Encyclopedia
Please note this is a comparison between Version 2 by Yvaine Wei and Version 1 by Valentin Djonov.
Macrophages are some of the first cells recruited to sites of radiation-induced injury where they can aid in tissue repair, propagate radiation-induced fibrogenesis and influence tumour dynamics. Radiation therapy (RT) is a vital component of multimodal cancer treatment, and its immunomodulatory effects are a major focus of current therapeutic strategies. Microbeam radiation therapy (MRT) is a unique, spatially fractionated radiation modality that has demonstrated exceptional tumour control and reduction in normal tissue toxicity, including fibrosis.
- microbeam radiotherapy
- DNA damage
- macrophages
- infiltration
1. Introduction
Despite recent advances in cancer therapy, the prognosis remains poor in many clinical cases. Radiotherapy (RT) has been the subject of technological innovation over the years as an essential component of treatment protocols for over 50% of all cancer patients [1]. Despite recent advances, the capabilities of RT in tumour control are still limited by normal tissue thresholds for radiation toxicity. In some cases, this prevents the appropriate dose escalation necessary to overcome radioresistance and limits its use for treating malignancies in particularly sensitive organs. Advances in the field of RT are therefore aimed at innovating the current clinical methodology in order to improve therapeutic outcomes.
Synchrotron microbeam radiation therapy (MRT) is a novel, pre-clinical RT modality that combines micrometric spatial fractionation and FLASH dose-rates to maximize the therapeutic ratio. Synchrotron-generated X-rays pass through a collimator to produce micro-planar beams of radiation at very high doses (up to thousands of Gray (Gy)) and dose-rates (up to 16,000 Gy/s). Microbeam widths range from 20 to 100 µm, while the spacing between them ranges from 50 to 500 µm. MRT provides enhanced tumour control relative to conventional RT, as the spatial fractionation and ultra-high dose rate allows for substantial dose escalation with significantly reduced collateral damage to normal tissues within the irradiation field [2,3][2][3]. The mechanisms behind the efficacy of MRT have not been fully elucidated, but some data have implicated macrophages as an important player in its therapeutic response [4,5,6,7][4][5][6][7].
Macrophages have an important pathophysiological role in normal tissue responses after radiation injury, a type of sterile injury. Studies have shown that ionizing radiation induces immunomodulatory effects [8] which can be either tolerogenic or immunogenic [9[9][10],10], depending on the distinct downstream signalling mechanisms that influence tissue repair [11,12][11][12]. Such responses involve a complex cascade of events that depend on a variety of factors, including the dose and fractionation, target site and volume of tissue irradiated. The type of cell death mechanisms induced by RT also influences the nature of the immune response [13]. The production of damage-associated molecular patterns (DAMPs) from irradiated cells elicits innate immune activation [12], with macrophages being one of the first immune cells to respond to sites of radiation-induced injury [14,15,16,17][14][15][16][17].
Normal tissue responses are particularly important to understand in order to avoid treatment-induced organ dysfunction. For example, the lung and liver are two of the most sensitive organs in terms of long-terms effects (e.g., radiation-induced fibrosis) and are often associated with collateral radiation toxicity in clinical RT regimens. Following conventional radiation of the lung, dynamic changes in immune cell populations at the site of irradiation are observed both in the parenchyma and alveoli [14]. Similarly, the liver is susceptible to radiation-induced disease and long-term toxicities [18]. Furthermore, all external irradiation regimens involve skin penetration and, therefore, the risk of cutaneous radiation injury. Overall, the radiation dose prescription is dictated by the radiation toxicity thresholds of such normal tissues, in some cases limiting its use entirely for primary malignancies that involve significant lung and liver exposure. It is therefore important when employing new RT modalities, such as MRT, to understand the influence of the modality on macrophage dynamics in normal tissues. It has been shown that exposure of the skin on the murine hind limb to a short pulse of MRT or broad beam (BB) prompted an increased frequency of macrophages in both the irradiated skin and out-of-field tissues [19]. An increase in tumour necrosis factor (Tnf) expression, a gene associated with inflammation, including promotion of macrophage activation, further underlines the role of macrophages in the normal tissue response to MRT [7].
It is now known that local immune responses also play an important role in MRT tumour control efficacy (reviewed in [20]). In tumours, endogenous cytokine and chemokine release from the tumour microenvironment attracts immune cells to the tumour, initiating a cascade of immunostimulatory responses [21]. Ionizing radiation attracts macrophages to irradiated tumours [22,23,24][22][23][24] in response to the radiation-mediated release of pro-inflammatory cytokines [25]. Tumour-associated macrophages (TAMs) form a substantial proportion of the infiltrating leucocytes in lung tumours and are an emerging target for lung cancer therapies [26]. TAMs in lung tumours can exhibit two opposite phenotypes: M1-like or M2-like, with the first displaying anti-inflammatory properties and the latter promoting tumour growth, invasion, and neo-angiogenesis [26,27][26][27].
Previous data have shown that MRT irradiation of a B16F10 melanoma resulted in an increase in chemokines associated with the migration and influx of inflammatory monocytes. These included monocyte chemoattractant protein (MCP)-1, macrophage inflammatory protein (MIP)-1α and MIP-1β, C-C motif ligand 5 (CCL5), as well as interleukin (IL)12p40 [4]. This was associated with an increased infiltration of CD68+ cells, especially at 9 days post-MRT [4].
2. The Macrophage Response following MRT
Exploiting the unique immunomodulatory cascade following RT is a point of great interest for adjuvant cancer treatment strategies. Based on previous studies investigating the outcome of tumour-bearing animals treated with MRT, macrophages were implicated as early responders to MRT-induced DNA damage in normal and tumour tissues [6,38,39][6][28][29]. Activated macrophages mobilised towards the site of injury and had a predominant phagocytic action against apoptotic cells in acute phases [40][30].
The sharp organization of F4/80+ macrophages densely populating the microbeam path in the liver are most likely resident liver macrophages, Kupffer cells, since recruited monocytes from the blood stream likely would not yet have differentiated into mature macrophages [40][30]. Nevertheless, this observation does not exclude the possibility of simultaneous immune cell recruitment, especially knowing that high peak-dose MRT provokes tumour vasculature disruption, creating a gate for the infiltration of different immune cells [41,42][31][32].
Macrophage populations of the liver consist of the resident Kupffer cells and bone-marrow derived monocytes, which migrate to the liver following injury [32][33]. Kupffer cells are strongly F4/80+ positive [27], account for the vast majority of hepatic macrophages [43][34] and are essential for repair following acute injury [44][35]. The type of injury, however, influences the inflammatory microenvironment, which in turn has a major impact on phenotypic changes of both resident and monocyte-derived macrophage populations, dictating their role in either resolving or aggravating tissue injury [45,46][36][37]. Typically, Kupffer cells are tolerogenic phagocytes, while bone marrow-derived populations are inflammatory [47][38]. It is apparent that resident macrophages of the liver respond to MRT-induced damage without eliciting a strong systemic immune response, as the recruitment of bone marrow-derived monocytes or peritoneal macrophages was minimal or absent, respectively. Interestingly, investigations into wound healing following focal sterile injury of the liver revealed a similar dynamic response by macrophages. Macrophages initially demarcated the site of injury within 48 h, where they then underwent a phenotypic conversion from a classical pro-inflammatory into non-classical macrophage populations that promoted tissue repair [48][39]. Therefore, the immune response to MRT-induced injury may not require the mobilization of monocytes from the bone marrow, as Kupffer cells may be sufficient scavengers of damaged cells induced by MRT, resulting in tissue-protective immunological tolerance and a non-inflammatory liver microenvironment [46][37]. Similarly, the lung also has resident macrophage populations residing in the alveolar and interstitial compartments [36][40], both expressing CD68 [49][41], with monocyte migration following injury contributing to inflammation and repair [50][42]. Thoracic irradiations have shown an increase in macrophage infiltration and pro-inflammatory activation in the lung [51][43]. Macrophages are responsible for the clearance of dying cells, and tissue-resident macrophages can efficiently clear large volumes of debris. When the injury is extensive, monocyte-derived alveolar macrophages support the onset of chronic inflammation [14] and subsequent fibrosis [52][44]. Since MRT only induces localized damage, it is, therefore, possible that resident macrophage populations of the normal liver and lung tissues can adequately clear the damaged cells confined to the microbeam path without the large and diffuse recruitment of potentially damaging inflammatory cells. The role of the macrophages following MRT may differ between normal and tumour tissues. Macrophages are heterogenous with pro- and anti-inflammatory polarization, with an anti-inflammatory phenotype associated with tumour progression (reviewed by [55][45]). While in normal tissues, it can be seen that the clear demarcation of the MRT beam path throughout the irradiated field, this localization is restricted to the periphery of the lung carcinoma samples. This may be explained by the presence of a necrotic core visible in the middle of the tumour. These necrotic areas are not vascularized and, as such, not supplied with oxygen and nutrients, and it is known that TAMs generally accumulate in pre-necrotic zones [56][46]. Although there are previous studies indicating a higher infiltration of TAMs in tumours after MRT compared to homogenous beam radiation [4[4][28],38], this specific immunostaining pattern has never been documented. Furthermore, tumour growth rates affect the maintenance of MRT geometry, with cellular migration disrupting the localization of peak- and valley-irradiated cells, making geometry demarcation at later timepoints post-irradiation difficult [57][47]. In the context of radiation, macrophage reprogramming is dependent on the dose and the type of radiation-induced cell death. The major factor in macrophage recruitment/activation following ionizing radiation depends on the type of signalling elicited by the tumour following exposure. This may depend on the type of radiation and the modality in which it is delivered. The production of cytokines and chemokines following ionizing radiation potentially varies with modality and may differentially influence macrophage polarization [22]. It has been shown that high doses (>10 Gy) of conventional irradiation increase M2-like, anti-inflammatory macrophage populations [58][48], while moderate doses (1–10 Gy) potentiate M1-like phenotypes [59][49]. The impact of radiation itself on macrophage function may be a key determinant of the tumour response to RT. Previous work has shown differential radiosensitivity between macrophage phenotypes with the M2, anti-inflammatory phenotype, having a greater degree of radiation resistance compared to the M1pheontype [60][50]. In contrast, during the acute phases following radiation, quiescent M1 macrophages were less sensitive to radiation-induced DNA damage and persisted over M2 populations [61][51]. In the context of spatially fractionated radiation therapy (SFRT), it has been shown that conventional-source SFRT is a powerful immune modulator [64][52]. Spatial fractionation may, therefore, have a dual effect on immunostimulatory responses, with high-dose deposition regions inducing immunogenic signalling, while at the same time preserving resident immune cells in the low-dose regions. Kanagavelu et al. [64][52] have shown that local ablative peak dose SFRT for LLC1 tumours induced increased systemic secretion of inflammatory cytokines, including IL-2, which is produced by macrophages and T-cells. MRT uses synchrotron-generated X-rays to deliver large doses in parallel microbeams and is therefore different to conventional RT due to its highly non-uniform dose distribution, ultra-high dose rates and kilovoltage rather than megavoltage photons.