Cancer Letters

Cancer Letters

Volume 370, Issue 1, 1 January 2016, Pages 117-124
Cancer Letters

Original Articles
Impact of the inflammatory microenvironment on T-cell phenotype in the progression from reflux oesophagitis to Barrett oesophagus and oesophageal adenocarcinoma

https://doi.org/10.1016/j.canlet.2015.10.019Get rights and content

Highlights

  • Profiled T-cells in oesophageal tissue across the GORD–BO–OAC sequence.

  • Assessed the impact of secreted factors from oesophageal tissue on T cell profile.

  • Higher proportion of CD8IL-4+ T cells in Barrett's tissue suggests Th2 profile.

  • Higher levels of secreted IL-6 in Barrett's environment reveal promising target.

  • Decreased T cell activation in OAC tissue suggests diminished immunity.

Abstract

The incidence of oesophageal adenocarcinoma (OAC), arising from reflux-induced Barrett oesophagus (BO), is increasing dramatically. T-cells have recently been implicated in the initiation of oesophagitis; however, their role in the progression from oesophagitis to BO and OAC has not been fully elucidated. Previous studies have examined the secreted cytokines from oesophageal tissue during disease progression but this study is the first to examine the activation phenotype and the inflammatory profile of CD4+ and CD8+ T-cells in human oesophagitis, BO and OAC tissue. Results demonstrated significantly higher levels of IL-4 producing CD4+ T-cells and secreted levels of IL-6, confirming a Th2 phenotype in BO. In OAC tissue, both pro- and anti-inflammatory cytokines were secreted, with significantly higher levels of IL-6, IL-1β, TNF-α, IFN-γ, IL-2 and IL-10 compared with normal oesophageal tissue. In addition, CD4+ T-cells infiltrating OAC tissue displayed a decreased activation profile, with significantly lower CD45RO and CD69 expression compared with normal tissue. Data from this study suggest that factors in the tissue microenvironment may alter T-cell phenotype and function early during oesophageal disease progression and may represent targets for immune intervention.

Introduction

The development of oesophageal adenocarcinoma (OAC) provides a well-defined model of an inflammatory driven neoplastic transformation, with chronic gastro-oesophageal reflux disease (GORD) and Barrett oesophagus (BO) being classified as major risk factors for OAC development [1], [2]. BO is a premalignant condition, characterised pathologically by specialised intestinal metaplasia of the distal oesophagus, and arises from a background of chronic GORD [3]. BO is a common finding in the general population with approximately a 1–2% incidence rate [4]. Patients suffering from GORD have a relative risk of 4.5 for developing OAC; this increases to 29.8 in patients who have progressed to BO [1]. However, the annual risk of developing OAC in a background of BO remains low, at just 0.12%, suggesting that an enhanced understanding of the underlying cellular biology could help identify those patients at heightened risk of disease progression [2].

The cellular pathogenesis of injury, inflammation, metaplasia and adenocarcinoma is unclear. In terms of GORD, it was initially assumed that cellular damage was a direct result of caustic injury at the luminal surface of the oesophagus due to chronic acid reflux [5], [6]. However, more recent evidence suggests that inflammation itself may be a key trigger, with an altered immune microenvironment providing a driving force for disease progression [7], [8], [9], [10]. OAC is a manifestation of prolonged inflammation and is thought to be influenced by immune cell infiltration at earlier stages of disease from oesophagitis and BO. T-cells, B cells, macrophages and dendritic cells have all been reported in high numbers in BO and OAC [11], [12], [13]; however, their profiles, functions and role in the inflammatory process are poorly understood. In-vivo models of reflux disease have demonstrated that T-cells infiltrate the sub-mucosal layers prior to tissue damage and prior to other immune cells, suggesting that they may initiate the inflammatory response and damage in GORD, making them pivotal players in oesophageal disease [9]. Studies have also demonstrated increased levels of IL-1β, IFN-γ and IL-8 mRNA in oesophagitis tissue compared to controls, whereas BO tissue displayed increased levels of IL-10 and IL-4 exclusively. This suggests that BO is characterised by a Th2 profile, while oesophagitis has a more pro-inflammatory cytokine profile [14]. An inflammatory shift to a more Th1 response is suggested to occur in OAC with levels of pro-inflammatory cytokines such as IFN-γ, IL-2, IL-1β and the chemokine IL-8 increased compared to healthy controls [15], [16]. While Th1 associated cytokines such as IFN-γ can stimulate potent anti-tumour immune responses [17], Th2 like cytokines such as IL-10 are considered immunosuppressive and pro-tumourigenic [18]. Therefore a shift to Th2 like responses during BO may promote tumourigenesis and a better understanding of such underlying immune mechanisms and the crosstalk between the tissue microenvironment and the immune system may reveal new immunotherapeutic targets.

For the first time, this phenotypic study directly assesses the profile of T-cells within the oesophageal tissue across the GORD–BO–OAC sequence and determines the impact of secreted factors from the oesophageal tissue microenvironment on CD4+ and CD8+ T-cells.

Section snippets

Acquisition of human oesophageal biopsies

All patients gave informed consent for use of their samples in this study, which was granted ethical approval by St James' Hospital/AMNCH Ethical Review Board. Biopsies were obtained from oesophagitis, Barrett oesophagus (BO) and oesophageal adenocarcinoma (OAC) patients undergoing endoscopy at St. James's Hospital. Biopsies from OAC patients were treatment naïve and obtained prior to initiation of any chemotherapy or radiotherapy. Control (normal) biopsies were taken from the mid-oesophagus of

T-cell activation is significantly diminished in oesophageal adenocarcinoma tissue

A detailed analysis of T-cell activation within oesophageal tissue was carried out using ex-vivo patient biopsies. Overall proportions of CD4+ and CD8+ cells did not differ between groups (data not shown). A significant reduction in the percentage of CD4+ T cells expressing CD45RO was observed in tumour tissue compared with normal (26.2 ± 10% vs. 69.4 ± 9.8%, p = 0.01) or Barrett tissue (72.2 ± 3.8%, p = 0.007) (Fig. 1A). Similarly, the percentage of CD4+ T cells expressing CD69 was

Discussion

This study aimed to carry out a detailed immunophenotyping of CD4+ and CD8+ T-cells in oesophagitis, Barrett and OAC tissue. While previous studies have proposed a Th1 type cytokine profile in oesophagitis, Th2 in Barrett and a return to Th1 again in OAC [11], [14], the actual T-cell profile in tissue has not been studied. This study directly assessed the profile of infiltrating T-cells within ex-vivo tissue samples across the GORD–BO–OAC sequence and examined how secreted factors from the

Funding

This work is funded by the Health Research Board (HRA_POR/2012/18).

Conflict of interest statement

We wish to confirm that there are no known conflicts of interest associated with this publication and there has been no significant financial support for this work that could have influenced its outcome.

Acknowledgements

The authors would also like to acknowledge the surgical and gastroenterology teams and patients at St. James's Hospital for partaking in the study.

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