Deferoxamine topical cream superior to patch in rescuing radiation-induced fibrosis of unwounded and wounded skin.

Topical patch delivery of deferoxamine (DFO) has been studied as a treatment for this fibrotic transformation in irradiated tissue. Efficacy of a novel cream formulation of DFO was studied as a RIF therapeutic in unwounded and excisionally wounded irradiated skin. C57BL/6J mice underwent 30 Gy of radiation to the dorsum followed by 4 weeks of recovery. In a first experiment, mice were separated into six conditions: DFO 50 mg cream (D50), DFO 100 mg cream (D100), soluble DFO injections (DI), DFO 1 mg patch (DP), control cream (Vehicle), and irradiated untreated skin (IR). In a second experiment, excisional wounds were created on the irradiated dorsum of mice and then divided into four treatment groups: DFO 100 mg Cream (W-D100), DFO 1 mg patch (W-DP), control cream (W-Vehicle), and irradiated untreated wounds (W-IR). Laser Doppler perfusion scans, biomechanical testing, and histological analysis were performed. In irradiated skin, D100 improved perfusion compared to D50 or DP. Both D100 and DP enhanced dermal characteristics, including thickness, collagen density and 8-isoprostane staining compared to untreated irradiated skin. D100 outperformed DP in CD31 staining, indicating higher vascular density. Extracellular matrix features of D100 and DP resembled normal skin more closely than DI or control. In radiated excisional wounds, D100 facilitated faster wound healing and increased perfusion compared to DP. The 100 mg DFO cream formulation rescued RIF of unwounded irradiated skin and improved excisional wound healing in murine skin relative to patch delivery of DFO.

Radiation-induced fibrosis: Mechanisms and therapeutic strategies from an immune microenvironment perspective.

Radiation-induced fibrosis (RIF) is a severe chronic complication of radiotherapy (RT) manifested by excessive extracellular matrix (ECM) components deposition within the irradiated area. The lung, heart, skin, jaw, pelvic organs and so on may be affected by RIF, which hampers body functions and quality of life. There is accumulating evidence suggesting that the immune microenvironment may play a key regulatory role in RIF. This article discussed the synergetic or antagonistic effects of immune cells and mediators in regulating RIF's development. Several potential preventative and therapeutic strategies for RIF were proposed based on the immunological mechanisms to provide clinicians with improved cognition and clinical treatment guidance.

Tissue fibrosis induced by radiotherapy: current understanding of the molecular mechanisms, diagnosis and therapeutic advances.

Cancer remains the leading cause of death around the world. In cancer treatment, over 50% of cancer patients receive radiotherapy alone or in multimodal combinations with other therapies. One of the adverse consequences after radiation exposure is the occurrence of radiation-induced tissue fibrosis (RIF), which is characterized by the abnormal activation of myofibroblasts and the excessive accumulation of extracellular matrix. This phenotype can manifest in multiple organs, such as lung, skin, liver and kidney. In-depth studies on the mechanisms of radiation-induced fibrosis have shown that a variety of extracellular signals such as immune cells and abnormal release of cytokines, and intracellular signals such as cGAS/STING, oxidative stress response, metabolic reprogramming and proteasome pathway activation are involved in the activation of myofibroblasts. Tissue fibrosis is extremely harmful to patients' health and requires early diagnosis. In addition to traditional serum markers, histologic and imaging tests, the diagnostic potential of nuclear medicine techniques is emerging. Anti-inflammatory and antioxidant therapies are the traditional treatments for radiation-induced fibrosis. Recently, some promising therapeutic strategies have emerged, such as stem cell therapy and targeted therapies. However, incomplete knowledge of the mechanisms hinders the treatment of this disease. Here, we also highlight the potential mechanistic, diagnostic and therapeutic directions of radiation-induced fibrosis.

Intraoral nasolabial flap for lower lip augmentation following post-radiation contracture.

The nasolabial flap (NLF) has been reported extensively for reconstruction of various intraoral and extraoral defects resulting from trauma or ablative surgery. However, it has not been described for post-radiation lip augmentation. Herein, we present the case of a 74-year-old female who previously underwent a subtotal glossectomy and free flap reconstruction followed by radiotherapy. While oncologically the patient did well, she developed a significant lower lip contracture which compromised oral intake, denture placement, lip excursion, and psychosocial well-being. The patient underwent release of the scar contracture and a NLF was utilized intraorally to act as a spacer between the gingiva and inner lip mucosa to augment the soft tissue deficit. The patient went on to regain oral intake and placement of her dentures, while reporting significant satisfaction with the post-procedural benefits.

Effects of Combination Treatments with Astaxanthin-Loaded Microparticles and Pentoxifylline on Intracellular ROS and Radiosensitivity of J774A.1 Macrophages.

Radiation-induced fibrosis (RIF) is a serious, yet incurable, complication of external beam radiation therapy for the treatment of cancer. Macrophages are key cellular actors in RIF because of their ability to produce reactive oxidants, such as reactive oxygen species (ROS) and inflammatory cytokines that, in turn, are the drivers of pro-fibrotic pathways. In a previous work, we showed that phagocytosis could be exploited to deliver the potent natural antioxidant astaxanthin specifically to macrophages. For this purpose, astaxanthin encapsulated into µm-sized protein particles could specifically target macrophages that can uptake the particles by phagocytosis. In these cells, astaxanthin microparticles significantly reduced intracellular ROS levels and the secretion of bioactive TGFß and increased cell survival after radiation treatments. Here we show that pentoxifylline, a drug currently used for the treatment of muscle pain resulting from peripheral artery disease, amplifies the effects of astaxanthin microparticles on J774A.1 macrophages. Combination treatments with pentoxifylline and encapsulated astaxanthin might reduce the risk of RIF in cancer patients.

Protective Effect of Low-Molecular-Weight Fucoidan on Radiation-Induced Fibrosis Through TGF-ß1/Smad Pathway-Mediated Inhibition of Collagen I Accumulation.

Radiation-induced fibrosis (RIF) occurs after radiation therapy in normal tissues due to excessive production and deposition of extracellular matrix proteins and collagen, possibly resulting in organ function impairment. This study investigates the effects of low-molecular-weight fucoidan (LMF) on irradiated NIH3T3 cells. Specifically, we quantified cellular metabolic activity, fibrosis-related mRNA expression, transforming growth factor beta-1 (TGF-ß1), and collagen-1 protein expression, and fibroblast contractility in response to LMF. LMF pre + post-treatment could more effectively increase cellular metabolic activity compared with LMF post-treatment. LMF pre + post-treatment inhibited TGF-ß1 expression, which mediates negative activation of phosphorylated Smad3 (pSmad3) and Smad4 complex formation and suppresses downstream collagen I accumulation. In addition, LMF pre + post-treatment significantly reduced actin-stress fibers in irradiated NIH3T3 cells. LMF, a natural substance obtained from brown seaweed, may be a candidate agent for preventing or inhibiting RIF.

Astragalus saponin attenuates the expression of fibrosis-related molecules in irradiated cardiac fibroblasts.

The main pathological change of radiation-induced heart disease is fibrosis. Emerging evidence has indicated that Astragalus membranaceus and its extractant, Astragalus saponin (AST), were used for treating fibrosis diseases. In the present study, the effects of AST on fibrosis damage induced by irradiation were determined. After being irradiated with 1 or 2-Gy X-rays, obvious changes of endoplasmic reticulum morphology were observed in cardiac fibroblasts (CFs), suggesting that its protein processing function was imbalanced, which indirectly indicated that fibrosis damage was caused by irradiating CFs. The expression levels of TGF-ß1 and collagen I (Col-1) were increased at 48-h post-irradiation. Administration of 20 µg/ml AST reduced the production of reactive oxygen species in irradiated CFs and decreased the expression of Col-1, TGF-ß1, and p-Smad2/3. Polymerase chain reaction (PCR)-array analysis showed that there were ~30 genes which were mainly classified into extracellular matrix, remodeling enzymes, inflammatory cytokines/chemokines, and TGF-ß superfamily, were up-regulated after treatment with 1-Gy X-ray, whereas most of these genes were down-regulated when pretreated with 20 µg/ml of AST. In addition, TIMP1 and Smad7 genes that were down-regulated after treatment with 1-Gy X-ray were up-regulated when pretreated with 20 µg/ml of AST. In conclusion, radiation-induced fibrosis damage was observed at a cellular level. AST attenuated this fibrosis damage effect in irradiated CFs and this anti-fibrosis effect may be closely related to its antioxidant action. The involvement of fibrosis-related molecules in irradiated CFs was systematically demonstrated by a PCR array for the first time. AST reversed the expression of the majority of genes changed by irradiation, which further confirmed its anti-fibrosis effect.

Sodium tanshinone IIA sulfonate attenuates radiation-induced fibrosis damage in cardiac fibroblasts.

The main pathological change in radiation-induced heart disease is fibrosis. Emerging evidence has indicated that sodium tanshinone IIA sulfonate (STS) was used for treating fibrosis diseases. The present study was undertaken to characterize the effect of STS on radiation-induced cardiac fibrosis (RICF) on cultured cardiac fibroblasts (CFs). CFs were irradiated with 1 or 2 Gy X-rays, and the expression of TGF-ß1 and collagen I (Col-1) increased, indicating that low-dose X-rays promoted fibrosis damage effect. The fibrosis damage was accompanied by morphologic changes in the endoplasmic reticulum (ER), as well as an increase in the expression of the ER stress-related molecules, GRP78 and CHOP. Administration of STS reduced ROS production and decreased the expression of Col-1, TGF-ß1, p-Smad2/3, GRP78, and CHOP in irradiated CFs, thus weakening the radiation-induced fibrosis damage and ER stress. Radiation-induced fibrosis damage was observed on a cellular level. The involvement of ER stress in radiation-induced fibrosis damage was demonstrated for the first time. STS attenuated the fibrosis damage effect in CFs and this effect may be related to its antioxidant action, and also related to its inhibition of ER stress and TGF-ß1-Smad pathway. These results suggest that STS shows a good prospect in clinical prevention and treatment of RICF.

Alpha-2-macroglobulin as a radioprotective agent: a review.

Radiation is an important modality in cancer treatment, and eighty percent of cancer patients need radiotherapy at some point during their clinical management. However, radiation-induced damage to normal tissues restricts the therapeutic doses of radiation that can be delivered to tumours and thereby limits the effectiveness of the treatment. The use of radioprotectors represents an obvious strategy to obtain better tumour control using a higher dose in radiotherapy. However, most of the synthetic radioprotective compounds studied have shown inadequate clinical efficacy owing to their inherent toxicity and high cost. Hence, the development of radioprotective agents with lower toxicity and an extended window of protection has attracted a great deal of attention, and the identification of alternative agents that are less toxic and highly effective is an absolute necessity. Recent studies have shown that alpha-2-macroglobulin (α2M) possesses radioprotective effects. α2M is a tetrameric, disulfide-rich plasma glycoprotein that functions as a non-selective inhibitor of different types of non-specific proteases and as a carrier of cytokines, growth factors, and hormones. α2M induces protein factors whose interplay underlies radioprotection, which supports the idea that α2M is the central effector of natural radioprotection in the rat. Pretreatment with α2M has also induced a significant reduction of irradiation-induced DNA damage and the complete restoration of liver and body weight. Mihailovic et al. concluded that the radioprotection provided by α2M was in part mediated through cytoprotection of new blood cells produced in the bone marrow; these authors also indicated that an important aspect of the radioprotective effect of amifostine was the result of the induction of the endogenous cytoprotective capability of α2M. The radioprotective effects of α2M are possibly due to antioxidant, anti-fibrosis, and anti-inflammatory functions, as well as the maintenance of homeostasis, and enhancement of the DNA repair and cell recovery processes. This review is the first to summarise the observations and elucidate the possible mechanisms responsible for the beneficial effects of α2M. The lacunae in the existing knowledge and directions for future research are also addressed.

Quantitative measurements of radiation-induced fibrosis for head and neck cancer: A narrative review.

Objectives: To provide a comprehensive summary of the different modalities available to measure soft tissue fibrosis after radiotherapy in head and neck cancer patients. Data Sources: PubMed, Scopus, and Web of Sciences. Review Methods: A search was conducted using a list of medical subject headings and terms related to head and neck oncology, radiation fibrosis, and quantitative measurements, including bioimpedance, MRI, and ultrasound. Original research related to quantitative measurement of neck fibrosis post-radiotherapy was included without time constraints, while reviews, case reports, non-English texts, and inaccessible studies were excluded. Discrepancies during the review were resolved by discussing with the senior author until consensus was reached. Results: A total of 284 articles were identified and underwent title and abstract screening. Seventeen articles had met our criteria for full-text review based on relevance, of which nine had met our inclusion criteria. Young's modulus (YM) and viscoelasticity measures have demonstrated efficacy in quantifying neck fibrosis, with fibrotic tissues displaying significantly higher YM values and altered viscoelastic properties such as increased stiffness rate-sensitivity and prolonged stress-relaxation post-radiation. Intravoxel incoherent motion offers detailed insights into tissue changes by assessing the diffusion of water molecules and blood perfusion, thereby differentiating fibrosed from healthy tissues. Shear wave elastography has proven to be an effective technique for quantifying radiation-induced fibrosis in the head and neck region by measuring shear wave velocity. Conclusion: There are various modalities to measure radiation-induced fibrosis, each with its unique strengths and limitations. Providers should be aware of these implications and decide on methodologies based on their specific clinical workflow. Level of Evidence: Step 5.

Single-cell transcriptional analysis of irradiated skin reveals changes in fibroblast subpopulations and variability in caveolin expression.

BACKGROUND: Radiation-induced fibrosis (RIF) is an important late complication of radiation therapy, and the resulting damaging effects of RIF can significantly impact reconstructive outcomes. There is currently a paucity of effective treatment options available, likely due to the continuing knowledge gap surrounding the cellular mechanisms involved. In this study, detailed analyses of irradiated and non-irradiated human skin samples were performed incorporating histological and single-cell transcriptional analysis to identify novel features guiding development of skin fibrosis following radiation injury. METHODS: Paired irradiated and contralateral non-irradiated skin samples were obtained from six female patients undergoing post-oncologic breast reconstruction. Skin samples underwent histological evaluation, immunohistochemistry, and biomechanical testing. Single-cell RNA sequencing was performed using the 10X single cell platform. Cells were separated into clusters using Seurat in R. The SingleR classifier was applied to ascribe cell type identities to each cluster. Differentially expressed genes characteristic to each cluster were then determined using non-parametric testing. RESULTS: Comparing irradiated and non-irradiated skin, epidermal atrophy, dermal thickening, and evidence of thick, disorganized collagen deposition within the extracellular matrix of irradiated skin were readily appreciated on histology. These histologic features were associated with stiffness that was higher in irradiated skin. Single-cell RNA sequencing revealed six predominant cell types. Focusing on fibroblasts/stromal lineage cells, five distinct transcriptional clusters (Clusters 0-4) were identified. Interestingly, while all clusters were noted to express Cav1, Cluster 2 was the only one to also express Cav2. Immunohistochemistry demonstrated increased expression of Cav2 in irradiated skin, whereas Cav1 was more readily identified in non-irradiated skin, suggesting Cav1 and Cav2 may act antagonistically to modulate fibrotic cellular responses. CONCLUSION: In response to radiation therapy, specific changes to fibroblast subpopulations and enhanced Cav2 expression may contribute to fibrosis. Altogether, this study introduces a novel pathway of caveolin involvement which may contribute to fibrotic development following radiation injury.

Swallowing-related muscle inflammation and fibrosis induced by a single dose of radiation exposure in mice.

BACKGROUND: Although radiotherapy is commonly used to treat head and neck cancer, it may lead to radiation-associated dysphagia (RAD). There are various causes of RAD, however, the mechanism has not yet been fully identified. Currently, the only effective treatment for RAD is rehabilitation. Additionally, there are few available animal models of RAD, necessitating the development of new models to establish and evaluate RAD treatments. We hypothesize that radiation-induced neck muscle fibrosis could be one of the causes of RAD due to impairment of laryngeal elevation. Therefore, in this study, we focused on the changes in inflammation and fibrosis of the strap muscles (Sternohyoid, Sternothyroid, and Thyrohyoid muscles) after a single-dose irradiation. This research aims to provide a reference animal model for future studies on RAD. RESULTS: Compared to control mice, those treated with 72-Gy, but not 24-Gy, irradiation had significantly increased tumor necrosis factor-α (TNF-α) (p < 0.01) and α-smooth muscle actin (αSMA) (p < 0.05) expression at 10 days and significantly increased expression levels of motif chemokine ligand-2 (CCL2), α-SMA, tumor growth factor-ß1 (TGF-ß1), type1 collagen, and interleukin-1ß (IL-1ß) (p < 0.05) in the muscles at 1 month by real-time PCR analysis. The results of immunohistochemistry showed that the deposition of type 1 collagen gradually increased in extracellular space after radiation exposure, and the positive area was significantly increased at 3 months compared to non-irradiated control. CONCLUSIONS: A single dose of 72-Gy irradiation induced significant inflammation and fibrosis in the strap muscles of mice at 1 month, with immunohistochemical changes becoming evident at 3 months. This cervical irradiation-induced fibrosis model holds potential for establishing an animal model for RAD in future studies. LEVEL OF EVIDENCE: N/A.

Assessment of pain as a sequela in patients treated with intracavitary brachytherapy accelerated partial breast irradiation (IBAPBI): preliminary results of a prospective study.

To assess pain rates and relationship to radiation-induced fibrosis (RIF) in patients treated with intracavitary brachytherapy accelerated partial breast irradiation (IBAPBI). Thirty-nine patients treated with IBAPBI were assessed prospectively for development of pain pretreatment, 1 month post-IBAPBI, and every 6 months thereafter. A qualitative subjective Late Effects of Normal Tissue-Subjective Objective Management Analytical (LENT-SOMA) questionnaire assessed pain. Use of pain medications was assessed as "no", "sometimes", or "regularly". A quantitative objective validated pressure threshold (PTH) measured pain in the site of IBAPBI breast (index) and its mirror-image in the nonirradiated breast (control). A validated tissue compliance meter (TCM) quantitatively assessed RIF in the index and control breasts at all time points. Mean ΔPTH(kg) and ΔTCM(mm) values reflected mean difference between the index and control breasts. Median follow-up is 44 months (range 5-59 months). According to LENT-SOMA, pain occurred in 89% at 1 and 24 months, 67% at 30 months, 30% at 36 months, 29% at 40 months, and 20% at 48 months. No patient used pain medication "regularly" but the use "sometimes" decreased over time: 61% at 1 month, 42% at 18 and 24 months, 13% at 36 months, and 10% at 40 months. Mean ΔPTH values, compared to Δ0 kg at baseline, peaked in absolute value by 1 month to -1.36 kg (p < 0.0001), persisted after 18 months at -0.99 kg (p < 0.0001) and 24 months at -0.73 kg (p < 0.0001), and returned to nearly baseline by 40 months at -0.11 kg (p < 0.57). Mean ΔPTH and ΔTCM correlated significantly with subjective patient reports of pain at each time point (p < 0.0001). To date, this is the first report to prospectively assess pain employing quantitative and qualitative inventories in patients treated with IBAPBI. Pain is experienced in the majority of patients experienced pain within the first 2 years, sometimes requiring a medication, and though subsides, it may persist 4 years after IBAPBI.

A Unique Adverse Event of Radiotherapy in a Patient With IgG4-related Disease: A Case Report.

BACKGROUND/AIM: IgG4-related disease (IgG4RD) is a rare autoimmune proinflammatory condition that mimics other cancers and has unique pathological findings. The effects of radiotherapy in patients with IgG4RD remain unknown. CASE REPORT: A male patient in his seventies who received radiotherapy (68 Gy/39 fr) for bladder cancer 5 months prior, presented to our hospital with fatigue and swelling in both legs. The patient had a history of IgG4-related sclerosing cholangitis, a subtype of IgG4RD. Leg edema gradually worsened despite treatment with a diuretic agent. Computed tomography showed hyperdense soft-tissue lesions in the irradiated area. The serum level of IgG4 increased to 1,380 mg/dl. One month after administration of a corticosteroid (10 mg per day) as an ex juvantibus treatment for IgG4RD, leg edema disappeared. Soft-tissue lesions in the irradiated area decreased in size. The adverse event was ultimately diagnosed as the recurrence of IgG4RD in the irradiated area. To the best of our knowledge, this is the first case report of an adverse event of radiotherapy for a patient with IgG4RD. CONCLUSION: We experienced a unique adverse event of radiotherapy in a patient with IgG4RD. Caution is advised on radiotherapy administration in patients with IgG4RD.

Deconstructing Fat to Reverse Radiation Induced Soft Tissue Fibrosis.

Adipose tissue is composed of a collection of cells with valuable structural and regenerative function. Taken as an autologous graft, these cells can be used to address soft tissue defects and irregularities, while also providing a reparative effect on the surrounding tissues. Adipose-derived stem or stromal cells are primarily responsible for this regenerative effect through direct differentiation into native cells and via secretion of numerous growth factors and cytokines that stimulate angiogenesis and disrupt pro-inflammatory pathways. Separating adipose tissue into its component parts, i.e., cells, scaffolds and proteins, has provided new regenerative therapies for skin and soft tissue pathology, including that resulting from radiation. Recent studies in both animal models and clinical trials have demonstrated the ability of autologous fat grafting to reverse radiation induced skin fibrosis. An improved understanding of the complex pathologic mechanism of RIF has allowed researchers to harness the specific function of the ASCs to engineer enriched fat graft constructs to improve the therapeutic effect of AFG.

Presentation of Eagle syndrome following radiation therapy to carcinoma of the larynx.

Eagle syndrome is a rare clinical condition that is characterized by either an elongated styloid process or a calcified stylohyoid ligament. This report describes the case of a 35-year-old woman who presented with Eagle syndrome following the treatment of recurrent laryngeal carcinoma with ionizing radiation.

Feasibility study for inducing the skeletal muscle fibrosis via irradiation using two mouse strains.

PURPOSE: Although the mechanism of onset and progression of radiation-induced fibrosis (RIF) has been studied, most studies to date have focused on pulmonary fibrosis. There are few studies on murine RIF in the skeletal muscle, and the pathogenic mechanism remains unclear. This pilot study aimed to evaluate the feasibility to create a murine model of RIF in the skeletal muscle and analyze strain differences in fibrosis sensitivity. MATERIALS AND METHODS: Two mouse strains, C57BL/6 and C3H/He, were used. Their right hind limbs were irradiated at a dose of 25 Gy once a week for three fractions. Gastrocnemius muscles were collected at day 4, and weeks 2, 4, 8, 12, and 24 after the third irradiation and subjected to histopathological examination and immunoblotting. RESULTS: In C57BL/6 mice, chronic inflammation and an increased expression of transforming growth factor-ß (TGF-ß) and fibronectin were observed 2 weeks after irradiation. A significant increase in fibrosis was detected after 8 weeks. However, in C3H/He mice, the expression of TGF-ß and fibronectin increased 8 weeks after irradiation, and fibrosis significantly increased after 12 weeks. Moreover, the degrees of inflammation and fibrosis were more remarkable in C57BL/6 mice than in C3H/He mice. CONCLUSION: The onset and degree of fibrosis may be associated with the expression of TGF-ß and fibronectin, and inflammation, in a strain-specific manner. Therefore, a murine model of RIF in the skeletal muscle could be created using the indicated method, suggesting that the C57BL/6 strain is more sensitive to fibrosis in the skeletal muscle, as well as the lung, than the C3H/He strain. Radiation-induced fibrosis in the skeletal muscle could be detected in C57BL/6 and C3H/He mice, with C57BL/6 mice being more sensitive to fibrosis in the skeletal muscle than C3H/He mice.

Radiation Fibrosis After Stereotactic Body Radiation Therapy for Osseous Metastases: A Case Report.

Radiation-induced fibrosis is a potentially severe late complication after high-dose radiotherapy. Over the last decade, there has been increasing use of stereotactic body radiation therapy (SBRT) to treat both primary and metastatic malignancies. While there has been evolving evidence of appropriate dose constraints for certain organs receiving hypofractionated radiotherapy, the risk, and appropriate dose constraints to limit the risk of radiation-induced muscle fibrosis are poorly defined. In this report, two patients are presented who underwent SBRT for osseous oligometastatic renal cell carcinoma. While the treatment was well-tolerated with no acute toxicities and complete local control of the metastasis, both patients experienced late toxicity of radiation-induced fibrosis in the adjacent musculature. In both cases, toxicity was nonresponsive to medical interventions and was severe enough to require surgical resection of the affected tissue. Following surgery, both patients reported improved pain relief and mobility. Further studies are needed to explore the dose constraints that may reduce the risk of radiation-induced muscle fibrosis in five-fraction treatment.

Gene Expression Profiles Reveal Extracellular Matrix and Inflammatory Signaling in Radiation-Induced Premature Differentiation of Human Fibroblast in vitro.

PURPOSE: Fibroblasts are considered to play a major role in the development of fibrotic reaction after radiotherapy and premature radiation-induced differentiation has been proposed as a cellular basis. The purpose was to relate gene expression profiles to radiation-induced phenotypic changes of human skin fibroblasts relevant for radiogenic fibrosis. MATERIALS AND METHODS: Exponentially growing or confluent human skin fibroblast strains were irradiated in vitro with 1-3 fractions of 4 Gy X-rays. The differentiated phenotype was detected by cytomorphological scoring and immunofluorescence microscopy. Microarray analysis was performed on Human Genome U133 plus2.0 microarrays (Affymetrix) with JMP Genomics software, and pathway analysis with Reactome R-package. The expression levels and kinetics of selected genes were validated with quantitative real-time PCR (qPCR) and Western blotting. RESULTS: Irradiation of exponentially growing fibroblast with 1 × 4 Gy resulted in phenotypic differentiation over a 5-day period. This was accompanied by downregulation of cell cycle-related genes and upregulation of collagen and other extracellular matrix (ECM)-related genes. Pathway analysis confirmed inactivation of proliferation and upregulation of ECM- and glycosaminoglycan (GAG)-related pathways. Furthermore, pathways related to inflammatory reactions were upregulated, and potential induction and signaling mechanisms were identified. Fractionated irradiation (3 × 4 Gy) of confluent cultures according to a previously published protocol for predicting the risk of fibrosis after radiotherapy showed similar downregulation but differences in upregulated genes and pathways. CONCLUSION: Gene expression profiles after irradiation of exponentially growing cells were related to radiation-induced differentiation and inflammatory reactions, and potential signaling mechanisms. Upregulated pathways by different irradiation protocols may reflect different aspects of the fibrogenic process thus providing a model system for further hypothesis-based studies of radiation-induced fibrogenesis.

Cervical Fibrosis as a Predictor of Dysphagia.

OBJECTIVE: Radiotherapy of head and neck cancer (HNCA) causes dysfunction through radiation-induced fibrosis (RIF). We hypothesize that the degree of cervical fibrosis is associated with swallowing dysfunction. This study evaluated the association between cervical fibrosis and swallowing dysfunction in patients after radiation therapy for HNCA. STUDY DESIGN: Cross sectional study. METHODOLOGY: A convenience sample of patients with dysphagia who were at least 1 year post radiation therapy for HNCA underwent simultaneous cervical ultrasound (US) and video-fluroscopic swallow study (VFSS). US determinants of fibrosis were measurements of sternocleidomastoid fascia (SCMF) thickness bilaterally at the level of the cricoid. Primary and secondary outcome variables on VFSS were pharyngeal constriction ratio, a validated measure of pharyngeal contractility, and penetration aspiration scale (PAS). A qualitative assessment of lateral neck rotation was performed as a functional measure of neck fibrosis. RESULTS: Simultaneous cervical US and VFSS examinations were performed on 18 patients with a history of radiotherapy for HNCA and on eight controls. The mean (±SD) age of the entire cohort (N = 26) was 66 (±10) years. Individuals with a history of radiation had significantly thinner mean SCMF (0.26 [±0.04 mm]) compared to controls (0.48 [±0.06 mm]; P < .05). Individuals with thinner SCMF were more likely to have moderate to severe restriction in lateral neck rotation, a higher PCR, and a higher PAS (P < .05). CONCLUSION: Thinner sternocleidomastoid fascia on ultrasound in patients having undergone radiotherapy for head and neck cancer was associated with reduced lateral neck movement, poorer pharyngeal constriction and greater penetration/aspiration scale. The data suggest that cervical fibrosis is associated with swallowing dysfunction in head and neck cancer survivors and support the notion that, "As the neck goes, so does the swallow." LEVEL OF EVIDENCE: 3. Laryngoscope, 131:548-552, 2021.