Nicotine-upregulated miR-30a arrests cell cycle in G1 phase by directly targeting CCNE2 in human periodontal ligament cells

2020 ◽  
Vol 98 (3) ◽  
pp. 354-361
Author(s):  
Lizheng Wu ◽  
Kuan Yang ◽  
Yajie Gui ◽  
Xiaojing Wang
Keyword(s):  
Cell Cycle ◽  
Periodontal Ligament ◽  
Luciferase Reporter ◽  
Pcr Analysis ◽  
Periodontal Ligament Cells ◽  
Periodontal Tissues ◽  
Human Pdl Cells ◽  
Pdl Cells ◽  
Reporter Assays

The consumption of nicotine via smoking tobacco has been reported to stimulate the occurrence and progression of periodontitis. Many studies have demonstrated that nicotine prevents the regeneration of periodontal tissues primarily by inhibiting the proliferation of human periodontal ligament (PDL) cells. However, the mechanisms underlying this process are still unclear. Therefore, we investigated whether nicotine-upregulated miR-30a inhibited the proliferation of human PDL cells by downregulating cyclin E2 (CCNE2), in vitro. Quantitative real-time PCR analysis revealed that nicotine upregulated the expression of miR-30a in human PDL cells. In addition, nicotine inhibited the proliferation of human PDL cells by inducing cell cycle arrest. To support this hypothesis, we showed that nicotine downregulated the expression of CCNE2 in human PDL cells, whereas inhibition of miR-30a restored CCNE2 expression that had been downregulated by nicotine. Furthermore, using luciferase reporter assays, we found that miR-30a directly interacts with the CCNE2 3′UTR. In conclusion, these findings indicate that nicotine-upregulated miR-30a inhibits the proliferation of human PDL cells by downregulating the expression of CCNE2.

scholarly journals Cyclic compression emerged dual effects on the osteogenic and osteoclastic status of LPS-induced inflammatory human periodontal ligament cells according to loading force

2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Ru Jia ◽  
Yingjie Yi ◽  
Jie Liu ◽  
Dandan Pei ◽  
Bo Hu ◽  
...  
Keyword(s):  
Periodontal Ligament ◽  
Osteoclast Differentiation ◽  
Cyclic Stress ◽  
Periodontal Ligament Cells ◽  
Mrna Levels ◽  
Bone Homeostasis ◽  
Periodontal Tissues ◽  
Human Periodontal Ligament Cells ◽  
Tnf Α

Abstract Background Appropriate mechanical stimulation is essential for bone homeostasis in healthy periodontal tissues. While the osteogenesis and osteoclast differentiation of inflammatory periodontal ligament cells under different dynamic loading has not been yet clear. The aim of this study is to clarify the inflammatory, osteogenic and pro-osteoclastic effects of different cyclic stress loading on the inflammatory human periodontal ligament cells (hPDLCs). Methods hPDLCs were isolated from healthy premolars and cultured in alpha minimum Eagle’s medium (α-MEM). Lipopolysaccharides (LPS) were used to induce the inflammation state of hPDLCs in vitro. Determination of LPS concentration for the model of inflammatory periodontium was based on MTT and genes expression analysis. Then the cyclic stress of 0, 0–50, 0–90 and 0–150 kPa was applied to the inflammatory hPDLCs for 5 days respectively. mRNA and protein levels of osteogenic, osteoclastic and inflammation-related markers were examined after the treatment. Results MTT and RT-PCR results showed that 10 μg/ml LPS up-regulated TNF-α, IL-1β, IL-6, IL-8 and MCP-1 mRNA levels (P < 0.05) and did not affect the cell viability (P > 0.05). The excessive loading of stress (150 kPa) with or without LPS strongly increased the expression of inflammatory-related markers TNF-α, IL-1β, IL-6, IL-8, MCP-1 (P < 0.05) and osteoclastic markers RANKL, M-CSF, PTHLH and CTSK compared with other groups (P < 0.05), but had no significant effect on osteogenic genes. While 0–90 kPa cyclic pressure could up-regulate the expression of osteogenic genes ALP, COL-1, RUNX2, OCN, OPN and OSX in the healthy hPDLSCs. Conclusions Collectively, it could be concluded that 0–150 kPa was an excessive stress loading which accelerated both inflammatory and osteoclastic effects, while 0–90 kPa may be a positive factor for the osteogenic differentiation of hPDLCs in vitro.

scholarly journals Pluripotency of Dental Pulp Cells and Periodontal Ligament Cells Was Enhanced through Cell-Cell Communication via STAT3/Oct-4/Sox2 Signaling

2021 ◽  
Vol 2021 ◽  
pp. 1-18
Author(s):  
Zhengjun Peng ◽  
Lu Liu ◽  
Wenyu Zhang ◽  
Xi Wei
Keyword(s):  
Dental Pulp ◽  
Periodontal Ligament ◽  
Cell Communication ◽  
Luciferase Reporter ◽  
Periodontal Ligament Cells ◽  
Dental Pulp Cells ◽  
Dental Tissue ◽  
Pulp Cells ◽  
Cell Cell

Alternation in culture environment due to cell-cell communications can rejuvenate the biological activity of aged/differentiated cells and stimulate the expression of pluripotency markers. Dental pulp cells (DPCs) and periodontal ligament cells (PDLCs) are promising candidates in dental tissue regeneration. However, the molecular network that underlies cell-cell communications between dental-derived cells and the microenvironment remains to be identified. To elucidate the signaling network that regulates the pluripotency of DPCs and PDLCs, proliferation, apoptosis, cell cycle, and the expression of Oct-4/Sox2/c-Myc in DPCs and PDLCs with indirect/direct coculture were examined. PCR arrays were constructed to identify genes that were differentially expressed, and the results were confirmed by a rat model with injury. Further research on the mechanism of the related signaling pathways was investigated by overexpression/silence of STAT3, ChIP, the dual-luciferase reporter assay, and EMSA. We found that the proliferation and apoptosis of DPCs and PDLCs were inhibited, and their cell cycles were arrested at the G0/G1 phase after coculture. Oct-4, Sox2, and STAT3 expression significantly increased and PAX5 expression decreased in the coculture systems. Oct-4/Sox2/STAT3/PAX5 was actively expressed in the rat defect model. Moreover, STAT3 was directly bound to the Oct-4 and Sox2 gene promoter regions and activated the expression of those genes. Our data showed that the pluripotency of DPCs and PDLCs was enhanced through cell-cell communication. STAT3 plays essential roles in regulating the pluripotency of DPCs and PDLCs by targeting Oct-4 and Sox2 both in vitro and in vivo.

scholarly journals LncRNA LINC00337 Sponges miR-1285-3p to Promote Proliferation and Metastasis of Lung Adenocarcinoma Cells Through Up-Regulating YTHDF1

2020 ◽  
Author(s):  
Ru-nan Zhang ◽  
Dong-mei Wu ◽  
Li-ping Wu ◽  
Guo-wei Gao
Keyword(s):  
Cell Cycle ◽  
Cell Proliferation ◽  
Lung Adenocarcinoma ◽  
Molecular Mechanisms ◽  
Luciferase Reporter ◽  
Gear Up ◽  
Knock Down ◽  
Reporter Assays

Abstract Background: Emerging studies have attested that long noncoding RNAs (lncRNAs) predominantly functioned in carcinogenesis of multiple developing human tumors. The current research aimed at probing the underlying participation and mechanisms of LINC00337 in lung adenocarcinoma.Methods: Here we analyzed TCGA and GTEx datasets and chose LINC00337 as research object. Cell proliferation, cell apoptosis, cell cycle, and invasion were detected in gain and loss experiment of LINC00337 both in vitro and in vivo. Moreover, RNA pull-down, luciferase reporter assays, western blotting analysis, rescue experiment were performed to investigate underlying molecular mechanisms of LINC00337 function. Results: LINC00337 was remarkably increased in lung adenocarcinoma. Also, LINC00337 knock-down was unraveled to repress cell invasion and proliferation as well as cell cycle, and gear up apoptosis in lung adenocarcinoma in vitro and in vivo. With respect to mechanism, LINC00337 knock-down boosted miR-1285-3p to be expressed and then restrained YTHDF1 to be expressed post-transcriptionally. Crucially, both miR-1285-3p decrement and YTHDF1 overexpression successfully countered the influence on cell proliferation, invasion and apoptosis caused by LINC00337 shRNA.Conclusions: These results suggest that LINC00337 acted as an oncogenic lncRNA, targeting miR-1285-3p and regulating YTHDF1 expression, to promote the progression of lung adenocarcinoma.

scholarly journals Cyclic compression emerged dual effects on the bone homeostasis of LPS-induced inflammatory human periodontal ligament cells according to loading force

2019 ◽  
Author(s):  
Ru Jia ◽  
Yingjie Yi ◽  
Jie Liu ◽  
Dandan Pei ◽  
Bo Hu ◽  
...  
Keyword(s):  
Periodontal Ligament ◽  
Cyclic Stress ◽  
Periodontal Ligament Cells ◽  
Mrna Levels ◽  
Bone Homeostasis ◽  
Protein Levels ◽  
Periodontal Tissues ◽  
Human Periodontal Ligament Cells ◽  
Tnf Α

Abstract Background: Appropriate mechanical stimulation is essential for bone homeostasis in healthy periodontal tissues. While the bone homeostasis of inflammatory periodontal tissues under different dynamic loading has not been yet clear. The aim of this study is to clarify the inflammatory, osteogenic and pro-osteoclastic effects of different cyclic stress loading on the inflammatory human periodontal ligament cells (hPDLCs). Methods: hPDLCs were isolated from healthy premolars and cultured in alpha minimum Eagle’s medium (α-MEM). Lipopolysaccharides (LPS) were used to induce the inflammation state of hPDLCs in vitro. Determination of LPS concentration for the model of inflammatory periodontium was based on MTT and genes expression analysis. Then the cyclic stress of 0, 0-50, 0-90 and 0-150 kPa was applied to the inflammatory hPDLCs for 5 days respectively. mRNA and protein levels of osteogenic, osteoclastic and inflammation-related markers were examined after the treatment. Results: MTT and RT-PCR results showed that 10 μg/ml LPS up-regulated TNF-α, IL-1β, IL-6, IL-8 and MCP-1 mRNA levels (P<0.05) and did not affect the cell viability (P>0.05). The excessive loading of stress (150 kPa) with or without LPS strongly increased the expression of inflammatory-related markers TNF-α, IL-1β, IL-6, IL-8, MCP-1 (P<0.05) and osteoclastic markers RANKL, PTHLH and CTSK compared with other groups (P<0.05), but had no significant effect on osteogenic genes. While 0-90 kPa cyclic pressure could up-regulate the expression of osteogenic genes ALP, COL-1 and RUNX2 in the healthy hPDLSCs. Conclusions: Collectively, it could be concluded that 0-150 kPa was an excessive stress loading which accelerated both inflammatory and osteoclastic effects, while 0-90 kPa may be a positive factor for the bone homeostasis of hPDLCs in vitro

scholarly journals Cyclic compression emerged dual effects on the osteogenic and osteoclastic status of LPS-induced inflammatory human periodontal ligament cells according to loading force

2019 ◽  
Author(s):  
Ru Jia ◽  
Yingjie Yi ◽  
Jie Liu ◽  
Dandan Pei ◽  
Bo Hu ◽  
...  
Keyword(s):  
Periodontal Ligament ◽  
Osteoclast Differentiation ◽  
Cyclic Stress ◽  
Periodontal Ligament Cells ◽  
Mrna Levels ◽  
Bone Homeostasis ◽  
Periodontal Tissues ◽  
Human Periodontal Ligament Cells ◽  
Tnf Α

Abstract Background: Appropriate mechanical stimulation is essential for bone homeostasis in healthy periodontal tissues. While the osteogenesis and osteoclast differentiation of inflammatory periodontal ligament cells under different dynamic loading has not been yet clear. The aim of this study is to clarify the inflammatory, osteogenic and pro-osteoclastic effects of different cyclic stress loading on the inflammatory human periodontal ligament cells (hPDLCs) . Methods: hPDLCs were isolated from healthy premolars and cultured in alpha minimum Eagle’s medium (α-MEM). Lipopolysaccharides (LPS) were used to induce the inflammation state of hPDLCs in vitro . Determination of LPS concentration for the model of inflammatory periodontium was based on MTT and genes expression analysis. Then the cyclic stress of 0, 0-50, 0-90 and 0-150 kPa was applied to the inflammatory hPDLCs for 5 days respectively. mRNA and protein levels of osteogenic, osteoclastic and inflammation-related markers were examined after the treatment. Results: MTT and RT-PCR results showed that 10 μg/ml LPS up-regulated TNF-α, IL-1β, IL-6, IL-8 and MCP-1 mRNA levels ( P <0.05) and did not affect the cell viability ( P >0.05). The excessive loading of stress (150 kPa) with or without LPS strongly increased the expression of inflammatory-related markers TNF-α , IL-1β , IL-6 , IL-8 , MCP-1 ( P <0.05) and osteoclastic markers RANKL , M-CSF , PTHLH and CTSK compared with other groups ( P <0.05), but had no significant effect on osteogenic genes. While 0-90 kPa cyclic pressure could up-regulate the expression of osteogenic genes ALP, COL-1 , RUNX2, OCN, OPN and OSX in the healthy hPDLSCs. Conclusions: Collectively, it could be concluded that 0-150 kPa was an excessive stress loading which accelerated both inflammatory and osteoclastic effects, while 0-90 kPa may be a positive factor for the osteogenic differentiation of hPDLCs in vitro.

Evaluation of Aloevera Gel as a Storage Medium in Maintaining the Viability of Periodontal Ligament Cells - An in Vitro Study

2016 ◽  
Vol 40 (1) ◽  
pp. 49-52 ◽  
Author(s):  
Punit Fulzele ◽  
Sudhindra Baliga ◽  
Nilima Thosar ◽  
Debaprya Pradhan
Keyword(s):  
Drinking Water ◽  
Periodontal Ligament ◽  
In Vitro Study ◽  
Periodontal Ligament Cells ◽  
Blue Dye ◽  
Storage Medium ◽  
Viable Cells ◽  
Pdl Cells ◽  
Significant Difference

Objective: To investigate the effectiveness of aloevera gel as a new storage medium in maintaining the viability of periodontal ligament cells. Study design: Premolars extracted for orthodontic reason were obtained. Confluent monolayers of fibroblasts were grown by cell culture method from the PDL cells isolated from the extracted teeth. One ml of this cell suspension was transferred to wells of culture plates, incubated for 24 hrs, followed by exposure to the three experimental media, Hank's balanced salt solution (HBSS), aloevera gel, and packaged drinking water. These plates were then assessed for viable cells using trypan blue dye exclusion test with haemocytometer after 15, 30, 60, 90 and 120 mins. The results obtained were statistically analysed using one-way analysis of variance (ANOVA). Results: At 15 min, HBSS presented maximum mean percentage of viable PDL cells (89%), followed by aloevera at 81% and packaged drinking water at 10%. Aloevera demonstrated 71%, 59%, 57% viable cells at 30, 60, 90 mins respectively. At 120 min, HBSS presented 57% viable cells followed by aloevera gel (45%) and packaged drinking water (3%). No statistical significant difference was observed between HBSS and aloevera gel. Conclusions: Within the parameters of this study, both aloevera gel and HBSS were effective in maintaining the viability of PDL cells. Hence, aloevera gel could be used as a storage media for avulsed tooth in situations where availability of HBSS is in question.

PTHrP Induces Notch Signaling in Periodontal Ligament Cells

2009 ◽  
Vol 88 (6) ◽  
pp. 551-556 ◽  
Author(s):  
A. Nakao ◽  
H. Kajiya ◽  
H. Fukushima ◽  
A. Fukushima ◽  
H. Anan ◽  
...  
Keyword(s):  
Bone Resorption ◽  
Notch Signaling ◽  
Periodontal Ligament ◽  
Alveolar Bone ◽  
Root Resorption ◽  
Tooth Eruption ◽  
Periodontal Ligament Cells ◽  
Dependent Manner ◽  
Human Pdl Cells ◽  
Pdl Cells

Periodontal ligament (PDL) cells are known to play important roles in tooth eruption and alveolar bone metabolism. We previously reported that PTHrP increases RANKL expression in human PDL cells, suggesting that it promotes odontoclastic root resorption during tooth eruption. While it is known that Notch-related genes play a key role during bone development, the role of the Notch signaling pathway in PDL cells during tooth and bone resorption is less clear. We hypothesized that PTHrP induces a Notch ligand in PDL cells and thereby regulates osteo- and odontoclastogenesis. We found that PTHrP increased Notch1 ligand Jagged1 expression in human PDL cells in a dose- and time-dependent manner. PTHrP-induced Jagged1 up-regulation was mediated by PKA activation, but not by PKC. Jagged1 also promoted RANKL-induced osteoclastogenesis. These results demonstrate that PTHrP induces Jagged1 expression in PDL cells, leading to osteo- and odontoclastogenesis, and thus likely promoting tooth and alveolar bone resorption.

scholarly journals CircMAP3K11 Contributes to Proliferation, Apoptosis and Migration of Human Periodontal Ligament Stem Cells in Inflammatory Microenvironment by Regulating TLR4 via miR-511 Sponging

2021 ◽  
Vol 12 ◽  
Author(s):  
Bohan Yu ◽  
Jiahui Hu ◽  
Qin Li ◽  
Fang Wang
Keyword(s):  
Stem Cells ◽  
Periodontal Ligament ◽  
Luciferase Reporter ◽  
Regenerative Capacity ◽  
Periodontal Ligament Stem Cells ◽  
Inflammatory Microenvironment ◽  
Periodontal Tissues ◽  
And Migration

Growing number of studies regarding the role of circRNAs in the development of various diseases have emerged in recent years, but the role of circRNAs in periodontitis pathogenesis remains obscure. Human periodontal ligament stem cells (hPDLSCs) play a critical role in periodontal remodeling, regeneration and repair processes, and their regenerative capacity could be prohibited in local periodontal inflammatory microenvironment. Herein, we sought to uncover the molecular mechanisms of periodontitis pathogenesis by investigating the role of circMAP3K11 (hsa_circ_002284) for regenerative capacity of hPDLSCs under an inflammatory condition. The hPDLSCs isolated from periodontitis patients were used as a cell model of inflammatory microenvironment to study the effect of the circMAP3K11/miR-511-3p/TLR4 axis on the proliferation, apoptosis and migration of hPDLSCs under inflammatory conditions. Compared to the periodontal tissues from normal subjects, those from periodontitis patients exhibited higher expression levels of circMAP3K11 and TLR4, and lower expression level of miR-511-3p. Both the expressions of circMAP3K11 and TLR4 were negatively correlated with the expressions of miR-511-3p in periodontitis. In vitro studies demonstrated that circMAP3K11 is capable of enhancing hPDLSCs proliferation and migration, and reducing the apoptosis of hPDLSCs. We also found that circMAP3K11 could up-regulate the expression of transcription factors that are closely related to periodontal regeneration (Runx2, OSX, ATF4, and BSP). RT-PCR and western blot showed that the inhibitory role of miR-511-3p on TLR4 expression could be reversed by circMAP3K11, which was in line with the results of bioinformatics tools and luciferase reporter assay. Meanwhile, both in vitro and in vivo studies indicated that circMAP3K11 could reverse the effects of miR-511-3p in periodontitis, which further confirmed that circMAP3K11 functioned as a ‘sponge’ of miR-511-3p to positively regulate the expression of TLR4. Taken together, our study preliminarily uncovered a circMAP3K11/miR-511-3p/TLR4 axis that regulates the function of hPDLSCs in periodontitis, providing novel insight and scientific base in the treatment of periodontal tissue regeneration based on stem cells.

PTH(1-34) Affects Osteoprotegerin Production in Human PDL Cells in vitro

2005 ◽  
Vol 84 (7) ◽  
pp. 634-638 ◽  
Author(s):  
S. Lossdörfer ◽  
W. Götz ◽  
A. Jäger
Keyword(s):  
Periodontal Ligament ◽  
Cell Number ◽  
Rankl Expression ◽  
Hormonal Stimulation ◽  
Human Pdl Cells ◽  
Pdl Cells ◽  
Maturation State ◽  
Osteocalcin Production ◽  
Intermittent Pth

Since periodontal ligament (PDL) cells exhibit several osteoblastic traits, we hypothesized that human PDL cells will respond to hormonal stimulation in an osteoblast-like manner. Confluent and pre-confluent PDL cells from six patients were challenged with PTH(1-34). Cell number, ALP, osteocalcin, osteoprotegerin, and RANKL expression were determined. Intermittent PTH(1-34) treatment of confluent PDL cells caused a significant increase in proliferation, whereas differentiation and osteoprotegerin production decreased significantly. In pre-confluent PDL cells, this treatment regimen induced a biphasic decrease in proliferation, but a biphasic increase in differentiation and osteoprotegerin production. Continuous PTH(1-34) exposure enhanced proliferation but inhibited osteocalcin production in confluent cells and stimulated osteoprotegerin production in pre-confluent PDL cells. RANKL was hardly detectable and unaffected by PTH(1-34) treatment. These results indicate that human PDL cells respond to PTH(1-34) in an osteoblast-like manner, and that the PTH(1-34) effect depends on the maturation state of the cells and on the mode of administration.

scholarly journals gga-miR-148a-5p-Targeting PDPK1 Inhibits Proliferation and Cell Cycle Progression of Avain Leukosis Virus Subgroup J (ALV-J)-Infected Cells

2020 ◽  
Vol 8 ◽  
Author(s):  
Heling Yu ◽  
Hengyong Xu ◽  
Chaoyang Yan ◽  
Shiliang Zhu ◽  
Xi Lan ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Progression ◽  
Luciferase Reporter ◽  
Sequencing Data ◽  
Functional Roles ◽  
Direct Target Gene ◽  
Reporter Assays ◽  
Infected Cells ◽  
Subgroup J

Avian leukosis virus subgroup J disease (ALV-J) is a contagious and immunosuppressive avian disease caused by ALV-J virus. Although miRNA participate in various biological processes of tumors, little is known about the potential role of miRNA in ALV-J. Our previous miRNA and RNA sequencing data showed that the expression of gga-miR-148a-5p was significantly different in ALV-J-infected chicken spleens compared with non-infected chickens. The aim of this study was to investigate the functional roles of gga-miR-148a-5p and identify downstream targets regulated by gga-miR-148a-5p in ALV-J-infected chickens. We found that the expression of gga-miR-148a-5p was significantly downregulated during ALV-J infection of chicken embryo fibroblasts (CEF). Dual luciferase reporter assays demonstrated that PDPK1 is a direct target gene of gga-miR-148a-5p. In vitro, overexpression of gga-miR-148a-5p significantly promoted ALV-J-infected CEF cell proliferation, included cell cycle, whereas inhibition of gga-miR-148a-5p had an opposite effect. Inhibition of PDPK1 promoted the proliferation of ALV-J-infected cells but had no effect on the activity of NF-κB. Together, these results suggested that gga-miR-148a-5p targets PDPK1 to inhibit the proliferation and cell cycle of ALV-J-infected CEF cells. Our study provides a new understanding for the tumor mechanism of ALV-J infection.

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