Time- and dose-dependent changes of intracellular cytokine and cytokine receptor profile of Ewing tumour subpopulations under the influence of ionizing radiation.

PURPOSE: Cytokines and their corresponding cell surface receptors are involved in intercellular signalling pathways and in the radioresistance of normal and malignant cells. The aim was the characterization of the expression of intracellular cytokines, their receptors and apoptosis-associated markers under the influence of radiation. MATERIAL AND METHODS: Two Ewing tumours were characterized in vitro before and 4, 24 and 72 h after radiation with 5 and 10 Gy, and in vivo 4, 6 and 15 days after radiation with 5 and 30 Gy by five parameter flow cytometry. Direct fluorescence-conjugated antibodies directed against intracellular cytokines (interferon-gamma, tumour necrosis factor [TNF]-alpha, interleukin 1) and their receptors (CD119, CD120a, CD121a) were used. Annexin V and 7-amino-actinomycin D were used to identify radiation-induced apoptosis. RESULTS: Inter- and intra-individual heterogeneities were identified by the expression of cytokine receptors and the intracellular cytokine profile before radiation. Time- and dose-dependent up-regulation of the cytokines TNF-alpha and interleukin 1 were found in vitro. In vivo, an up-regulation of CD120a and CD121a was detectable on tumour cell subpopulations. For interferon-gamma and CD119, no changes were seen. CONCLUSIONS: The observed radiation-induced changes of cytokine and receptor profile are an indication for complex intercellular interactions in view of radioresistance-associated mechanisms between cell populations within one individual tumour. The observed heterogeneous response on radiation might have therapeutic implications for an individualized therapy based on combined radiation and cytokine modulation, defined by flow cytometric characterization of markers potentially informative for radioresistance.

Effect of radiation on Ewing tumour subpopulations characterized on a single-cell level: intracellular cytokine, immunophenotypic, DNA and apoptotic profile.

PURPOSE: Adhesion molecules, cytokines and their corresponding cell-surface receptors are involved in intercellular signalling pathways, radioresistance and metastasis-mediating mechanisms of malignant cells. The aim was the characterization of changes in the marker profile of Ewing tumour cell subpopulations under the influence of radiation. MATERIALS AND METHODS: Three Ewing tumours were characterized in vitro and in vivo in a xenograft model before and after radiation by five-parameter flow cytometry. Antibodies directed against cell surface and intracellular antigens, apoptosis-associated markers and the DNA dye 7-aminoactinomycin D were used. RESULTS: Tumour cell subpopulations were identified by expression of adhesion molecules and cytokine receptors, intracellular cytokines, apoptotic markers and DNA content. Heterogeneous changes of flow cytometric profile were identified on tumour cell subpopulations after radiation. CONCLUSIONS: The changed profile of tumour cells under radiation might be associated with biological changes of tumour subpopulations in view of radioresistance and metastatic potential and might be useful to identify intercellular regulation mechanisms and to define parameters being predictive for a response to therapy.

Limited effects of selenium substitution in the prevention of radiation-associated toxicities. results of a randomized study in head and neck cancer patients.

OBJECTIVE: The substitution of selenium activates the selenium-dependent enzyme glutathione peroxidase, which is important for scavenging free radicals. To date, only limited data are available about the clinical impact of selenium regarding the toxicities due to free radical producing therapies, e.g. irradiation or chemotherapy, and therefore the objective of this study was to investigate the clinical impact of selenium in such therapies. PATIENTS AND METHODS: 39 patients (8 female, 31 male) with advanced head and neck cancer were included in a randomised phase II study. The mean age was 63.52+/-9.31 years. Tumour localizations: oral cavity 15 patients, oropharynx 19 patients, hypopharynx 5 patients, carcinoma of unknown primary 1 patient. Group A (n=22) received 500 microg sodium selenite on the days of radiotherapy and 300 microg sodium selenite on days without radiotherapy. Group B (17) was irradiated without any selenium substitution. Both groups were well balanced according to age, gender, localization and stage of the tumour. The RTOG grade of radiation-associated toxicities was evaluated once per week. RESULTS: The following serious toxicities were observed (group A vs. group B): dysphagia 22.7% vs. 35.3%, loss of taste 22.7% vs. 47.1%, dry mouth 22.7% vs. 23.5%, and stomatitis 36.4% vs. 23.5%. A statistical trend (Fisher's exact test) was only seen for the loss of taste (p=0.172). The weekly patient analysis (Student's t-test) showed a significant reduction of dysphagia in the selenium group (Group 1) at the last week of irradiation. CONCLUSION: This small randomised trial showed limited effects of selenium in the prevention of ageusia (loss of taste) and dysphagia due to radiotherapy of head and neck cancer.

Heterogeneity of radiation induced apoptosis in Ewing Tumor cell lines characterized on a single cell level.

The objective of this study was to investigate heterogeneity of radiation induced apoptosis on a single cell level. Two Ewing tumor cell lines were characterized in vitro before and 24 and 72 h after radiation with 5 Gy by multiparametric flow cytometry. Annexin V, 7-AAD and fluorescence conjugated antibodies that were directed against HLA-ABC, CD11a and CD62L were used. Based on these markers radiation induced apoptosis was quantified, multiple apoptotic subpopulations were identified and a characteristic individual apoptotic profile was characterized. The characterization of HLA-ABC, CD11a and CD62L was informative to detect subpopulations of apoptotic cells. The observed heterogeneity and the identification of multiple apoptotic subpopulations reflect the complexity and diversity of biology of radiation induced cell death. This might be an indication for co-existing apoptotic pathways or it might represent sequential steps of the apoptotic cascade.

Documentation of radiation-induced oral mucositis. Scoring systems.

BACKGROUND: Radiation therapy of tumors in the head and neck region is frequently associated with severe side effects in the oral mucosa which often necessitate interruption of the prescribed treatment protocol. In order to compare therapeutic strategies and, more important, in order to perform multi center studies, generally accepted scoring systems have to be applied for uniform documentation of the oral mucosal response. METHODS: Different scoring protocols are found in the literature. The scoring protocols most widely accepted are the CTC classification and the RTOG/EORTC classification. These are compared with more detailed systems. RESULTS: In the CTC classification, grading of stomatitis is included in the responses of the gastrointestinal tract and emphasizes dietary effects. For effects of radiation alone or of radiochemotherapy, the RTOG/EORTC system, focusing on therapeutic interventions, has been established. However, there are only minor differences in the grading of mucositis between these 2 protocols. Based on the RTOG/EORTC classification, Maciejewski et al. introduced a classification system with inclusion of the area affected, but also changed the sensitivity of the scores. The latter may be confusing if the source of the system used is not cited in a report. An alternative system was proposed by Dische, which in addition to objective morphologic criteria also includes the symptoms induced by the mucosal response, and hence includes some subjective aspects reported by the patient. CONCLUSIONS: For routine documentation of acute radiation side effects in the oral cavity, the German version of the RTOG/EORTC classification can be recommended. In studies with particular interest in oral mucositis, a more sensitive scoring system may be applied. In any publication concerning mucositis, a table or a detailed description of the system used should be included.

Photographic documentation of acute radiation-induced side effects of the oral mucosa.

BACKGROUND: Radiotherapy in cancer of the head and neck induces cutaneous and mucosal reactions. These must be carefully assessed and documentated to control the accuracy of individual treatment, the overall toxicity of particular treatment schedules, the efficacy of prophylaxis and treatment and to determine the adequate therapy of treatment sequelae depending on the severity of the reactions. The accurate classification of lesions according to internationally accepted schedules (WHO/RTOG/CTC) is indispensable for the comparison of radiotherapy treatment results and efficacy of supportive care. METHODS: While the treatment of cancer depends on tumor stage and medical circumstances of the patient and is more or less standardized, prophylaxis and treatment of side-effects is highly variable. Discussing an optimized prophylaxis and therapy of oral mucositis, the problem of accurate classification and documentation emerged. The verbal description of mucosal lesions is open to many subjective interpretations. Photographic documentation seems a suitable method to optimize the grading of toxicity. RESULTS: A photographic survey of typical lesions for each grade of toxicity is a tool to reach several aims in one step. Toxicity of an individual patient may be compared with representative photographic examples in daily routine to decide quickly on the grade of toxicity. Subjective differences due to intra- and interpersonal variability of the evaluating radiooncologist will be reduced. The efficacy of treatment can be proven by accurate documentation. Randomized clinical studies concerning prophylaxis and treatment of oral mucositis will provide more reliable results if evaluation of toxicity grading is standardized by photographs. CONCLUSIONS: Photographic documentation of lesions of the oral mucosa might be the best means to reduce interindividual subjectivity in grading. It is a valuable appendix to standard classification systems and only concerns the visible signs of mucosal lesions. However, the exact grading of mucositis is only possible with additional clinical information about pain and nutritional situation.

[Chronic radiation effects on dental hard tissue (radiation caries). Classification and therapeutic strategies]. / Chronische Strahlenfolgen an den Zahnhartgeweben ("Strahlenkaries"). Klassifikation und Behandlungsansätze.

OBJECTIVES: Since the first description of rapid destruction of dental hard tissues following head and neck radiotherapy 80 years ago, "radiation caries" is an established clinical finding. The internationally accepted clinical evaluation score RTOG/EORTC however is lacking a classification of this frequent radiogenic alteration. MATERIAL AND METHODS: Medical records, data and images of radiation effects on the teeth of more than 1,500 patients, who underwent periradiotherapeutic care, were analyzed. Macroscopic alterations regarding the grade of late lesions of tooth crowns were used for a classification into 4 grades according to the RTOG/EORTC guidelines. RESULTS: No early radiation effects were found by macroscopic inspection. In the first 90 days following radiotherapy 1/3 of the patients complained of reversible hypersensitivity, which may be related to a temporary hyperemia of the pulp. It was possible to classify radiation caries as a late radiation effect on a graded scale as known from RTOG/EORTC for other organ systems. This is a prerequisite for the integration of radiation caries into the international nomenclature of the RTOG/EORTC classification. CONCLUSIONS: The documentation of early radiation effects on dental hard tissues seems to be neglectable. On the other hand the documentation of Late radiation effects has a high clinical impact. The identification of an initial lesion at the high-risk areas of the neck and incisal part of the tooth can lead to a successful therapy as a major prerequisite for orofacial rehabilitation. An internationally standardized documentation is a basis for the evaluation of the side effects of radiooncotic therapy as well as the effectiveness of protective and supportive procedures.