The objective of this thesis was to develop new methodologies to assess upper gastro-intestinal function in health and disease. Several different technologies were studied in a range of upper gastro-intestinal diseases and adapted to try and provide more meaningful insights. The thesis has three main sections. In the first section, High Resolution Oesophageal Manometry (HRM) was used to assess unexplained upper gastro-intestinal symptoms in a group of patients referred to a tertiary centre. 46 patients were diagnosed with rumination syndrome following HRM. A retrospective review was completed of these patients case notes and HRM data. The predominant aim of this section was to identify if common mechanisms exist within rumination and its variations and to establish if the variety of presenting symptoms is due to different underlying problems or a common behavioural response to a variety of stimuli, with symptoms being dependent on the circumstance the behaviour exists in. This would support a generic biofeedback technique being useful regardless of presenting complaint. Comparing the variety of symptoms, exhibited behaviour and manometric findings, a new classification system for rumination was then developed; 1. Primary or “classical” rumination a. Increase in abdominal strain with corresponding rise in intra-gastric pressure and return of gastric contents to the mouth 2. Secondary or reflux-related rumination a. Reflux event causes the patient to respond with increase in intra-abdominal muscle strain and subsequent rumination 3. Supra-gastric belching independent of meals. a. Rise in intra-gastric pressure whilst a closed gastro-oesophageal junction, therefore producing rapid belching of air from the oesophagus without any return of gastric contents Generic biofeedback therapy was used (regardless of presenting symptoms) to control the abnormal behavioural response to symptoms. 20/46 patients reported full resolution of their symptoms and a further 13 / 46 reported improvement in their symptoms with this, while underlying mechanisms were targeted e.g. reflux with proton pump inhibitors, pain in functional dyspepsia. In the second main section of this thesis, gastro-oesophageal reflux disease (GORD) is considered. GORD is currently diagnosed by 24 hour pH studies. These are often difficult for patients to tolerate and require time off medication. A more attractive method would be for diagnosis to occur at the same time as gastroscopy. A novel instrument is the EndoFLIP® device. This measures cross-sectional area (CSA) and distensibility at the gastro-oesophageal junction (GOJ) via a long catheter with a balloon at the end that straddles the GOJ. It has been hypothesised that these measurements will be increased in those with GORD, as the GOJ is more distensible, allowing more retrograde movement of gastric contents. The aim of this section of the thesis was to establish if GOJ CSA and distensibility differentiate between healthy volunteers (HV) and GORD patients based on i) symptoms and ii) prolonged oesophageal acid exposure. 21 HV and 18 patients with GORD (based on symptoms) had EndoFLIP® measurements and wireless pH studies to assess this. 14% of HV and 50% GORD patients had pathological acid exposure. CSA and distensibility were both significantly higher in the HV’s compared to GORD patients. However, there was an inverse correlation between CSA and body mass index (BMI) which was significantly higher in the patient population. This may explain differences seen due to corresponding higher intra-abdominal pressure in those individuals with a high BMI, sub-sequentially affecting the CSA and distensibility. The complex structure of the GOJ and multiple factors involved in the pathogenesis of GORD present difficulties in using EndoFLIP® to diagnose GORD. It may find applications in other areas, such as serial measurements in single patients. In the final section of this thesis, gastric emptying is the focus and its pathogenesis in functional dyspepsia (FD). Current gastric emptying studies only find abnormalities in approximately 40% of patients with FD. Gamma scintigraphy is used in routine clinical practice for gastric emptying studies. Magnetic resonance imaging (MRI) is emerging as a modality in gastric emptying assessment and potentially provides additional information. This thesis hypothesised that standard gastric emptying studies may not be measuring the parameters reflective of underlying pathophysiology in FD. Also, most have a relatively small meal size that may be too small to trigger dysfunction. MRI may provide additional insights as can assess gastric contents and surrounding structure (unlike GS). To investigate these a 400ml test meal was utilised and gastric emptying parameters i) gastric contents volume at time 0 (GCV0, representative of early emptying), ii) gastric emptying rate at the time taken for half the meal volume to empty (GE rate @T50, representative of later emptying) and the more traditional measurement iii) time taken for half the gastric contents to empty (T50) in bopth GS and MRI studies. The hypothesis of this study is that early emptying is more rapid in FD due to impaired accommodation (therefore a lower GCV0) leading to a slower later emptying (therefore a lower GE rate @ T50). Following validation studies in a large healthy population (n=53), GS and magnetic resonance imaging (MRI) studies with a test meal of 400ml were used in 8 FD patients and 24 matched HV (from the pool of HV) . FD had a significantly lower BMI. Early emptying (represented by gastric contents volume after ingestion of meal (GCV0)) was significantly lower in GS for FD patients but higher in MRI. Time for half the meal to empty (T50) and gastric emptying rate at T50 (GE rate @T50) were similar. The difference between the two modalities was thought to be due to increased secretion production in the patients, which is measureable in MRI but not in GS. A further study with a solid component of 12 non-nutrient agar beads in addition to the liquid component was completed. 24 HV’s, 17 FD patients and 11 gastro-oesophageal reflux disease (GORD) patients were studied. FD patients and GORD patients had rapid early gastric emptying in comparison to HV in gamma scintigraphy (represented by GCV0) but higher GCV0 in MRI (significantly so between HV and GORD), suggesting increased secretion production is present in both conditions. These findings do support impaired fundal accommodation within the FD population but that other factors, such as secretion production and the rate of this in comparison to gastric emptying are important in the later stages of emptying. Further work is ongoing within the MRI department to quantify and measure the emptying of these secretions. This thesis explores how existing and new technologies can be applied to clinical conditions to identify possible pathophysiology and potential targets for treatment. Only by these ongoing efforts can we endeavour to improve the care we deliver to our patients.

Pioglitazone (PGZ) is a peroxisome proliferator-activated receptor (PPAR)-γ agonist that improves peripheral insulin sensitivity and reduces hepatocellular injury/ inflammation in non-alcoholic steatohepatitis (NASH). However, the underlying hepatic mechanism of action is not clearly understood with PGZ treatment. Therefore, liver biopsies were used to study genes, protein and immunohistochemistry expression of hepatocyte and hepatic stellate cell markers. PGZ decreased both αSMA and PPAR β expression in stellate cell and PPAR α hepatocyte expression, hence inhibit both stellate cell activation and β-oxidation in hepatocytes. PGZ also inhibit cell proliferation by reducing both PCNA and Ki67 in the liver. Up regulation of PPAR β, PXR, LXRα, IkBα and TNFRSF1B were observed (using Taqman Low density array gene expression), which indicates that PGZ exerts an anti-inflammatory and anti-fibrotic effect. PPAR β, PGC1α, ACADVL and UCP2 up regulation lead to increase β-oxidation and reduced reactive oxygen species (ROS) production. LXRα, ChREBP and SREBP1C activation leads to lipogenesis in the liver was also observed in PGZ treatment group. In vitro study was also conducted in freshly isolated human hepatic stellate cells, where these cells were incubated in vehicle and 5μM of PGZ for 72 hours consecutively. After 24 hours, 15ng/ml of PDGF-BB was incubated for 48 consecutive hours, followed by cell proliferation and gene expression analysis. PGZ up regulate the adipogenic genes expression followed by reduction in stellate cells marker expression and cell proliferation induced by PDGF-BB. PPAR β elevation after PGZ treatment in human liver and HSCs culture are novel findings in this study, but the role of PPAR β is clearly unknown in the liver and stellate cells. Therefore, similar treatment was performed using PDGF-BB on freshly isolated human HSCs, followed by treatment with PPAR β agonist (GW0742). GW0742 restores the adipogenic genes expression maintaining the quiescent phenotype of the stellate cell in contrast to previous study where PPAR β has been reported to cause stellate cells activation and proliferation. Overall, PGZ improved injury and fibrosis, inhibiting cell proliferation, increased both lipogenesis and β-oxidation in NASH patients. PGZ also inhibit stellate cells activation and proliferation and up regulated the adipogenic genes.

No description available.

No description available.

Phytic acid (PA) reduces nutrient digestibility in pigs and poultry, and has been shown to increase endogenous nutrient losses (ENL) in poultry. However, there is lack of information on the effect of PA on ENL in pigs, and mechanisms by which PA increases ENL. Three experiments were conducted to determine the effects of PA on ENL in pigs and to establish mechanisms by which PA increases the ENL. The first experiment investigated the effect of PA on ileal digestibility and ileal endogenous nutrient flows. Phytic acid decreased the apparent ileal sodium digestibility to a negative value (-18%). The second experiment investigated the effect of PA on gut enzyme activities, histomorphology and sodium-dependent glucose transporter 1 (SGLT1) gene expression. Phytic acid did not affect the gut villous height, villous height to crypt depth ratio, and jejunal SGLT1 gene expression, but decreased gastric pepsin activity and tended to decrease jejunal Na-K-ATPase activity. In the third experiment, the effect of PA on piglet performance and ion uptake in jejunum mounted in Ussing chamber, and jejunal SGLT1 protein level was evaluated. Phytic acid did not affect jejunal SGLT1 protein expression, but lowered piglet performance and jejunal active ion uptake. In conclusion, results from this study show that PA can reduce the apparent ileal digestibility of sodium to a negative value, indicating that PA can increase ileal endogenous sodium loss. The results also show that PA can reduce the pepsin activity and ion uptake in the gut. The reduced pepsin activity implies increased secretion of the enzyme plus hydrochloric acid and hence increased secretion of sodium bicarbonate that neutralizes the acid. The reduced ion uptake by PA implies reduced nutrient absorption. Because sodium is absorbed partly by co-transportation with other nutrients, the reduced ion uptake by PA implies reduced sodium absorption. Thus, it appears that PA increases ileal endogenous sodium flow partly through reduced pepsin activity and ion uptake in the small intestine. Overall, the results show that phytase (a PA-hydrolysing enzyme), which is added in pig diets to improve phosphorus availability, does not only improve phosphorus availability, but alleviates ant-nutritional effects of PA as well.

With the advent of DGHAL and PPH, treatments that purport to work by disrupting the arterial supply of haemorrhoids, there has been resurgence in interest in the vascular theory of pathogenesis of haemorrhoids. Despite uncertainty surrounding recurrence and complication rates there has been significant uptake of the new surgical approach due to decreased post-operative pain. However this has not been matched by discussion or evaluation of how haemorrhoidal disease and successful outcome should be evaluated. This thesis evaluates different approaches to the measurement of the burden of haemorrhoidal disease to the patient. A patient reported outcome measure was designed, administered and evaluated by the investigator. Reliability, reproducibility, validity, responsiveness and acceptability have been demonstrated. Three-dimensional ultrasound was used to acquire volumetric data and power Doppler angiography from the anal canal, which was shown to be reliable. Measures of power Doppler angiography were shown to be significantly lower in healthy volunteers than in patients. This technique represents promising value as an outcome measure of haemorrhoidal disease. A dual isotope-surgical nuclear probe technique attempted to measure change in volume of haemorrhoids following rubber band ligation, however consistent results were not obtained. Magnetic resonance imaging was able to demonstrate anal cushions and haemorrhoids, and the feasibility of this method has been demonstrated.

The incidence of oesophageal adenocarcinoma (OA) has increased exponentially in the western world over the past few decades. Barrett's oesophagus (BO) is a well known precursor of OA with a risk approximately 20 times more than that of background population. Regular endoscopic surveillance in patients with BO is recommended by most of the national gastroenterological societies. The advantage of Barrett's surveillance is to identify early subtle lesions which could then be managed early to avoid symptomatic and advanced cancers. The detection of such early lesions are challenging as they could be flat and inconspicuous on routine endoscopic examination. In the absence of any lesions, four quadrant biopsies every 1-2 cm of the whole length of Barrett's oesophagus is advised. This technique would map only 5-10% of the surface area of Barrett's segment and hence it is associated with significant sampling error. The improvement in electronics over the past decade has led to the production of endoscopes with better charged coupled devices and image enhancement techniques by altering the spectrum of light. This thesis examines the role of multi modal imaging in Barrett's oesophagus with a focus on detecting dysplasia and early cancer (EC). Firstly, the role of high definition (HD) imaging in routine clinical setting was studied using data from patients who have undergone Barrett's· surveillance. The yield of dysplasia by HD endoscopy was compared to standard definition (SD) endoscopy in this study. The role of narrow band imaging (NBI) with magnification in characterising abnormal lesions detected during BO surveillance was evaluated by performing a meta- analysis of clinical studies. The role of autofluorescence imaging (AFI) in Barrett's oesophagus was examined in detail with a view to understand the biological basis of autofluorescence and to improve the specificity of this technique as it is associated with significant false positive results in clinical studies. A meta-analysis was performed to identify whether AFI has a clinical advantage over white light endoscopy in detecting Barrett's dysplasia and the inter-observer reliability of this technology was studied using AFI expert and AFI non-expert endoscopists. An objective method of measuring the autofluorescence intensity was proposed as a ratio of the red to the green colour tone (AF ratio) of the area of interest. When the AF ratio of the lesion was divided by the AF ratio of the background mucosa, an AF index is obtained. A pilot study was performed to identify a cut-off value of AF index to differentiate high grade dysplasia (HGD) and EC from non-dysplastic BO. Finally, the biological basis of AF intensity was examined using APCmin mouse colonic models. This study looked into the AF ratio of the colonic mucosal lesions and correlated it with the amount of collagen and elastin in the submucosal tissue. Collagen and elastin are known to be the strongest fluorophores of the gastrointestinal tract and the question addressed is whether the low AF intensity associated with dysplastic lesions is due to the thickening of mucosa or to a reduction of collagen and elastin.

Phytic acid (PA) reduces nutrient digestibility in pigs and poultry, and has been shown to increase endogenous nutrient losses (ENL) in poultry. However, there is lack of information on the effect of PA on ENL in pigs, and mechanisms by which PA increases ENL. Three experiments were conducted to determine the effects of PA on ENL in pigs and to establish mechanisms by which PA increases the ENL. The first experiment investigated the effect of PA on ileal digestibility and ileal endogenous nutrient flows. Phytic acid decreased the apparent ileal sodium digestibility to a negative value (-18%). The second experiment investigated the effect of PA on gut enzyme activities, histomorphology and sodium-dependent glucose transporter 1 (SGLT1) gene expression. Phytic acid did not affect the gut villous height, villous height to crypt depth ratio, and jejunal SGLT1 gene expression, but decreased gastric pepsin activity and tended to decrease jejunal Na-K-ATPase activity. In the third experiment, the effect of PA on piglet performance and ion uptake in jejunum mounted in Ussing chamber, and jejunal SGLT1 protein level was evaluated. Phytic acid did not affect jejunal SGLT1 protein expression, but lowered piglet performance and jejunal active ion uptake. In conclusion, results from this study show that PA can reduce the apparent ileal digestibility of sodium to a negative value, indicating that PA can increase ileal endogenous sodium loss. The results also show that PA can reduce the pepsin activity and ion uptake in the gut. The reduced pepsin activity implies increased secretion of the enzyme plus hydrochloric acid and hence increased secretion of sodium bicarbonate that neutralizes the acid. The reduced ion uptake by PA implies reduced nutrient absorption. Because sodium is absorbed partly by co-transportation with other nutrients, the reduced ion uptake by PA implies reduced sodium absorption. Thus, it appears that PA increases ileal endogenous sodium flow partly through reduced pepsin activity and ion uptake in the small intestine. Overall, the results show that phytase (a PA-hydrolysing enzyme), which is added in pig diets to improve phosphorus availability, does not only improve phosphorus availability, but alleviates ant-nutritional effects of PA as well.

Introduction. The anti-idiotypic monoclonal antibody 10SAD7 mimics the tumour associated antigen 791T/ gp72, present on approximately 80% of colorectal cancer cells. A Phase I study using 10SAD7 in 13 patients with advanced colorectal cancer has shown that it is nontoxic, and conferred a survival advantage on patients who received it [Denton GWL 1994]. Aim. There were two aims of this work. The first was to assess whether. the survival advantage seen in the Phase I study was reproducible in a Phase II study. The second was to immunise patients with primary colorectal cancer, in a non-randomised adjuvant study, and explore further the immune responses generated. Materials and Methods. Patients with advanced colorectal cancer were recruited to a randomised, double-blind, placebo controlled survival study. The first patient was recruited to this Phase II study in April 1994, and the last in October 1996. Four trial centres were used- Nottingham, Hull, Leeds, and Newcastle. Eligible patients had a life expectancy of 3 months, and none had received radiotherapy or chemotherapy in the preceding 1 and 3 months, respectively. Patients attended on 3 occasions, 6 weeks apart, receiving 10µg of 10SAD7/alum i.d. followed by 100µg i.m. Venous blood was assayed for blood count and differential, liver function, urea and electrolytes, and CEA. Chest X-rays and CT scans were performed at trial entry and week 12 where possible. Dates of death were recorded following consultation with General Practitioner or referring clinician. In addition, patients with primary colorectal cancer were recruited to a non-randomised adjuvant study, whereby they received 10SAD7 before surgery. Venous blood samples were taken between immunisation and operation, and assayed for lymphocyte subsets. Samples taken from resection specimens were analysed immunohistochemically. Fresh tumours were in addition disaggregated, and separated TIL labelled with a panel of monoclonal antibodies, and analysed by flow cytometry. Control tumours were similarly labelled. All analysis was performed blind. Results. 162 patients were randomised to the Phase II study, between April 1994 and October 1996. 85 received 105AD7 and 77 placebo. The mean ages and sex-ratios of the two groups were comparable, as was the time from diagnosis of advanced disease to trial entry (172v179 days). Median survival from date on study was 124 and 184 days, in 105AD7 and placebo arms, respectively (p=O.38). Survival from date of diagnosis of advanced disease was 456 and 486 days (p=O.82). Chemotherapy and radiotherapy all prolonged survival in a multivariate analysis. Only one serious adverse event was seen in the 105AD7 arm, and this was felt unlikely to be attributable to the vaccine. Twenty-four patients were recruited to the adjuvant study. Immunohistochemical analysis of tumour sections from 16 patients showed increased infiltration of CD4 and CD8 expressing lymphocytes, relative to a well matched control group (p<O.05). Infiltration of CD4, CD8 and CD56 expressing lymphocytes combined was significantly higher, as was that of the mitochondrial antigen 7A6, expressed on cells undergoing apoptosis (p<O.005). The activation marker CD25 was also significantly increased (p<O.05). Flow cytometric analysis of disaggregated tumours from 16 trial and 22 control patients, confirmed the increased expression of CD25 on TIL in the 105AD7 group (p<O.01). Peripheral blood phenotyping failed to show any significant increase in any lymphocyte subset, following immunisation. A separate analysis was performed comparing 2 year survival and recurrence in 23 patients immunised by the previous CRC Fellow, with 97 matched controls from the Trent Audit. No significant difference was seen between the two groups. Discussion. No survival difference was seen between patients receiving l05AD7 and placebo, in the Phase II study. This suggests that any immune responses generated by l05AD7 are insufficient to have a significant effect on tumour growth, in patients with advanced disease. Work has therefore focused on immunising patients with primary colorectal cancer. Patients receiving l05AD7 prior to resection of their primary tumours, showed an increased number of activated lymphocytes, and apoptosis, at the tumour site, relative to a well-matched control group. The numbers in the survival analysis based on patients recruited by the previous CRC fellow, are insufficient to show whether any of these immunological changes confer a survival advantage. This question can only be answered in a large, prospective, placebo-controlled study in patients with primary colorectal cancer.

The incidence of the idiopathic inflammatory bowel diseases ulcerative colitis and Crohn’s disease appears to have risen markedly during the 20th century. These diseases now account for a considerable proportion of the workload of gastroenterologists in the developed world, and may affect as much as 1% of the population at some point in their lives. The aetiology of these diseases has been subject of much research over a number of decades and it is clear that both genetic and environmental factors are involved. The certain knowledge of environmental risk factors however remains scant. Similarly although inflammatory bowel diseases cause considerable morbidity and a small amount of mortality for their sufferers directly there is little agreement as to their overall impact once indirect effects are accounted for. This thesis contains studies contributing to the knowledge of both these areas using routinely or semi-routinely collected data. It examines two hypotheses relating to the aetiology of IBD (that risk is related to the season of birth, and that it is related to antibiotic use), and two areas of the impact of the diseases (overall mortality and fracture risk). With regard to aetiology the studies described show no variation in the risk of IBD with season of birth. They do show an increase in risk associated with the use of antibiotics, but since this is not specific (it is seen to occur with other groups of drugs also) it is far from clear that the association is causal. With regard to the indirect impact of the diseases a significant excess in overall mortality is demonstrated which is greater in Crohn’s disease than in ulcerative colitis, and is greatest in relative terms in the young but in absolute terms in the elderly. An excess is also shown for hip fractures in those with inflammatory bowel diseases, which is only partially explained by the use of corticosteroids.