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Acidic pH environment induces autophagy in osteoblasts

View Article: PubMed Central - PubMed

ABSTRACT

Osteoblasts (OBs) play an important role in bone fracture healing, yet the extreme adverse microenvironment in fracture sites has a negative impact on the survival of OBs. Therefore, it is important to study how OBs behave in the complex fracture microenvironment. Studies have shown that autophagy plays a pivotal role in maintaining cellular homeostasis and defending the cell against adverse microenvironments. In this study we found the induction of autophagy in OBs at femoral bone fracture sites, which may be a result of ischemia, oxidative stress and hypoxia within the local area. At fracture sites a low pH environment also developed. Until now it has been unclear whether the induction of autophagy in osteoblasts is triggered by the acidic pH environment. Therefore, we cultured OBs in vitro in media of different pH values, and found both autophagy and apoptosis increased in OBs in acidic conditions. However, when autophagy inhibitor chloroquine (CQ) was used, apoptosis increased significantly compared with that without CQ. Thus indicating that inhibition of autophagy may promote apoptosis in OBs in an acidic environment, which may provide a new therapeutic strategy to decrease cell apoptosis in OBs through the use of drugs that modulate the autophagic state.

No MeSH data available.


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Autophagy is induced in OBs near fracture sites in the group 36 h after fracture.Longitudinal sections of rat fractured bones were detected. (A) Immunohistological staining showed the expression of LC3 and P62 in OBs at fractured sites (arrows). Size bar = 100 μm. As showed in figures e and f were control groups, a and b were experiment groups. a and c were from the same section with different magnification which showed the expression of LC3. b and d were from the same section with different magnification which showed the expression of P62. (B,D) Immunofluorescence analysis noted the distinct punctate pattern of LC3 and P62 expression (arrows) in OBs near fracture sites after 36 h. Size bar = 100 μm. (C,E) Quantified analysis of LC3 and P62 expression in osteoblasts in bones from the control and fracture group. *P < 0.05 vs.control. Values are the mean ± sd, *P < 0.05 in comparison to the control by one-way ANOVA.
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f1: Autophagy is induced in OBs near fracture sites in the group 36 h after fracture.Longitudinal sections of rat fractured bones were detected. (A) Immunohistological staining showed the expression of LC3 and P62 in OBs at fractured sites (arrows). Size bar = 100 μm. As showed in figures e and f were control groups, a and b were experiment groups. a and c were from the same section with different magnification which showed the expression of LC3. b and d were from the same section with different magnification which showed the expression of P62. (B,D) Immunofluorescence analysis noted the distinct punctate pattern of LC3 and P62 expression (arrows) in OBs near fracture sites after 36 h. Size bar = 100 μm. (C,E) Quantified analysis of LC3 and P62 expression in osteoblasts in bones from the control and fracture group. *P < 0.05 vs.control. Values are the mean ± sd, *P < 0.05 in comparison to the control by one-way ANOVA.

Mentions: Positive immunohistological or immunofluorescent staining was not found in group A(6 h) and group B(24 h). Immunohistological examination of femora 36 hours post fracture in group C, showed that LC3 immunoreactivity could be detected in a punctate pattern in OBs at fracture sites, which was not observed in un-fractured controls (Fig. 1A). Similarly, in group C immunofluorescent staining of LC3 also showed high expression in OBs around the bone marrow cavity in the fractured areas compared with the control femora, suggesting that autophagy was induced in OBs at fracture sites (Fig. 1B).


Acidic pH environment induces autophagy in osteoblasts
Autophagy is induced in OBs near fracture sites in the group 36 h after fracture.Longitudinal sections of rat fractured bones were detected. (A) Immunohistological staining showed the expression of LC3 and P62 in OBs at fractured sites (arrows). Size bar = 100 μm. As showed in figures e and f were control groups, a and b were experiment groups. a and c were from the same section with different magnification which showed the expression of LC3. b and d were from the same section with different magnification which showed the expression of P62. (B,D) Immunofluorescence analysis noted the distinct punctate pattern of LC3 and P62 expression (arrows) in OBs near fracture sites after 36 h. Size bar = 100 μm. (C,E) Quantified analysis of LC3 and P62 expression in osteoblasts in bones from the control and fracture group. *P < 0.05 vs.control. Values are the mean ± sd, *P < 0.05 in comparison to the control by one-way ANOVA.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5382697&req=5

f1: Autophagy is induced in OBs near fracture sites in the group 36 h after fracture.Longitudinal sections of rat fractured bones were detected. (A) Immunohistological staining showed the expression of LC3 and P62 in OBs at fractured sites (arrows). Size bar = 100 μm. As showed in figures e and f were control groups, a and b were experiment groups. a and c were from the same section with different magnification which showed the expression of LC3. b and d were from the same section with different magnification which showed the expression of P62. (B,D) Immunofluorescence analysis noted the distinct punctate pattern of LC3 and P62 expression (arrows) in OBs near fracture sites after 36 h. Size bar = 100 μm. (C,E) Quantified analysis of LC3 and P62 expression in osteoblasts in bones from the control and fracture group. *P < 0.05 vs.control. Values are the mean ± sd, *P < 0.05 in comparison to the control by one-way ANOVA.
Mentions: Positive immunohistological or immunofluorescent staining was not found in group A(6 h) and group B(24 h). Immunohistological examination of femora 36 hours post fracture in group C, showed that LC3 immunoreactivity could be detected in a punctate pattern in OBs at fracture sites, which was not observed in un-fractured controls (Fig. 1A). Similarly, in group C immunofluorescent staining of LC3 also showed high expression in OBs around the bone marrow cavity in the fractured areas compared with the control femora, suggesting that autophagy was induced in OBs at fracture sites (Fig. 1B).

View Article: PubMed Central - PubMed

ABSTRACT

Osteoblasts (OBs) play an important role in bone fracture healing, yet the extreme adverse microenvironment in fracture sites has a negative impact on the survival of OBs. Therefore, it is important to study how OBs behave in the complex fracture microenvironment. Studies have shown that autophagy plays a pivotal role in maintaining cellular homeostasis and defending the cell against adverse microenvironments. In this study we found the induction of autophagy in OBs at femoral bone fracture sites, which may be a result of ischemia, oxidative stress and hypoxia within the local area. At fracture sites a low pH environment also developed. Until now it has been unclear whether the induction of autophagy in osteoblasts is triggered by the acidic pH environment. Therefore, we cultured OBs in vitro in media of different pH values, and found both autophagy and apoptosis increased in OBs in acidic conditions. However, when autophagy inhibitor chloroquine (CQ) was used, apoptosis increased significantly compared with that without CQ. Thus indicating that inhibition of autophagy may promote apoptosis in OBs in an acidic environment, which may provide a new therapeutic strategy to decrease cell apoptosis in OBs through the use of drugs that modulate the autophagic state.

No MeSH data available.


Related in: MedlinePlus