Table.1 Time to Occlusion of carotid artery in four genotype mice with four different periods of cigarette smoke exposure
Unit: minutes
Mean ± SEM. N=5 or 6. The alphabet a and b showed significant p value<0.05. Control (CON). Acute cigarette smoke exposure (ACE).
Prolonged cigarette smoke exposure 5 weeks (PCE 5). Prolonged cigarette smoke exposure 8 weeks (PCE8).
TXAS+/+TP+/+ TXAS-/-TP+/+ TXAS+/+TP-/- TXAS-/-TP
-/-CON 31.4±4.9b 41.6±5.2b 38.2±4.4b 34.4±6.2b
ACE 14.2±1.6a 17.3±3.4a 27.3±3.7ab 16.8±2.7a
PCE 5 11.6±1.3a 16.7±1.7a 19.0±5.0a 17.3±2.1a PCE 8 12.3±2.1a 15.8±1.5a 20.0±4.0a 12.3±0.9a
Figure. 1 Whole-body smoke exposure model
Figure. 2 The carotid artery blood flow of control group in the four genotypes mice
Figure. 3 The carotid artery blood flow of ACE group in the four
genotypes mice
Figure. 4 The carotid artery blood flow of PCE 5 treated group in the
four genotypes mice
Figure. 5 The carotid artery blood flow of PCE 8 treated group in the
four genotypes mice
(a)
(b)
Figure. 6 Comparisons of Time to Occlusion of carotid artery in mice based on cigarrette smoke exposure periods and genotypes
(a) The TTO across mice genotypes with different CS exposure duration.
(Mean ± SEM.; Mice number=5 or 6; *, compare to CON of the same genotype p<0.05; **, p<0.01.) (b) The TTO across different CS exposure duration with mice genotypes. (Mean ± SEM.; Mice number=5 or 6)
(a)
(b)
0"s 100"s 200"s 300"s
400"s 500"s 600"s 700"s
FP
FP-FeCl3 TXAS+/+TP+/+
300 m"
0"s 100"s 200"s 300"s
400"s 500"s 600"s 700"s
FP
FP-FeCl3 TXAS%/%TP+/+
300 m"
(c)
(e)
Figure. 7 Platelet adhesiveness in mesentaric artery of different genotype mice
We used FP and FeCl3 induced endothelial injury to test platelet adhesiveness. (a) TXAS+/+TP+/+, (b) TXAS-/-TP+/+, (c) TXAS+/+TP-/- and (d) TXAS-/-TP-/- was exhibited individually. (e) The brightness of each genotype platelet (Intensity/area, cd/μm2 , Mean ± SEM.; N=3; *, compare to wild type p<0.05)
(a)
(b)
Figure. 8 Comparisons of luminol-induced blood H
2O
2activites in mice based on cigarette smoke exposure periods and genotypes
(a) The Luminol-induced H2O2 CL counts across mice genotypes with different CS exposure duration. (Mean ± SEM.; N=3 or 4; *, compare to CON of the same genotype p<0.05; **, p<0.01) (b) The Luminol-induced H2O2 CL counts across different CS exposure duration with mice genotypes.
(Mean ± SEM.; N=3 or 4; *, compare to TXAS+/+TP+/+ mice of the same duration)
(a) (b)
(c) (d)
Figure. 9 Reactive oxygen species levels in bronchoalveolar lavage fluid
(a) The CL counts in Lucigenin induced O2˙- across mice genotypes with different CS exposure duration. (Mean ± SEM.; N=3 or 4; *, compare to control of the same genotype p<0.05; **, p<0.01) (b) The CL counts in Lucigenin induced H2O2 across different CS exposure duration with mice genotypes. (Mean ± SEM.; N=3 or 4; *, compare to TXAS+/+TP+/+ mice of the same duration p<0.05; **, p<0.01) (c) The CL counts in Limnol induced O2˙- across mice genotypes with different CS exposure duration. (Mean ± SEM.; N=3 or 4; *, compare to control of the same genotype p<0.05; **, p<0.01) (d) The CL counts in Luminol induced H2O2 across different CS exposure duration with mice genotypes. (Mean ± SEM.; N=3 or 4; *, compare to TXAS+/+TP+/+ mice of the same duration p<0.05; **, p<0.01)(a)
(b)
Figure. 10 Plasma MDA concentration
(a) The plasma MDA concentration across mice genotypes with different CS exposure duration. (b) The plasma MDA concentration across different CS exposure duration with mice genotypes. (Mean ± SEM.; N=3 or 4, no significance)
(a)
(b) (c)
Figure. 11 Pulmonary artery thickness histological feture and relative thickness
(a) The pulmonary artery histological structure showed increment of pulmonary artery thickness while treatment with CS exposure. (H&E stain, x 400) (b) The relative thickness across mice genotypes with different CS exposure duration. (Mean ± SEM.; N=3 or 4; *, compare to control of the
same genotype p<0.05; **, p<0.01) (c) The relative thickness across different CS exposure duration with mice genotypes. (Mean ± SEM.; N=3 or 4) There was no interaction between mice genotypes and exposure treatment.
(a)
(b)
(c) (d)
Figure. 12 Histological feture of lung
(a) The lung histological structure. Emphysema was shown in CS exposure treatment groups (orange arrows) (H&E stain, x 100) (b) The lung histological structure. Arrows are pointed to alveolus where were bleeding or full of leukocytes. (H&E stain, x 400) (c) The numbers of leukocyptes across mice genotypes. (Mean ± SEM.; N=3; *, compare to control of the same genotype p<0.05; **, p<0.01) (d) The numbers of leukocyptes across exposure treatment. (Mean ± SEM.; N=6; *, compare to TXAS+/+TP+/+ mice of the same duration p<0.05; **, p<0.01)
(a)
(b) (c)
Figure. 13 Immunohistochemistry of 4-HNE
(a) The lung histological structure showed increment expression of 4-HNE while treatment with CS exposure. 4-HNE expressed in CS exposure treatment groups (orange arrow). (800 x) (b) The ratio of 4-HNE stain across mice genotypes with different CS exposure duration. (Mean ± SEM.;
N=6; **, compare to control of the same genotype p<0.01) (c) The ratio of 4-HNE stain across different CS exposure duration with mice genotypes.
(Mean ± SEM.; N=6; *, compare to TXAS+/+TP+/+ mice of the same duration p<0.05; **, p<0.01)
(a)
(b) (c)
Figure. 14 Immunohistochemistry of VWF
(a) The lung histological structure showed increment expression of VWF while treatment with CS exposure. VWF expressed in CS exposure treatment groups (orange arrow). (800 x) (b) The ratio of VWF stain across mice genotypes with different CS exposure duration. (Mean ± SEM.; N=6;
*, compare to control of the same genotype p<0.05; **, p<0.01) (c) The
ratio of VWF stain across different CS exposure duration with mice genotypes. (Mean ± SEM.; N=6; **, compare to TXAS+/+TP+/+ mice of the same duration p<0.01)
(a)
(b) (c)
Figure. 15 Immunohistochemistry of IL-1β
(a) The lung histological structure showed increment expression of IL-1β while treatment with CS exposure. IL-1β expressed in CS exposure treatment groups (orange arrow). (800 x) (b) The ratio of IL-1β stain across mice genotypes with different CS exposure duration. (Mean ± SEM.; N=6;
*, compare to control of the same genotype p<0.05; **, p<0.01) (c) The
ratio of IL-1β stain across different CS exposure duration with mice genotypes. (Mean ± SEM.; N=6; **, compare to TXAS+/+TP+/+ mice of the same duration p<0.01)
(a)
(b) (c)
Figure. 16 Immunohistochemistry of PARP-1
(a) The lung histological structure showed increment expression of PARP-1 while treatment with CS exposure. PARP-1 expressed in CS exposure treatment groups (orange arrow). (800 x) (b) The ratio of PARP-1 stain across mice genotypes with different CS exposure duration. (Mean ± SEM.;
N=6; *, compare to control of the same genotype p<0.05; **, p<0.01) (c) The ratio of PARP-1 stain across different CS exposure duration with mice
genotypes. (Mean ± SEM.; N=6; **, compare to TXAS+/+TP+/+ mice of the same duration p<0.01)
(a)
(b) (c)
Figure. 17 Immunohistochemistry of Beclin-1
(a) The lung histological structure showed increment expression of Beclin-1 while treatment with CS exposure. Beclin-1 expressed in CS exposure treatment groups (orange arrow). (800 x) (b) The ratio of Beclin-1 stain across mice genotypes with different CS exposure duration. (Mean ± SEM.;
N=6; *, compare to control of the same genotype p<0.05; **, p<0.01) (c) The ratio of Beclin-1 stain across different CS exposure duration with mice
genotypes. (Mean ± SEM.; N=6; *, compare to TXAS+/+TP+/+ mice of the same duration p<0.05; **, p<0.01)
(a)
(b) (c)
Figure. 18 Terminal deoxynucleotide transferase dUTP Nick End Labeling stain
(a) The lung histological structure showed increment expression of apoptotic cells while treatment with CS exposure. Apoptotic cells expressed in CS exposure treatment groups (orange arrow). (400 x) (b) The ratio of TUNEL stain across mice genotypes with different CS exposure duration.
(Mean ± SEM.; N=6; *, compare to control of the same genotype p<0.05;
**, p<0.01) (c) The ratio of TUNEL stain across different CS exposure duration with mice genotypes. (Mean ± SEM.; N=6; *, compare to TXAS+/+TP+/+ mice of the same duration p<0.05; **, p<0.01)
(a)
PARP-1 β-actin
(b) (c)
Figure. 19 Western blot of PARP-1
Western blot of PARP-1 in lung. (a) Expression of PARP-1 in different CS treatment during four genotypes. (b) The ratio of PARP-1/β-actin across mice genotypes with different CS exposure duration. (c) The ratio of PARP-1/β-actin across different CS exposure duration with mice genotypes.
(Mean ± SEM.; N=3 or 4)
(a)
Bcl-2 β-actin Bax
(b) (c)
(d) (e)
(f) (g)
Figure. 20 Western blot of Bcl-2 and Bax
Western blot of Bcl-2 and Bax in lung. (a) Expression of Bcl-2 and Bax in different CS treatment during four genotypes. (b) The ratio of bcl-2/β-actin across mice genotypes with different CS exposure duration. (c) The ratio of bcl-2/β-actin across different CS exposure duration with mice genotypes. (d) The ratio of bax/β-actin across mice genotypes with different CS exposure duration. (e) The ratio of bax/β-actin across different CS exposure duration with mice genotypes. (f) The ratio of bax/bcl-2 across mice genotypes with different CS exposure duration. (g) The ratio of bax/bcl-2 across different CS exposure duration with mice genotypes. (Mean ± SEM.; N=3 or 4, **, compare to same genotype control, p<0.01)
(a)
LC3 β β-actin
(b) (c)
Figure. 21 Western blot of LC3 β
Western blot of LC3 β in lung. (a) Expression of LC3 β in different CS treatment during four genotypes. (b) The ratio of LC3 β/β-actin across mice genotypes with different CS exposure duration. (c) The ratio of LC3 β/β-actin across different CS exposure duration with mice genotypes. (Mean
± SEM.; N=5 or 6, *, compare to same genotype control, p<0.05)
(a)
Beclin-1 β-actin
(b) (c)
Figure. 22 Western blot of Beclin-1
Western blot of Beclin-1 in lung. (a) Expression of Beclin-1 in different CS treatment during four genotypes. (b) The ratio of Beclin-1/β-actin across mice genotypes with different CS exposure duration. (c) The ratio of Beclin-1/β-actin across different CS exposure duration with mice genotypes.
(Mean ± SEM.; N=5 or 6, *, compare to same genotype control, p<0.05)
(a)
4-HNE β-actin
(b) (c)
Figure. 23 Western blot of 4-HNE
Western blot of 4-HNE in lung. (a) Expression of 4-HNE in different CS treatment during four genotypes. (b) The ratio of 4-HNE/β-actin across mice genotypes with different CS exposure duration. (c) The ratio of 4-HNE/β-actin across different CS exposure duration with mice genotypes.
(Mean ± SEM.; N=3 or 4)
(a)
VWF β-actin
(b) (c)
Figure. 24 Western blot of VWF
Western blot of VWF in lung. (a) Expression of VWF in different CS treatment during four genotypes. (b) The ratio of VWF/β-actin across mice genotypes with different CS exposure duration. (c) The ratio of VWF/β-actin across different CS exposure duration with mice genotypes.
(Mean ± SEM.; N=3 or 4)
(a) treatment during four genotypes. (b) The ratio of NF-κB/β-actin across mice genotypes with different CS exposure duration. (c) The ratio of NF-κB/β-actin across different CS exposure duration with mice genotypes.
(Mean ± SEM.; N=5 or 6)
(a)
ATF-6 β-actin
(b) (c)
Figure. 26 Western blot of ATF-6
Western blot of ATF-6 in lung. (a) Expression of ATF-6 in different CS treatment during four genotypes. (b) The ratio of ATF-6/β-actin across mice genotypes with different CS exposure duration. (c) The ratio of ATF-6/β-actin across different CS exposure duration with mice genotypes.
(Mean ± SEM.; N=3 or 4)
(a)
p-JNK β-actin
(b) (c)
Figure. 27 Western blot of p-JNK
Western blot of p-JNK in lung. (a) Expression of p-JNK in different CS treatment during four genotypes. (b) The ratio of p-JNK/β-actin across mice genotypes with different CS exposure duration. (c) The ratio of p-JNK/β-actin across different CS exposure duration with mice genotypes.
(Mean ± SEM.; N=3 or 4)