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Expression of PHB2 in rat brain cortex following traumatic brain injury.

Xu T, Fan X, Tan Y, Yue Y, Chen W, Gu X - Int J Mol Sci (2014)

Bottom Line: Knocking down PHB2 by siRNA partly increased the apoptosis level of PC12 stimulated by H2O2.While the PHB2 was interrupted by siRNA, the proliferation level of primary cultured astrocytes was inhibited notably than that in the control group.Together with our data, we hypothesized that PHB2 might play an important role in CNS pathophysiology after TBI.

View Article: PubMed Central - PubMed

Affiliation: The Center Laboratory of Huai'an First People's Hospital Nanjing Medical University, Huai'an 223300, China. xuting-1988@163.com.

ABSTRACT
Prohibitin2 (PHB2) is a ubiquitous, evolutionarily strongly conserved protein. It is one of the components of the prohibitin complex, which comprises two highly homologous subunits, PHB1 and PHB2. PHB2 is present in various cellular compartments including the nucleus and mitochondria. Recent studies have identified PHB2 as a multifunctional protein that controls cell proliferation, apoptosis, cristae morphogenesis and the functional integrity of mitochondria. However its distribution and function in the central nervous system (CNS) are not well understood. In this study, we examined PHB2 expression and cellular localization in rats after acute traumatic brain injury (TBI). Western Blot analysis showed PHB2 level was significantly enhanced at five days after injury compared to control, and then declined during the following days. The protein expression of PHB2 was further analyzed by immunohistochemistry. In comparison to contralateral cerebral cortex, we observed a highly significant accumulation of PHB2 at the ipsilateral brain. Immunofluorescence double-labeling showed that PHB2 was co-expressed with NeuN, GFAP. Besides, PHB2 also colocalized with activated caspase-3 and PCNA. To further investigate the function of PHB2, primary cultured astrocytes and the neuronal cell line PC12 were employed to establish a proliferation model and an apoptosis model, respectively, to simulate the cell activity after TBI to a certain degree. Knocking down PHB2 by siRNA partly increased the apoptosis level of PC12 stimulated by H2O2. While the PHB2 was interrupted by siRNA, the proliferation level of primary cultured astrocytes was inhibited notably than that in the control group. Together with our data, we hypothesized that PHB2 might play an important role in CNS pathophysiology after TBI.

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The staining changes of PHB2 immunoreactivity in rat brain cortex after TBI. We performed immunohistochemistry with anti-PHB2 rabbit polyclonal antibody on transverse cryosections of brain cortex and investigated the temporal pattern of PHB2. We could see that the immunostaining of PHB2 was widely distributed throughout the rat brain with a relatively lower level staining in the contralateral and sham brain, and the major cellular morphology of PHB2-positive cells appeared as neuron (A,B,E); While the expression pattern was different, that expression was intensive and the cellular morphology appeared as both neurons and astrocytes at day five after TBI (C,D); Quantitative analysis documented that there was a dramatic elevation of PHB2-positive cells after TBI (G); (F) Immunostaining of negative control for PHB2. Error bars represent SEM, Scale bars: 200 μm (A,C), 50 μm (B,D–F), * p < 0.05 (n = 3).
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f2-ijms-15-03299: The staining changes of PHB2 immunoreactivity in rat brain cortex after TBI. We performed immunohistochemistry with anti-PHB2 rabbit polyclonal antibody on transverse cryosections of brain cortex and investigated the temporal pattern of PHB2. We could see that the immunostaining of PHB2 was widely distributed throughout the rat brain with a relatively lower level staining in the contralateral and sham brain, and the major cellular morphology of PHB2-positive cells appeared as neuron (A,B,E); While the expression pattern was different, that expression was intensive and the cellular morphology appeared as both neurons and astrocytes at day five after TBI (C,D); Quantitative analysis documented that there was a dramatic elevation of PHB2-positive cells after TBI (G); (F) Immunostaining of negative control for PHB2. Error bars represent SEM, Scale bars: 200 μm (A,C), 50 μm (B,D–F), * p < 0.05 (n = 3).

Mentions: We could see that the immunostaining of PHB2 staining was widely distributed throughout the rat brain in the contralateral and sham brain but with a relatively lower level staining, and the major cellular morphology of PHB2-positive cells appeared as neuron (Figure 2A,B,E), while the expression pattern was different that expression was intensive and the cellular morphology appeared as both neurons and astrocytes at day 5 after TBI (Figure 2C,D). Quantitative analysis documented that there was a dramatic elevation of PHB2 positive cells after TBI (Figure 2G).


Expression of PHB2 in rat brain cortex following traumatic brain injury.

Xu T, Fan X, Tan Y, Yue Y, Chen W, Gu X - Int J Mol Sci (2014)

The staining changes of PHB2 immunoreactivity in rat brain cortex after TBI. We performed immunohistochemistry with anti-PHB2 rabbit polyclonal antibody on transverse cryosections of brain cortex and investigated the temporal pattern of PHB2. We could see that the immunostaining of PHB2 was widely distributed throughout the rat brain with a relatively lower level staining in the contralateral and sham brain, and the major cellular morphology of PHB2-positive cells appeared as neuron (A,B,E); While the expression pattern was different, that expression was intensive and the cellular morphology appeared as both neurons and astrocytes at day five after TBI (C,D); Quantitative analysis documented that there was a dramatic elevation of PHB2-positive cells after TBI (G); (F) Immunostaining of negative control for PHB2. Error bars represent SEM, Scale bars: 200 μm (A,C), 50 μm (B,D–F), * p < 0.05 (n = 3).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3958913&req=5

f2-ijms-15-03299: The staining changes of PHB2 immunoreactivity in rat brain cortex after TBI. We performed immunohistochemistry with anti-PHB2 rabbit polyclonal antibody on transverse cryosections of brain cortex and investigated the temporal pattern of PHB2. We could see that the immunostaining of PHB2 was widely distributed throughout the rat brain with a relatively lower level staining in the contralateral and sham brain, and the major cellular morphology of PHB2-positive cells appeared as neuron (A,B,E); While the expression pattern was different, that expression was intensive and the cellular morphology appeared as both neurons and astrocytes at day five after TBI (C,D); Quantitative analysis documented that there was a dramatic elevation of PHB2-positive cells after TBI (G); (F) Immunostaining of negative control for PHB2. Error bars represent SEM, Scale bars: 200 μm (A,C), 50 μm (B,D–F), * p < 0.05 (n = 3).
Mentions: We could see that the immunostaining of PHB2 staining was widely distributed throughout the rat brain in the contralateral and sham brain but with a relatively lower level staining, and the major cellular morphology of PHB2-positive cells appeared as neuron (Figure 2A,B,E), while the expression pattern was different that expression was intensive and the cellular morphology appeared as both neurons and astrocytes at day 5 after TBI (Figure 2C,D). Quantitative analysis documented that there was a dramatic elevation of PHB2 positive cells after TBI (Figure 2G).

Bottom Line: Knocking down PHB2 by siRNA partly increased the apoptosis level of PC12 stimulated by H2O2.While the PHB2 was interrupted by siRNA, the proliferation level of primary cultured astrocytes was inhibited notably than that in the control group.Together with our data, we hypothesized that PHB2 might play an important role in CNS pathophysiology after TBI.

View Article: PubMed Central - PubMed

Affiliation: The Center Laboratory of Huai'an First People's Hospital Nanjing Medical University, Huai'an 223300, China. xuting-1988@163.com.

ABSTRACT
Prohibitin2 (PHB2) is a ubiquitous, evolutionarily strongly conserved protein. It is one of the components of the prohibitin complex, which comprises two highly homologous subunits, PHB1 and PHB2. PHB2 is present in various cellular compartments including the nucleus and mitochondria. Recent studies have identified PHB2 as a multifunctional protein that controls cell proliferation, apoptosis, cristae morphogenesis and the functional integrity of mitochondria. However its distribution and function in the central nervous system (CNS) are not well understood. In this study, we examined PHB2 expression and cellular localization in rats after acute traumatic brain injury (TBI). Western Blot analysis showed PHB2 level was significantly enhanced at five days after injury compared to control, and then declined during the following days. The protein expression of PHB2 was further analyzed by immunohistochemistry. In comparison to contralateral cerebral cortex, we observed a highly significant accumulation of PHB2 at the ipsilateral brain. Immunofluorescence double-labeling showed that PHB2 was co-expressed with NeuN, GFAP. Besides, PHB2 also colocalized with activated caspase-3 and PCNA. To further investigate the function of PHB2, primary cultured astrocytes and the neuronal cell line PC12 were employed to establish a proliferation model and an apoptosis model, respectively, to simulate the cell activity after TBI to a certain degree. Knocking down PHB2 by siRNA partly increased the apoptosis level of PC12 stimulated by H2O2. While the PHB2 was interrupted by siRNA, the proliferation level of primary cultured astrocytes was inhibited notably than that in the control group. Together with our data, we hypothesized that PHB2 might play an important role in CNS pathophysiology after TBI.

Show MeSH
Related in: MedlinePlus