An investigation into the role of interleukin-6 in linking systemic and local inflammatory responses in patients with colorectal cancer

Colorectal cancer is the second most common cause of cancer death in the UK. It is accepted that both tumour and host factors are important determinants of disease progression and survival. While systemic and local inflammatory responses are increasingly recognized to be of particular importance the understanding of the mechanisms linking these important inflammatory processes remains unclear. This thesis examines the prognostic importance of measures of systemic and local inflammation and proposes a hypothesis for a link between tumour necrosis, systemic and local inflammatory responses in patients with colorectal cancer. Chapter 3 reports the comparison of the prognostic value of longitudinal measurements of systemic inflammation in patients undergoing curative resection of colorectal cancer. In Chapter 3 the results demonstrate that there was no significant overall change in either mGPS or NLR from pre- to post- operatively. This study highlighted the associations between pre- and post- operative mGPS and NLR and T-stage (p<0.001), TNM stage (p<0.005) and cancer-specific survival. The relationships between pre-operative measurements were examined using multivariate analysis. For pre-operative measurement both mGPS and NLR were associated with cancer-specific survival while when post-operative measures were examined only mGPS was specifically associated with cancer-specific survival (HR 4.81, CI 2.13-10.83, P<0.001). Chapter 4 examines the prognostic value of the Klintrup-Makinen scoring method and the existing limitations with regard to its clinical utility. An automated scoring method using commercially available image analysis software was developed and compared with manual scoring of tumour inflammatory infiltrates. This study demonstrated that both manual K-M scoring (p<0.001) and automated K-M scoring (p<0.05) had prognostic value in patients who had undergone potentially curative resection of colorectal cancer, and that the novel automated method may provide an objective method of assessment of tumour inflammatory infiltrates using routinely stained haematoxylin and eosin sections of tumour samples. In chapter 5 a hypothesis was proposed that Interleukin-6 may link tumour necrosis and systemic and local inflammatory responses in patients with colorectal cancer. This chapter examined the basis for this hypothesis, which is presented in figure 5.1. In addition, in chapter 5 the importance of this potential link is examined. In chapter 6, the hypothesis outlined in chapter 5 was examined in a cohort of patients who had undergone attempted curative resection of colorectal cancer. This study examined the inter-relationships between circulating mediators, in particular IL-6, tumour necrosis and systemic and local inflammatory responses. This results of this study demonstrated that IL-6 was associated with tumour necrosis (<0.001) and mGPS (<0.001) independent of T-stage. Thus adding weight to the hypothesis that elevated circulating concentrations of IL-6 may play a role in modulating both the systemic and local inflammatory responses in patients with cancer. Chapter 7 further develops the hypothesis that IL-6 signalling may be important in modulating systemic and local inflammatory responses in patients with colorectal cancer. Further, in chapter 7 the basis for the role of trans-signalling in this signaling pathway was examined. In this study, we reported that neither expression of the soluble IL-6 receptor or soluble gp130 were associated with systemic or local inflammatory responses. As a result the possible reasons for these findings were explored and future work suggested. A prospective database of patients undergoing attempted curative resection of colorectal cancer in Glasgow Royal Infirmary was used throughout this thesis. This database was created and is maintained regularly by successive research fellows at the Royal Infirmary. The work presented in this thesis highlights the importance of the host response in the form of systemic and local inflammation in patients with colorectal cancer and proposes a link between these responses and tumour necrosis. In addition, this work adds weight to the body of evidence suggesting that assessment of these host responses may improve stratification to treatment for patients with colorectal cancer. Further, this work proposes a mechanistic link, between tumour necrosis, systemic and local inflammatory responses through Interleukin-6, that merits further investigation.

MRI indices of functional recovery following intracerebral implantation of a human neural stem cell line in a pre-clinical model of ischaemic stroke

Stem cell therapy for ischaemic stroke is an emerging field in light of an increasing number of patients surviving with permanent disability. Several allogenic and autologous cells types are now in clinical trials with preliminary evidence of safety. Some clinical studies have reported functional improvements in some patients. After initial safety evaluation in a Phase 1 study, the conditionally immortalised human neural stem cell line CTX0E03 is currently in a Phase 2 clinical trial (PISCES-II). Previous pre-clinical studies conducted by ReNeuron Ltd, showed evidence of functional recovery in the Bilateral Asymmetry test up to 6 weeks following transplantation into rodent brain, 4 weeks after middle cerebral artery occlusion. Resting-state fMRI is increasingly used to investigate brain function in health and disease, and may also act as a predictor of recovery due to known network changes in the post-stroke recovery period. Resting-state methods have also been applied to non-human primates and rodents which have been found to have analogous resting-state networks to humans. The sensorimotor resting-state network of rodents is impaired following experimental focal ischaemia of the middle cerebral artery territory. However, the effects of stem cell implantation on brain functional networks has not previously been investigated. Prior studies assessed sensorimotor function following sub-cortical implantation of CTX0E03 cells in the rodent post-stroke brain but with no MRI assessments of functional improvements. This thesis presents research on the effect of sub-cortical implantation of CTX0E03 cells on the resting- state sensorimotor network and sensorimotor deficits in the rat following experimental stroke, using protocols based on previous work with this cell line. The work in this thesis identified functional tests of appropriate sensitivity for long-term dysfunction suitable for this laboratory, and investigated non-invasive monitoring of physiological variables required to optimize BOLD signal stability within a high-field MRI scanner. Following experimental stroke, rats demonstrated expected sensorimotor dysfunction and changes in the resting-state sensorimotor network. CTX0E03 cells did not improve post-stroke functional outcome (compared to previous studies) and with no changes in resting-state sensorimotor network activity. However, in control animals, we observed changes in functional networks due to the stereotaxic procedure. This illustrates the sensitivity of resting-state fMRI to stereotaxic procedures. We hypothesise that the damage caused by cell or vehicle implantation may have prevented functional and network recovery which has not been previously identified due to the application of different functional tests. The findings in this thesis represent one of few pre-clinical studies in resting-state fMRI network changes post-stroke and the only to date applying this technique to evaluate functional outcomes following a clinically applicable human neural stem cell treatment for ischaemic stroke. It was found that injury caused by stereotaxic injection should be taken into account when assessing the effectiveness of treatment.