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Bone Regeneration Effect of Cassia occidentalis Linn. Extract and Its Isolated Compounds
Published in Brijesh Kumar, Vikas Bajpai, Vikaskumar Gond, Subhashis Pal, Naibedya Chattopadhyay, Phytochemistry of Plants of Genus Cassia, 2021
Brijesh Kumar, Vikas Bajpai, Vikaskumar Gond, Subhashis Pal, Naibedya Chattopadhyay
The anti-oxidant effect of kaempferol in osteoblasts has been studied. 2-Deoxy-D-ribose (dRib) is a sugar with a high reducing capacity and induces oxidative stress and kaempferol mitigates dRib-induced oxidative stress in MC3T3-E1 cells by reducing malondialdehyde content (Suh et al., 2009). Kaempferol is also known to activate both ERα- and ERβ transactivation and promote the expression of osteogenic genes (Guo et al., 2012; Tang et al., 2008). In MC3T3-E1 cells, induction of differentiation by kaempferol was accompanied by autophagy assessed by the expression of beclin-1, sequestosome-1, and conversion of LC3-II to LC3-I (Kim et al., 2016). Kaempferol also protects osteoblasts against dexamethasone-induced apoptosis through the activation of the ERK pathway (Adhikary et al., 2018). Together the data suggest that kaempferol promotes osteogenic differentiation, protects against oxidative stress, and drug-induced apoptosis. The signaling mechanisms are varied including ER, BMP, mTOR and autophagy pathways.
Methods for Characterization of Bioactivity Using Confocal Microscopy *
Published in Mary Anne S. Melo, Designing Bioactive Polymeric Materials for Restorative Dentistry, 2020
Jirun Sun, Joy P. Dunkers, Sheng Lin-Gibson, Nancy J. Lin
The MC3T3-E1 subclone 4 murine pre-osteoblast cell line was purchased from the American Type Culture Collection and maintained in alpha Minimum Essential Medium Eagle (α-MEM) supplemented with 10% (volume fraction) fetal bovine serum, 2 mmol/L L-glutamine, and 1% (volume fraction) penicillin/streptomycin at 37°C, 5% (by volume) CO2. Cells of passages 3–6 were used in this study. Scaffolds were sterilized using ethylene oxide, degassed for at least three days, and hydrated through an ethanol series with moderate vacuum to facilitate wetting throughout the scaffold pores. Scaffolds were then transferred to a 48-well plate and prewetted with growth medium for 1 h prior to seeding. For scaffold bars, 1.4 × 106 cells were evenly distributed along the bar. Cylindrical scaffolds were each seeded with 1 × 105 or 5 × 105 cells. (The seeding density for the scaffold bars is equivalent to 2.5 × 105 cells on the cylindrical scaffolds.) The drop-wise seeding protocol for the cylindrical scaffolds was optimized in preliminary studies. Briefly, after removing the medium used for prewetting, half of the cells were suspended in 250 µL and added to the scaffold by pipetting up and down. After 1 h incubation at 37°C, the scaffolds were turned bottom-up and seeded dropwise (without pipetting) with the remaining cells suspended in 125 µL medium. After 1 h incubation, scaffolds were placed top-up in new 12-well plates, covered with 3 mL medium, and returned to the incubator. The growth medium was changed every three days.
Animal Models of Bone Defect Repair
Published in Yuehuei H. An, Richard J. Friedman, Animal Models in Orthopaedic Research, 2020
Yuehuei H. An, Richard J. Friedman
Bone grafting is aimed to provide the missing elements necessary for bone formation in a bone defect and thereby restore the bone integrity. Using cell-seeding to a substrate to make an implantable graft is not a new concept.147,148 Recently, a few groups reported some preliminary and very important data on the use of cell-seeded implants for repairing osseous or chondral defects.93,149-151 In 1991, Frayssinet et al. reported that bone cells from canine humeri were grown on HA granules and the cell-HA composite was placed in a bioreactor and implanted into a canine ulna defect. Osteogenesis was seen in active bioreactors three weeks after implantation.93 Cultured chondrocytes bound to a HA block were implanted to repair a rabbit ulna defect.70 Osteoblast-like cells (MC3T3-E1) were also used to study the potential of bioabsorbable polymers and ceramics to support osteoblastic growth for a bone-polymer composite in bone repair.150
Acteoside Derived from Cistanche Improves Glucocorticoid-Induced Osteoporosis by Activating PI3K/AKT/mTOR Pathway
Published in Journal of Investigative Surgery, 2023
Shumei Li, Yajie Cui, Min Li, Wenting Zhang, Xiaoxue Sun, Zhaoxu Xin, Jing Li
We assessed the potential molecular mechanisms to further explore the protective role of ACT on bone formation and differentiation in GIOP Zhou et al. screened potential therapeutic targets for osteoporosis by informatics methods, and identified mTOR [34]. HIF-1α regulates GIOP through the AKT/mTOR signaling pathway [35]. More importantly, ACT derived from Radix Rehmanniae was shown to inhibit bone loss and structural deterioration and promote bone formation by acting on the PI3K/AKT/mTOR pathway [19]. Erythrina Sativa activates autophagy via the PI3K/AKT/mTOR pathway through involvement in rat osteoblast differentiation [22]. Resveratrol protects from OP by regulating the SIRT1 and PI3K/AKT/mTOR pathways [25]. Therefore, we focused on the PI3K/AKT/mTOR pathway in this study. We confirmed that Dex inhibited the levels of key proteins in these signaling pathways. In contrast, ACT attenuated this inhibition, and the protective functions of ACT on osteoblast viability, apoptosis, osteogenic differentiation, and mineralization were reversed with the addition of a signaling pathway inhibitor. This study in fact has some limitations to consider when explaining the findings. Firstly, it is unfortunate that HE histology was not provided to confirm our experimental results, which was a flaw in the initial design of our experiments. Furthermore, an osteoblast-like cell line, MC3T3-E1 cells, which are also commonly used in osteoporosis, was used in this study, and we will use primary osteoblasts or bone marrow stromal cells for in vitro studies in subsequent more in-depth studies.
Morroniside ameliorates glucocorticoid-induced osteoporosis and promotes osteoblastogenesis by interacting with sodium-glucose cotransporter 2
Published in Pharmaceutical Biology, 2023
Hou-Zhi Yang, Runbei Dong, Yutao Jia, Yuqiao Li, Gan Luo, Tianhao Li, Yao Long, Shuang Liang, Shanshan Li, Xin Jin, Tianwei Sun
We then evaluated the effects of MOR on osteogenic differentiation and alkaline phosphatase (ALP) activity by using ARS and ALP staining, respectively. After seven days of culture with MOR, ARS and ALP staining demonstrated that 20 µM of MOR significantly increased the differentiation and ALP activity of MC3T3-E1 cells (Figure 1(D,F)). The qRT-PCR results revealed that the mRNA levels of ALP (Figure 2(A)), Col I (Figure 2(B)) and BMP2 (Figure 2(C)) were significantly increased in the MOR treatment groups. Furthermore, compared to the controls, the protein levels of principal osteogenic markers, such as OPN and Runx2, were significantly elevated in the 10 and 20 µM MOR groups (Figure 2(E,F)). These results indicated that 20 µM MOR remarkably promoted the proliferation and differentiation of MC3T3-E1 cells.
LncRNA LINC00963 promotes osteogenic differentiation of hBMSCs and alleviates osteoporosis progression by targeting miRNA-760/ETS1 axis
Published in Autoimmunity, 2021
Lirong Ren, Limin Guo, Nannan Kou, Jia Lv, Zhihua Wang, Kaishun Yang
Human bone marrow mesenchymal stem cells (hBMSCs) were isolated from discarded femoral head tissues of postmenopausal women with OP fractures or non-OP fractures (control groups) as previously described [23]. HEK 293T cells and a mouse calvaria-derived preosteoblast cell line, MC3T3-E1 were purchased from the Chinese Academy of Sciences Cell Bank (Shanghai, China). Cells were cultured with the phenol-red-free α-MEM medium containing 10% FBS-HI, 100 U/mL penicillin and 100 μg/mL streptomycin at 37 °C with 5% CO2. For osteogenic induction, the concentration of FBS-HI was decreased from 10% to 1%, meanwhile 50 μg/mL ascorbic acid, 10 mM β-glycerophosphate, and 0.1 μg/mL dexamethasone (osteogenic medium, OM) was added to cells and incubated for 21 d. The morphological identification of cultured hBMSCs was performed using transmission electron microscopy (TEM) as previously described [24].