The Role of Autocrine Growth Factors in Radiation Damage to the Epiphyseal Growth Plate

Department of Orthopaedic Surgery, University of Rochester School of Medicine and Dentistry, Rochester, New York, USA.
Radiation Research (Impact Factor: 2.91). 07/2001; 155(6):847-57. DOI: 10.1667/0033-7587(2001)155[0847:TROAGF]2.0.CO;2
Source: PubMed


Radiation therapy plays an important role as part of the multimodality treatment for a number of childhood malignancies. Dose-limiting complications of radiotherapy include skeletal abnormalities and disturbances in skeletal development within the irradiated field. The current study was undertaken to investigate the molecular mechanisms involved in radiation-induced arrest of bone growth. Our hypotheses were: (1) Expression of autocrine growth factors that regulate chondrocyte proliferation is inhibited by radiation in a specific pattern; (2) the disparity in radiosensitivity of growth plate chondrocytes and epiphyseal chondrocytes is due to differential modulation of autocrine growth factor expression by radiation. Given the important role these cells play in skeletal growth and development, we examined the comparative effects of radiation on expression of specific mitogenic growth factors in growth plate chondrocytes. The effect of radiation on the expression of autocrine/paracrine growth factors was examined in an established avian model of epiphyseal growth plate maturation. Exposure of growth plate chondrocytes to radiation resulted in a specific pattern of biochemical and morphological alterations that were dependent on dose and were progressive over time. While radiation did not affect the mRNA expression of some of the autocrine and paracrine factors important in endochondral ossification (such as FGF2 and TGFB isoforms), it did lead to a decrease in the mRNA expression of PTHrP, a critically important mitogen in growth plate chondrocytes, and a dose-dependent decrease in the PTH/PTHrP receptor mRNA. Interestingly, PTHrP mRNA levels were not affected in irradiated epiphyseal chondrocytes, the main source of PTHrP. Given evidence indicating a role for intracellular calcium levels in regulating PTHrP expression, basal calcium levels in irradiated growth plate chondrocytes and epiphyseal chondrocytes were examined 24 h after treatment. While cytosolic calcium levels were significantly higher in irradiated growth plate chondrocytes, they were not significantly affected in irradiated epiphyseal chondrocytes. The importance of calcium in mediating radiation damage to growth plate chondrocytes was further demonstrated by the finding that the addition of 4.0 mM EGTA (a calcium chelator) to the cell cultures before irradiation prevented the decrease in PTHrP mRNA levels. Since PTHrP up-regulates BCL2 levels and prevents growth plate chondrocyte maturation and apoptosis, BCL2 mRNA levels were examined in irradiated growth plate chondrocytes, and a dose-dependent decrease was found. An increase in apoptosis was further confirmed by a fivefold increase in caspase 3 levels in irradiated growth plate chondrocytes. The results of the current study suggest that radiation may interfere with proliferation of growth plate chondrocytes in part by causing an increase in cytosolic calcium levels which in turn leads to a decrease in PTHrP mRNA. Growth plate chondrocyte PTHrP receptor mRNA expression is also inhibited by radiation, further decreasing PTHrP signaling. Despite subtle differences between the chick and mammalian growth plates, further studies should provide an enhanced understanding of the mechanism(s) of radiation injury to the growth plate, as well as possibilities for new therapeutic strategies to protect the growing skeleton from the detrimental effects of radiotherapy.

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    • "Growth plate is situated at both ends of long bones, which is composed of three distinct zones: the resting, proliferative, and hypertrophic zones. Bone growth begins as progenitor cells at resting zone are activated and enter the cell cycle at the proliferative zone [2] and produce extracellular matrix rich in collagen-II and aggrecan [1]. The hypertrophic chondrocytes secrete matrix rich in collagen-X and direct mineralisation of their surrounding matrix while undergoing apoptosis [3]. "
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    ABSTRACT: The advancement and intensive use of chemotherapy in treating childhood cancers has led to a growing population of young cancer survivors who face increased bone health risks. However, the underlying mechanisms for chemotherapy-induced skeletal defects remain largely unclear. Methotrexate (MTX), the most commonly used antimetabolite in paediatric cancer treatment, is known to cause bone growth defects in children undergoing chemotherapy. Animal studies not only have confirmed the clinical observations but also have increased our understanding of the mechanisms underlying chemotherapy-induced skeletal damage. These models revealed that high-dose MTX can cause growth plate dysfunction, damage osteoprogenitor cells, suppress bone formation, and increase bone resorption and marrow adipogenesis, resulting in overall bone loss. While recent rat studies have shown that antidote folinic acid can reduce MTX damage in the growth plate and bone, future studies should investigate potential adjuvant treatments to reduce chemotherapy-induced skeletal toxicities.
    BioMed Research International 03/2011; 2011:903097. DOI:10.1155/2011/903097 · 2.71 Impact Factor
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    • "Once it has been traumatized either by accidents, radiation (Bakker et al. 2003, Pateder et al. 2001) or unknown, internal regulating incidents, it hardly ever recovers (Davis and Green 1976, Farnum et al. 2000). This generally leads to premature closure of the growth plate such that ossification sets in and longitudinal growth is abruptly stopped (Bakker et al. 2003, Canadell and de Pablos 1985, Davis and Green 1976). "
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    ABSTRACT: Periosteal transection and elevation is a standard treatment for angular limb deformities in foals. It is used to correct axis deviations in the limbs at an early age to assure that the foals grow up with straight limbs to improve their chances to reach their full potential as future athletes. Although clinically proven, its mechanisms of action were never elucidated on a more basic scientific level. In this experimental study the molecular response to periosteal stripping was investigated within the growth plate and adjacent perichondrium. The study was based on the hypothesis that a growth restraining feedback loop related to Indian hedgehog (lhh), parathyroid hormone related protein (PTHrP) and parathyroid hormone receptors (PTHR) was responsible for the corrective effect of periosteal stripping. Twelve 3 months old lambs underwent periosteal stripping of the distal lateral radius and tibia on one side. The contralateral side served as non-operated controls. Two animals each group were sacrificed at 2, 6, 10, 14, 18 and 21 days after surgery and the growth plates with minimal adjacent bone tissue were harvested for histological investigations. After decalcification, paraffin-embedded sections with routine hematoxylin-eosin stains were prepared to assess morphology and length of growth plates, whereas immunohistochemistry of lhh, PTHrP, PTHR and the two cytokines fibroblast- (FGF) and transforming growth factor (TGF) was performed to study different protein expression between operated limbs and controls. The results indicate that periosteal stripping caused an up-regulation of lhh in the early pre- and hypertrophic zone of the growth plate, followed by an increase of PTHrP mainly in the perichondrium, while an increase of PTHR was noticed in all zones, although highest in the perichondrium and hypertrophic zones. The growth factors FGF and TGF were upregulated in all zones, but FGF in response to periosteal stripping was more intensely expressed in the proliferative zone and the highest peak of TGF was found in the perichondrium. Length measurements of the various growth zones revealed significant negative correlations between the proliferative and pre-and hypertrophic zones, indicating that indeed a negative feed back loop after periosteal stripping exists coupled by the Ihh/PTHrP/PTHR cascade.The hypothesis that periosteal stripping had an effect on the Ihh/PTHrP/PTHR related feedback loop in epiphyseal growth was confirmed in this experimental study in lambs. Since these mechanisms are very basic and similar in most species, it can be safely assumed that the effects in foals are similar. In fact, the asymmetric mechanical load in animals suffering from axis deviation may even increase the enhancing effect of length correction.
    Pferdeheilkunde 07/2010; 26(4). DOI:10.5167/uzh-42239 · 0.19 Impact Factor
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    • "In addition to phosphate, adverse effects have been seen by glucocorticoids and radiation, which yields and increase in apoptosis [52] [53] [54] [55]. In animals treated with a ten-day course of glucocorticoids, an increase of apoptosis in the hypertrophic chondrocytes yields a growth plate with a reduced width was revealed [53]. Both, PTHrH and parathyroid hormone are the main stimulators of the perichondrium. "
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    ABSTRACT: The epiphyseal growth plate develops from the cartilaginous-orientated mesenchymal cells that express SOX family genes. This multilayer structure is formed by the proliferation and hypertrophy of cells that synthesize the extracellular matrix composed of collagen (mainly type II, IX, X, XI) and proteoglycans (aggrecan, decorin, annexin II, V and VI). The resting zone is responsible for protein synthesis and maintaining a germinal structure. In the proliferative zone, cells rapidly duplicate. The subsequent morphological changes take place in the transformation zone, divided into the upper and lower hypertrophic layers. In the degenerative zone, the mineralization process becomes intensive due to increased release of alkaline phosphate, calcium and matrix vesicles by terminally differentiated chondrocytes and some other factors e.g., metaphyseal ingrowth vessels. At this level, as well as in the primary and secondary spongiosa zones, chondrocytes undergo apoptosis and are physiologically eliminated. Unlike adult cartilage, in fetal and early formed growth plates, unusual forms such as authophagal bodies, paralysis and dark chondrocytes were also observed. Their ultrastructure differs greatly from apoptotic and normal cartilage cells. Chondrocyte proliferation and differentiation are regulated by various endocrine, paracrine, and autocrine agents such as growth, thyroid and sex hormones, beta-catenin, bone morphogenetic proteins, insulin-like growth factor, iodothyronine deiodinase, leptin, nitric oxide, transforming growth factor beta and vitamin D metabolites. However, the most significant factor is parathyroid hormone-related protein (PTHrP) which is synthesized in the perichondrium by terminally differentiated chondrocytes. Secondary to activation of PTH/PTHrP receptors, PTHrP stimulates cell proliferation by G protein activation and delays their transformation into prehypertrophic and hypertrophic chondrocytes. When proliferation is completed, chondrocytes release Indian hedgehog (Ihh), which stimulates PTHrP synthesis via a feedback loop. Any disturbances of the epiphyseal development and its physiology result in various skeletal abnormalities known as dysplasia.
    Folia Histochemica et Cytobiologica 02/2009; 47(1):5-16. DOI:10.2478/v10042-009-0007-1 · 1.36 Impact Factor
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