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Conditional ablation of macrophages disrupts ovarian vasculature.

Turner EC, Hughes J, Wilson H, Clay M, Mylonas KJ, Kipari T, Duncan WC, Fraser HM - Reproduction (2011)

Bottom Line: As macrophage ablation progressed, increasing amounts of ovarian haemorrhage were observed affecting both luteal and thecal tissue associated with significant endothelial cell depletion, increased erythrocyte accumulation and increased follicular atresia by 16  h.These events were followed by necrosis and profound structural damage.These results show that macrophages play a critical role in maintaining ovarian vascular integrity.

View Article: PubMed Central - PubMed

Affiliation: MRC Human Reproductive Sciences Unit, Queen's Institute of Medical Research, Centre for Reproductive Biology Obstetrics and Gynaecology, University of Edinburgh, 47 Little France Crescent, Edinburgh, EH16 4TJ, UK.

ABSTRACT
Macrophages are the most abundant immune cell within the ovary. Their dynamic distribution throughout the ovarian cycle and heterogenic array of functions suggest the involvement in various ovarian processes, but their functional role has yet to be fully established. The aim was to induce conditional macrophage ablation to elucidate the putative role of macrophages in maintaining the integrity of ovarian vasculature. Using the CD11b-diphtheria toxin receptor (DTR) mouse, in which expression of human DTR is under the control of the macrophage-specific promoter sequence CD11b, ovarian macrophages were specifically ablated in adult females by injections of diphtheria toxin (DT). CD11b-DTR mice were given DT treatment or vehicle and ovaries collected at 2, 8, 16, 24 and 48  h. Histochemical stains were employed to characterise morphological changes, immunohistochemistry for F4/80 to identify macrophages and the endothelial cell marker CD31 used to quantify vascular changes. In normal ovaries, macrophages were detected in corpora lutea and in the theca layer of healthy and atretic follicles. As macrophage ablation progressed, increasing amounts of ovarian haemorrhage were observed affecting both luteal and thecal tissue associated with significant endothelial cell depletion, increased erythrocyte accumulation and increased follicular atresia by 16  h. These events were followed by necrosis and profound structural damage. Changes were limited to the ovary, as DT treatment does not disrupt the vasculature of other tissues likely reflecting the unique cyclical nature of the ovarian vasculature and heterogeneity between macrophages within different tissues. These results show that macrophages play a critical role in maintaining ovarian vascular integrity.

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H&E-stained sections showing (A) healthy luteal tissue from control mouse ovary, (B) luteal haemorrhage in 16 h post-DT ovary surrounded by mostly healthy luteal cells, although some pyknotic cells observed, (C) by 24 h, the haemorrhage is more extensive being associated with visible reduction in healthy luteal cells, increased incidence of pyknosis and ovarian necrosis, (D) by 48 h, the extent of haemorrhage has resulted in profound ovarian damage with a dearth of healthy luteal cells. RBC, red blood cells. Bar =20 μm.
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fig2: H&E-stained sections showing (A) healthy luteal tissue from control mouse ovary, (B) luteal haemorrhage in 16 h post-DT ovary surrounded by mostly healthy luteal cells, although some pyknotic cells observed, (C) by 24 h, the haemorrhage is more extensive being associated with visible reduction in healthy luteal cells, increased incidence of pyknosis and ovarian necrosis, (D) by 48 h, the extent of haemorrhage has resulted in profound ovarian damage with a dearth of healthy luteal cells. RBC, red blood cells. Bar =20 μm.

Mentions: Macroscopically, ovaries of PBS-treated CD11b-DTR mice or DT-treated FVB/nj controls appeared normal (Fig. 1A), although increasing incidence of haemorrhage was observed with time following DT treatment, beginning at 16 h (Fig. 1B). Ovarian sections stained with haematoxylin and eosin (H&E) showed the presence of healthy corpora lutea and luteal tissue from previous cycles in all controls (Figs 1C and 2A). DT treatment of CD11b-DTR mice had no effect on morphology in ovaries collected 2 h following the DT treatment when compared with controls (not shown). By 8 h, there appeared to be focal areas of haemorrhage within the regressing luteal tissue, possibly indicating the early signs of vascular disruption. At 16 h, the changes were striking, with haemorrhage affecting a large proportion of ovarian interstitial and luteal tissue (Fig. 1D). Within the areas of luteal haemorrhage, although some cells showed signs of pyknosis, the majority of cells appeared to be healthy with nuclei retaining morphologically normal appearance (Fig. 2).


Conditional ablation of macrophages disrupts ovarian vasculature.

Turner EC, Hughes J, Wilson H, Clay M, Mylonas KJ, Kipari T, Duncan WC, Fraser HM - Reproduction (2011)

H&E-stained sections showing (A) healthy luteal tissue from control mouse ovary, (B) luteal haemorrhage in 16 h post-DT ovary surrounded by mostly healthy luteal cells, although some pyknotic cells observed, (C) by 24 h, the haemorrhage is more extensive being associated with visible reduction in healthy luteal cells, increased incidence of pyknosis and ovarian necrosis, (D) by 48 h, the extent of haemorrhage has resulted in profound ovarian damage with a dearth of healthy luteal cells. RBC, red blood cells. Bar =20 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3101494&req=5

fig2: H&E-stained sections showing (A) healthy luteal tissue from control mouse ovary, (B) luteal haemorrhage in 16 h post-DT ovary surrounded by mostly healthy luteal cells, although some pyknotic cells observed, (C) by 24 h, the haemorrhage is more extensive being associated with visible reduction in healthy luteal cells, increased incidence of pyknosis and ovarian necrosis, (D) by 48 h, the extent of haemorrhage has resulted in profound ovarian damage with a dearth of healthy luteal cells. RBC, red blood cells. Bar =20 μm.
Mentions: Macroscopically, ovaries of PBS-treated CD11b-DTR mice or DT-treated FVB/nj controls appeared normal (Fig. 1A), although increasing incidence of haemorrhage was observed with time following DT treatment, beginning at 16 h (Fig. 1B). Ovarian sections stained with haematoxylin and eosin (H&E) showed the presence of healthy corpora lutea and luteal tissue from previous cycles in all controls (Figs 1C and 2A). DT treatment of CD11b-DTR mice had no effect on morphology in ovaries collected 2 h following the DT treatment when compared with controls (not shown). By 8 h, there appeared to be focal areas of haemorrhage within the regressing luteal tissue, possibly indicating the early signs of vascular disruption. At 16 h, the changes were striking, with haemorrhage affecting a large proportion of ovarian interstitial and luteal tissue (Fig. 1D). Within the areas of luteal haemorrhage, although some cells showed signs of pyknosis, the majority of cells appeared to be healthy with nuclei retaining morphologically normal appearance (Fig. 2).

Bottom Line: As macrophage ablation progressed, increasing amounts of ovarian haemorrhage were observed affecting both luteal and thecal tissue associated with significant endothelial cell depletion, increased erythrocyte accumulation and increased follicular atresia by 16  h.These events were followed by necrosis and profound structural damage.These results show that macrophages play a critical role in maintaining ovarian vascular integrity.

View Article: PubMed Central - PubMed

Affiliation: MRC Human Reproductive Sciences Unit, Queen's Institute of Medical Research, Centre for Reproductive Biology Obstetrics and Gynaecology, University of Edinburgh, 47 Little France Crescent, Edinburgh, EH16 4TJ, UK.

ABSTRACT
Macrophages are the most abundant immune cell within the ovary. Their dynamic distribution throughout the ovarian cycle and heterogenic array of functions suggest the involvement in various ovarian processes, but their functional role has yet to be fully established. The aim was to induce conditional macrophage ablation to elucidate the putative role of macrophages in maintaining the integrity of ovarian vasculature. Using the CD11b-diphtheria toxin receptor (DTR) mouse, in which expression of human DTR is under the control of the macrophage-specific promoter sequence CD11b, ovarian macrophages were specifically ablated in adult females by injections of diphtheria toxin (DT). CD11b-DTR mice were given DT treatment or vehicle and ovaries collected at 2, 8, 16, 24 and 48  h. Histochemical stains were employed to characterise morphological changes, immunohistochemistry for F4/80 to identify macrophages and the endothelial cell marker CD31 used to quantify vascular changes. In normal ovaries, macrophages were detected in corpora lutea and in the theca layer of healthy and atretic follicles. As macrophage ablation progressed, increasing amounts of ovarian haemorrhage were observed affecting both luteal and thecal tissue associated with significant endothelial cell depletion, increased erythrocyte accumulation and increased follicular atresia by 16  h. These events were followed by necrosis and profound structural damage. Changes were limited to the ovary, as DT treatment does not disrupt the vasculature of other tissues likely reflecting the unique cyclical nature of the ovarian vasculature and heterogeneity between macrophages within different tissues. These results show that macrophages play a critical role in maintaining ovarian vascular integrity.

Show MeSH
Related in: MedlinePlus