RESUMO
More than half of cancer patients are treated with radiation therapy. Despite its high therapeutic index, radiation therapy can cause disabling injuries to normal tissues, especially in long-term survivors. Thus, one of the great challenges of modern radiation therapy is to increase tolerance of normal tissue to ionizing radiation in order to improve the quality of life of cancer survivors and/or enhance local control using dose escalation. The physiopathological aspects of normal tissue toxicity have been widely explored; however, none of these descriptive findings has led to the development of effective therapeutic strategies. Several empirical treatments have also been used in clinical trials (superoxide dismutase, pentoxifylline-tocopherol); however, the results are still controversial, and their mechanisms of action have not been clearly defined. The recent development of high-throughput biological approaches will contribute greatly to the characterization of the molecular pathways associated with normal tissue toxicity and the identification of specific and effective molecular targets for therapeutic interventions using already known or new pharmacological compounds. In this paper, we will discuss recent advances made in the characterization of one of the most serious complications of radiation therapy, late intestinal toxicity, using molecular profiling. We will focus on the involvement of the Rho/ROCK pathway in the development and maintenance of late radiation enteropathy. The role of the Rho/ROCK pathway in tissue response to radiation injury will be reviewed, as well as therapeutic perspectives.
Assuntos
Intestinos/efeitos da radiação , Peptídeos e Proteínas de Sinalização Intracelular/fisiologia , Proteínas Serina-Treonina Quinases/fisiologia , Lesões por Radiação/fisiopatologia , Animais , Fator de Crescimento do Tecido Conjuntivo , Fibrose/etiologia , Fibrose/fisiopatologia , Humanos , Proteínas Imediatamente Precoces/fisiologia , Peptídeos e Proteínas de Sinalização Intercelular/fisiologia , Intestinos/patologia , Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Lesões por Radiação/tratamento farmacológico , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Quinases Associadas a rhoRESUMO
Type I interferon (IFN) is shown to control the reversible quiescence of a primitive human bone marrow mesenchymal stem cell (MSC) subpopulation. A 24 h pre-treatment of Stro1+/GlycoA- or CD45-/GlycoA- subpopulations with a monoclonal antibody (mAb) against the IFNAR1 chain of the human type I IFN receptor (64G12), or with a polyclonal anti-IFNalpha antibody, resulted in a marked increase in the number of very large colonies (CFU-F >3000 cells) obtained in the presence of low, but necessary, concentrations of bFGF. Over a 2-month culture period, this short activation promoted a faster and greater amplification of mesenchymal progenitors for adipocytes and osteoblasts. Activation correlated with inhibition of STAT1 and STAT2 phosphorylation and of STAT1 nuclear translocation. A non-neutralizing anti-IFNAR1 mAb was ineffective. We demonstrate that control and activated MSCs express ST3GAL3, a sialyltransferase necessary to produce the embryonic antigens SSEA-3 and -4. Interestingly, activated MSC progeny expressed SSEA-3 and -4 at a higher level than control cultures, but this was not correlated with a significant expression of other embryonic markers. As MSCs represent an essential tool in tissue regeneration, the use of 64G12, which rapidly recruits a higher number of primitive cells, might increase amplification safety for cell therapy.
Assuntos
Interferon-alfa/fisiologia , Interferon beta/fisiologia , Células-Tronco Mesenquimais/citologia , Células da Medula Óssea/citologia , Técnicas de Cultura de Células , Diferenciação Celular , Divisão Celular/efeitos dos fármacos , Ensaio de Unidades Formadoras de Colônias , Primers do DNA , Matriz Extracelular/fisiologia , Humanos , Imunofenotipagem , Cinética , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/imunologia , Reação em Cadeia da Polimerase , Fator de Crescimento Transformador beta1/farmacologiaRESUMO
BACKGROUND: Late radiation enteritis in humans is associated with accumulation of extracellular matrix and increased connective tissue growth factor (CTGF) expression that may involve intestinal muscular layers. AIMS: We investigated the molecular pathways involved in maintenance of radiation induced fibrosis by gene profiling and postulated that alteration of the Rho pathway could be associated with radiation induced fibrogenic signals and CTGF sustained expression. PATIENTS AND METHODS: Ileal biopsies from individuals with and without radiation enteritis were analysed by cDNA array, and primary cultures of intestinal smooth muscle cells were established. Then, the effect of pharmacological inhibition of p160 Rho kinase, using Y-27632, was studied by real time reverse transcription-polymerase chain reaction, western blot, and electrophoretic mobility shift assay. RESULTS: Molecular profile analysis of late radiation enteritis showed alterations in expression of genes coding for the Rho proteins. To investigate further the involvement of the Rho pathway in intestinal radiation induced fibrosis, primary intestinal smooth muscle cells were isolated from radiation enteritis. They retained their fibrogenic differentiation in vitro, exhibited a typical cytoskeletal network, a high constitutive CTGF level, increased collagen secretory capacity, and altered expression of genes coding for the Rho family. Rho kinase blockade induced a simultaneous decrease in the number of actin stress fibres, alpha smooth muscle actin, and heat shock protein 27 levels. It also decreased CTGF levels, probably through nuclear factor kappaB inhibition, and caused decreased expression of the type I collagen gene. CONCLUSION: This study is the first showing involvement of the Rho/Rho kinase pathway in radiation fibrosis and intestinal smooth muscle cell fibrogenic differentiation. It suggests that specific inhibition of Rho kinase may be a promising approach for the development of antifibrotic therapies.