Proceedings of The Physiological Society

Physiology 2016 (Dublin, Ireland) (2016) Proc Physiol Soc 37, PCA213

Poster Communications

Melatonin and alendronate synergistically preserved bone matrix and increased trabecular thickness in rats with overiectomy

E. B. GURLER1, I. Peker2, O. T. Cilingir Kaya3, M. Akkiprik2, F. Ercan3, B. C. Yegen1

1. Physiology, Marmara University, Istanbul, Turkey. 2. Medical Biology and Genetics, Marmara University, Istanbul, Turkey. 3. Histology, Marmara University, Istanbul, Turkey.

BACKGROUND: Post-menopausal osteoporosis is frequently treated by bisphosphonates (e.g. alendronate) to maintain bone mass. Anti-inflammatory agent melatonin was suggested to have a regulatory role in bone physiology. The aim was to evaluate the possible additive anti-osteoporotic effect of melatonin. METHODS: Under anaesthesia with ketamine (100 mg/kg, intraperitoneally) and chlorpromazine (0.75mg/kg, intraperitoneally), female Sprague-Dawley rats (n=56) underwent bilateral ovariectomy (OVX), while control group had sham-surgery (n=8). Four weeks after the surgery, OVX rats were treated with saline, alendronate (70 µg/kg/week, subcutaneously), melatonin (25 mg/kg/day, orally), melatonin+alendronate, melatonin+melatonin receptor antagonist (luzindole, 10 µg/kg/day, intraperitoneally) or alendronate+melatonin+luzindole for 8 weeks. Rats were euthanized at the end of 12th week, while an additional saline-treated OVX group (n=8) was euthanized at the end of 4th week. Runx2 expression in bone marrow was determined using real-time polymerase chain reaction. Excised tibiae, fixed in formalin, were treated with decalcifier. Paraffin sections were stained with TUNEL kit for evaluation of apoptotic cells and with Masson's trichrome to evaluate bone matrix, mineralization and trabecular thickness. Statistical analysis was performed using Kruskal-Wallis and ANOVA tests. RESULTS: Runx2 expression was depressed in all OVX groups. Serum oestrogen level in the saline-treated groups was decreased at both the 4th and 12th weeks following OVX (p<0.05), while melatonin abolished this reduction. At the 12th week, saline-treated OVX group presented an extreme decrease in calcified area along with increased un-mineralized area and reduced thickness of the trabecular bone with separation of lamellae, while these alterations were milder at 4th week. In melatonin- or alendronate-treated groups, trabecular bones were mostly calcified, with new bone formation in some regions and less separated lamellae. In alendronate+melatonin-treated group, quite regular, mostly calcified trabecular bones were present, while trabecular thickness was similar to sham-operated group. Moderate decreases in calcified areas and in trabecular thickness, increased decalcified areas with severe separation of lamellae in trabecular bones were observed in melatonin + luzindole-treated group, while these histopathological alterations were milder in melatonin+luzindole+alendronate-treated group. Quantitative TUNEL analysis also revealed significant decreases in both alendronate-treated and melatonin-treated groups. CONCLUSION: Similar to alendronate, melatonin has direct anti-apoptotic and bone-mass-preserving effects without any additive action. The stimulatory effect of melatonin on trabecular thickness appears to be receptor-mediated.

Where applicable, experiments conform with Society ethical requirements