Background:
Regulation of mechanical stress plays a key role in maintainability of bone volume in dental prosthetics treatment, especially in dental implants treatment. In bone remodeling, consisting of osteoclast breaking down bone matrix and the osteoblast forming new bone, osteoblast differentiation is determined by a complex array of growth factors and signaling pathways. Mechanical stress is transformed into biological signals, which ultimately direct bone formation and/or bone resorption. However, the intimate mechanism of bone formation and resorption by mechanical stress has not been fully elucidated. To date, accumulated evidence has indicated that P2X receptors, as the family of trimeric ion channels, enhance osteoblast differentiation and bone formation. ERK1/2 is well known signaling molecule that is indispensable for osteoblast differentiation. Nevertheless, the relationship between mechanical stimulation and ERK pathway remains unclear.
Aim/Hypothesis:
Our purpose of this study is to uncover the mechanosensor that mediates osteogenic ERK signal in bone mechanobiology and to discover the molecular link between mechanical stress and anabolic responses of osteoblasts, which have clinical relevance in bone regenerative therapy.
Material and Methods:
(1)To study the effect of mechanical strain on differentiation of osteoblast, MC3T3-E1 mouse clonal osteogenic cells were subjected to 0%, 2%, 6% and 12% mechanical stretch, elongation for 60min, at a frequency of 10 cycles per minute, by using ST-140 cell cyclic stretcher system (Strex). Immunoblot analysis was used to study the effects of mechanical strain on ERK activity; Reverse transcription polymerase chain reaction (RT-PCR) was used to examine mRNA level of collagen Ia (ColIa) and osteopontin (OPN). (2)To illuminate the mechanisms of the upstream receptors of ERK signaling in mechanotransdution activated by low-magnitude mechanical stress, MC3T3-E1 osteoblastic cells were subjected to 2% mechanical stretch elongation for 0min, 15min, 30min and 60min, at a frequency of 10 cycles per minute, by using ST-140 cell cyclic stretcher system. RNA interference (RNAi) was used to analyses the role of P2X family receptors. MC3T3-E1 cells were transfected with the small interfering RNAs (siRNAs) for P2X1~P2X7 using Lipofectamine RNAiMAX. To evaluate the specificity of the siRNA, non-operative siRNA was used in parallel. Mechanical stretch was applied 72 hours after transfection.
Results:
The study showed that (1) Upon different magnitude of mechanical stretch loading, ERK pathway was strongly activated by low-magnitude of mechanical stretch (2%). RT-PCR analysis referred that expression of ColIa and OPN was consistently induced by 2% mechanical stretch while ColIa and OPN expression was not enhanced upon 12% mechanical stretch. (2)After screening the role of P2X family factors by using RNA interference, we found that knockdown of P2X7 receptor inhibited the ERK pathway activity, suggesting that P2X7 receptor is a highly sensitive P2X receptor which can mediate osteogenic ERK signal under low-magnitude of mechanical stress in osteoblasts.
Conclusions and clinical implications:
From this study, we concluded that low-magnitude of mechanical stretch preferentially activated ERK pathway which induced the expression of two osteogenic genes, ColIa and OPN. P2X7 is a highly sensitive P2X family receptor, which mediates low-magnitude of mechanical stress-induced ERK activation in osteoblasts.
Background:
Regulation of mechanical stress plays a key role in maintainability of bone volume in dental prosthetics treatment, especially in dental implants treatment. In bone remodeling, consisting of osteoclast breaking down bone matrix and the osteoblast forming new bone, osteoblast differentiation is determined by a complex array of growth factors and signaling pathways. Mechanical stress is transformed into biological signals, which ultimately direct bone formation and/or bone resorption. However, the intimate mechanism of bone formation and resorption by mechanical stress has not been fully elucidated. To date, accumulated evidence has indicated that P2X receptors, as the family of trimeric ion channels, enhance osteoblast differentiation and bone formation. ERK1/2 is well known signaling molecule that is indispensable for osteoblast differentiation. Nevertheless, the relationship between mechanical stimulation and ERK pathway remains unclear.
Aim/Hypothesis:
Our purpose of this study is to uncover the mechanosensor that mediates osteogenic ERK signal in bone mechanobiology and to discover the molecular link between mechanical stress and anabolic responses of osteoblasts, which have clinical relevance in bone regenerative therapy.
Material and Methods:
(1)To study the effect of mechanical strain on differentiation of osteoblast, MC3T3-E1 mouse clonal osteogenic cells were subjected to 0%, 2%, 6% and 12% mechanical stretch, elongation for 60min, at a frequency of 10 cycles per minute, by using ST-140 cell cyclic stretcher system (Strex). Immunoblot analysis was used to study the effects of mechanical strain on ERK activity; Reverse transcription polymerase chain reaction (RT-PCR) was used to examine mRNA level of collagen Ia (ColIa) and osteopontin (OPN). (2)To illuminate the mechanisms of the upstream receptors of ERK signaling in mechanotransdution activated by low-magnitude mechanical stress, MC3T3-E1 osteoblastic cells were subjected to 2% mechanical stretch elongation for 0min, 15min, 30min and 60min, at a frequency of 10 cycles per minute, by using ST-140 cell cyclic stretcher system. RNA interference (RNAi) was used to analyses the role of P2X family receptors. MC3T3-E1 cells were transfected with the small interfering RNAs (siRNAs) for P2X1~P2X7 using Lipofectamine RNAiMAX. To evaluate the specificity of the siRNA, non-operative siRNA was used in parallel. Mechanical stretch was applied 72 hours after transfection.
Results:
The study showed that (1) Upon different magnitude of mechanical stretch loading, ERK pathway was strongly activated by low-magnitude of mechanical stretch (2%). RT-PCR analysis referred that expression of ColIa and OPN was consistently induced by 2% mechanical stretch while ColIa and OPN expression was not enhanced upon 12% mechanical stretch. (2)After screening the role of P2X family factors by using RNA interference, we found that knockdown of P2X7 receptor inhibited the ERK pathway activity, suggesting that P2X7 receptor is a highly sensitive P2X receptor which can mediate osteogenic ERK signal under low-magnitude of mechanical stress in osteoblasts.
Conclusions and clinical implications:
From this study, we concluded that low-magnitude of mechanical stretch preferentially activated ERK pathway which induced the expression of two osteogenic genes, ColIa and OPN. P2X7 is a highly sensitive P2X family receptor, which mediates low-magnitude of mechanical stress-induced ERK activation in osteoblasts.
Background:
Regulation of mechanical stress plays a key role in maintainability of bone volume in dental prosthetics treatment, especially in dental implants treatment. In bone remodeling, consisting of osteoclast breaking down bone matrix and the osteoblast forming new bone, osteoblast differentiation is determined by a complex array of growth factors and signaling pathways. Mechanical stress is transformed into biological signals, which ultimately direct bone formation and/or bone resorption. However, the intimate mechanism of bone formation and resorption by mechanical stress has not been fully elucidated. To date, accumulated evidence has indicated that P2X receptors, as the family of trimeric ion channels, enhance osteoblast differentiation and bone formation. ERK1/2 is well known signaling molecule that is indispensable for osteoblast differentiation. Nevertheless, the relationship between mechanical stimulation and ERK pathway remains unclear.
Aim/Hypothesis:
Our purpose of this study is to uncover the mechanosensor that mediates osteogenic ERK signal in bone mechanobiology and to discover the molecular link between mechanical stress and anabolic responses of osteoblasts, which have clinical relevance in bone regenerative therapy.
Material and Methods:
(1)To study the effect of mechanical strain on differentiation of osteoblast, MC3T3-E1 mouse clonal osteogenic cells were subjected to 0%, 2%, 6% and 12% mechanical stretch, elongation for 60min, at a frequency of 10 cycles per minute, by using ST-140 cell cyclic stretcher system (Strex). Immunoblot analysis was used to study the effects of mechanical strain on ERK activity; Reverse transcription polymerase chain reaction (RT-PCR) was used to examine mRNA level of collagen Ia (ColIa) and osteopontin (OPN). (2)To illuminate the mechanisms of the upstream receptors of ERK signaling in mechanotransdution activated by low-magnitude mechanical stress, MC3T3-E1 osteoblastic cells were subjected to 2% mechanical stretch elongation for 0min, 15min, 30min and 60min, at a frequency of 10 cycles per minute, by using ST-140 cell cyclic stretcher system. RNA interference (RNAi) was used to analyses the role of P2X family receptors. MC3T3-E1 cells were transfected with the small interfering RNAs (siRNAs) for P2X1~P2X7 using Lipofectamine RNAiMAX. To evaluate the specificity of the siRNA, non-operative siRNA was used in parallel. Mechanical stretch was applied 72 hours after transfection.
Results:
The study showed that (1) Upon different magnitude of mechanical stretch loading, ERK pathway was strongly activated by low-magnitude of mechanical stretch (2%). RT-PCR analysis referred that expression of ColIa and OPN was consistently induced by 2% mechanical stretch while ColIa and OPN expression was not enhanced upon 12% mechanical stretch. (2)After screening the role of P2X family factors by using RNA interference, we found that knockdown of P2X7 receptor inhibited the ERK pathway activity, suggesting that P2X7 receptor is a highly sensitive P2X receptor which can mediate osteogenic ERK signal under low-magnitude of mechanical stress in osteoblasts.
Conclusions and clinical implications:
From this study, we concluded that low-magnitude of mechanical stretch preferentially activated ERK pathway which induced the expression of two osteogenic genes, ColIa and OPN. P2X7 is a highly sensitive P2X family receptor, which mediates low-magnitude of mechanical stress-induced ERK activation in osteoblasts.
Background:
Regulation of mechanical stress plays a key role in maintainability of bone volume in dental prosthetics treatment, especially in dental implants treatment. In bone remodeling, consisting of osteoclast breaking down bone matrix and the osteoblast forming new bone, osteoblast differentiation is determined by a complex array of growth factors and signaling pathways. Mechanical stress is transformed into biological signals, which ultimately direct bone formation and/or bone resorption. However, the intimate mechanism of bone formation and resorption by mechanical stress has not been fully elucidated. To date, accumulated evidence has indicated that P2X receptors, as the family of trimeric ion channels, enhance osteoblast differentiation and bone formation. ERK1/2 is well known signaling molecule that is indispensable for osteoblast differentiation. Nevertheless, the relationship between mechanical stimulation and ERK pathway remains unclear.
Aim/Hypothesis:
Our purpose of this study is to uncover the mechanosensor that mediates osteogenic ERK signal in bone mechanobiology and to discover the molecular link between mechanical stress and anabolic responses of osteoblasts, which have clinical relevance in bone regenerative therapy.
Material and Methods:
(1)To study the effect of mechanical strain on differentiation of osteoblast, MC3T3-E1 mouse clonal osteogenic cells were subjected to 0%, 2%, 6% and 12% mechanical stretch, elongation for 60min, at a frequency of 10 cycles per minute, by using ST-140 cell cyclic stretcher system (Strex). Immunoblot analysis was used to study the effects of mechanical strain on ERK activity; Reverse transcription polymerase chain reaction (RT-PCR) was used to examine mRNA level of collagen Ia (ColIa) and osteopontin (OPN). (2)To illuminate the mechanisms of the upstream receptors of ERK signaling in mechanotransdution activated by low-magnitude mechanical stress, MC3T3-E1 osteoblastic cells were subjected to 2% mechanical stretch elongation for 0min, 15min, 30min and 60min, at a frequency of 10 cycles per minute, by using ST-140 cell cyclic stretcher system. RNA interference (RNAi) was used to analyses the role of P2X family receptors. MC3T3-E1 cells were transfected with the small interfering RNAs (siRNAs) for P2X1~P2X7 using Lipofectamine RNAiMAX. To evaluate the specificity of the siRNA, non-operative siRNA was used in parallel. Mechanical stretch was applied 72 hours after transfection.
Results:
The study showed that (1) Upon different magnitude of mechanical stretch loading, ERK pathway was strongly activated by low-magnitude of mechanical stretch (2%). RT-PCR analysis referred that expression of ColIa and OPN was consistently induced by 2% mechanical stretch while ColIa and OPN expression was not enhanced upon 12% mechanical stretch. (2)After screening the role of P2X family factors by using RNA interference, we found that knockdown of P2X7 receptor inhibited the ERK pathway activity, suggesting that P2X7 receptor is a highly sensitive P2X receptor which can mediate osteogenic ERK signal under low-magnitude of mechanical stress in osteoblasts.
Conclusions and clinical implications:
From this study, we concluded that low-magnitude of mechanical stretch preferentially activated ERK pathway which induced the expression of two osteogenic genes, ColIa and OPN. P2X7 is a highly sensitive P2X family receptor, which mediates low-magnitude of mechanical stress-induced ERK activation in osteoblasts.