Fig 1.
Histology of swine coronary artery and SEV tissue sections.
The H&E staining of uninjured coronary artery (A, B, C) shows normal histology, whereas 16 weeks after balloon angioplasty neointimal hyperplasia (arrows) is noted in post PTCA-coronary arteries (D, E, F). H&E staining of the uninjured SEV showed normal histology (G, H, I). SEV that was used as a graft (J, K, L) developed neointimal hyperplasia (arrows). All images were taken in 4x, 10x and 20x objective. This is a representative of tissue histology in 2–5 individual swine in each experimental group. L: lumen, M: medial layer, NI: neointima.
Fig 2.
Double immunofluorescence showing the expression of PLK1 and SMemb in post PTCA- coronary arteries, SEV-grafts, and their uninjured counterparts in porcine model.
There is less immunopositivity to PLK1 (red) and SMemb (green) expression in uninjured coronary artery at low (20x) (A) and high (40x) magnification (B,C,D). The immunostaining of PLK1 (red) and SMemb (green) is noted in restenotic post PTCA- coronary arteries (arrows) at low (20x) (E) and high (40x) magnification (F, G, H). Similarly, less immunostaining of PLK1 (red) and SMemb (green) expression is noted in the uninjured SEV at low (20x) (I) and high (40x) magnification (J, K, L) compared to SEV post CABG, where many cells showed immunopositivity to PLK1 (red) and SMemb (green) at low (20x) (M) and high (40x) (N,O,P) magnification. This is a representative of tissue histology in 2–5 individual swine in each experimental group. MFI measured in arbitrary units (AU) in post-PTCA coronary arteries (Q) and SEV-grafts (R) compared to their uninjured counterparts. Graphs showing number of PLK1- positive (+) cells in the neointima of the post-PTCA restenotic coronary artery (S) and SEV-grafts (T) compared to their uninjured counterparts. There was significantly increased immunopositivity to SMemb and PLK1 in the neointima of the post-PTCA restenotic coronary artery (arrows) compared to uninjured coronary artery. SEV-graft had significantly increased immunopositivity to SMemb and PLK1 in the neointima compared to uninjured SEV. Data are shown as mean ± SEM of values of at least three measurements in 2–5 individual swine in each experimental group. *P < 0.05. L: lumen, M: medial layer, NI: neointima.
Fig 3.
Double immunofluorescence showing the expression of pPLK1 and SMemb in post- PTCA coronary arteries, SEV-grafts, and their uninjured counterparts in swine.
There is less immunopositivity to pPLK1 (green) and SMemb (red) expression in uninjured coronary artery at low (20x) (A) and high (40x) magnification (B,C,D). The immunostaining of pPLK1 (green) and SMemb (red) is noted in restenotic post PTCA- coronary arteries (arrows) at low (20x) (E) and high (40x) magnification (F, G, H). Similarly, less immunostaining of pPLK1 (green) and SMemb (red) expression is noted in the uninjured SEV shown at low (20x) (I) and high (40x) magnification (J, K, L) compared to SEV post CABG, where many cells showed immunopositivity to pPLK1 (green) and SMemb (red) at low (20x) (M) and high (40x) (N, O, P) magnification. This is a representative of tissue histology in 2–5 individual swine in each experimental group. MFI measured in arbitrary units (AU) in post-PTCA coronary arteries (Q) and SEV-grafts (R) compared to their uninjured counterparts. Graphs showing number of pPLK1- positive (+) cells in the neointima of the post-PTCA restenotic coronary artery (S) and SEV-grafts (T) compared to their uninjured counterparts. There was significantly increased immunopositivity to SMemb and pPLK1 in the restenotic post-PTCA coronary arteries compared to uninjured coronary arteries. SEV-graft had significantly increased immunopositivity to SMemb and pPLK1 in the neointima compared to uninjured SEV. Data are shown as mean ± SEM of values of at least three measurements in 2–5 individual swine in each experimental group. *P < 0.05. L: lumen, M: medial layer, NI: neointima.
Fig 4.
Double immunofluorescence showing expression of pHistone H3 at Ser-10 and SMemb in thin sections of uninjured swine coronaries, post-PTCA coronary arteries, uninjured swine SEV and SEV-graft.
There is less immunopositivity to pHistone (green) and SMemb (red) expression in uninjured coronary artery shown at low (20x) (A) and high (40x) magnification (B, C, D). The immunostaining of pHistone (green) and SMemb (red) is noted in restenotic post PTCA- coronary arteries (arrows) at low (20x) (E) and high (40x) magnification (F, G, H). Similarly, less immunostaining of pHistone (green) and SMemb (red) expression is noted in the uninjured SEV shown at low (20x) (I) and high (40x) magnification (J, K, L) compared to SEV post CABG, where many cells showed immunopositivity to pHistone (green) and SMemb (red) at low (20x) (M) and high (40x) (N,O,P) magnification. This is a representative of tissue histology in 2–5 individual swine in each experimental group. MFI measured in arbitrary units (AU) in post-PTCA coronary arteries (Q) and SEV-grafts (R) compared to their uninjured counterparts. Graphs showing number of pHistone- positive (+) cells in the neointima of the post-PTCA restenotic coronary artery (S) and SEV-grafts (T) compared to their uninjured counterparts. Strong immunopositivity to pHistone H3 was found in the neointima of the post PTCA-coronary arteries and in SEV-graft. Many cells in the neointima of the injured vessels were immunopositive for both, pHistone H3 and SMemb, especially around the pseudo lumen in SEV-graft. Data are shown as mean ± SEM of values of at least three measurements in each group in 2–5 individual swine in each experimental group. *P < 0.05. L: lumen, M: medial layer, NI: neointima.
Fig 5.
Double immunofluorescence showing expression of smooth muscle α-actin (α-SMC) and SMemb in thin sections of uninjured coronaries, post PTCA-coronary arteries, uninjured SEV and SEV-grafts in swine.
Expression of α-SMC (green) and SMemb (red) are noted in uninjured coronary artery at low (20x) (A) and high (40x) magnification (B, C, D). Less expression of α-SMC (green) and increased expression of SMemb (red) are found in the hyperplastic region post PTCA- coronary arteries (arrows) at low (20x) (E) and high (40x) magnification (F, G, H). Similarly, immunopositivity to α-SMC (green) and SMemb (red) are observed in the uninjured SEV at low (20x) (I) and high (40x) magnification (J, K, L) compared to SEV post CABG, where few cells are immunopositive to α-SMC (green) and many cells showed immunopositivity to SMemb (red) at low (20x) (M) and high (40x) (N, O, P) magnification. Increased density of SMemb-immunopositive cells are found at the neointima in post PTCA- coronary arteries and SEV-grafts compared to that of normal uninjured coronary artery and vein, respectively. Strong expression of α-SMA is noted in all the vessels, injured or uninjured. This is a representative of tissue histology in 2–5 individual swine in each experimental group. L: lumen, M: medial layer, NI: neointima.
Fig 6.
Double immunofluorescence showing expression of IFNγ (green) and SMemb (red) in thin sections of uninjured coronaries, post PTCA-coronary arteries, uninjured SEV and SEV-graft.
Less immunopositivity to IFNγ (green) and SMemb (red) are noted in uninjured coronary artery at low (20x) (A) and high (40x) magnification (B, C, D). Strong expression of IFNγ (green) and SMemb (red) are found in the hyperplastic region post PTCA- coronary arteries (arrows) at low (20x) (E) and high (40x) magnification (F, G, H). Similarly, less immunostaining of IFNγ (green) and SMemb (red) expression is noted in the uninjured SEV at low (20x) (I) and high (40x) magnification (J, K, L) compared to SEV post CABG, where many cells showed immunopositivity to IFNγ (green) and SMemb (red) at low (20x) (M) and high (40x) (N,O,P) magnification. This is a representative of tissue histology in 2–5 individual swine in each experimental group. MFI measured in arbitrary units (AU) in post-PTCA coronary arteries (Q) and SEV-grafts (R) compared to their uninjured counterparts. Graphs showing number of IFNγ - positive (+) cells in the neointima of the post-PTCA restenotic coronary artery (S) and SEV-grafts (T) compared to their uninjured counterparts. Many cells in the neointima of the injured vessels were immunopositive for both, IFNγ and SMemb, especially around the pseudo lumen in SEV-graft. Data are shown as mean ± SEM of values of at least three measurements in each group in 2–5 individual swine in each experimental group. *P < 0.05. L: lumen, M: medial layer, NI: neointima.
Fig 7.
Double immunofluorescence showing expression of pStat-3 (green) and PLK1 (red) in thin sections of uninjured coronaries, post PTCA-coronary arteries, uninjured SEV and SEV-graft in swine.
Less immunopositivity to pStat-3 (green) and PLK1 (red) are noted in uninjured coronary artery at low (20x) (A) and high (40x) magnification (B, C, D). Strong expression of pStat-3 (green) and PLK1 (red) are found in the hyperplastic region post PTCA- coronary arteries (arrows) at low (20x) (E) and high (40x) magnification (F, G, H). Similarly, less immunostaining of pStat-3 (green) and PLK1 (red) expression is noted in the uninjured SEV at low (20x) (I) and high (40x) magnification (J, K, L) compared to SEV post CABG, where many cells showed immunopositivity to pStat-3 (green) and PLK1 (red) at low (20x) (M) and high (40x) (N,O,P) magnification. This is a representative of tissue histology in 2–5 individual swine in each experimental group. MFI measured in arbitrary units (AU) in post-PTCA coronary arteries (Q) and SEV-grafts (R) compared to their uninjured counterparts. Graphs showing number of pStat-3—positive (+) cells in the neointima of the post-PTCA restenotic coronary artery (S) and SEV-grafts (T) compared to their uninjured counterparts. Increased immunopositivity towards pStat-3 and PLK1 are found in the neointimal region of the injured vessels compared to the uninjured vessels. This is a representative of tissue histology in 2–5 individual swine in each experimental group. L: lumen, M: medial layer, NI: neointima.