L-methionine protects against nephrotoxicity induced by methotrexate through modulation of redox status and inflammation

Figures & data

Figure 1. Changes in body weight (A) and kidney weight (B) of rats treated with MTX with or without L-Met. Values are presented as mean ± SEM (n = 6). Significant differences (P < 0.05) between groups were indicated by different letters above the error bars. Similar letters above the error bars indicate no significant differences. Abbreviations: L-Met, L-methionine; MTX, methotrexate.

Table 1. Changes in serum biochemical parameters related to kidney function and injury under the influence of MTX with and without L-Met.

Group	KIM-1 (pg/ml)	BUN (mg/dl)	Cr (mg/dl)	UA (mg/dl)	Protein (mg/dl)	Alb (mg/dl)
Control	108.300^a ± 10.99	27.583^a ± 1.20	0.480^a ± 0.05	3.483^a ± 0.34	7.600^a ± 0.68	4.550^a ± 0.52
L-Met400	104.267^a ± 6.19	27.100^a ± 2.06	0.447^a ± 0.04	3.067^a ± 0.35	7.417^a,b ± 0.77	4.750^a± 0.43
MTX	326.783^b ± 16.88	69.350^b ± 3.40	2.117^b ± 0.36	11.250^b ± 0.71	5.267^b ± 0.53	2.450^b ± 0.44
MTX-Met300	244.725^c ± 17.48	46.200^c ± 1.86	1.650^b,c ± 0.22	7.750^c ± 0.64	6.233^a,b ± 0.53	2.850^b,c ± 0.25
MTX-Met400	160.867^d ± 10.12	41.717^c ± 2.67	1.152^c ± 0.18	5.450^a,c ± 0.37	6.617^a,b ± 0.51	3.650^a,b ± 0.41
One Way ANOVA F (p)	52.04 (<0.001)	33.258 (<0.001)	24.595 (<0.001)	37.93 (<0.001)	3.083 (<0.05)	7.723 (<0.001)

Notes: Values are presented as mean ± SEM (n = 6). The mean values within the same column that have different superscripts (a–d) are significantly different (P < 0.05). Abbreviations: Alb, albumin; BUN, blood urea nitrogen; Cr, creatinine; KIM-1, kidney injury molecule -1; L-Met, L-methionine; MTX, methotrexate; UA, uric acid.

Figure 2. Oxidative markers in rats treated with MTX with and without L-Met. Renal ROS (A); MDA (B); NO (C) and PC (D) levels were evaluated for different groups. Values are presented as mean ± SEM (n = 6). Significant differences (P < 0.05) between groups were indicated by different letters above the error bars. Similar letters above the error bars indicate no significant differences. Abbreviations: L-Met, L-methionine; MTX, methotrexate; MDA, malondialdehyde; NO, nitric oxide; PC, protein carbonyl; ROS, reactive oxygen species.

Figure 3. Antioxidant markers in rats treated with MTX with and without L-Met. Different groups were evaluated for their level of renal GSH (A); GSSG (B), GSH/GSSG ratio (C), the activities of GPx (D); CAT (E); and SOD (F). Values are presented as mean ± SEM (n = 6). Significant differences (P < 0.05) between groups were indicated by different letters above the error bars. Similar letters above the error bars indicate no significant differences. Abbreviations: CAT, catalase; GPx, glutathione peroxidase; GSH, reduced glutathione; GSSG, oxidized glutathione; L-Met, L-methionine; MTX, methotrexate; SOD, superoxide dismutase.

Figure 4. Inhibitory effect of L-Met on MTX-induced inflammatory cytokines in rats. Different groups were assessed for their levels of IL-1β (A); IL-6 (B); and TNF-α (C). Values are presented as mean ± SEM (n = 6). Significant differences (P < 0.05) between groups were indicated by different letters above the error bars. Similar letters above the error bars indicate no significant differences. Abbreviations: IL, interleukin; L-Met, L-methionine; MTX, methotrexate; TNF, tumor necrosis factor.

Table 2. Changes in rats’ blood cell counts under the influence of MTX with and without L-Met.

Group	RBCs	WBCs	Platelets
Control	7.350^a ± 0.59	9.367 ± 0.75	480.550^a ± 30.55
L-Met400	7.867^a,b ± 0.50	10.017 ± 0.69	495.483^a,b ± 33.42
MTX	6.167^a,c ± 0.28	8.133 ± 0.44	305.517^c ± 33.42
MTX-Met300	6.200^a,b,c ± 0.29	8.533 ± 0.70	338.750^a,c ± 30.39
MTX-Met400	7.050^a,b,c ± 0.39	9.067 ± 0.76	408.767^a,b,c ± 32.15
One Way ANOVA F (p)	3.141 (<0.05)	1.502 (0.233)	6.462 (<0.001)

Notes: Values are presented as mean ± SEM (n = 6). The mean values within the same column that have different superscripts (a, b, c) are significantly different (P < 0.05). Abbreviations: L-Met, L-methionine; MTX, methotrexate; RBCs, red blood cells; WBCs, white blood cells.

Table 3. Changes in the differential leucocytic count of rats under the influence of MTX with and without L-Met.

Group	Lymphocytes %	Monocytes %	Neutrophils %	Basophils %	Eosinophils %
Control	46.133 ± 1.51	5.267 ± 0.45	40.950^a ± 1.13	0.652^a ± 0.09	2.120 ± 0.13
L-Met400	45.483 ± 2.54	5.700 ± 0.50	42.000^a ± 1.30	0.692^a,b ± 0.05	2.250 ± 0.20
MTX	38.050 ± 1.51	4.100 ± 0.49	53.067^b ± 2.75	0.400^a,c ± 0.06	1.617 ± 0.15
MTX-Met300	39.717 ± 1.25	4.517 ± 0.47	51.583^b ± 2.79	0.475^a,b,c ± 0.06	1.733 ± 0.19
MTX-Met400	43.317 ± 2.48	5.033 ± 0.52	45.183^a,b ± 2.51	0.600^a,b,c ± 0.04	1.917 ± 0.20
One Way ANOVA F (p)	2.897 (<0.05)	1.723 (0.178)	5.539 (<0.01)	4.135 (<0.05)	2.280 (<0.01)

Notes: Values are presented as mean ± SEM (n = 6). The mean values within the same column that have different superscripts (a–c) are significantly different (P < 0.05). Abbreviations: L-Met, L-methionine; MTX, methotrexate.

Data availability statement

The data used to support the findings of this study are available from the corresponding author upon request.