Effects of Parathyroid Hormone on Osteoporotic Fracture Healing in Mice via Non-Phospholipases C-Dependent Protein Kinase C Signaling Pathway

2021 ◽  
Vol 11 (5) ◽  
pp. 903-911
Author(s):  
Lei Wang ◽  
Linjuan Liu ◽  
Sixin Sun ◽  
Li Xiao ◽  
Qinyi Jiang ◽  
...  
Keyword(s):  
Parathyroid Hormone ◽  
Fracture Healing ◽  
Signaling Pathway ◽  
Osteoporotic Fracture ◽  
Expression Level ◽  
Enzyme Linked Immunosorbent Assay ◽  
Mineral Density ◽  
Pkc Inhibitor ◽  
Pkc Signaling

Objectives: This study was aimed to explore the effects of parathyroid hormone (PTH) on osteoporotic fracture healing in mice and the underlying mechanisms. Methods: Microarray analysis was conducted to analyze the gene expression level in MC3T3-E1 cells. Carboxyfluorescein succinimidyl ester (CFSE) staining and flow cytometry was adopted to analyze the proliferation and apopto-sis of MC3T3-E1 cells. qRT-PCR was used to analyze the mRNA expression level. Fluorescence resonance energy transfer (FRET) assay was conducted to detect PKC activity. The bone mineral density (BMD) and bone volume (BV)/total volume (TV) were determined via enzyme-linked immunosorbent assay (ELISA) and microscopic computed tomography (micro-CT). Results: ERK1/2 was abnormally expressed in MC3T3-E1 cells after GlylArg19hPTH (1-34) + KT5720 treatment. GlylArg19hPTH (1-34)+ KT5720 treatment promoted cell proliferation, inhibited cell apoptosis, and upregulatedthe expression of osteogenesis-related genes (ALP, OPN, Runx2 and OPG) in MC3T3-E1 cells, which were due to the activation of the non-PLC-dependent PKC signaling pathway and can be blocked by PKC inhibitor Go6983 or ERK1/2 inhibitor BVD-523. Moreover, the activity of PKC in MC3T3-E1 cells treated with GlylArg19hPTH (1-34) + KT5720 + Go6983 was alleviated by ERK1/2 inhibitor BVD-523. In vivo, specific activation of the non-PLC-dependent PKC signaling pathway increased the serum levels of APL and OPG in mice with osteoporotic fracture, which were reversed by PKC inhibitor Go6983 and ERK1/2 inhibitor BVD-523. Moreover, PKC inhibitor Go6983 and ERK1/2 inhibitor BVD-523 suppressed the elevation of BV/TV and BMD induced by specific activation of the non-PKC-dependent signaling pathway. Conclusions: Taken together, PTH stimulates osteoporotic fracture healing in mice through the non-PLC-dependent PKC signaling pathway in which ERK1/2 exerts a vital role.

scholarly journals DNA methylation mediated RSPO2 to promote follicular development in mammals

2021 ◽  
Vol 12 (7) ◽  
Author(s):  
Xiaofeng Zhou ◽  
Yingting He ◽  
Nian Li ◽  
Guofeng Bai ◽  
Xiangchun Pan ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Wnt Signaling ◽  
Signaling Pathway ◽  
Methylation Level ◽  
Molecular Mechanisms ◽  
Cpg Island ◽  
Wnt Signaling Pathway ◽  
Follicular Development ◽  
Expression Level

AbstractIn female mammals, the proliferation, apoptosis, and estradiol-17β (E2) secretion of granulosa cells (GCs) have come to decide the fate of follicles. DNA methylation and RSPO2 gene of Wnt signaling pathway have been reported to involve in the survival of GCs and follicular development. However, the molecular mechanisms for how DNA methylation regulates the expression of RSPO2 and participates in the follicular development are not clear. In this study, we found that the mRNA and protein levels of RSPO2 significantly increased during follicular development, but the DNA methylation level of RSPO2 promoter decreased gradually. Inhibition of DNA methylation or DNMT1 knockdown could decrease the methylation level of CpG island (CGI) in RSPO2 promoter and upregulate the expression level of RSPO2 in porcine GCs. The hypomethylation of −758/−749 and −563/−553 regions in RSPO2 promoter facilitated the occupancy of transcription factor E2F1 and promoted the transcriptional activity of RSPO2. Moreover, RSPO2 promoted the proliferation of GCs with increasing the expression level of PCNA, CDK1, and CCND1 and promoted the E2 secretion of GCs with increasing the expression level of CYP19A1 and HSD17B1 and inhibited the apoptosis of GCs with decreasing the expression level of Caspase3, cleaved Caspase3, cleaved Caspase8, cleaved Caspase9, cleaved PARP, and BAX. In addition, RSPO2 knockdown promoted the apoptosis of GCs, blocked the development of follicles, and delayed the onset of puberty with decreasing the expression level of Wnt signaling pathway-related genes (LGR4 and CTNNB1) in vivo. Taken together, the hypomethylation of −758/−749 and −563/−553 regions in RSPO2 promoter facilitated the occupancy of E2F1 and enhanced the transcription of RSPO2, which further promoted the proliferation and E2 secretion of GCs, inhibited the apoptosis of GCs, and ultimately ameliorated the development of follicles through Wnt signaling pathway. This study will provide useful information for further exploration on DNA-methylation-mediated RSPO2 pathway during follicular development.

scholarly journals Guhong Injection promotes fracture healing by activating Wnt/beta-catenin signaling pathway in vivo and in vitro

2019 ◽  
Vol 120 ◽  
pp. 109436 ◽  
Author(s):  
Zhizhen Sun ◽  
Hongting Jin ◽  
Huifen Zhou ◽  
Li Yu ◽  
Haitong Wan ◽  
...  
Keyword(s):  
Fracture Healing ◽  
Signaling Pathway ◽  
Beta Catenin

Osteogenic activity of silymarin through enhancement of alkaline phosphatase and osteocalcin in osteoblasts and tibia-fractured mice

2012 ◽  
Vol 237 (4) ◽  
pp. 417-428 ◽  
Author(s):  
Jung-Lye Kim ◽  
Sin-Hye Park ◽  
Daewon Jeong ◽  
Ju-Suk Nam ◽  
Young-Hee Kang
Keyword(s):  
Alkaline Phosphatase ◽  
Fracture Healing ◽  
Bone Strength ◽  
Nodule Formation ◽  
Signal Pathways ◽  
Dependent Manner ◽  
Mineral Density ◽  
Alp Activity ◽  
Collagen Secretion

Bone-remodeling imbalance induced by increased bone resorption and osteoclast formation is known to cause skeletal diseases such as osteoporosis. There has been growing interest in the anabolic natural agents that enhance bone formation. Silymarin is flavonolignans extracted from blessed milk thistle. Several studies suggest that silymarin possesses antihepatotoxic properties and anticancer effects against carcinoma cells. This study investigated promoting effects of silymarin on differentiation and mineralization of osteoblastic MC3T3-E1 mouse cells and on bone mineral density (BMD) by in vivo fracture experiments. Osteoblasts were treated with 1–20 μmol/L silymarin for 15 days in a differentiating medium. In addition, this study explored signaling pathways implicated in the osteoblastogenesis of silymarin. It was found that silymarin stimulated alkaline phosphatase (ALP) activity and calcium nodule formation in a dose-dependent manner with a substantial effect on osteoblast proliferation. Silymarin treatment enhanced collagen secretion, osteocalcin transcription and bone morphogenetic protein (BMP) expression. The BMP inhibitor noggin suppressed the silymarin-promoted ALP activity in differentiated osteoblasts, suggesting that its osteoblastogenic actions entail the BMP pathway. This was proved by increased SMAD1/5/8 phosphorylation and runt-related transcription factor 2 (Runx2) expression in the presence of silymarin. In 21-day fracture-healing experiments, fractured and silymarin (10 mg/kg)-treated C57BL/6 mice showed better bone healing than fractured mice. Silymarin supplementation improved tibial bone strength with elevated BMD and serum levels of osteogenic ALP and osteocalcin. Taken together, these results demonstrate, for the first time, that silymarin has a potential to enhance osteoblastogenesis through accelerating BMP/SMAD/Runx2 signal pathways and to improve fracture healing and bone strength in mouse tibiae.

scholarly journals Paeoniflorin Attenuates Inflammatory Pain by Inhibiting Microglial Activation and Akt-NF-κB Signaling in the Central Nervous System

2018 ◽  
Vol 47 (2) ◽  
pp. 842-850 ◽  
Author(s):  
Bo Hu ◽  
Guangtao Xu ◽  
Xiaomin Zhang ◽  
Long Xu ◽  
Hong Zhou ◽  
...  
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Signaling Pathway ◽  
Inflammatory Pain ◽  
Microglial Activation ◽  
Enzyme Linked Immunosorbent Assay ◽  
Pain Model ◽  
The Central Nervous System

Background/Aims: Paeoniflorin (PF) is known to have anti-inflammatory and paregoric effects, but the mechanism underlying its analgesic effect remains unclear. The aim of this study was to clarify the effect of PF on Freund’s complete adjuvant (CFA)-induced inflammatory pain and explore the underlying molecular mechanism. Methods: An inflammatory pain model was established by intraplantar injection of CFA in C57BL/6J mice. After intrathecal injection of PF daily for 8 consecutive days, thermal and mechanical withdrawal thresholds, the levels of inflammatory factors TNF-α, IL-1β and IL-6, microglial activity, and the expression of Akt-NF-κB signaling pathway in the spinal cord tissue were detected by animal ethological test, cell culture, enzyme-linked immunosorbent assay, immunofluorescence histochemistry, and western blot. Results: PF inhibited the spinal microglial activation in the CFA-induced pain model. The production of proinflammatory cytokines was decreased in the central nervous system after PF treatment both in vivo and in vitro. PF further displayed a remarkable effect on inhibiting the activation of Akt-NF-κB signaling pathway in vivo and in vitro. Conclusion: These results suggest that PF is a potential therapeutic agent for inflammatory pain and merits further investigation.

scholarly journals Reduced H3K27me3 Suppresses Wnt/β-catenin Signaling by S-adenosylmethionine Deficiency in Neural Tube Development

2021 ◽  
Author(s):  
Li Zhang ◽  
Xiuwei Wang ◽  
Rui Cao ◽  
Dandan Li ◽  
Yufei Wang ◽  
...  
Keyword(s):  
Incidence Rate ◽  
Signaling Pathway ◽  
Neural Tube ◽  
Underlying Mechanism ◽  
Enzyme Linked Immunosorbent Assay ◽  
The Novel ◽  
Marker Proteins ◽  
Neural Tube Development ◽  
Gsk 3Β

Abstract Background: S-adenosylmethionine as a major methyl donor play a key role in methylation modification in vivo, and its disorder was closely related to neural tube defects. However, the underlying mechanism between SAM deficiency and NTDs remained unclear.Methods: we investigated the association between histone methylation modification and Wnt/β-catenin signaling pathway in NTDs induced by SAM deficiency. The levels of SAM and SAH were determined by enzyme linked immunosorbent assay. The expressions of H3K27me3 and Wnt/β-catenin signaling pathway specific markers were demonstrated by western blotting, reverse transcription, and quantitative PCR and immunofluorescence in ethionine induced E11.5 mouse NTDs and NSCs models. Results: we found that the incidence rate of NTDs induced by ethionine were 46.2%, post treatment of ethionine combined with SAM, the incidence rate of NTDs was reduced to 26.2%. The level of SAM was significantly decreased (P<0.05) and a reduction in the SAM/SAH ratio was observed. The SAM depletion caused the reduction of both H3K27me3 modifications and UTX activity, and inhibited the marker proteins (β-catenin, TCF-4, Axin-2, p-GSK-3β, CyclinD1, and C-myc) in Wnt/β-catenin signaling pathway (P<0.05). The differentiations of neural stem cells into neurons and oligodendrocytes were inhibited under SAM deficiency (P<0.05).Conclusions: These results indicated that the depletion of SAM led to reduced H3K27me3 modifications, prevented the activation of Wnt/β-catenin signaling pathway and NSCs differentiation, which provided an understanding of the novel function of epigenetic regulation in NTDs.

scholarly journals The Recombinant Protein EphB4-Fc Changes the Ti Particle-Mediated Imbalance of OPG/RANKL via EphrinB2/EphB4 Signaling Pathway and Inhibits the Release of Proinflammatory Factors In Vivo

2020 ◽  
Vol 2020 ◽  
pp. 1-15
Author(s):  
Yu-Wei Ge ◽  
Kai Feng ◽  
Xiao-Liang Liu ◽  
Hong-Fang Chen ◽  
Zhen-Yu Sun ◽  
...  
Keyword(s):  
Recombinant Protein ◽  
Inflammatory Response ◽  
Signaling Pathway ◽  
Osteoclast Differentiation ◽  
Wear Particle ◽  
Enzyme Linked Immunosorbent Assay ◽  
Tartrate Resistant Acid Phosphatase ◽  
Wear Particles ◽  
Proinflammatory Factors

Aseptic loosening caused by wear particles is one of the common complications after total hip arthroplasty. We investigated the effect of the recombinant protein ephB4-Fc (erythropoietin-producing human hepatocellular receptor 4) on wear particle-mediated inflammatory response. In vitro, ephrinB2 expression was analyzed using siRNA-NFATc1 (nuclear factor of activated T-cells 1) and siRNA-c-Fos. Additionally, we used Tartrate-resistant acid phosphatase (TRAP) staining, bone pit resorption, Enzyme-linked immunosorbent assay (ELISA), as well as ephrinB2 overexpression and knockdown experiments to verify the effect of ephB4-Fc on osteoclast differentiation and function. In vivo, a mouse skull model was constructed to test whether the ephB4-Fc inhibits osteolysis and inhibits inflammation by micro-CT, H&E staining, immunohistochemistry, and immunofluorescence. The gene expression of ephrinB2 was regulated by c-Fos/NFATc1. Titanium wear particles activated this signaling pathway to the promoted expression of the ephrinB2 gene. However, ephrinB2 protein can be activated by osteoblast membrane receptor ephB4 to inhibit osteoclast differentiation. In in vivo experiments, we found that ephB4 could regulate Ti particle-mediated imbalance of OPG/RANKL, and the most important finding was that ephB4 relieved the release of proinflammatory factors. The ephB4-Fc inhibits wear particle-mediated osteolysis and inflammatory response through the ephrinB2/EphB4 bidirectional signaling pathway, and ephrinB2 ligand is expected to become a new clinical drug therapeutic target.

Parathyroid hormone increases the expression level of matrix metalloproteinase-13 in vivo

2001 ◽  
Vol 19 (4) ◽  
pp. 207-212 ◽  
Author(s):  
Motoyuki Uchida ◽  
Hideyuki Yamato ◽  
Yumiko Nagai ◽  
Hiroshi Yamagiwa ◽  
Tadashi Hayami ◽  
...  
Keyword(s):  
Parathyroid Hormone ◽  
Matrix Metalloproteinase ◽  
Expression Level ◽  
Matrix Metalloproteinase 13

scholarly journals Zymosan-a Protects the Hematopoietic System from Radiation-Induced Damage by Targeting TLR2 Signaling Pathway

2017 ◽  
Vol 43 (2) ◽  
pp. 457-464 ◽  
Author(s):  
Jicong Du ◽  
Ying Cheng ◽  
Suhe Dong ◽  
Pei Zhang ◽  
Jiaming Guo ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Cell Viability ◽  
Signaling Pathway ◽  
Enzyme Linked Immunosorbent Assay ◽  
Hematopoietic System ◽  
Zymosan A ◽  
Radiation Induced ◽  
Total Body ◽  
Tlr2 Signaling

Background/Aims: The hematopoietic system is vulnerable to ionizing radiation and is often severely damaged by radiation. Molecules affecting radioresistance include Toll-like receptor 2. We investigated whether Zymosan-A, a novel TLR2 agonist, can protect the hematopoietic system from radiation-induced damage after total body irradiation. Methods: Mice were exposed to total body radiation after treatment with Zymosan-A or normal saline, and their survival was recorded. Tissue damage was evaluated by hematoxylin–eosin staining. The number of nucleated cells in bone marrow was determined by flow cytometry. Cell viability and apoptosis assay were determined by CCK-8 assay and flow cytometry assay. Enzyme-linked immunosorbent assay was used to detect the level of cytokines. Results: Zymosan-A protected mice from radiation-induced death and prevented radiation-induced hematopoietic system damage. Zymosan-A also promoted cell viability and inhibited cell apoptosis caused by radiation, induced radioprotective effects via TLR2, upregulated IL-6, IL-11, IL-12, and TNF-α in vivo. Conclusion: Zymosan-A can provide protection against radiation-induced hematopoietic system damage by targeting the TLR2 signaling pathway. Thus, Zymosan-A can be potentially effective radioprotectant.

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