Limits...
Endothelial Antioxidant-1: a Key Mediator of Copper-dependent Wound Healing in vivo

View Article: PubMed Central - PubMed

ABSTRACT

Copper (Cu), an essential nutrient, promotes wound healing, however, target of Cu action and underlying mechanisms remain elusive. Cu chaperone Antioxidant-1 (Atox1) in the cytosol supplies Cu to the secretory enzymes such as lysyl oxidase (LOX), while Atox1 in the nucleus functions as a Cu-dependent transcription factor. Using mouse cutaneous wound healing model, here we show that Cu content (by X-ray Fluorescence Microscopy) and nuclear Atox1 are increased after wounding, and that wound healing with and without Cu treatment is impaired in Atox1−/− mice. Endothelial cell (EC)-specific Atox1−/− mice and gene transfer of nuclear-target Atox1 in Atox1−/− mice reveal that Atox1 in ECs as well as transcription factor function of Atox1 are required for wound healing. Mechanistically, Atox1−/− mice show reduced Atox1 target proteins such as p47phox NADPH oxidase and cyclin D1 as well as extracellular matrix Cu enzyme LOX activity in wound tissues. This in turn results in reducing O2− production in ECs, NFkB activity, cell proliferation and collagen formation, thereby inhibiting angiogenesis, macrophage recruitment and extracellular matrix maturation. Our findings suggest that Cu-dependent transcription factor/Cu chaperone Atox1 in ECs plays an important role to sense Cu to accelerate wound angiogenesis and healing.

No MeSH data available.


Related in: MedlinePlus

Atox1 is required for ECM maturation and Cu enzyme LOX activation.(A,B) Masson’s Trichrome staining, scale bars = 500 μm (A) and LOX activity (B) in wound tissues at day 7 after wounding in WT and Atox1−/− mice. In (A) boxed regions are shown at higher magnification to the right, scale bars = 100 μm. Blue color indicates the collagen deposition; light red or pink for keratin, muscle or cytoplasm; and dark brown or black for cell nuclei. W: wound area; WE: wound edge In (B) a graph represents mean ± SE for LOX activity and a western blot represents Pro-LOX protein expression in wound tissues at days 0 and 7 (n = 3. **p < 0.01 vs. WT). (C) Schematic diagram showing the essential role of Cu-dependent transcription factor and Cu chaperone function of Atox1 in Cu-dependent wound healing.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC5036036&req=5

f9: Atox1 is required for ECM maturation and Cu enzyme LOX activation.(A,B) Masson’s Trichrome staining, scale bars = 500 μm (A) and LOX activity (B) in wound tissues at day 7 after wounding in WT and Atox1−/− mice. In (A) boxed regions are shown at higher magnification to the right, scale bars = 100 μm. Blue color indicates the collagen deposition; light red or pink for keratin, muscle or cytoplasm; and dark brown or black for cell nuclei. W: wound area; WE: wound edge In (B) a graph represents mean ± SE for LOX activity and a western blot represents Pro-LOX protein expression in wound tissues at days 0 and 7 (n = 3. **p < 0.01 vs. WT). (C) Schematic diagram showing the essential role of Cu-dependent transcription factor and Cu chaperone function of Atox1 in Cu-dependent wound healing.

Mentions: Masson’s Trichrome staining in Fig. 9A revealed that ECM maturation via collagen deposition was markedly reduced in the healing wound after 7 days in Atox1−/− mice as compared to WT mice. There appears to be an increase in the amount of muscle tissue or keratin presumably due to compensatory response to impaired ECM maturation and angiogenic responses in Atox1−/− wound tissues. Since cytosolic Atox1 functions as a Cu chaperone for secretory ECM Cu enzyme LOX which is involved in ECM maturation during wound healing192022 and VEGF-induced angiogenesis in ECs25, we next examined a role of Atox1 for LOX activity in wound tissues. Figure 9B showed that wound injury-induced LOX activity was almost completely abolished in Atox1−/− mice without affecting precursor pro-LOX protein expression. These results suggest that Cu chaperone function of Atox1 is required for ECM maturation by activating LOX which catalyses the cross links and acts collagen deposition or fibrosis during wound healing.


Endothelial Antioxidant-1: a Key Mediator of Copper-dependent Wound Healing in vivo
Atox1 is required for ECM maturation and Cu enzyme LOX activation.(A,B) Masson’s Trichrome staining, scale bars = 500 μm (A) and LOX activity (B) in wound tissues at day 7 after wounding in WT and Atox1−/− mice. In (A) boxed regions are shown at higher magnification to the right, scale bars = 100 μm. Blue color indicates the collagen deposition; light red or pink for keratin, muscle or cytoplasm; and dark brown or black for cell nuclei. W: wound area; WE: wound edge In (B) a graph represents mean ± SE for LOX activity and a western blot represents Pro-LOX protein expression in wound tissues at days 0 and 7 (n = 3. **p < 0.01 vs. WT). (C) Schematic diagram showing the essential role of Cu-dependent transcription factor and Cu chaperone function of Atox1 in Cu-dependent wound healing.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5036036&req=5

f9: Atox1 is required for ECM maturation and Cu enzyme LOX activation.(A,B) Masson’s Trichrome staining, scale bars = 500 μm (A) and LOX activity (B) in wound tissues at day 7 after wounding in WT and Atox1−/− mice. In (A) boxed regions are shown at higher magnification to the right, scale bars = 100 μm. Blue color indicates the collagen deposition; light red or pink for keratin, muscle or cytoplasm; and dark brown or black for cell nuclei. W: wound area; WE: wound edge In (B) a graph represents mean ± SE for LOX activity and a western blot represents Pro-LOX protein expression in wound tissues at days 0 and 7 (n = 3. **p < 0.01 vs. WT). (C) Schematic diagram showing the essential role of Cu-dependent transcription factor and Cu chaperone function of Atox1 in Cu-dependent wound healing.
Mentions: Masson’s Trichrome staining in Fig. 9A revealed that ECM maturation via collagen deposition was markedly reduced in the healing wound after 7 days in Atox1−/− mice as compared to WT mice. There appears to be an increase in the amount of muscle tissue or keratin presumably due to compensatory response to impaired ECM maturation and angiogenic responses in Atox1−/− wound tissues. Since cytosolic Atox1 functions as a Cu chaperone for secretory ECM Cu enzyme LOX which is involved in ECM maturation during wound healing192022 and VEGF-induced angiogenesis in ECs25, we next examined a role of Atox1 for LOX activity in wound tissues. Figure 9B showed that wound injury-induced LOX activity was almost completely abolished in Atox1−/− mice without affecting precursor pro-LOX protein expression. These results suggest that Cu chaperone function of Atox1 is required for ECM maturation by activating LOX which catalyses the cross links and acts collagen deposition or fibrosis during wound healing.

View Article: PubMed Central - PubMed

ABSTRACT

Copper (Cu), an essential nutrient, promotes wound healing, however, target of Cu action and underlying mechanisms remain elusive. Cu chaperone Antioxidant-1 (Atox1) in the cytosol supplies Cu to the secretory enzymes such as lysyl oxidase (LOX), while Atox1 in the nucleus functions as a Cu-dependent transcription factor. Using mouse cutaneous wound healing model, here we show that Cu content (by X-ray Fluorescence Microscopy) and nuclear Atox1 are increased after wounding, and that wound healing with and without Cu treatment is impaired in Atox1&minus;/&minus; mice. Endothelial cell (EC)-specific Atox1&minus;/&minus; mice and gene transfer of nuclear-target Atox1 in Atox1&minus;/&minus; mice reveal that Atox1 in ECs as well as transcription factor function of Atox1 are required for wound healing. Mechanistically, Atox1&minus;/&minus; mice show reduced Atox1 target proteins such as p47phox NADPH oxidase and cyclin D1 as well as extracellular matrix Cu enzyme LOX activity in wound tissues. This in turn results in reducing O2&minus; production in ECs, NFkB activity, cell proliferation and collagen formation, thereby inhibiting angiogenesis, macrophage recruitment and extracellular matrix maturation. Our findings suggest that Cu-dependent transcription factor/Cu chaperone Atox1 in ECs plays an important role to sense Cu to accelerate wound angiogenesis and healing.

No MeSH data available.


Related in: MedlinePlus