J Vet Sci.  2014 Mar;15(1):51-60. 10.4142/jvs.2014.15.1.51.

Protective effects of silymarin on fumonisin B1-induced hepatotoxicity in mice

• Sozmen M
• Devrim AK
• Tunca R
• Bayezit M
• Dag S
• Essiz D

• KMID: 1737610
• DOI: http://doi.org/10.4142/jvs.2014.15.1.51

Abstract

The present study was conducted to investigate the effect of silymarin on experimental liver toxication induced by Fumonisin B1 (FB1) in BALB/c mice. The mice were divided into six groups (n = 15). Group 1 served as the control. Group 2 was the silymarin control (100 mg/kg by gavage). Groups 3 and 4 were treated with FB1 (Group 3, 1.5 mg/kg FB1, intraperitoneally; and Group 4, 4.5 mg/kg FB1). Group 5 received FB1 (1.5 mg/kg) and silymarin (100 mg/kg), and Group 6 was given a higher dose of FB1 (4.5 mg/kg FB1) with silymarin (100 mg/kg). Silymarin treatment significantly decreased (p < 0.0001) the apoptotic rate. FB1 administration significantly increased (p < 0.0001) proliferating cell nuclear antigen and Ki-67 expression. Furthermore, FB1 elevated the levels of caspase-8 and tumor necrosis factor-alpha mediators while silymarin significantly reduced (p < 0.0001) the expression of these factors. Vascular endothelial growth factor (VEGF) and fibroblast growth factor-2 (FGF-2) expressions were significantly elevated in Group 4 (p < 0.0001). Silymarin administration alleviated increased VEGF and FGF-2 expression levels (p < 0.0001). In conclusion, silymarin ameliorated toxic liver damage caused by FB1 in BALB/c mice.

Keyword

caspase-8; fumonisin B1; fibroblast growth factor-2; galectin-3; silymarin

MeSH Terms

Animals
Antioxidants/*pharmacology
Apoptosis/drug effects
Cell Proliferation/drug effects
Female
Fibroblast Growth Factor 2/genetics/metabolism
Fumonisins/*toxicity
Gene Expression Regulation/drug effects
Hepatocytes/*drug effects
Ki-67 Antigen/metabolism
Liver/drug effects
Mice
Mice, Inbred BALB C
Mycotoxins/*toxicity
Neovascularization, Physiologic/drug effects
Proliferating Cell Nuclear Antigen/metabolism
Silymarin/*pharmacology
Tumor Necrosis Factor-alpha/metabolism
Vascular Endothelial Growth Factor A/genetics/metabolism
Antioxidants
Fibroblast Growth Factor 2
Fumonisins
Ki-67 Antigen
Mycotoxins
Proliferating Cell Nuclear Antigen
Silymarin
Tumor Necrosis Factor-alpha
Vascular Endothelial Growth Factor A

Figure

• 

  Fig. 1 H&E, TUNEL, and caspase-8-specific immunohistochemistry in hepatocytes from the experimental and control groups. (A) Megalocytosis in mouse liver (arrow; Group 1A). (B) Mitosis in mouse hepatocytes (arrow; Group 3A). (C) An apoptotic hepatocyte with a pycnotic nucleus (arrow; Group 5A). (D) Eosinophilic, intranuclear inclusion body in a mouse hepatocyte (arrow; Group 6A). (E) Cytoplasm and nucleus of two non-pyknotic hepatocytes exhibiting a TUNEL-positive reaction (arrows; Group 3A). (F) Cytoplasmic reaction in mouse liver. The non-pyknotic nucleus is negative (arrow; Group 6A). (G) Free apoptotic bodies in the liver parenchyma (arrows; Group 5A). (H) Hepatocytes exhibiting cytoplasmic staining for caspase-8 (Group 3B). H&E staining (A~D), TUNEL assay results (E~G), Harris hematoxylin counterstaining (E), methylene blue counterstaining (F), and Mayer's hematoxylin counterstain (G). Scale bars = 6 µm (B inset and D inset), 8.5 µm (C inset), 13 µm (A), 17 µm (C), 22 µm (B, D~F, and H), and 44 µm (G).

• 

  Fig. 2 Caspase-8, Ki-67, and Gal-3 immunohistochemistry in hepatocytes from the experimental and control groups. (A) Caspase-8 expression was observed in some periacinar hepatocytes (arrows; Group 6A). (B) In this panel, only the cytoplasm (long arrow) or both cytoplasm and nucleus (short arrow) are positive for Ki-67 (Group 5C). (C) Only the nucleus was positive for Ki-67 (short arrows). Periacinar hepatocyte cytoplasms were distinctively positive (long arrow) for Ki-67 expression (Group 3A). (D) Diffuse Ki-67-specific staining in mouse liver (Group 4B). (E) Hepatocytes were negative for the Ki-67 on the 21st day (Group 6C). (F) Weak cytoplasmic staining specific for Gal-3 in mouse liver (arrows; Group 3A). (G) Kupffer cells were intensely stained with the Gal-3 antibody (arrows). The hepatocytes were negative (Group 3A). (H) A limited number of hepatocytes showed weak cytoplasmic staining for the Gal-3 antibody (arrows; Group 5A). Scale bars = 15 µm (E), 22 µm (A~C and F~H), and 44 µm (D).

• 

  Fig. 3 PCNA, TNFα, and FGF-2 immunohistochemistry in hepatocytes from the experimental and control groups. (A) Only the nuclei are positive for PCNA (white arrows). Only cytoplasmic PCNA staining was observed in some hepatocytes while the nucleus was negative (long black arrow). Both cytoplasmic and nuclear staining with the PCNA antibody (short black arrow; Group 6B) is shown. (B) Cytoplasmic PCNA-specific staining in periacinar hepatocytes (Group 4B). (C) Intense cytoplasmic PCNA staining of the periacinar hepatocytes on the 14th day (Group 4A). (D) Decreased PCNA expression in the hepatocytes (arrows; Group 5A). (E) Diffuse cytoplasmic staining specific for TNFα; the nucleus is negative (Group 3C). (F) Cytoplasmic TNFα-specific staining of some periacinar hepatocytes (arrow; Group 5C). (G) Hepatocytes surrounding the central vein were positive for FGF-2 (arrow; Group 1A). (H) Cytoplasmic FGF-2 expression (Group 4A). Scale bars = 18 µm (G), 22 µm (B ~ F and H), and 44 µm (A).

• 

  Fig. 4 VEGF expression in hepatocytes from the experimental and control groups. (A) Cytoplasmic VEGF immunolabeling (Group 4B). (B) VEGF staining shown in panel B was significantly decreased compared that observed in panel A (Group 6C). Scale bars = 44 µm.

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