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Protosappanin A induces immunosuppression of rats heart transplantation targeting T cells in grafts via NF-κB pathway

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Abstract

Protosappanin A as one major and effective ingredient from Caesalpinia sappan L. exhibited antirejection activity obviously in heart-transplanted rat. The present study was designed to screen out the potential target genes of protosappanin A with microarray technology and reveal some molecular mechanism of immunosuppressive effect. Rats performed with ectopic peritoneal heart transplantation were randomized into three groups receiving different treatments for 7 days: protosappanin A group (25 mg kg−1), cyclosporine A group (10 mg kg−1), and control group. The differentially expressed genes responding to protosappanin A were analyzed with microarrays. Among common differentially expressed genes, the ones of interest were selected for further evaluation by real-time quantitative reverse transcriptase polymerase chain reaction (qRT-PCR), Western blot, immunochemistry, immunofluorescence, and ELISA. Among the 146 common differentially expressed genes, NF-κB and related genes like IκBa, IFN-r, and IP10 were selected for verification. The results of qRT-PCR, Western blot, immunochemistry, and ELISA showed that protosappanin A significantly reduced the expression of NF-κB, IFN-r, and IP10 (p < 0.05) and increased IκBa expression (p < 0.05) in graft. Moreover, the immunochemistry staining of NF-κB and IκBa was mainly observed in infiltrating mononuclear cells. Strikingly, immunofluorescent staining localized NF-κB to the TCR-positive T cells in graft. Furthermore, protosappanin A exhibited inhibitory effect on T cell proliferation in recipients after 7-day treatment. In conclusion, protosappanin A might act on T cells through inhibiting NF-κB activation and downstream gene expressions of IFN-r and IP10, meanwhile reducing T cell proliferation responding to alloantigen, so as to induce immunosuppressive effect. The results encourage a potential therapeutic evaluation of protosappanin A for clinical organ transplantation or other T cell-mediated immune disorders. Additionally, our study also verified the feasibility of microarray utilization in Chinese herb research to explore molecular mechanism and promote development of scientific theories.

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Abbreviations

PrA:

Protosappanin A

CsA:

Cyclosporin A

NF-κB:

Nuclear factor kappa B

IκBa:

Inhibitor of nuclear factor kappa B alpha

IFN-r:

Interferon-gamma

IP10:

Interferon-gamma-inducible protein 10

qRT-PCR:

Real-time quantitative reverse transcriptase polymerase chain reaction

FITC:

Fluorescein isothiocyannate

TRITC:

Tetramethylrhodamine isothiocyannate

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Acknowledgments

Our projects are supported by the National Nature Science Foundation of China (C03020504), the Heilongjiang Province Nature Science Foundation (D200660), the Heilongjiang Province Nature Foundation of Great Subject (ZJY03-7), and Heilongjiang Province Scientific Technique Project (GC02C122).

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Authors and Affiliations

  1. Department of Cardiology, The Second Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang Province, China

    Jian Wu, Maomao Zhang, Haibo Jia, Xingtao Huang, Qi Zhang, Jingbo Hou & Yu Bo

  2. The Cardiovascular Department, The Second Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang Province, 150081, China

    Jingbo Hou

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  1. Jian Wu
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  2. Maomao Zhang
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  3. Haibo Jia
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  4. Xingtao Huang
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  5. Qi Zhang
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  6. Jingbo Hou
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  7. Yu Bo
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Corresponding author

Correspondence to Jingbo Hou.

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J. Wu and M. Zhang contributed equally in the study.

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Wu, J., Zhang, M., Jia, H. et al. Protosappanin A induces immunosuppression of rats heart transplantation targeting T cells in grafts via NF-κB pathway. Naunyn-Schmied Arch Pharmacol 381, 83–92 (2010). https://doi.org/10.1007/s00210-009-0461-5

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  • DOI: https://doi.org/10.1007/s00210-009-0461-5

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