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Title: Investigation of effects of novel selenium nanoparticles functionalized with cordyceps sinensis on osteoblastogenesis
Authors: Chan, Cheuk Hin
Degree: M.Phil.
Issue Date: 2018
Abstract: Osteoporosis is a skeletal disease characterized by low bone mass and density as well as deterioration in bone microarchitecture, leading to high risk of bone fracture. With an increase in aging population, osteoporosis has become one of the major public health issues nowadays, causing significant medical and socioeconomic burdens. Selenium is an essential trace mineral to human health. Substantial evidences have demonstrated that selenium deficiency is detrimental to bone microarchitecture and even associated with osteoporosis, suggesting its crucial role in bone metabolism. Recently, selenium nanoparticles (SeNPs) have become the new research target, since they were found to possess relatively low toxicity and remarkable anti-tumor efficacy compared to other organic and inorganic selenocompounds. Nevertheless, scientific research concerning their effects on bone health is currently very limited. By using our patented nanotechnology [US patent no.: 9,072,669], our research team has successfully prepared novel SeNPs (Cs4-SeNPs) using the mushroom polysaccharide isolated from Cordyceps sinensis. We discovered that Cs4-SeNPs existed as well-dispensed spherical particles in water with an average diameter of 73.2 ± 3.49nm and was highly stable without significant increase in size after 6 weeks under 4°C. Further characterization using TEM also showed that the particle size of SeNPs in Cs4-SeNPs was around 30nm. For individual SeNP, the clear lattice fringes (3.35Å), SAED pattern as well as EDX spectrum obtained by HR-TEM-EDX collectively indicated that the resulting nanoparticle possessed a polycrystalline structure with high level of Se (83.9%), implying a successfully fabrication of SeNPs using mushroom polysaccharides. Endocytosis has been widely reported as the major cellular uptake mechanism for nanoparticles. Our study found that coumarin-6 labelled Cs4-SeNPs mainly localized in lysosomes of the murine preosteoblast MC3T3-E1 subclone 4 cells (bone forming progenitor cells) as early as 12 min after cellular internalization. Besides, substantial previous findings have demonstrated that promoting bone formation is one of the effective strategies to prevent and/or manage postmenopausal osteoporosis. Unlike the common food source of organic (e.g. selenomethionine) and inorganic selenocompounds (e.g. selenite), our study indicated that Cs4-SeNPs was found to exhibit a significant dose-dependent proliferation effect (range from 1.35 - 1.45 folds) on the MC3T3-E1 cells for 24, 48 and 72 hrs with the most effect dosage of 10µM. More importantly, Cs4-SeNPs (10µM) were found to markedly induce both osteoblast differentiation and bone mineral formation of the MC3T3-E1 cells as evidenced by a significant increase in ALP activity (3.05 ± 0.15 folds), as well as an enhancement of bone nodule formation (Von Kossa and Alizarin Red S staining). Further investigation on their possible interaction with osteoclasts (bone resorption cells) also showed that Cs4-SeNPs could significantly up-regulate the gene expression ratio of the bone remodeling markers OPG/RANKL (2.56 ± 0.24 folds), suggesting their indirect inhibition effect on osteoclastogenesis in addition to promoting osteoblastogenesis.
Osteoblast differentiation is a crucial step of bone formation. Previous studies have demonstrated that BMP-2 signaling via Smad-dependent and/or Smad-independent pathway(s) is one of the most important signaling transduction cascades for regulating osteoblast differentiation. Interestingly, our study discovered that Cs4-SeNPs could trigger both BMP-2/Smad dependent and independent pathways simultaneously as evidenced by a significant upregulation of BMP-2 protein expression (1.46 ± 0.06 folds) as well as phosphorylation of Smad 1/5/8 (2.00 ± 0.25 folds) and p38 (1.60 ± 0.10 folds) proteins in the MC3T3-E1 cells after 3-day treatment. Besides, Cs4-SeNPs were found to significantly up-regulate the gene expression of those major downstream biomarkers in bone remodeling such as Dlx5 (1.71 ± 0.15 folds), Runx2 (1.48 ± 0.06 folds), Osx (1.46 ± 0.08 folds), ALP (1.77 ± 0.11 folds), and OCN (2.64 ± 0.14 folds), further supporting their active role in promotion of osteoblast differentiation and bone mineralization. Our long-term goal is to develop an evidence-based bone-forming agent for promoting the bone health of postmenopausal patients in our community.
Subjects: Hong Kong Polytechnic University -- Dissertations
Selenium -- Research
Cordyceps -- Therapeutic use
Pages: xiv, 78 pages : color illustrations
Appears in Collections:Thesis

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