scholarly journals Tussilagone Inhibits Osteoclastogenesis and Periprosthetic Osteolysis by Suppressing the NF-κb and p38 MAPK Signaling Pathways

2019 ◽  
Author(s):  
Xuantao Hu ◽  
Zhengxiao Ouyang ◽  
Dan Peng ◽  
Ziqing Yin ◽  
Xia Chen ◽  
...  
Keyword(s):  
Bone Resorption ◽  
Signaling Pathways ◽  
P38 Mapk ◽  
Western Blotting ◽  
Mapk Signaling ◽  
Luciferase Reporter ◽  
Periprosthetic Osteolysis ◽  
Prosthetic Loosening ◽  
Mapk Signaling Pathways

Abstract Backgroud: Aseptic prosthetic loosening is one of main factor producing poor prognosis of limb function after joint replacement and requiring troublesome revision surgery. It is featured by wear particle–induced periprosthetic osteolysis mediated by excessive osteoclasts activated in inflammatory cell context. In our previous study, some natural compounds showing anti-osteoclast trait with high cost-efficiency and few side effects. Tussilagone (TUS), as the main functional extract from Tussilago Farfara precedently used for relieving cough, asthma and eliminating phlegm in traditional medicine, has been proved to appease several RAW264.7-mediated inflammatory diseases via suppressing osteoclast-related signaling cascades. However, whether and how TUS can improve aseptic prosthetic loosening via modulating osteoclast-mediated bone resorption still need to be answered. Methods: We established a murine calvarial osteolysis model to detect the preventative effect of TUS on osteolysis in vivo. Micro-CT scanning and histomorphometric analysis were used to determine the variation of bone resorption and osteoclastogenesis in samples. The anti-osteoclast-differentiation and anti-bone-resorption bioactivities of TUS in vitro were investigated using bone slice resorption pit evaluation and interference caused by cytotoxicity of TUS was excluded according to CCK-8 assay. Quantitative PCR analysis was applied to prove the decreased expression of osteoclast-specific genes after TUS treatment. The inhibition effect of TUS on NF-κb and p38 MAPK signaling pathways was testified by western blotting and NF-κB-linked luciferase reporter gene assay.Results: TUS demonstrated bone protective effect against osteolysis in murine calvarial osteolysis model with reduced osteoclasts compared to the control group. Following studies in vitro witnessed that TUS exert anti-osteoclastogenesis and anti-bone-resorption effects in both BMMs and RAW264.7 cells, as evidenced by the decline of osteoclast specific genes according to quantative PCR. Western blotting revealed that TUS-treated demonstrated inhibited IκBα degradation and p38 phosphorylation.Conclusions: Collectively, for the first time our studies prove that TUS inhibits osteoclastogenesis by suppressing the NF-κb and p38 MAPK signaling pathways, therefore serving as a potential natural compound to treat periprosthetic osteolysis-induced aseptic prosthetic loosening.

scholarly journals Cimifugin Suppresses NF-κB Signaling to Prevent Osteoclastogenesis and Periprosthetic Osteolysis

2021 ◽  
Vol 12 ◽  
Author(s):  
Juan Duan ◽  
Xuantao Hu ◽  
Tao Li ◽  
Gen Wu ◽  
Pengcheng Dou ◽  
...  
Keyword(s):  
Bone Resorption ◽  
Western Blot ◽  
Signaling Pathway ◽  
Osteoclast Differentiation ◽  
Mapk Signaling ◽  
Luciferase Reporter ◽  
Pcr Analysis ◽  
Periprosthetic Osteolysis

Background: Aseptic loosening of prosthesis (ALP) is one of the most common long-term complications of knee and hip arthroplasty. Wear particle-induced osteoclastogenesis and subsequent periprosthetic osteolysis account for the morbidity of ALP. Here, we investigate the potential of cimifugin (CIM), a natural extract from Cimicifuga racemosa and Saposhnikovia divaricata, as a bone-protective drug in the treatment of ALP.Method: First, we performed cell viability and osteoclast formation assays to assess the effect of noncytotoxic CIM on osteoclast differentiation in vitro. Bone slice resorption and F-actin ring immunofluorescence assays were adopted to assess the effects of CIM on bone-resorption function. Then, quantitative real-time polymerase chain reaction (qRT–PCR) analysis was performed to further assess the repressive effects of CIM on osteoclastogenesis at the gene expression level. To elucidate the mechanisms underlying the above findings, Western blot and luciferase reporter gene assays were used to assess the regulatory effects of CIM on the NF-κB and MAPK signaling pathways. Moreover, a Ti particle-induced murine calvarial osteolysis model and subsequent histomorphometric analysis via micro-CT and immunohistochemical staining were used to elucidate the effect of CIM on periprosthetic osteolysis in vivo.Result: CIM dose-dependently inhibited both bone marrow-derived macrophage (BMM)- and RAW264.7 cell-derived osteoclastogenesis and bone resorption pit formation in vitro, which was further supported by the reduced expression of F-actin and osteoclast-specific genes. According to the Western blot analysis, inhibition of IκBα phosphorylation in the NF-κB signaling pathway, not the phosphorylation of MAPKs, was responsible for the suppressive effect of CIM on osteoclastogenesis. Animal experiments demonstrated that CIM alleviated Ti particle-induced bone erosion and osteoclast accumulation in murine calvaria.Conclusion: The current study suggested for the first time that CIM can inhibit RANKL-induced osetoclastogenesis by suppressing the NF-κB signaling pathway in vitro and prevent periprosthetic osteolysis in vivo. These findings suggest the potential of CIM as a therapeutic in ALP.

scholarly journals Hsp90 Modulates Human Sperm Capacitation via the Erk1/2 and p38 MAPK Signaling Pathways

2020 ◽  
Author(s):  
Peibei Sun ◽  
Yayan Wang ◽  
Tian Gao ◽  
Kun Li ◽  
Dongwang Zheng ◽  
...  
Keyword(s):  
Signaling Pathways ◽  
P38 Mapk ◽  
Sperm Motility ◽  
Western Blotting ◽  
Protein Complex ◽  
Acrosome Reaction ◽  
Human Sperm ◽  
Mapk Signaling ◽  
Sperm Capacitation ◽  
Mapk Signaling Pathways

Abstract Background: Heat shock protein 90 (Hsp90) is a highly abundant eukaryotic molecular chaperone that plays important roles in client protein maturation, protein folding and degradation, and signal transduction. Previously, we found that both Hsp90 and its co-chaperone cell division cycle protein 37 (Cdc37) were expressed in human sperm. Hsp90 is known to be involved in human sperm capacitation via unknown underlying mechanism(s). As Cdc37 was a kinase-specific co-chaperone of Hsp90, Hsp90 may regulate human sperm capacitation via other kinases. It has been reported that two major mitogen-activated protein kinases (MAPKs), extracellular signal-regulated kinase 1/2 (Erk1/2) and p38, are expressed in human sperm in the same locations as Hsp90 and Cdc37. Phosphorylated Erk1/2 has been shown to promote sperm hyperactivated motility and acrosome reaction, while phosphorylated p38 inhibits sperm motility. Therefore, in this study we explored whether Hsp90 modulates human sperm capacitation via the Erk1/2 and p38 MAPK signaling pathways. Methods: Human sperm was treated with the Hsp90-specific inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG) during capacitation. Computer-assisted sperm analyzer (CASA) was used to detect sperm motility and hyperactivation. The sperm acrosome reaction was analyzed by using fluorescein isothiocyanate-conjugated Pisum sativum agglutinin (PSA-FITC) staining. The interactions between Hsp90, Cdc37, Erk1/2 and p38 were assessed using co-immunoprecipitation (Co-IP) experiments. Western blotting analysis was used to evaluate the levels of protein expression and phosphorylation. Results: Human sperm hyperactivation and acrosome reaction were inhibited by 17-AAG, suggesting that Hsp90 is involved in human sperm capacitation. In addition, Co-IP experiments revealed that 17-AAG reduced the interaction between Hsp90 and Cdc37, leading to the dissociation of Erk1/2 from the Hsp90-Cdc37 protein complex. Western blotting analysis revealed that levels of Erk1/2 and its phosphorylated form were subsequently decreased. Decreasing of Hsp90-Cdc37 complex also affected the interaction between Hsp90 and p38. Nevertheless, p38 dissociated from the Hsp90 protein complex and was activated by autophosphorylation. Conclusions: Taken together, our findings indicate that Hsp90 is involved in human sperm hyperactivation and acrosome reaction. In particular, Hsp90 and its co-chaperone Cdc37 form a protein complex with Erk1/2 and p38 to regulate their kinase activity. These results suggest that Hsp90 regulates human sperm capacitation via the Erk1/2 and p38 MAPK signaling pathways.

scholarly journals Hsp90 modulates human sperm capacitation via the Erk1/2 and p38 MAPK signaling pathways

2021 ◽  
Author(s):  
Peibei Sun ◽  
Yayan Wang ◽  
Tian Gao ◽  
Kun Li ◽  
Dongwang Zheng ◽  
...  
Keyword(s):  
Signaling Pathways ◽  
P38 Mapk ◽  
Sperm Motility ◽  
Western Blotting ◽  
Protein Complex ◽  
Acrosome Reaction ◽  
Human Sperm ◽  
Mapk Signaling ◽  
Sperm Capacitation ◽  
Mapk Signaling Pathways

Abstract Background: Heat shock protein 90 (Hsp90) is a highly abundant eukaryotic molecular chaperone that plays important roles in client protein maturation, protein folding and degradation, and signal transduction. Previously, we found that both Hsp90 and its co-chaperone cell division cycle protein 37 (Cdc37) were expressed in human sperm. Hsp90 is known to be involved in human sperm capacitation via unknown underlying mechanism(s). As Cdc37 was a kinase-specific co-chaperone of Hsp90, Hsp90 may regulate human sperm capacitation via other kinases. It has been reported that two major mitogen-activated protein kinases (MAPKs), extracellular signal-regulated kinase 1/2 (Erk1/2) and p38, are expressed in human sperm in the same locations as Hsp90 and Cdc37. Phosphorylated Erk1/2 has been shown to promote sperm hyperactivated motility and acrosome reaction, while phosphorylated p38 inhibits sperm motility. Therefore, in this study we explored whether Hsp90 modulates human sperm capacitation via the Erk1/2 and p38 MAPK signaling pathways. Methods: Human sperm was treated with the Hsp90-specific inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG) during capacitation. Computer-assisted sperm analyzer (CASA) was used to detect sperm motility and hyperactivation. The sperm acrosome reaction was analyzed by using fluorescein isothiocyanate-conjugated Pisum sativum agglutinin (PSA-FITC) staining. The interactions between Hsp90, Cdc37, Erk1/2 and p38 were assessed using co-immunoprecipitation (Co-IP) experiments. Western blotting analysis was used to evaluate the levels of protein expression and phosphorylation. Results: Human sperm hyperactivation and acrosome reaction were inhibited by 17-AAG, suggesting that Hsp90 is involved in human sperm capacitation. In addition, Co-IP experiments revealed that 17-AAG reduced the interaction between Hsp90 and Cdc37, leading to the dissociation of Erk1/2 from the Hsp90-Cdc37 protein complex. Western blotting analysis revealed that levels of Erk1/2 and its phosphorylated form were subsequently decreased. Decreasing of Hsp90-Cdc37 complex also affected the interaction between Hsp90 and p38. Nevertheless, p38 dissociated from the Hsp90 protein complex and was activated by autophosphorylation. Conclusions: Taken together, our findings indicate that Hsp90 is involved in human sperm hyperactivation and acrosome reaction. In particular, Hsp90 and its co-chaperone Cdc37 form a protein complex with Erk1/2 and p38 to regulate their kinase activity. These results suggest that Hsp90 regulates human sperm capacitation via the Erk1/2 and p38 MAPK signaling pathways.

scholarly journals Hsp90 modulates human sperm capacitation via the Erk1/2 and p38 MAPK signaling pathways

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Peibei Sun ◽  
Yayan Wang ◽  
Tian Gao ◽  
Kun Li ◽  
Dongwang Zheng ◽  
...  
Keyword(s):  
Signaling Pathways ◽  
P38 Mapk ◽  
Sperm Motility ◽  
Western Blotting ◽  
Protein Complex ◽  
Acrosome Reaction ◽  
Human Sperm ◽  
Mapk Signaling ◽  
Sperm Capacitation ◽  
Mapk Signaling Pathways

Abstract Background Heat shock protein 90 (Hsp90) is a highly abundant eukaryotic molecular chaperone that plays important roles in client protein maturation, protein folding and degradation, and signal transduction. Previously, we found that both Hsp90 and its co-chaperone cell division cycle protein 37 (Cdc37) were expressed in human sperm. Hsp90 is known to be involved in human sperm capacitation via unknown underlying mechanism(s). As Cdc37 was a kinase-specific co-chaperone of Hsp90, Hsp90 may regulate human sperm capacitation via other kinases. It has been reported that two major mitogen-activated protein kinases (MAPKs), extracellular signal-regulated kinase 1/2 (Erk1/2) and p38, are expressed in human sperm in the same locations as Hsp90 and Cdc37. Phosphorylated Erk1/2 has been shown to promote sperm hyperactivated motility and acrosome reaction, while phosphorylated p38 inhibits sperm motility. Therefore, in this study we explored whether Hsp90 modulates human sperm capacitation via the Erk1/2 and p38 MAPK signaling pathways. Methods Human sperm was treated with the Hsp90-specific inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG) during capacitation. Computer-assisted sperm analyzer (CASA) was used to detect sperm motility and hyperactivation. The sperm acrosome reaction was analyzed by using fluorescein isothiocyanate-conjugated Pisum sativum agglutinin (PSA-FITC) staining. The interactions between Hsp90, Cdc37, Erk1/2 and p38 were assessed using co-immunoprecipitation (Co-IP) experiments. Western blotting analysis was used to evaluate the levels of protein expression and phosphorylation. Results Human sperm hyperactivation and acrosome reaction were inhibited by 17-AAG, suggesting that Hsp90 is involved in human sperm capacitation. In addition, Co-IP experiments revealed that 17-AAG reduced the interaction between Hsp90 and Cdc37, leading to the dissociation of Erk1/2 from the Hsp90-Cdc37 protein complex. Western blotting analysis revealed that levels of Erk1/2 and its phosphorylated form were subsequently decreased. Decreasing of Hsp90-Cdc37 complex also affected the interaction between Hsp90 and p38. Nevertheless, p38 dissociated from the Hsp90 protein complex and was activated by autophosphorylation. Conclusions Taken together, our findings indicate that Hsp90 is involved in human sperm hyperactivation and acrosome reaction. In particular, Hsp90 and its co-chaperone Cdc37 form a protein complex with Erk1/2 and p38 to regulate their kinase activity. These results suggest that Hsp90 regulates human sperm capacitation via the Erk1/2 and p38 MAPK signaling pathways.

scholarly journals TRF-Val-CAC-016 Modulates the Transduction of CACNA1d-Mediated MAPK Signaling Pathways to Suppress the Proliferation of Gastric Carcinoma

2022 ◽  
Author(s):  
Weiguo Xu ◽  
Junyu Zheng ◽  
Xiao Wang ◽  
Bin Zhou ◽  
Huanqiu Chen ◽  
...  
Keyword(s):  
Gastric Carcinoma ◽  
Signaling Pathways ◽  
Signaling Pathway ◽  
Mapk Signaling ◽  
Mapk Signaling Pathway ◽  
Luciferase Reporter ◽  
Trna Derivatives ◽  
Mapk Signaling Pathways

Abstract Background: As a new kind of non-coding RNAs (ncRNAs), tRNA derivatives play an important role in gastric carcinoma (GC). Nevertheless, the underlying mechanism tRNA derivatives were involved in was rarely illustrated. Methods: We screened out the tRNA derivative, tRF-Val-CAC-016, based on the tsRNA sequencing and demonstrated the effect tRF-Val-CAC-016 exerted on GC proliferation in vitro and in vivo. We applied Dual-luciferase reporter assay, RIP assay, and bioinformatic analysis to discover the downstream target of tRF-Val-CAC-016. Then CACNA1d was selected, and the oncogenic characteristics were verified. Subsequently, we detected the possible regulation of the canonical MAPK signaling pathway to further explore the downstream mechanism of tRF-Val-CAC-016. Results: As a result, we found that tRF-Val-CAC-016 was low-expressed in GC, and upregulation of tRF-Val-CAC-016 could significantly suppress the proliferation of GC cell lines. Meanwhile, tRF-Val-CAC-016 regulated the canonical MAPK signaling pathway by targeting CACNA1d. Conclusions: tRF-Val-CAC-016 modulates the transduction of CACNA1d-mediated MAPK signaling pathways to suppress the proliferation of gastric carcinoma. This study discussed the function and mechanism of tRF-Val-CAC-016 in GC for the first time. The pioneering work has contributed to our present understanding of tRNA derivative, which might provide an alternative mean for the targeted therapy of GC.

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