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Immunohistochemical study of NF-kB activity and IL-8 abundance in oesophageal adenocarcinoma; a useful strategy for monitoring these biomarkers
  1. Gareth JS Jenkins (g.j.jenkins{at}swansea.ac.uk)
  1. Swansea School of Medicine, United Kingdom
    1. Jane Mikhail (j.mikhail{at}ntlworld.com)
    1. Swansea School of Medicine, United Kingdom
      1. Ali Alhamdani (doctorali2000{at}hotmail.com)
      1. Swansea School of Medicine, United Kingdom
        1. Tim Brown (brown320{at}aol.com)
        1. Morriston Hospital, United Kingdom
          1. Scott Caplin (scott.caplin{at}swansea-tr.wales.nhs.uk)
          1. Morriston Hospital, United Kingdom
            1. James Manson (james.manson{at}swansea-tr.wales.nhs.uk)
            1. Singleton Hospital, United Kingdom
              1. Rhian Bowden (rhian.kift{at}swansea-tr.wales.nhs.uk)
              1. Singleton Hospital, United Kingdom
                1. Nassima Toffazal (nassima.toffazal{at}swansea-tr.wales.nhs.uk)
                1. Singleton Hospital, United Kingdom
                  1. Paul Griffiths (paul.griffiths{at}swansea-tr.wales.nhs.uk)
                  1. Morriston Hospital, United Kingdom
                    1. James Parry (jmp{at}swansea.ac.uk)
                    1. Swansea School of Medicine, United Kingdom
                      1. John Baxter (john.baxter{at}swansea-tr.wales.nhs.uk)
                      1. Morriston Hospital, United Kingdom

                        Abstract

                        Aims: To determine if immunohistochemistry (IHC) could be used to monitor NF-kB activity in oesophageal adenocarcinoma and pre-malignant (Barrett¡¦s) oesophageal tissues, relative to normal oesophageal mucosa. The pro-inflammatory cytokine IL-8, a transcriptional target of NF-kB, was also studied to better understand NF-kB functionality and its RNA and protein levels were assessed in oesophageal tissues. Methods: IHC was employed using an antibody against the nuclear localisation sequence (NLS) of the p65 subunit as well as an antibody against IL-8. To assess NF-kB function, we also assessed changes in gene expression of NF-kB controlled genes (IL-8 and I-kB) in the histological sequence using real-time PCR. More global expression changes were also studied using membrane arrays. Results We found that IHC was effective at monitoring overall NF-kB activity and IL-8 abundance. This method also allowed NF-kB activity and IL-8 abundance to be pin-pointed in specific cell-types. ,We saw significant increases in nuclear NF-kB activity and IL-8 abundance across the histological series by IHC analysis. Gene expression analysis also showed consistent up-regulation of IL-8 confirming the IHC data and demonstrating enhanced transcriptional NF-kB activity. I-kB (another NF-kB target) showed down-regulation in dysplastic and adenocarcinoma tissues. Down-regulation of I-kB gene expression may partly explain increased NF-kB activity. Conclusion We suggest that IHC using antibodies against the NLS of p65 may be useful in monitoring overall NF-kB activity in oesophageal tissues. As IHC is amenable to highthroughput screening (whereas traditional EMSA methods are not), this may lead to the development of a better screening tool for early cancer risk.

                        • Barrett's oesophagus
                        • IL-8
                        • Immunohistochemistry
                        • NF-kB
                        • biomarker

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