Ionizing radiation reduces the capacity of activated macrophages to induce T-cell proliferation, but does not trigger dendritic cell-mediated non-targeted effects.

PURPOSE: Previous investigations revealed influences of irradiation up to 2Gy on the cytokine secretion profile of inflammatory and peritoneal mouse macrophages (pMФ). This raised the question if those alterations impact on dendritic cells and consecutive T-cell responses. Further, the impact of irradiation directly on pMФ capacity to induce T-cell responses was analyzed. MATERIALS AND METHODS: pMФ were LPS-activated, irradiated and the expression of activation markers was assessed. Treated pMФ were co-incubated with T-cells to investigate proliferation. To verify modulating properties of pMФ supernatants isolated 24 h after irradiation, bone marrow-derived dendritic cells (BMDC) were co-incubated with supernatants and activation markers as well as the BMDC-induced proliferation of T-cells were measured. RESULTS: pMФ showed a highly significantly decreased major histocompatibility complexII (MHCII) expression within a dose range from 0.7-2Gy. Further, the proliferation rate of cluster of differentiation 4+ (CD4+) T-cells was decreased after co-incubation particularly with 2 Gy irradiated pMФ. The co-incubation of BMDC with supernatants of activated, irradiated pMФ significantly reduced the CD40 expression, but did not impact on the BMDC-derived induction of T-cell proliferation. CONCLUSIONS: Inflammatory macrophages being exposed to irradiation have the potential to modulate consecutive adaptive immune reactions. But supernatants of irradiated macrophages do not influence the dendritic cells (DC)-mediated induction of T cell proliferation.

Low-Dose Radiotherapy Ameliorates Advanced Arthritis in hTNF-α tg Mice by Particularly Positively Impacting on Bone Metabolism.

Inflammation and bone erosion are central in rheumatoid arthritis (RA). Even though effective medications for control and treatment of RA are available, remission is only seen in a subset of patients. Treatment with low-dose radiotherapy (LD-RT) which has been already successfully used for amelioration of symptoms in benign diseases should be a promising approach to reduce pain, inflammation, and particularly bone erosion in patients with RA. Even though anti-inflammatory effects of LD-RT are already described with non-linear dose response relationships, and pain-reducing effects have been clinically observed, the underlying mechanisms are widely unknown. Besides immune cells many other cell types, such as fibroblast-like synoviocytes (FLS), osteoclasts, and osteoblast are present in the affected joint and might be modulated by LD-RT. For this study, these cell types were obtained from human tumor necrosis factor-α transgenic (hTNF-α tg) mice and were consecutively exposed to different doses of ionizing radiation (0.1, 0.5, 1.0, and 2.0 Gy, respectively) in vitro. In order to study the in vivo effects of LD-RT within the arthritic joint, hind paws of arthritic hTNF-α tg mice were locally irradiated with 0.5 Gy, a single dose per fraction that is known for good clinical responses. Starting at a dose of 0.5 Gy, proliferation of FLS was reduced and apoptosis significantly enhanced with no changes in necrosis. Further, expression of RANK-L was slightly reduced following irradiation with particularly 0.5 Gy. Starting from 0.5 Gy, the numbers of differentiated osteoclasts were significantly reduced, and a lower bone resorbing activity of treated osteoclasts was also observed, as monitored via pit formation and Cross Laps presence. LD-RT had further a positive effect on osteoblast-induced mineralization in a discontinuous dose response relationship with 0.5 Gy being most efficient. An increase of the gene expression ratio of OPG/RANK-L at 0.1 and 0.5 Gy and of production of OPG at 0.5 and 1.0 Gy was observed. In vivo, LD-RT resulted in less severe arthritis in arthritic hTNF-α tg mice and in significant reduction of inflammatory and erosive area with reduced osteoclasts and neutrophils. Locally applied LD-RT can, therefore, induce a beneficial micro-environment within arthritic joints by predominantly positively impacting on bone metabolism.

Decrease of Markers Related to Bone Erosion in Serum of Patients with Musculoskeletal Disorders after Serial Low-Dose Radon Spa Therapy.

Musculoskeletal disorders (MSDs) are the most frequent cause of disability in Europe. Reduced mobility and quality of life of the patients are often associated with pain due to chronic inflammation. The inflammatory process, accompanied by a destruction of the cartilage and bone tissue, is discussed as a result of (A) the infiltration of immune cells into the joints, (B) an altered homeostasis of the joint cavity (synovium) with a critical role of bone remodeling cells, and (C) release of inflammatory factors including adipokines in the arthritic joint. In addition to the classical medication, low-dose radiation therapy using photons or radon spa treatments has shown to reduce pain and improve the mobility of the patients. However, the cellular and molecular mechanisms of anti-inflammatory effects of radon are yet poorly understood. We analyzed blood and serum samples from 32 patients, suffering from MSDs, who had been treated in the radon spa in Bad Steben (Germany). Before and after therapy, we measured the levels of markers related to bone metabolism (collagen fragments type-1, cartilage oligomeric matrix protein, receptor activator of NFκB ligand, and osteoprotegerin) in the serum of patients. In addition, adipokines related to inflammation (visfatin, leptin, resistin, and adiponectin) were analyzed. Some of these factors are known to correlate with disease activity. Since T cells play an important role in the progression of the disease, we further analyzed in blood samples the frequency of pro- and anti-inflammatory T cell subpopulations (CD4+IL17+ T cells and CD4+FoxP3+ regulatory T cells). Overall, we found a decrease of collagen fragments (CTX-I), indicating decreased bone resorption, presumably by osteoclasts, in the serum of MSD patients. We also observed reduced levels of visfatin and a consistent trend toward an increase of regulatory T cells in the peripheral blood, both indicating attenuation of inflammation. However, key proteins of bone metabolism were unchanged on a systemic level, suggesting that these factors act locally after radon spa therapy of patients with MSDs.

Primary glioblastoma multiforme tumors and recurrence : Comparative analysis of the danger signals HMGB1, HSP70, and calreticulin.

PURPOSE: Glioblastoma multiforme (GBM) is the most common and aggressive brain tumor. Despite improved multimodal therapies, the tumor recurs in most cases. Diverging patient survival suggests great tumor heterogeneity and different therapy responses. Danger signals such as high-mobility group box protein 1 (HMGB1), heat shock protein 70 (HSP70), and calreticulin (CRT) are biomarker candidates, due to their association with tumor progression versus induction of antitumor immune responses. Overexpression of these danger signals has been reported for various types of tumors; however, their role in GBM is still elusive. A direct comparison of their expression in the primary tumor versus the corresponding relapse is still lacking for most tumor entities. PATIENTS AND METHODS: We therefore performed an expression analysis by immunohistochemistry of the danger signals HMGB1, HSP70, and CRT in primary tumors and the corresponding relapses of 9 patients with de novo GBM. RESULTS: HMGB1 was highly expressed in primary tumors with a significant reduction in the respective relapse. The extracellular HSP70 expression was significantly increased in the relapse compared to the primary tumor. CRT was generally highly expressed in the primary tumor, with a slight increase in the relapse. CONCLUSION: The combination of a decreased expression of HMGB1, an increased expression of extracellular HSP70, and an increased expression of CRT in the relapse seems to be beneficial for patient survival. HMGB1, extracellular HSP70, and CRT could be taken into concerted consideration as potential biomarkers for the prognosis of patients with GBM.

Reduced secretion of the inflammatory cytokine IL-1ß by stimulated peritoneal macrophages of radiosensitive Balb/c mice after exposure to 0.5 or 0.7 Gy of ionizing radiation.

Since the beginning of the 20th century, low dose radiotherapy (LD-RT) has been practiced and established as therapy of inflammatory diseases. Several clinical studies already have proven the anti-inflammatory effect of low doses of ionizing irradiation (LDR). However, further research is inevitable to reveal the underlying immune-biological mechanisms. Focus has been set on the modulation of activated macrophages by LDR, since they participate in both, initiation and resolution of inflammation. Here we examined with an ex vivo peritoneal mouse macrophage model how LDR modulates the secretion of the inflammatory cytokines IL-1ß and TNF-α by activated macrophages and whether the basal radiosensitivity of the immune cells has influence on it. Peritoneal macrophages of Balb/c mice responded to exposure of 0.5 or 0.7 Gy of ionizing irradiation (X-ray) with significant decreased release of IL-1ß and slightly, but not significantly, reduced release of TNF-α. Macrophages of the less radiosensitive C57BL/6 mice did not show this anti-inflammatory reaction. This was observed in both wild type and human TNF-α transgenic animals with C57BL/6 background. We conclude that only the inflammatory phenotype of more radiosensitive macrophages is reduced by LDR and that ex vivo and in vivo models with primary cells should be applied to examine how the immune system is modulated by LDR.

How does ionizing irradiation contribute to the induction of anti-tumor immunity?

Radiotherapy (RT) with ionizing irradiation is commonly used to locally attack tumors. It induces a stop of cancer cell proliferation and finally leads to tumor cell death. During the last years it has become more and more evident that besides a timely and locally restricted radiation-induced immune suppression, a specific immune activation against the tumor and its metastases is achievable by rendering the tumor cells visible for immune attack. The immune system is involved in tumor control and we here outline how RT induces anti-inflammation when applied in low doses and contributes in higher doses to the induction of anti-tumor immunity. We especially focus on how local irradiation induces abscopal effects. The latter are partly mediated by a systemic activation of the immune system against the individual tumor cells. Dendritic cells are the key players in the initiation and regulation of adaptive anti-tumor immune responses. They have to take up tumor antigens and consecutively present tumor peptides in the presence of appropriate co-stimulation. We review how combinations of RT with further immune stimulators such as AnnexinA5 and hyperthermia foster the dendritic cell-mediated induction of anti-tumor immune responses and present reasonable combination schemes of standard tumor therapies with immune therapies. It can be concluded that RT leads to targeted killing of the tumor cells and additionally induces non-targeted systemic immune effects. Multimodal tumor treatments should therefore tend to induce immunogenic tumor cell death forms within a tumor microenvironment that stimulates immune cells.