J Vet Sci.  2008 Sep;9(3):257-266. 10.4142/jvs.2008.9.3.257.

Plasma haptoglobin and immunoglobulins as diagnostic indicators of deoxynivalenol intoxication

• Kim EJ
• Jeong SH
• Cho JH
• Ku HO
• Pyo HM
• Kang HG
• Choi KH

• KMID: 1718578
• DOI: http://doi.org/10.4142/jvs.2008.9.3.257

Abstract

This study aimed to discover potential biomarkers for dioxynivalenol (DON) intoxication. B6C3F1 male mice were rally exposed to 0.83, 2.5 and 7.5 mg/kg body weight (bw) DON for 8 days and the differential protein expressions in their blood plasma were determined by SELDI - Time-of-Flight/Mass Spectrometry (TOF/MS) and the immunoglobulins (Igs) G, A, M and E in the serum were investigated. 11.7 kDa protein was significantly highly expressed according to DON administration and this protein was purified by employing a methyl ceramic HyperD F column with using optimization buffer for adsorption and desorption. The purified protein was identified as a haptoglobin precursor by peptide mapping with using LC/Q-TOF/MS and MALDI-TOF/MS and this was confirmed by western blotting and ELISA. IgG and IgM in serum were decreased in a dose-dependent manner and IgA was decreased at 7.5 mg/kg bw DON administration, but the IgE level was not changed. To compare the expressions of haptoglobin and the Igs patterns between aflatoxin B1 (AFB1), zearalenone (ZEA) and DON intoxications, rats were orally administered with AFB1 1.0, ZEA 240 and DON 7.5 mg/kg bw for 8 days. Haptoglobin was increased only at DON 7.5 mg/kg bw, while it was slightly decreased at ZEA 240 mg/kg bw and it was not detected at all at AFB1 1.0 mg/kg bw. IgG and IgA were decreased by DON, but IgG, IgA, IgM and IgE were all increased by AFB1. No changes were observed by ZEA administration. These results show that plasma haptoglobin could be a diagnostic biomarker for DON intoxication when this is combined with examining the serum Igs.

Keyword

biomarker; deoxynivalenol; haptoglobin; immunoglobulins; SELDI-TOF/MS

MeSH Terms

Aflatoxin B1/toxicity
Animals
Blood Proteins/drug effects/*metabolism
Enzyme-Linked Immunosorbent Assay
Haptoglobins/drug effects/*metabolism
Immunoglobulins/*blood/drug effects
Male
Mass Spectrometry
Mice
Mice, Inbred Strains
Rats
Rats, Wistar
Trichothecenes/*toxicity
Zearalenone/toxicity

Figure

• 

  Fig. 1 Protein profiles of plasma on the H50, IMAC30 and CM-low ProteinChip array surfaces. The mice were orally administered with vehicle control (D.W.), DON 0.83, 2.5 or 7.5 mg/kg bw for 8 days, respectively. All the mass peaks were normalized by the total ion current (TIC) normalization function. **p < 0.01.

• 

  Fig. 2 Column optimization of the binding and elution conditions for the 11.7 kDa hydrophobic protein. The protein peaks were those of the plasma on the gold chip before loading onto the column (A) and those of the elutes after sample adsorption onto the activated column (B) and after the first and second desorption (C and D).

• 

  Fig. 3 SDS-PAGE analysis of the purified plasma 11.7 kDa protein. The flow-through fraction from the Methyl Ceramic HyperD F spin column was run on a SDS-PAGE gel and the proteins were stained with Coomassie blue R-250. Lane A: molecular marker, Lane B: Control, Lane C: DON 7.5 mg/kg bw.

• 

  Fig. 4 The 11.7 kDa protein (haptoglobin precursor) was reconfirmed by immunoblotting with using a polyclonal antibody of chicken haptoglobin. Lane A: molecular marker, Lane B: Control, Lane C: DON 7.5 mg/kg bw.

• 

  Fig. 5 Changes of haptoglobin in the plasma of the B6C3F1 male mice that were orally exposed to DON (0, 0.83, 2.5 or 7.5 mg/kg bw, respectively) for 8 days. The data is presented as mean ± SD (N = 6). **p < 0.01.

• 

  Fig. 6 Changes of haptoglobin in the plasma of rats orally exposed to DON 7.5 mg/kg, AFB1 1.0 mg/kg and ZEA 240 mg/kg for 8 days. The data is presented as mean ± SD (N = 6). *p < 0.05 . **p < 0.01. ND: not detected.

• 

  Fig. 7 Changes of the immunoglobulins content in serum by the administration of DON in B6C3F1 male mice. The values are the mean ± SD of 5 replicas. **p < 0.01.

• 

  Fig. 8 Changes of the immunoglobulin content in the serum of Wistar male rats by the administration of DON, AFB1 and ZEA. The rats were administered with DON (7.5 mg/kg), AFB1 (1.0 mg/kg) or ZEA (240 mg/kg) by gavage once per day for 8 days. The values are the mean ± SD of 5 replicas. *p < 0.05, **p < 0.01.

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