International Journal of Cardiology 176 (2014) 1225–1226
Contents lists available at ScienceDirect
International Journal of Cardiology journal homepage: www.elsevier.com/locate/ijcard
Letter to the Editor
Vitamin D supplementation intensifies cardiac remodeling after experimental myocardial infarction☆ Andrea F. Gonçalves a, Priscila P. Santos a, Bruna P. Rafacho a, Diego F. Batista a, Paula S. Azevedo a, Marcos F. Minicucci a, Bertha F. Polegato a, Fernanda Chiuso-Minicucci b, Katashi Okoshi a, Ana Angélica Fernandes c, Sergio A. Paiva a, Leonardo A. Zornoff a,⁎ a b c
Internal Medicine Department, Botucatu Medical School, UNESP — Univ Estadual Paulista, Botucatu, Brazil Department of Microbiology and Immunology, Institute of Biosciences, UNESP — Univ Estadual Paulista, Botucatu, Brazil Chemistry and Biochemistry Department, Institute of Biosciences, UNESP — Univ Estadual Paulista, Botucatu, Brazil
a r t i c l e
i n f o
Article history: Received 1 July 2014 Accepted 27 July 2014 Available online 5 August 2014 Keywords: Remodeling Cardiac function Apoptosis Vitamin D
In recent years, vitamin D has emerged as an important nutrient for heart health, and its deficiency has been linked to cardiac dysfunction. Thus, the present study tested the hypothesis that vitamin D supplementation would attenuate cardiac remodeling induced by coronary occlusion. The protocol of this study was approved by the Ethics Committee on Animal Experiments of our institution. Wistar rats were allocated into four groups: 1) sham control (SC): animals that received standard chow; 2) sham vitamin D (SD): sham animals that received standard chow and cholecalciferol (3000 IU/kg chow); 3) infarction control (IC): infarcted animals that received standard chow; and 4) infarction vitamin D (ID): infarcted animals that received standard chow and cholecalciferol. The tests were performed as previously described [1–3]. Comparison between groups was performed by two-way ANOVA and the post-hoc Holm–Sidak test.
☆ Funding: This work was supported by FAPESP (12/14500-2) and CNPq (301945/ 2010-1). ⁎ Corresponding author at: Botucatu Medical School, Botucatu, SP 18618-970, Brazil. Tel.: +55 14 38222969. E-mail address: [email protected] (L.A. Zornoff).
http://dx.doi.org/10.1016/j.ijcard.2014.07.217 0167-5273/© 2014 Elsevier Ireland Ltd. All rights reserved.
Considering our results, after three months, the infarct size did not differ between the infarcted groups (IC = 46.7 ± 6.5%, ID = 45.8 ± 6.3% p = 0.593). Serum concentrations of 25[OH]D3 were higher in supplemented groups than non-supplemented groups (SC = 8.48 ± 1.27 ng/mL, SD = 14.94 ± 1.73, IC = 11.88 ± 1.28, ID = 13.47 ± 1.76; p(I) = 0.610, p(VD) = 0.034, p(I × VD) = 0.173). Vitamin D increased the left ventricular systolic and diastolic areas and decreased the fractional area change and posterior wall shortening velocity associated with increased E/E′ ratio. In addition, vitamin D decreased phosphofructokinase activity (Table 1). Conversely, vitamin D did not affect interstitial collagen volume fraction, vitamin D3-upregulated protein 1 (VDUP-1), myocardial nuclear factor erythroidderived 2 (Nrf-2), catalase, superoxide dismutase, glutathione peroxidase, myocardial interleukin 10 (IL-10), intercellular adhesion molecule 1 (ICAM-1), tissue inhibitor of metalloproteinase 1 (TIMP-1), or myocardial metalloproteinase-2 (MMP-2) or MMP-9 activity (Table 2). Caspase 3 expression increased and Bcl-2 expression decreased in the group of infarcted animals that received vitamin D supplementation (Fig. 1). In conclusion, contrary to our hypothesis, vitamin D supplementation intensified the cardiac remodeling process following coronary occlusion in rats, which is associated with impaired systolic and diastolic ventricular functions. In addition, this phenomenon was associated with increased apoptosis and impaired cardiac energy metabolism, without an effect on oxidative stress, VDUP-1, inflammation, or extracellular matrix. Conflict of interest The authors report no relationships that could be construed as a conflict of interest. References [1] Minicucci MF, Azevedo PS, Martinez PF, et al. Critical infarct size to induce ventricular remodeling, cardiac dysfunction and heart failure in rats. Int J Cardiol 2011;151: 242–3. [2] Ardisson LP, Minicucci MF, Azevedo PS, et al. Influence of AIN-93 diet on mortality and cardiac remodeling after myocardial infarction in rats. Int J Cardiol 2012;156: 265–9. [3] Assalin HB, Rafacho BP, Dos Santos PP, et al. Impact of the length of vitamin d deficiency on cardiac remodeling. Circ Heart Fail 2013;6:809–16.
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A.F. Gonçalves et al. / International Journal of Cardiology 176 (2014) 1225–1226
Table 1 Biochemical and echocardiographic data. Variables
SC (n = 25)
SD (n = 24)
IC (n = 18)
ID (n = 20)
p(I)
p(VD)
p(I × VD)
Calcium (mg/dL) Phosphorus (mg/dL) Phosphofructokinase (nmol/g) LVWT (mm/s) FAC E/E′ SA (mm2) DA (mm2)
8.53 6.44 94.4 36.3 74.4 19.2 12.3 47.5
8.95 7.04 104.6 37.1 73.3 17.8 12.3 45.9
8.9 6.38 129.8 33.8 52 20.4 31.5 62
8.6 5.82 92.8 28.9 39.6 25.3 53.4 84.9
0.849 0.107 0.291 b0.001 b0.001 0.001 b0.001 b0.001
0.669 0.958 0.213 0.159 0.011 0.18 b0.001 b0.001
0.172 0.146 0.041 0.049 0.031 0.016 b0.001 b0.001
± ± ± ± ± ± ± ±
0.21 0.35 10A 1.18 2.1b 0.87 2.4 A 2.5 A
± ± ± ± ± ± ± ±
0.22 0.37 12.2 1.2 2.21b,B 0.87 2.4B 2.5B
± ± ± ± ± ± ± ±
0.26 0.42 10a,A 1.78a 3.2a 1.95a 3.67ª,A 3.82ª,A
± ± ± ± ± ± ± ±
0.23 0.39 10a 1.39a 2.55a,B 0.97a 2.87ª,B 2.99ª,B
SC = sham animals that received standard diet; SD = sham animals that received diet with supplementation; IC = infarcted animals that received standard diet; and ID = infarcted animals that received diet with supplementation. LVWT: posterior wall shortening velocity; FAC: fractional area change; SA: systolic area; and DA: diastolic area. Data are expressed as the mean ± standard error of the mean. n = number of rats. p(I) = p value of the I effect; p(VD) = p value of the vitamin D effect; p(I × VD) = p value of their interaction. Different superscript letters in the same row denote significant differences. Comparisons for factor infarction: (a denotes IC ≠ ID; b denotes SC ≠ SD). Comparisons for factor Vitamin D: (A denotes IC ≠ SC; B denotes ID ≠ SD).
Table 2 Biochemical and molecular data. Variables
SC (n = 7)
SD (n = 7)
IC (n = 7)
ID (n = 7)
p(I)
p(VD)
p(I × VD)
IL-10 (pg/mg protein) ICAM-1 (pg/mg protein) TIMP-1 (pg/mg protein) MMP-2 activity MMP-9 activity Nrf-2 Catalase (μmol/g) Superoxide dismutase (nmol/mg)
84.6 137.7 4.12 1.21 0.9 0.72 75.1 9.55
60 101.6 3.7 1.67 1.03 0.82 82.5 8.15
82.6 139.2 4.18 0.9 1.26 0.66 57.8 15.4
67.8 ± 10.7 136 ± 17.8 4.14 ± 0.16 1.04 0.96 ± 0.11 0.48 ± 0.099 68.9 ± 6.76 11.2 ± 0.88
0.8 0.268 0.2 0.18 0.26 0.043 0.045 b0.001
0.09 0.228 0.23 0.395 0.492 0.620 0.213 0.008
0.66 0.31 0.329 0.647 0.093 0.146 0.798 0.149
± ± ± ± ± ± ± ±
10.7 15 0.16 0.37 0.12 0.101 6.76 0.88
± ± ± ± ± ± ± ±
10.7 15 0.16 0.31 0.12 0.082 8.2 1.08
± ± ± ± ± ± ± ±
12.6 17.8 0.19 0.34 0.13 0.089 6.76 0.88
SC = sham animals that received standard diet; SD = sham animals that received diet with supplementation; IC = infarcted animals that received standard diet; ID = infarcted animals that received diet with supplementation. Data are expressed as the mean ± standard error of mean. p(I) = p value of the I effect; p(VD) = p value of the vitamin D effect; p(IxVD) = p value of their interaction. Diferent letters denote significant differences. Comparisons for factor Infarction: (a denotes IC ≠ ID; b denotes SC ≠ SD). Comparisons for factor Vitamin D: (A denotes IC ≠ SC; B denotes ID ≠ SD).
Fig. 1. A: Caspase 3 expression. SC = sham animals that received standard diet; SD = sham animals that received diet with supplementation; IC = infarcted animals that received standard diet; and ID = infarcted animals that received diet with supplementation. B: Bcl-2 expression. SC = sham animals that received standard diet; SD = sham animals that received diet with supplementation; IC = infarcted animals that received standard diet; and ID = infarcted animals that received diet with supplementation.*p b 0.05.
Contents lists available at ScienceDirect
International Journal of Cardiology journal homepage: www.elsevier.com/locate/ijcard
Letter to the Editor
Vitamin D supplementation intensifies cardiac remodeling after experimental myocardial infarction☆ Andrea F. Gonçalves a, Priscila P. Santos a, Bruna P. Rafacho a, Diego F. Batista a, Paula S. Azevedo a, Marcos F. Minicucci a, Bertha F. Polegato a, Fernanda Chiuso-Minicucci b, Katashi Okoshi a, Ana Angélica Fernandes c, Sergio A. Paiva a, Leonardo A. Zornoff a,⁎ a b c
Internal Medicine Department, Botucatu Medical School, UNESP — Univ Estadual Paulista, Botucatu, Brazil Department of Microbiology and Immunology, Institute of Biosciences, UNESP — Univ Estadual Paulista, Botucatu, Brazil Chemistry and Biochemistry Department, Institute of Biosciences, UNESP — Univ Estadual Paulista, Botucatu, Brazil
a r t i c l e
i n f o
Article history: Received 1 July 2014 Accepted 27 July 2014 Available online 5 August 2014 Keywords: Remodeling Cardiac function Apoptosis Vitamin D
In recent years, vitamin D has emerged as an important nutrient for heart health, and its deficiency has been linked to cardiac dysfunction. Thus, the present study tested the hypothesis that vitamin D supplementation would attenuate cardiac remodeling induced by coronary occlusion. The protocol of this study was approved by the Ethics Committee on Animal Experiments of our institution. Wistar rats were allocated into four groups: 1) sham control (SC): animals that received standard chow; 2) sham vitamin D (SD): sham animals that received standard chow and cholecalciferol (3000 IU/kg chow); 3) infarction control (IC): infarcted animals that received standard chow; and 4) infarction vitamin D (ID): infarcted animals that received standard chow and cholecalciferol. The tests were performed as previously described [1–3]. Comparison between groups was performed by two-way ANOVA and the post-hoc Holm–Sidak test.
☆ Funding: This work was supported by FAPESP (12/14500-2) and CNPq (301945/ 2010-1). ⁎ Corresponding author at: Botucatu Medical School, Botucatu, SP 18618-970, Brazil. Tel.: +55 14 38222969. E-mail address: [email protected] (L.A. Zornoff).
http://dx.doi.org/10.1016/j.ijcard.2014.07.217 0167-5273/© 2014 Elsevier Ireland Ltd. All rights reserved.
Considering our results, after three months, the infarct size did not differ between the infarcted groups (IC = 46.7 ± 6.5%, ID = 45.8 ± 6.3% p = 0.593). Serum concentrations of 25[OH]D3 were higher in supplemented groups than non-supplemented groups (SC = 8.48 ± 1.27 ng/mL, SD = 14.94 ± 1.73, IC = 11.88 ± 1.28, ID = 13.47 ± 1.76; p(I) = 0.610, p(VD) = 0.034, p(I × VD) = 0.173). Vitamin D increased the left ventricular systolic and diastolic areas and decreased the fractional area change and posterior wall shortening velocity associated with increased E/E′ ratio. In addition, vitamin D decreased phosphofructokinase activity (Table 1). Conversely, vitamin D did not affect interstitial collagen volume fraction, vitamin D3-upregulated protein 1 (VDUP-1), myocardial nuclear factor erythroidderived 2 (Nrf-2), catalase, superoxide dismutase, glutathione peroxidase, myocardial interleukin 10 (IL-10), intercellular adhesion molecule 1 (ICAM-1), tissue inhibitor of metalloproteinase 1 (TIMP-1), or myocardial metalloproteinase-2 (MMP-2) or MMP-9 activity (Table 2). Caspase 3 expression increased and Bcl-2 expression decreased in the group of infarcted animals that received vitamin D supplementation (Fig. 1). In conclusion, contrary to our hypothesis, vitamin D supplementation intensified the cardiac remodeling process following coronary occlusion in rats, which is associated with impaired systolic and diastolic ventricular functions. In addition, this phenomenon was associated with increased apoptosis and impaired cardiac energy metabolism, without an effect on oxidative stress, VDUP-1, inflammation, or extracellular matrix. Conflict of interest The authors report no relationships that could be construed as a conflict of interest. References [1] Minicucci MF, Azevedo PS, Martinez PF, et al. Critical infarct size to induce ventricular remodeling, cardiac dysfunction and heart failure in rats. Int J Cardiol 2011;151: 242–3. [2] Ardisson LP, Minicucci MF, Azevedo PS, et al. Influence of AIN-93 diet on mortality and cardiac remodeling after myocardial infarction in rats. Int J Cardiol 2012;156: 265–9. [3] Assalin HB, Rafacho BP, Dos Santos PP, et al. Impact of the length of vitamin d deficiency on cardiac remodeling. Circ Heart Fail 2013;6:809–16.
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A.F. Gonçalves et al. / International Journal of Cardiology 176 (2014) 1225–1226
Table 1 Biochemical and echocardiographic data. Variables
SC (n = 25)
SD (n = 24)
IC (n = 18)
ID (n = 20)
p(I)
p(VD)
p(I × VD)
Calcium (mg/dL) Phosphorus (mg/dL) Phosphofructokinase (nmol/g) LVWT (mm/s) FAC E/E′ SA (mm2) DA (mm2)
8.53 6.44 94.4 36.3 74.4 19.2 12.3 47.5
8.95 7.04 104.6 37.1 73.3 17.8 12.3 45.9
8.9 6.38 129.8 33.8 52 20.4 31.5 62
8.6 5.82 92.8 28.9 39.6 25.3 53.4 84.9
0.849 0.107 0.291 b0.001 b0.001 0.001 b0.001 b0.001
0.669 0.958 0.213 0.159 0.011 0.18 b0.001 b0.001
0.172 0.146 0.041 0.049 0.031 0.016 b0.001 b0.001
± ± ± ± ± ± ± ±
0.21 0.35 10A 1.18 2.1b 0.87 2.4 A 2.5 A
± ± ± ± ± ± ± ±
0.22 0.37 12.2 1.2 2.21b,B 0.87 2.4B 2.5B
± ± ± ± ± ± ± ±
0.26 0.42 10a,A 1.78a 3.2a 1.95a 3.67ª,A 3.82ª,A
± ± ± ± ± ± ± ±
0.23 0.39 10a 1.39a 2.55a,B 0.97a 2.87ª,B 2.99ª,B
SC = sham animals that received standard diet; SD = sham animals that received diet with supplementation; IC = infarcted animals that received standard diet; and ID = infarcted animals that received diet with supplementation. LVWT: posterior wall shortening velocity; FAC: fractional area change; SA: systolic area; and DA: diastolic area. Data are expressed as the mean ± standard error of the mean. n = number of rats. p(I) = p value of the I effect; p(VD) = p value of the vitamin D effect; p(I × VD) = p value of their interaction. Different superscript letters in the same row denote significant differences. Comparisons for factor infarction: (a denotes IC ≠ ID; b denotes SC ≠ SD). Comparisons for factor Vitamin D: (A denotes IC ≠ SC; B denotes ID ≠ SD).
Table 2 Biochemical and molecular data. Variables
SC (n = 7)
SD (n = 7)
IC (n = 7)
ID (n = 7)
p(I)
p(VD)
p(I × VD)
IL-10 (pg/mg protein) ICAM-1 (pg/mg protein) TIMP-1 (pg/mg protein) MMP-2 activity MMP-9 activity Nrf-2 Catalase (μmol/g) Superoxide dismutase (nmol/mg)
84.6 137.7 4.12 1.21 0.9 0.72 75.1 9.55
60 101.6 3.7 1.67 1.03 0.82 82.5 8.15
82.6 139.2 4.18 0.9 1.26 0.66 57.8 15.4
67.8 ± 10.7 136 ± 17.8 4.14 ± 0.16 1.04 0.96 ± 0.11 0.48 ± 0.099 68.9 ± 6.76 11.2 ± 0.88
0.8 0.268 0.2 0.18 0.26 0.043 0.045 b0.001
0.09 0.228 0.23 0.395 0.492 0.620 0.213 0.008
0.66 0.31 0.329 0.647 0.093 0.146 0.798 0.149
± ± ± ± ± ± ± ±
10.7 15 0.16 0.37 0.12 0.101 6.76 0.88
± ± ± ± ± ± ± ±
10.7 15 0.16 0.31 0.12 0.082 8.2 1.08
± ± ± ± ± ± ± ±
12.6 17.8 0.19 0.34 0.13 0.089 6.76 0.88
SC = sham animals that received standard diet; SD = sham animals that received diet with supplementation; IC = infarcted animals that received standard diet; ID = infarcted animals that received diet with supplementation. Data are expressed as the mean ± standard error of mean. p(I) = p value of the I effect; p(VD) = p value of the vitamin D effect; p(IxVD) = p value of their interaction. Diferent letters denote significant differences. Comparisons for factor Infarction: (a denotes IC ≠ ID; b denotes SC ≠ SD). Comparisons for factor Vitamin D: (A denotes IC ≠ SC; B denotes ID ≠ SD).
Fig. 1. A: Caspase 3 expression. SC = sham animals that received standard diet; SD = sham animals that received diet with supplementation; IC = infarcted animals that received standard diet; and ID = infarcted animals that received diet with supplementation. B: Bcl-2 expression. SC = sham animals that received standard diet; SD = sham animals that received diet with supplementation; IC = infarcted animals that received standard diet; and ID = infarcted animals that received diet with supplementation.*p b 0.05.
