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The antioxidant effect exerted by TGF-1β-stimulated hyaluronan production reduced NF-kB activation and apoptosis in human fibroblasts exposed to FeSo4 plus ascorbate

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Abstract

Previous studies suggest that Transforming growth factor-1beta (TGF-1β) administration in human fibroblasts exposed to oxidative stress is able to modulate hyaluronan synthases (HASs). HAS modulation in turn increases high molecular weight (Hyaluronan) HA concentration. Nuclear factor kB (NF-kB) is a response transcription factor involved in inflammation and acts by enabling the expression of certain detrimental molecules. Caspases are specific proteases responsible for regulating and programming cell death. HA at medium molecular weight together with chondroitin-4-sulphate proved to be effective on NF-kB and caspases. We investigated whether the protective effect afforded by the high molecular weight HA produced by TGF-1β treatment has any effect on NF-kB and apoptosis activation in fibroblast cultures exposed to oxidative stress. Generation of free radicals gives rise to cell death, increases lipid peroxidation, activates NF-kB, reduces its cytoplasmic inhibitor IkBα, augments caspase-3 and caspase-7 gene expression and their relative protein activity, and depletes catalase (CAT) and glutathione peroxidase (GPx). Treatment of fibroblasts with TGF-1β 12 h before inducing oxidative stress greatly increased HA levels, ameliorated cell survival, inhibited lipid peroxidation, blunted NF-kB translocation, normalized IkBα protein, reduced caspase gene expression and protein levels, and restored the endogenous antioxidants CAT and GPx. Since it was previously reported that antioxidants can work as inhibitors of NF-kB and apoptosis induction we can hypothesize that endogenous HA, by inhibiting lipid peroxidation, may block a step whereby free radical activity converges in the signal transduction pathway leading to NF-kB and caspase activation.

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Abbreviations

BHT:

Butylated hydroxytoluene

CAT:

Catalase

DMEM:

Dulbecco’s modified Eagle’s medium

ECM:

Extracellular matrix

EDTA:

Ethylenediaminetetraacetic acid

GAGs:

Glycosaminoglycans

FBS:

Foetal bovine serum

HA:

Hyaluronic acid

HAE:

Hydroxyalkenals

HASs:

Hyaluronan synthases

IFN-γ:

Interferon gamma

IFN-β:

Interferon beta

NADH:

Reduced nicotinamide adenine dinucleotide

NADPH:

Reduced nicotinamide adenine dinucleotide phosphate

PBS:

Phosphate buffered saline

PMNs:

Polymorphonuclear leukocytes

SDS:

Sodium dodecylsulphate

ROS:

Reactive oxygen species

SD:

Standard deviation

TNF-α:

Tumour necrosis factor alpha

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Acknowledgements

This study was supported in part by a grant ex 40% (COFIN 2004) from the MIUR, Italy and in part by a grant PRA (Research Athenaeum Project 2003) from the University of Messina, Italy.

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Authors and Affiliations

  1. Department of Biochemical, Physiological and Nutritional Sciences, School of Medicine, University of Messina, Policlinico Universitario, Torre Biologica, 5° piano, Via C. Valeria, 98125, Messina, Italy

    Giuseppe M. Campo, Angela Avenoso, Salvatore Campo, Angela D’Ascola, Paola Traina, Dario Samà & Alberto Calatroni

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  1. Giuseppe M. Campo
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  2. Angela Avenoso
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  3. Salvatore Campo
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Correspondence to Giuseppe M. Campo.

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Campo, G.M., Avenoso, A., Campo, S. et al. The antioxidant effect exerted by TGF-1β-stimulated hyaluronan production reduced NF-kB activation and apoptosis in human fibroblasts exposed to FeSo4 plus ascorbate. Mol Cell Biochem 311, 167–177 (2008). https://doi.org/10.1007/s11010-008-9707-7

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