial for optimizing radiotherapy. Herein, leukocyte subpopulations (T-CD4+, T-CD8+, B-cells, NK-cells, neutrophils, monocytes) during and after brain irradiation (using X-rays or Protons) in tumor-free mice were used to compute ALC, LMR, and NLR, on which radiation parameter influence was assessed by principal component analysis (PCA). NLR kinetics were further examined using modeling. Leukocyte subpopulations interplays and their response to radiation parameters were examined using PCA and correlation analysis. Under X-rays, ALC and LMR decreased, with ALC recovered to baseline after irradiation, but not LMR. Both X-rays and protons increased the NLR during irradiation, recovering in protons but not X-rays. Both irradiation volume and dose rate had a pronounced effect on the NLR. Leukocyte subpopulation interplay was observed under X-rays and protons, normalizing in the proton group by day 28. Lymphopenia was observed in all lymphocyte subpopulations under X-ray irradiation but not protons. The recovery patterns varied among the subpopulations. Neutrophil counts increased during irradiation, with the recovery of protons, but not X-rays, by day 28. Interplays between NK-cells and myeloid subpopulations were evident under X-rays but not protons. Importantly, no interplay was detected between myeloid cells and T/B-cells, indicating that LMR and NLR variations were primarily due to independent responses to brain irradiation. A tumor-free experimental mouse model was used to study the effects of brain radiotherapy on systemic immunity. When administering fractionated irradiation with a total dose of 20 Gy using a vertical beam to either the whole brain or hemi-brain, proton irradiation had fewer adverse impacts on the immune system compared to X-rays in tumor-free rodents.

Lymphocyte radiosensitivity: An extension to the linear-quadratic model?

BACKGROUND AND PURPOSE: The linear-quadratic (LQ) model has been pivotal for evaluating the effects of radiation on cells, but it is primarily characterized by linear responses, which has exhibited limitations when applied to lymphocyte data. The present research aims to address these limitations and to explore an alternative model extended from the conventional LQ model. MATERIALS AND METHODS: Literature providing lymphocyte counts from assays investigating apoptosis and survival after in vitro irradiation was selected. To address the nonlinearity in lymphocyte responses to radiation, we developed a saturation model characterized by a negative exponential relationship between radiation dose and cellular response. We compared the performance of this saturation model against that of conventional models, including the LQ model and its variants (linear model LM and linear-quadratic-cubic model LQC), as well as the repair-misrepair (RMR) model. The models were evaluated based on prediction-residual plots, residual standard errors, and the Akaike information criterion (AIC). We applied the saturation model to two additional datasets: (1) a dataset from the existing literature that assessed stimulated and unstimulated human lymphocytes exposed to gamma irradiation in vitro and (2) a novel dataset involving T lymphocytes from rodent spleens after exposure to various radiation types (X-rays and protons). RESULTS: The literature (n = 15 out of 2342) showed that lymphocyte apoptosis varies with dose, time and experimental conditions. The saturation model had a lower AIC of 718 compared to the LM, LQ, LQC and RMR models (AIC of 728, 720, 720 and 734, respectively). The saturation model had a lower residual error and more consistent error distribution. Integrating time as a covariate, the saturation model also had a better AIC for demonstrating time-dependent variations in lymphocyte responses after irradiation. For datasets involving unstimulated lymphocytes before irradiation, the saturation model provided a more accurate fit than did the LM, LQ, and RMR models. In these cases, the fit of the saturation model was comparable to that of the LQC model but offered an advantage when extrapolating to higher doses, where the LQC model might underestimate survival. For stimulated lymphocytes, which are radioresistant, all the models approximated the LM. Both the LQ and saturation models indicated greater radiosensitivity to protons in vitro. CONCLUSION: The new "saturation model" performed better than the LQ model in quantifying lymphocyte apoptosis and survival, estimating time dependency and assessing the role of radiation modalities or lymphocyte stimulation. Further experiments are warranted to experimentally explore the validity of the saturation model as a promising alternative in the clinical setting.

Early effects of different brain radiotherapy modalities on circulating leucocyte subpopulations in rodents.

PURPOSES: Lymphopenia is extensively studied, but not circulating leucocyte subpopulations, which however have distinct roles in tumor tolerance. Proton therapy has been shown to have a lesser impact on the immune system than conventional X-ray radiotherapy through lower dose exposure to healthy tissues. We explored the differential effects of brain X-ray and proton irradiation on circulating leucocyte subpopulations. MATERIALS AND METHODS: Leucocyte subpopulation counts from tumor-free mice were obtained 12 hours after 4 fractions of 2.5 Gy. The relationships between irradiation type (X-rays or protons), irradiated volume (whole-brain/hemi-brain) and dose rate (1 or 2 Gy/min) with circulating leucocyte subpopulations (T-CD4+, T-CD8+, B, and NK-cells, neutrophils, and monocytes) were investigated using linear regression and tree-based modeling approaches. Relationships between dose maps (brain, vessels, lymph nodes (LNs)) and leucocyte subpopulations were analyzed and applied to construct the blood dose model, assessing the hypothesis of a direct lymphocyte-killing effect in radiation-induced lymphopenia. RESULTS: Radiation-induced lymphopenia occurred after X-ray but not proton brain irradiation in lymphoid subpopulations (T-CD4+, T-CD8+, B, and NK-cells). There was an increase in neutrophil counts following protons but not X-rays. Monocytes remained unchanged under both X-rays and protons. Besides irradiation type, irradiated volume and dose rate had a significant impact on NK-cell, neutrophil and monocyte levels but not T-CD4+, T-CD8+, and B-cells. The dose to the blood had a heterogeneous impact on leucocyte subpopulations: neutrophil counts remained stable with increasing dose to the blood, while lymphocyte counts decreased with increasing dose (T-CD8+-cells > T-CD4+-cells > B-cells > NK-cells). Direct cell-killing effect of the dose to the blood mildly contributed to radiation-induced lymphopenia. LN exposure significantly contributed to lymphopenia and partially explained the distinct impact of irradiation type on circulating lymphocytes. CONCLUSIONS: Leucocyte subpopulations reacted differently to X-ray or proton brain irradiation. This difference could be partly explained by LN exposure to radiation dose. Further researches and analyses on other biological processes and interactions between leucocyte subpopulations are ongoing. The various mechanisms underlying leucocyte subpopulation changes under different irradiation modalities may have implications for the choice of radiotherapy modalities and their combination with immunotherapy in brain cancer treatment.

Hepatic abscess due to Candida species in neonates: Case reports in Vietnam.

Neonatal hepatic abscess (NHA) is a fatal condition in neonates. NHA can be caused by many organisms including bacteria, parasites, and fungi. Fungal NHA is a rare but troublesome cause in terms of diagnosis and treatment. We present three cases of fungal NHA caused by Candida. In these three cases, different underlying problems associated with NHA had been found.

Beyond lymphopenia, unraveling radiation-induced leucocyte subpopulation kinetics and mechanisms through modeling approaches.

Leucocyte subpopulations in both lymphoid and myeloid lineages have a significant impact on antitumor immune response. While radiation-induced lymphopenia is being studied extensively, radiation effects on lymphoid and myeloid subtypes have been relatively less addressed. Interactions between leucocyte subpopulations, their specific radiation sensitivity and the specific kinetics of each subpopulation can be modeled based on both experimental data and knowledge of physiological leucocyte depletion, production, proliferation, maturation and homeostasis. Modeling approaches of the leucocyte kinetics that may be used to unravel mechanisms underlying radiation induced-leucopenia and prediction of changes in cell counts and compositions after irradiation are presented in this review. The approaches described open up new possibilities for determining the influence of irradiation parameters both on a single-time point of acute effects and the subsequent recovery of leukocyte subpopulations. Utilization of these approaches to model kinetic data in post-radiotherapy states may be a useful tool for further development of new treatment strategies or for the combination of radiotherapy and immunotherapy.

Multisite light-induced phosphorylation of the transcription factor PIF3 is necessary for both its rapid degradation and concomitant negative feedback modulation of photoreceptor phyB levels in Arabidopsis.

Plants constantly monitor informational light signals using sensory photoreceptors, which include the phytochrome (phy) family (phyA to phyE), and adjust their growth and development accordingly. Following light-induced nuclear translocation, photoactivated phy molecules bind to and induce rapid phosphorylation and degradation of phy-interacting basic Helix Loop Helix (bHLH) transcription factors (PIFs), such as PIF3, thereby regulating the expression of target genes. However, the mechanisms underlying the signal-relay process are still not fully understood. Here, using mass spectrometry, we identify multiple, in vivo, light-induced Ser/Thr phosphorylation sites in PIF3. Using transgenic expression of site-directed mutants of PIF3, we provide evidence that a set of these phosphorylation events acts collectively to trigger rapid degradation of the PIF3 protein in response to initial exposure of dark-grown seedlings to light. In addition, we show that phyB-induced PIF3 phosphorylation is also required for the known negative feedback modulation of phyB levels in prolonged light, potentially through codegradation of phyB and PIF3. This mutually regulatory intermolecular transaction thus provides a mechanism with the dual capacity to promote early, graded, or threshold regulation of the primary, PIF3-controlled transcriptional network in response to initial light exposure, and later, to attenuate global sensitivity to the light signal through reductions in photoreceptor levels upon prolonged exposure.