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Up-regulation of stomatin expression by hypoxia and glucocorticoid stabilizes membrane-associated actin in alveolar epithelial cells.

Chen JC, Cai HY, Wang Y, Ma YY, Song LN, Yin LJ, Cao DM, Diao F, Li YD, Lu J - J. Cell. Mol. Med. (2013)

Bottom Line: Stomatin is an important lipid raft-associated protein which interacts with membrane proteins and plays a role in the membrane organization.In this study we found that hypoxia and dexamethasone (dex), a synthetic GC not only up-regulated the expression of stomatin alone, but also imposed additive effect on the expression of stomatin in A549 cells, primary AEC and lung of rats.Moreover, we found that hypoxia exposure did not affect membrane-associated actin, but decreased actin in cytoplasm in A549 cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathophysiology, The Second Military Medical University, Shanghai, China.

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Stomatin stabilized membrane-associated actin. (A) A549 cells were transiently transfected with blank vectors pEGFP-N3 or pEGFP-N3-stom. Images of stomatin tagged by EGFP (green) and actin filaments labelled by rhodamine-phalloidine (red) were visualized and captured using confocal microscopy. (B) A549 cells were cultured in normoxia or exposed to hypoxia or treated with 100 nM dex for 12 hrs. Actin filaments were viewed using confocal microscopy (100×). (C) Stomatin protein in A549 cells (control) or cells transfected with negative control siRNAs (control siRNA) or with 1 or 3 nM stomatin siRNAs (siRNA-stom) were analysed by Western blot. β-actin was detected as an internal control. (D) A549 cells transfected with 3 nM siRNA-stom or control siRNA were cultured either in normoxic or hypoxic environment or treated with 100 nM dex. Actin filaments were viewed using confocal microscopy (60×).
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fig06: Stomatin stabilized membrane-associated actin. (A) A549 cells were transiently transfected with blank vectors pEGFP-N3 or pEGFP-N3-stom. Images of stomatin tagged by EGFP (green) and actin filaments labelled by rhodamine-phalloidine (red) were visualized and captured using confocal microscopy. (B) A549 cells were cultured in normoxia or exposed to hypoxia or treated with 100 nM dex for 12 hrs. Actin filaments were viewed using confocal microscopy (100×). (C) Stomatin protein in A549 cells (control) or cells transfected with negative control siRNAs (control siRNA) or with 1 or 3 nM stomatin siRNAs (siRNA-stom) were analysed by Western blot. β-actin was detected as an internal control. (D) A549 cells transfected with 3 nM siRNA-stom or control siRNA were cultured either in normoxic or hypoxic environment or treated with 100 nM dex. Actin filaments were viewed using confocal microscopy (60×).

Mentions: As actin cytoskeleton plays an important role in alveolar epithelial barrier and in many cellular functions, we examined the relationship between stomatin and actin cytoskeleton. We first studied the cellular localization of actin and stomatin in A549 cells transiently transfected with blank vector (pEGFP-N3) or EGFP-tagged stomatin expression vector (pEGFP-N3-stom). The results showed that EGFP-tagged stomatin protein was clustered in the membrane and around the perinucleus. Stomatin proteins in the membrane were co-localized with membrane-associated actin filaments labelled by rhodamine-phalloidine in A549 cells (Fig. 6A).


Up-regulation of stomatin expression by hypoxia and glucocorticoid stabilizes membrane-associated actin in alveolar epithelial cells.

Chen JC, Cai HY, Wang Y, Ma YY, Song LN, Yin LJ, Cao DM, Diao F, Li YD, Lu J - J. Cell. Mol. Med. (2013)

Stomatin stabilized membrane-associated actin. (A) A549 cells were transiently transfected with blank vectors pEGFP-N3 or pEGFP-N3-stom. Images of stomatin tagged by EGFP (green) and actin filaments labelled by rhodamine-phalloidine (red) were visualized and captured using confocal microscopy. (B) A549 cells were cultured in normoxia or exposed to hypoxia or treated with 100 nM dex for 12 hrs. Actin filaments were viewed using confocal microscopy (100×). (C) Stomatin protein in A549 cells (control) or cells transfected with negative control siRNAs (control siRNA) or with 1 or 3 nM stomatin siRNAs (siRNA-stom) were analysed by Western blot. β-actin was detected as an internal control. (D) A549 cells transfected with 3 nM siRNA-stom or control siRNA were cultured either in normoxic or hypoxic environment or treated with 100 nM dex. Actin filaments were viewed using confocal microscopy (60×).
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
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fig06: Stomatin stabilized membrane-associated actin. (A) A549 cells were transiently transfected with blank vectors pEGFP-N3 or pEGFP-N3-stom. Images of stomatin tagged by EGFP (green) and actin filaments labelled by rhodamine-phalloidine (red) were visualized and captured using confocal microscopy. (B) A549 cells were cultured in normoxia or exposed to hypoxia or treated with 100 nM dex for 12 hrs. Actin filaments were viewed using confocal microscopy (100×). (C) Stomatin protein in A549 cells (control) or cells transfected with negative control siRNAs (control siRNA) or with 1 or 3 nM stomatin siRNAs (siRNA-stom) were analysed by Western blot. β-actin was detected as an internal control. (D) A549 cells transfected with 3 nM siRNA-stom or control siRNA were cultured either in normoxic or hypoxic environment or treated with 100 nM dex. Actin filaments were viewed using confocal microscopy (60×).
Mentions: As actin cytoskeleton plays an important role in alveolar epithelial barrier and in many cellular functions, we examined the relationship between stomatin and actin cytoskeleton. We first studied the cellular localization of actin and stomatin in A549 cells transiently transfected with blank vector (pEGFP-N3) or EGFP-tagged stomatin expression vector (pEGFP-N3-stom). The results showed that EGFP-tagged stomatin protein was clustered in the membrane and around the perinucleus. Stomatin proteins in the membrane were co-localized with membrane-associated actin filaments labelled by rhodamine-phalloidine in A549 cells (Fig. 6A).

Bottom Line: Stomatin is an important lipid raft-associated protein which interacts with membrane proteins and plays a role in the membrane organization.In this study we found that hypoxia and dexamethasone (dex), a synthetic GC not only up-regulated the expression of stomatin alone, but also imposed additive effect on the expression of stomatin in A549 cells, primary AEC and lung of rats.Moreover, we found that hypoxia exposure did not affect membrane-associated actin, but decreased actin in cytoplasm in A549 cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathophysiology, The Second Military Medical University, Shanghai, China.

Show MeSH
Related in: MedlinePlus