Evaluation of Pinus plant species from North Western Himalayas for Osteoporosis

Abstract

newline xvi newlineAbstract newlineBackground: newlinePinus plant species are found in most of Himalayan states of which Pinus roxburghii and Pinus wallichiana are used traditionally as a paste to treat bone related disorders and fractures. Osteoporosis is a skeletal disorder associated with low bone mass and strength leading to bone fractures. It is highly prevalent especially in geriatric populations. For several years estrogen therapy was used clinically for treatment and management of osteoporosis, However their prolong use are associated with various side effects including breast cancer and cardiovascular diseases. The aim of the present study has been designed to evaluate the pharmacological activity of stem bark of Pinus plants extracts and fractions in surgically induced osteoporosis in female rats. The bioactive fraction was further developed into a tablet formulation. newlineMethods: newlineThe plant extracts and fractions are subjected for the phytochemical screening. The presence of gallic acid, quercetin, ascorbic acid, tannic acid, caffeic acid and catechin was quantified in plant extracts and in potent bioactive fraction using HPLC and HPTLC study. The three Pinus extracts Pinus roxburghii, Pinus wallichiana, Pinus gerardiana and further their four fractions prepared by using the solvents of graded polarity was subjected for antioxidant activity using DPPH, nitric oxide and hydrogen peroxide radicals assays. Anti-inflammatory activity was carried out using albumin denaturation and HRBC membrane stabilization assays. In-vitro osteoblastic proliferation was assessed on UMR-106 cell lines. Acute toxicity study of prepared extracts was conducted as per OECD423 guidelines. In vivo antiosteoporotic activity was done on ovariectomized female rats. The most active fraction was then formulated into tablet. newlineResults: newlineThe qualitative phytochemical analysis of hydro-alcoholic stem bark extracts of three plant species revealed the presence of various biochemical compounds such as alkaloids, flavonoids, glycosides, triterpenoids and saponins. Quantitative phytochemical analysis of plant extracts showed the presence of phenolics, flavonoids, tannins. Plant extracts of three Pinus species showed significant antioxidant activity against DPPH, nitric oxide and H2O2 radicals. In in-vitro antiand#8208;inflammatory investigation P. roxburghii and P. gerardiana newlinexvii newlineexhibited with percentage protection 86.54±1.85; 82.03±2.67.In in-vivo acute toxicity study studies Pinus plant extract revealed no sign of toxicity or mortality in rodents on. Administration of Pinus extracts and bioactive fractions prevented bone loss and showed marked efficacy against osteoporosis condition by modulating parameters, body weight change, biochemical parameters; serum phosphorus, calcium, estradiol, ALP and hydroxyproline levels. Biomechanical bone breaking strength for the tibia, 4th lumber, and femur bones, BMD, Pinus extract and bioactive fractions reversed the increased mRNA expression of RANKL, cathepsin K and decreased OPG expression in tibia bone tissue. Bone tissue histology showed the restoration of the disrupted trabecular network. The presence of gallic acid, quercetin and ascorbic acid in HPLC and HPTLC study signify their contribution to treating osteoporosis. The presence of phytoconstituents may be inhibiting bone loss, oxidative stress and suppress the release of inflammatory mediators. These may be responsible for the protective effect of extracts and fraction against the experimental model of osteoporosis. The most active n butanolic fraction of P.gerardiana was formulated into tablet and evaluation was done by testing the hardness, friability, disintegration time. newlineConclusion: The findings from present investigation may conclude that the Pinus plant species has efficacy to ameliorate the pathological state of osteoporosis via modulation of estrogen, RANK and cathepsin signaling. This effect may be due to the presence of quercetin, gallic acid, ascorbic acid, catechin, caffeic acid and tannic acid phytoconstituents. newlineKeywords: Osteoporosis, Pinus roxburghii, Pinus wallichiana, Pinus gerardiana, Bone, HPLC, HPTLC