Arqoub, H.A.; (2007) Modulation of haem oxygenase-1 by phytochemicals: a novel stratagem in the mitigation of cardiovascular tissue dysfunction. Doctoral thesis , University of London.

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Abstract

Background: The haem oxygenase-1 (HO-1) pathway plays a key role in the preservation of tissue integrity against damaging insults such as oxidative stress and inflammatory injuries. Thus, HO-1 is regarded as a potential therapeutic target in conditions which are mediated by oxidative stress and inflammation, e.g. ischaemic heart disease, septic shock, and ischaemia reperfusion injury.;Aims: The objective of the study was to investigate if the up-regulation of the HO-1 expression by two plant-derived polyphenolic compounds, i.e. curcumin and 2'-hydroxychalcone (2-HC) offers protection to against stress-induced cellular damage in cardiovascular tissue. And to elucidate the molecular and cellular signalling mechanisms involved.;Methods: Parallel experiments were conducted using 1) Lipopolysaccharide (LPS)-induced model of inflammation in RAW 264.7 murine macrophages, the inflammatory response was assessed using nitrite assay, Western blot for the determination of the inducible nitric oxide synthase (iNOS), HO-1 protein expression and the activation of the transcriptional factor NF-KB. The role of HO-1 in this process was elucidated using small interfering RNA (siRNA). The involvement of the mitogen-activated protein kinase (MAPK), the phosphoinositide 3-kinase pathway (PI3K) pathway and the nuclear transcription factor (Nrf2) was also studied. 2) We also utilised Carbon Monoxide-Releasing Molecules (CO-RMs) as a method to study the effects of CO, one of the products of HO-1 enzymatic activity, in vitro model of inflammation. 3) In vitro model of H202-induced oxidative stress, using human cardiac myoblasts. 4) In vitro model of ischaemia-reperfusion injury and cold preservation in cardiac myoblasts. HO-1 mRNA expression during hypothermia was determined using RT-PCR.;Results: The major findings indicated that up-regulation of HO-1 gene and protein expression resulted in the protection of cells against oxidative and inflammatory injuries. We also found that 2-HC is a potent inducer of HO-1 protein expression in bovine aortic endothelial cells and in macrophages, in mechanisms which involve the activation of PI3K pathway. Treatment of macrophages with 2-HC resulted in the reduction of pro-inflammatory cytokines (TNF-a), and down regulation of iNOS expression but did not inhibit the activation of NF-KB. Using HO-1 siRNA, we confirmed that HO-1 mediated the anti-inflammatory actions of 2-HC. We also identified two novel CO-RMs (CORM-43 and CORM-319) as potent anti-inflammatory compounds. We also found that curcumin mitigated the cellular dysfunction-mediated by H2O2. This study also demonstrated that cold storage of human cardiac myoblasts caused marked increase in cytotoxicity primarily due to loss of cell membrane integrity as a result of necrosis. The addition of curcumin to the cold preservation solution (Celsior solution) resulted in preservation of cellular membrane integrity as well as cellular metabolism in cells exposed to cold storage and rewarming.;Conclusion: up-regulation of HO-1 is an effective therapeutic strategy to ameliorate cardiovascular injury and protect cardiac tissue during pathological conditions mediated by oxidative and inflammatory stresses.

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