Provider: - Institution: - Data provided by Europeana Collections- Kosti su stalno u procesu trošenja i obnavljanja, a ravnoteža razgradnje i izgradnje preduvjet je za zdravo koštano tkivo. Slabljenje kvalitete kosti posljedica je poremećaja te ravnoteže. Za razgradnju su odgovori osteoklasti dok su za izgradnju odgovorni osteoblasti. Osteopenija je stanje smanjene gustoće kosti, a do nje može doći i zbog dugotrajne primjene glukokortikosteroida. Glukokortikosteroidi su protuupalni lijekovi koji se, između ostalog, koriste za sprečavanje upale dišnih puteva kod astme. Budući je astma česta kronična bolest u djece potrebno je pratiti metaboličke i koštane učinke glukokortikosteroida tijekom terapije. Cilj ovog rada je uspostava modela hipoplazije fize izazvane glukokortikosteroidima i odabir optimalnih biljega njihovih metaboličkih i koštanih učinaka. Istražen je učinak 3 glukokortikosteroida na rast i promjenu metabolizma kosti te na ostale sustavne učinke u štakora. Beklometazon dipropionat, prednizolon i ciklezonid, davani mladim muškim Sprague-Dawley štakorima 7 dana u dozama od 0,3-10 mg/kg dnevno, s.c., su ovisno o dozi inhibirali indeks tjelesne mase timusa (za 57%, 44% i 76% s 3 mg/kg). Ciklezonid i manje učinkovit prednizolon su utjecali na ploču rasta glave femura inhibirajući rast femura (za 41% i 18% s 10 mg/kg), značajno smanjujući povećanje tjelesne mase (oboje za 100% s 10 mg/kg), te serumske koncentracije kisele fosfataze i tartarat rezistentne kisele fosfataze (za >30% s 10 mg/kg); oba su povećala serumske razine glukoze i triglicerida. Beklometazon dipropionat je imao slab učinak na ove dodatne varijable. Ciklezonid pokazuje izraženo inhibirajuće djelovanje na rast kosti u štakora. Možemo zaključiti da je ovo dobar model za ispitivanje utjecaja glukokortikosteroida na metabolizam kosti.- Bone in children is structurally different from adult bone. It is weaker but less brittle. Bone growth starts with cartilage formation. Then vessels invade the cartilage, delivering pluripotent stem cells, which initiate the formation of a primary center of ossification. Secondary ossification centers are formed at each end of long bone, and between the primary and secondary ossification centers the growth plate, or physis, develops. Bone grows as secondary and primary ossification centers unite. Bone tissue is in dinamic process of constant deteriorating and regeneration. Weakening of bone quality is a result of imbalance in a process of bone remodelling. Bone remodelling has two stages: bone resorption with the osteoclasts, bone cells which resorb the bone, and bone formation with osteoblasts, bone cells which form the bone. The bone remodelling cycle ends with bone mineralisation. The content of mineral in bones is defined as bone density. Osteopenia is a condition with decreased bone density. Apart being a sign of normal aging, osteopenia can be induced with prolonged use of glucocorticosteroids. Glucocorticosteroids are antiinflammatory medications prescribed, among others, for reducing and prevention inflammation of respiratory pathways in asthma. Since asthma is the most common chronic disease in children, need for monitoring metabolic and bone effects of glucocorticosteroids during therapy is appearing. Although inhaled glucocorticoids are known to have systemic effects on bone metabolism, there is little comparative information on their relative potencies. The goal of this work is establishment of glucocorticosteroid induced hypoplasia of the physis and finding the optimal markers of their metabolic and bone effects. The effects of three standard glucocorticoids, beclomethasone dipropionate, prednisolone and ciclesonide, in causing changes in bone metabolism and growth were investigated in relation to other systemic effects in the rat. Male, specific pathogen-free, Sprague-Dawley rats, 4,5–5,5 weeks old (at the beginning of the experiments), were used in the study. The rat femur model of glucocorticosteroid-induced hypoplasia of the physis was established according to Belvisi et al., using subcutaneous (s.c.) drug administration to allow for future parenteral comparison with novel compounds. Briefly, rats were randomly assigned to experimental groups of 8 animals each. In total, three experiments were performed for each glucocorticoid; beclomethasone dipropionate, prednisolone and ciclesonide, at doses of 0,3– 10 mg/kg daily for 7 days. Animals in the control groups received s.c. the volume of 10 ml/kg of vehicle (4% DMSO in 0,125% CMC) daily, for 7 days. Twenty-four hours after the last treatment, animals were anaesthetized with sodium thiopental and the blood was collected at exsanguination in order to obtain serum. Also, thymus weights were recorded and the femoral bones removed (for measurement of the thickness of the proliferating zone). Animal body weights were correspondingly documented at the beginning and at the end of each experiment. Body weight gain was calculated as the change in body weight from day 1 until 24 h after treatment on day 7. Biochemical analyses were performed on rat sera. Serum concentrations of glucose and triglycerides, alkaline and acid phosphatases, and tartrate-resistant acid phosphatase were determined on the biochemical analyzer. Concentration of osteocalcin, as a biochemical marker for bone formation, and TRACP 5b, as a biochemical marker for bone resorption, were also determined using ELISA. The thymus was dissected free of connective tissue and immediately weighed. Thymus body mass index (BMI) was calculated according to the following formula: BMI (thymus)=thymus weight (mg)/body mass (mg). The left femur was exposed and removed with the head intact in the acetabulum by cutting through the pelvic girdle and through the femur shaft above the knee joint. The tissue was then fixed in 10% neutral buffered formalin for histological assessment. For the purpose of quantitative histology of the femoral head proliferating zone femurs were fixed, decalcified and processed to paraffin using the unit for tissue processing. Threemicrometer- thick sections were cut in a way to include femoral head and stained. The femoral head growth plate was examined under a light microscope. Images of the growth plate were captured onto a computer. One image was captured from each tissue section, five measurements of the growth plate width being obtained from each calibrated image. Measurements involved drawing a line perpendicular to the growth plate between the edge of the hypertrophic zone, distal to the articular cartilage and the end of the proliferating zone. Daily treatment for 7 days with standard glucocorticoids resulted in significant increases in serum glucose and triglycerides concentrations at the highest doses (10 mg/kg) of prednisolone and ciclesonide. The most pronounced changes were observed with ciclesonide, which also significantly increased serum triglycerides at a daily dose of 3 mg/kg. None of the standard glucocorticoids had any significant effect on serum ALP, over the tested dose range, given daily for 7 days. However, at the highest dose (10 mg/kg), prednisolone and ciclesonide significantly inhibited both serum ACP and TRACP. The most pronounced changes were observed with ciclesonide, which also significantly decreased serum ACP and TRACP at a daily dose of 3 mg/kg. Beclomethasone dipropionate was less effective, causing a slight but significant decrease in serum ACP (but not TRACP) at doses of 0,3 and 1 mg/kg. The comparative potency of the three glucocorticoids in influencing non-specific serum parameters of bone metabolism was also reflected in their effects on the proliferating zone thickness of the femoral bone head. While beclomethasone dipropionate had no significant effect, ciclesonide and prednisolong decreased physeal growth plate width. Prednisolone reduced median bone growth by 18% at the highest dose (10 mg/kg) and ciclesonide caused a dosedependent reduction in median bone growth, with significant inhibition of up to 41% over the whole tested dose range (0,3–10 mg/kg). All three standard glucocorticoids exerted significant, dose related inhibitory effects on median body weight gain and thymus body mass indices after daily treatment for 7 days. Beclomethasone dipropionate was the least growth inhibitory; although it caused statistically significant inhibition of thymus BMIs by 50% at dose of 1 mg/kg per day, it caused statistically significant 23% inhibition of median body weight gain only at 10 mg/kg per day. Prednisolone affected thymus BMIs causing statistically significant inhibition by 44% at dose of 3 mg/kg per day and body weight gain causing statistically significant inhibition by 33% at dose of 1 mg/kg per day. Ciclesonide exerted the most pronounced inhibition of body weight gain and thymus BMIs, causing statistically significant inhibition by 34% and 55%, respectively, already at the lowest dose of 0,3 mg/kg per day. In order to see how examined glucocorticoids effected bone markers results showed that prednisolone had no statistical effect on bone formation while ciclesonide significantly reduced osteocalcin concentration in doses of 3 and 10 mg/kg per day (157 ng/mL and 88 ng/mL, respectively) vs. control (453 ng/mL). On the other hand, ciclesonid showed no significant effect on bone resorption while prednisolone significantly reduced TRACP 5b concentration at the highest dose of 10 mg/kg per day compared to negative control group (10,6±0,9 U/L vs. 19,7±3,1 U/L). Ciclesonide, although a pro-drug, still has potent systemic activity in the rat, causing typical glucocorticoid effects, including inhibition of bone growth. Prednisolone exhibits a similar, though less potent, spectrum of systemic activity, while beclomethasone dipropionate has weak activity in causing systemic metabolic effects, but retains thymus inhibiting potency. However, although the distinction between the effect of glucocorticoids on bone growth was observed in this stud