GROWTH PLATE RADIATION RESPONSE--MECHANISMS AND THERAPY

University Of Rochester, Rochester NY

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Abstract

Disturbance or arrest of bone growth has long been recognized as a complication of the treatment of childhood milignancies with radiotherapy. However. There has been almost no work on the molecular mechanism of this phenomenon in the growth plate. This lack of information constitutes a glaring gap in our knowledge about this serious clinical problem, and precludes development of any therapeutic strategies to ameliorate radiation-induced growth plate injury. In the present proposal we present preliminary data on possible mechanisms by which radiation may disrupt specific aspects of normal cytokine and growth factor function in growth plate cartilage. The long term goals of the research are (1) to understand the derangements of growth factor function which contribute to radiation-induced bone growth arrest, and (2) to translate this information to clinically applicable strategies using nontoxic pharmacologic agents to potentially ameliorate gorwth plate injury in children undergoing radiotherapy for malignancies. Preliminary data using chick and rat growth plate chondrocyte culture and in vivo models indicate a specific pattern of the cellular and molecular events following irradiation, among which are the selective suppression of TGFbeta and PTHrP, the major mitogens driving proliferation, and inappropriate expression of TNFalpha, which may lead to premature hypertrophy and apoptosis. The obliteration of the autocrine mitogenic stimulus and evidence of premature apoptosis are consistent with the observed histology. The specific aims of the proposed reserch include: (1A) use our chick chondrocyte in vitro model to initially characterize and determine the pattern of disruptio of 3 specific mitogenic growth factors (bFGF.TFGbeta.PTHrP), and the innappropriate activation of one key cytokine (TNFalpha) following irradiation. (1B) Use this initial study of avian chondrocytes to help guide our shift to study of a more levelvant mammalian in vitro model of radiation effects on devloping rodent chondrocytes, using pellet cultures which recapitulate the tissue arechitecture and differentiation cascade of normal rat growth plate. (2A) Identify the signaling mechanisms leading to growth factor and TNFalpha derangement, through investigation of radiation effects on 2 specific second messengers: cytosolic calcium and cAMP/protein kinase A, both of which appear to be involved in mitogen suppression and stimulation of apoptosis. (2B) In this in vitro model of radiation damage, we will study response modification using retinoic acid to: suppress cytosolic calcium, TNFalpha expression, and apoptosis; and to stimulate bFGF, TFGbeta, and chondrocyte proliferation. (2C) We will study pentoxifylline as an additinal response modifier to: decrease cytosolic calcium and TNFalpha expression; and to stimulate cAMP and proliferation. (3A) Correlate in vitro findings with findings in a rat in vivo model using tissue-based approaches such as immunocytochemistry and in situ hybridization to conform mechanisms of radiation injury to the physis, as well as new clinically aplicable therapeutic strategies.