Long-term histological effects of high-dose prednisolone administration on the mitral valve in normal Beagle dogs

Figures

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  Fig. 1.. Image showing each layer (atrialis, spongiosa, and fibrosa layer) used for measurement in a HE-stained sample in the proximal of the AML. (A) Atrialis layer; (B) spongiosa layer; and (C) fibrosa layer. Scale bar: 100 μm.

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  Fig. 2.. HE-stained sample images of the MV (AML and PML) in the control and prednisolone groups. C group: Control group; P group: Prednisolone group; blue bar: atrialis layer; yellow bar: spongiosa layer; green bar: fibrosa layer; *: in univariable linear regression, the percentage (%) of the total layer occupied by the spongiosa layer was higher in group P than in group C at the proximal (p = 0.018) and middle AML (p = 0.011). In contrast, the percentage of the fibrosa layer thickness to the total layer thickness (%) was lower in group P than in group C in the middle PML region (p = 0.047) (Table 2); †: in univariable linear regression, cell counts in the fibrosa layer were lower in group P than in group C at the proximal AML (p = 0.033), in the spongiosa layer (p = 0.046), and overall (p = 0.049) at the middle AML region (Table 4). Scale bar: 100 μm.

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  Fig. 3.. AB-stained sample images of the MV (AML and PML) in the control and prednisolone groups. The light blue stained areas indicate the presence of acidic sulfated mucosubstances. C group: Control group; P group: Prednisolone group; blue bar: atrialis layer; yellow bar: spongiosa layer; green bar: fibrosa layer; *: in univariable linear regression, the percentage (%) of the positively stained area was lower in group P than in group C in the fibrosa layer of the middle AML (p = 0.031) and the total layer of the middle AML (p = 0.023) (Table 6). Scale bar: 100 μm.

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  Fig. 4.. MT-stained sample images of the MV (AML and PML) in the control and prednisolone groups. The blue-stained areas indicate the presence of collagen-rich deposits. C group: Control group; P group: Prednisolone group; blue bar: atrialis layer; yellow bar: spongiosa layer; green bar: fibrosa layer; *: univariable linear regression analysis of the percentage of the positively stained area (%) was lower in group P than in group C in the spongiosa layer and total layers of the proximal AML (p = 0.032 and 0.004, respectively); in the total layer of the middle of the AML (p = 0.004); in the fibrosa layer and total layers of the proximal PML (p = 0.006 and 0.004, respectively); in the fibrosa layer and total layers in the middle PML (p = 0.009 and 0.002, respectively); and in the fibrosa layer and total layers in the distal of the PML (p = 0.041, 0.034) (Table 8). Scale bar: 100 μm.

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  Fig. 5.. ADN-immunohistochemistry images of the MV (AML and PML) in the control and prednisolone groups. Arrowheads show ADN-positive areas in the cytoplasm of mature adipocytes in the spongiosa layer. C group: Control group; P group: Prednisolone group; blue bar: atrialis layer; yellow bar: spongiosa layer; green bar: fibrosa layer; *: in univariable linear regression the percentage of the ADN-positive areas in the spongiosa layer was higher in group P than in group C in the middle of the AML (p = 0.016) (Table 9). Scale bar: 100 μm.

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  Fig. 6.. GCR immunohistochemistry images of the MV (AML and PML) in the control and prednisolone groups. Arrowheads show GCR-positive nuclei in the spongiosa (mainly expressed in adipocytes and cardiomyocytes) and fibrosa layer. C group: Control group; P group: Prednisolone group; blue bar: atrialis layer; yellow bar: spongiosa layer; green bar: fibrosa layer; *: univariable linear regression analysis of the percentage of GCR-positive nuclei (%) was lower in group P than in group C in the fibrosa layer in the proximal AML (p = 0.039); in the spongiosa layer in the distal AML (p = 0.034); in the fibrosa and total layers in the proximal PML (p = 0.031 and 0.031, respectively) (Table 12). GCR was expressed in the spongiosa and fibrosa layers of the AML and PML in groups C and P. Scale bar: 100 μm.