Folic acid ameliorates celecoxib cardiotoxicity in a doxorubicin heart failure rat model

Figures & data

Table

Group-I	Control: 0.5 mL/kg/d, 1% carboxymethylcellulose (CMC), p.o. for 28 days +0.5 mL/kg normal saline, intraperitoneal (i.p.), over a period of 2 weeks (3^rd and 4^th week)
Group-II	DOX: 2.5 mg/kg in six equal i.p. injections over a period of 2 weeks (total cumulative dose of 15 mg/kg)
Group-III	CEL: 100 mg/kg/d, p.o. suspended in 1% CMC for 28 days
Group-IV	CEL + DOX: 100 mg/kg/d, p.o for 28 days +2.5 mg/kg, six equal i.p. injections over a period of 2 weeks (3^rd and 4^th weeks)
Group-V	FA: 10 mg/kg/d, p.o. suspended in 1% CMC for 28 days
Group-VI	CEL + FA: 100 mg/kg/d, p.o. for 28 days +10 mg/kg/d, p.o. for 28 days
Group-VII	CEL + FA + DOX: 100 mg/kg/d, p.o. for 28 days +10 mg/kg/d, p.o. for 28 days +2.5 mg/kg in six equal i.p injections over a period of 2 weeks (3^rd and 4^th weeks)

Table 1. Effect of CEL and its combination with FA on the LDH and CK-MB levels.

Group	Treatment	LDH (IU/L)	CK-MB (IU/L)
I	CONTROL	83.67 ± 4.32	67.17 ± 8.07
II	DOX	213.34 ± 12.06***	181.20 ± 8.02***
III	CEL	105.25 ± 13.34†††	91.02 ± 9.91†††
IV	CEL + DOX	249.60 ± 11.65###	202.67 ± 17.70###
V	FA	87.63 ± 4.68	75.98 ± 4.42
VI	CEL + FA	91.18 ± 2.54*	73.79 ± 6.18*
VII	CEL + FA + DOX	235.33 ± 9.83†	186.60 ± 5.99†

Values are expressed as the mean ± SD and analyzed by one-way ANOVA followed by the Tukey–Kramer multiple comparison test.

* p < 0.05 (VI vs. III).

† p < 0.05 (VII vs. IV).

*** p < 0.001 (II vs. I).

††† p < 0.001(III vs. I)

### p < 0.001(IV vs. II), DOX: Doxorubicin; CEL: Celecoxib; FA: Folic acid; DOX: Doxorubicin.

Table 2. Effect of CEL and its combination with FA on the Tn-T and TNF-α levels.

Group	Treatment	Tn-T (pg/ml)	TNF-α (pg/ml)
I	CONTROL	10.89 ± 0.94	0.044 ± 0.01
II	DOX	196.09 ± 26.58***	1740.91 ± 148.68***
III	CEL	138.27 ± 11.92†††	1109.67 ± 91.71†††
IV	CEL + DOX	389.77 ± 70.11###	2743.78 ± 188.27###
V	FA	9.57 ± 0.81	0.042 ± 0.01
VI	CEL + FA	68.17 ± 12.08**	683.86 ± 115.47§§§
VII	CEL + FA + DOX	204.55 ± 18.71‡‡‡	1892.41 ± 244.61‡‡‡

Values are expressed as the mean ± SD and analyzed by one-way ANOVA followed by the Tukey–Kramer multiple comparison test.

** p < 0.01 (VI vs. III).

*** p < 0.001 (II vs. I).

††† p < 0.001(III vs. I).

### p < 0.001 (IV vs. II).

§§§ p < 0.001 (VI vs. III), and

‡‡‡ p < 0.001 (VII vs. IV).

CEL: Celecoxib; FA: Folic acid; DOX: Doxorubicin.

Table 3. Effect of CEL and its combination with FA on the cardiac TBARS and GSH levels.

Group	Treatment	TBARS (nmMDA/mg protein)	GSH (μg/mg of protein)
I	CONTROL	2.24 ± 0.55	22.36 ± 2.39
II	DOX	8.31 ± 2.61***	10.63 ± 1.59***
III	CEL	4.89 ± 0.83**	18.74 ± 2.43*
IV	CEL + DOX	12.32 ± 2.28†††	8.22 ± 1.76
V	FA	1.95 ± 0.57	23.25 ± 2.52
VI	CEL + FA	2.51 ± 0.63†	20.54 ± 2.44
VII	CEL + FA + DOX	9.54 ± 1.25††	17.09 ± 1.55###

Values are expressed as the mean ± SD and analyzed by one-way ANOVA followed by the Tukey–Kramer multiple comparison test.

* p < 0.05(III vs. I).

† p < 0.05 (VI vs. III).

** p < 0.01(III vs. I).

†† p < 0.01 (VII vs. IV).

*** p < 0.001 (II vs. I).

††† p < 0.001 (IV vs. II), and

### p < 0.001(VII vs. IV).

CEL: Celecoxib; FA: Folic acid; DOX: Doxorubicin.

Figure 1. Effect of CEL and its combination with FA on SBP. *p < 0.05 CEL + FA (3rd and 4th weeks) vs. Baseline, ***p < 0.001 CEL (3rd and 4th weeks) vs. Baseline; DOX (3rd week) vs. Baseline; and CEL + DOX (3rd week) vs. Baseline. †p < 0.05 CEL + FA (3rd week) vs. CEL, CEL + DOX (4th week) vs. DOX; ††p < 0.01 CEL (2nd week) vs. Control, †††p < 0.001 CEL (3rd and 4th weeks) vs. Control; CEL + FA (4th week) vs. CEL; DOX (3rd week) vs. Control; CEL + DOX (3rd week) vs. DOX; and CEL + FA + DOX (3rd and 4th weeks) vs. CEL + DOX. DOX: Doxorubicin; CEL: Celecoxib; FA: Folic acid; SBP: systolic blood pressure.

Figure 2. Effect of CEL and its combination with FA on heart rate. **p < 0.01 CEL (2nd week) vs. Baseline, ***p < 0.001 CEL (3rd and 4th weeks) vs. Baseline; CEL + FA (4th week) vs. Baseline; DOX (3rd week) vs. baseline; DOX (3rd vs. 4th week); CEL + DOX (3rd week) vs. Baseline; and CEL + DOX (3rd vs. 4th week). ††p < 0.01 CEL + DOX (3rd week) vs. DOX, †††p < 0.001 CEL (4th week) vs. Control; CEL + FA (4th week) vs. CEL; DOX (3rd and 4th weeks) vs. Control; and CEL + FA + DOX (3rd and 4th weeks) vs. CEL + DOX. DOX: Doxorubicin; CEL: Celecoxib; FA: Folic acid; BPM: Beats per min.

Figure 3. Transmission electron microscopy of cardiomyocytes. (a) Normal control group showing well-defined Z-bands (Z), myofibrils (MF) arranged in regular rows and mitochondria (MT). (b) DOX-treated group showing disarray of the myofibrillar arrangement, swollen mitochondria, and a pyknotic nucleus (N) with chromatin margination. (c) CEL-treated group showing loss of myofibrils, mild mitochondrial swelling, and condensed chromatin in the nucleus. (d) CEL + DOX treated group showing severe disruption revealing loss of myofibrils and mitochondrial swelling. (e) FA-treated group showing well-defined Z-Bands, myofibrils, and mitochondria. (f) CEL + FA-treated group showing normal myofibrils, well-defined Z-Bands, and mitochondria. (g) DOX + CEL + FA-treated group showing maintenance of myofibrillar arrangement with mild mitochondrial swelling and margination of chromatin in the nucleus (N). DOX: Doxorubicin; CEL: Celecoxib; FA: Folic acid.