NSAIDs, which act as COX antagonists, are widely used in veterinary practice to provide long-term pain relief, however a major limiting complication of their use in dogs is gastric ulceration. Gastric ulceration may result from either direct toxic effects on gastric epithelial cells or systemic action, including the inhibition of gastric epithelial cell migration. The role of cell migration in this effect and the specific signalling pathways involved are not known. COX-derived prostaglandins are known to have an important role in gastric defence and cytoprotection through the promotion of gastric mucosal blood flow and mucus secretion and the inhibition of gastric acid secretion. Given the importance of gastric epithelial cell migration in re-establishing gastric mucosal integrity following injury, the aim of this investigation was to test the hypothesis that paracrine PGE2 signalling modulates gastric epithelial cell migration. In order to address this hypothesis, a primary cell culture model which incorporates intact canine gastric glands that spread to form monolayer cell islands, was first characterised and then used. The effects of non-selective and COX-2 selective antagonism on cell spreading in this model and wound healing in immortalised cell monolayers was assessed. Furthermore, the involvement of subtype-specific EP receptor signalling in PGE2-mediated modulation of epithelial cell migration was investigated through treatment with specific agonists and antagonists. Both non-selective and COX-2 selective antagonism inhibited PGE2 production and epithelial cell migration, thus providing evidence that COX-2-derived PGE2 is important for the modulation of epithelial cell migration. Furthermore, non-selective and COX-2 selective antagonism inhibited cellular protrusive activity. The effects of PGE2 on epithelial cell migration were shown to be mediated through EP3 and EP4 receptor signalling. Expression of COX-2, EP3 and EP4 was found to be readily induced in response to stressors. Interestingly, COX-2 expression was up-regulated in patients infected with spiral bacteria. These findings provide evidence that COX-2-derived PGE2 stimulates epithelial cell migration and the formation of cellular protrusions. Thus, reduced PGE2 production in the gastric mucosa may inhibit gastric epithelial migration and contribute to the ulcerogenic effects associated with COX antagonist therapy.

The enteric nervous system (ENS) arises mainly from cells exiting the vagal neural crest, entering the bowel and migrating caudally. A failure in this migratory process is thought to result in the clinical entity Hirschsprung’s disease (HSCR), which is typically characterised by the absence of enteric ganglia in the colon and rectum (aganglionosis). Untreated HSCR may present with life-threatening bowel obstruction in the first few days of life. However, despite recent surgical advances children and adults still suffer significant life-long post-operative morbidity. ENS progenitor cells (ENSPC) have been shown to persist in the postnatal bowel, thereby stimulating research into the potential of manipulating or transplanting these cells to improve long-term outcomes. Early results have been promising, ENSPC have been isolated from children with HSCR and clonally expanded in cultured neurospheres, after which they have been transplanted into aganglionic embryonic mouse gut ex-vivo and shown to restore a normal pattern of contractility. However, questions still remain about the functional potential and the safety implications of transplanting these cells back into humans. Therefore, this thesis is aimed at improving our understanding of the properties of these cells and the mechanisms controlling their behaviour when they are brought into culture and after subsequent transplantation. The first part of this thesis focuses on the proliferation of ENSPC within the in-vitro environment of cultured mouse and human neurospheres. The main findings were that cells undergoing proliferation are predominantly found at the neurosphere periphery rather than being evenly distributed throughout the neurosphere. Subsequently, post-mitotic peripheral cells were found to be distributed throughout the neurosphere. Furthermore, using embryonic and postnatal mouse tissue it was demonstrated that the proportion of cells undergoing of proliferation decreased with increasing maturity of the source tissue. The second part of the thesis identifies the requirement for Notch signalling pathway in the self-renewal of undifferentiated human ENSPC. Inhibition of the Notch signalling pathway with both chemical inhibitors and siRNA knockdown resulted in decreased ENSPC proliferation together with increased neuronal differentiation. Manipulation of this pathway may therefore improve both the effectiveness and safety of any future cell-based therapy. The next section of the thesis describes the development of an ex-vivo human colonic smooth muscle model to investigate the behaviour of ENSPC in an environment closer to that of the human colon. The chapter describes the development and maintenance of human colonic smooth muscle cells in long-term culture and demonstrates contractility, thus providing a model in which ENSPC behaviour can be investigated in a more physiologically relevant environment. During the routine culture of human aganglionic smooth muscle preliminary observations were made consistent with the presence of cells that could give rise to neurospheres, despite the absence of an ENS in this region of the bowel. The final part of this thesis confirms the presence of ENSPC within the aganglionic bowel and characterises their behaviour. These progenitors are demonstrated to differentiate into mature ENS specific neuronal phenotypes, and importantly have the capability to restore a normal pattern of contractility in the embryonic aganglionic gut model. Taken together the work in this thesis furthers our understanding of the control and behaviour of human ENSPC. Furthermore, demonstration of the existence of ENSPC in the bowel of children with HSCR raises important questions regarding both the aetiology HSCR and the potential to utilise the cells in future autologous cell-­‐based therapies.

Energy homeostasis is a tightly regulated system that is vital for survival involving anorectic and orexigenic signals. Obesity is a maladaptive response where the balance becomes disrupted. Obesity is one of the most concerning health problems of our time. It is no longer considered a consequence of a western lifestyle, with more developing countries now reporting an increased incidence of obesity and associated illnesses. While obesity itself can be debilitating and decrease quality of life, it is the associated comorbidities that are the main cause for concern; including type two diabetes, cancer and thrombo-occlusive diseases. One of the molecules thought to be responsible for occlusive events is plasminogen activator inhibitor (PAI)-1. This inhibitor of the plasminogen system is also reported to be up to 5 fold higher in obese subjects in plasma, and similar to leptin, is released from adipose tissue. PAI-1 is considered to play a protective role in circumstances of gastric mucosal attack, thus a transgenic mouse (PAI-1HKβ) was generated, with targeted expression of PAI-1 to the gastric parietal cells, to study this. However, an unexpected phenotype emerged, most notably hyperphagia and increased body weight, which formed the basis of these present studies. The gut-brain axis is a major and well-studied regulator of energy homeostasis and this was the focus of this project. The PAI-1HKβ mice when compared to wild-type had decreased brain stem responses to the satiety hormone, Cholecystokinin (CCK). Brainstem responses were also attenuated in wild types pre-treated with exogenous PAI-1. Furthermore, it was shown that the urokinase plasminogen activator (uPA) receptor by which PAI-1 binds, was required to influence the observed decrease in brainstem responses. CCK also has other physiological functions in the role of energy homeostasis, including gastric emptying. While delayed gastric emptying was observed following a protein rich liquid test meal in C57BL/6 mice, PAI-1HKβ mice had a blunted response. Blockade of the CCK1 receptor in C57BL/6 mice also attenuated the delay in gastric emptying. Moreover, exogenous PAI-1 attenuated CCK-mediated inhibition of gastric emptying. The PAI-1HKβ mice had an attenuated inhibition of gastric emptying of a non-nutrient containing liquid test meal in response to CCK. Treatment with gastrin was shown to increase plasma PAI-1 and attenuated delayed gastric emptying in C57BL/6 mice. Food intake is stimulated by orexigens, most notably ghrelin, working via appetite-stimulating neurons in the arcuate nucleus. While ghrelin stimulated feeding in fed ad libitum C57BL/6 mice, PAI-1 increased feeding in previously fasted C57BL/6 mice only. This response to ghrelin and PAI-1 was also replicated in PAI-1 -/- mice, suggesting PAI-1 is not required for the orexigenic effect of ghrelin. Moreover, intrapertoneal (ip.) administered ghrelin increased fos expression in arcuate neurons of both C57BL/6 and PAI-1 -/- mice, whereas ip. PAI-1 did not. Weight loss in the PAI-1HKβ mice appeared to reverse the insensitivity to CCK in terms of gastric emptying. PAI-1HKβ mice were also found to be insensitive to other gut-derived satiety hormones, suggesting gastric PAI-1 is an anti-satiety factor. However, mice null for wild-type gastric PAI-1 responded normally to CCK prior to feeding, indicating that wild type is necessary for CCK insensitivity in the PAI-1HKβ mice. The current findings demonstrate that PAI-1 plays a role in the control of food intake. PAI-1 is an example of a novel anti-satiety factor that can modulate gut-brain signalling via the vagus nerve in order to preserve nutrient intake. This work provides a platform for future investigations into novel pathways implicated in the development and treatment of obesity.

Colorectal cancer is a potentially curable disease if diagnosed at its earliest stages. However, as the current tools for early diagnosis of colorectal cancer are suboptimal, this condition is still a major health issue in the UK and the whole world. Each year nearly 40,000 new cases are diagnosed in the UK. Approximately half of these patients have advanced disease at the time of diagnosis and this is associated with a less favourable prognosis. Therefore, there is still an urgent need for better colorectal cancer screening tools. Moreover, it has long been suggested that APC deletion is an early and major event in the initiation of more than 80% of all colorectal cancers. However, the molecular events that occur following APC deletion are yet to be fully understood. Working on an acute Apc deletion animal model, our group has identified several candidate biomarker proteins. It is postulated that studying these proteins will reveal new aspects about early colorectal tumourigenesis. We hypothesised that these proteins are upregulated very early following Apc deletion and are involved in various aspects of colorectal cancer development, therefore they are potential biomarkers and/or therapeutic targets for this disease. Most of the project’s aims were addressed using immunohistochemical assessment of the expression of several candidate biomarkers in a novel in vivo model of acute Apc deletion (AhCre+Apcfl/fl mouse), a model of established early intestinal neoplasia (ApcMin/+ mouse) and a model of invasive disease (AhCreERT+Apcfl/+Ptenfl/fl mouse) as well as clinical samples from patients with early and advanced colorectal cancer. Mechanistic studies were carried out using the human colonic adenocarcinoma cell lines, HCT116 and HT29. In the animal models, deletion of Apc resulted in an early activation of the Wnt signalling pathway as indicated by nuclear localisation of Beta catenin. Six (NAP1L1, RPL6, SFRS2, PHB, FABP6 and NCL) out of the nine candidate biomarker proteins tested and two proposed partner molecules (Cyclin E and CDC5L), demonstrated obvious differential patterns of expression in areas where Apc loss had induced Wnt pathway activation. Specific knockdown studies of selected members of this protein list in human colon adenocarcinoma cell lines using siRNA identified important effects of these proteins on critical cellular functions such as proliferation and apoptosis. NAP1L1 and RPL6 knockdown had an inhibitory effect on cell proliferation and survival and caused a simultaneous increase in apoptosis. SFRS2 knockdown caused increased abundance of nuclear CDC5L and vice versa. SFRS2 down regulation had marginal effects on cell proliferation and cell survival and resulted in a small reduction in the amount of apoptosis. In contrast, CDC5L knockdown caused a dramatic inhibition of cellular proliferation, loss of the G2 cell cycle peak and a significant increase in the amount of apoptosis and caspase 8 activity. HCT116 cells with a p53 deletion were more sensitive to toxicity of experimental (transfection) reagents and SFRS2 knockdown in these cells caused obvious inhibition of cell proliferation with increased apoptosis. Immunohistochemical staining of the candidate biomarker proteins in human samples of colorectal cancer generally supported the results shown in animal studies for early stages of the disease. Moreover, it demonstrated reduced nuclear expression of Beta catenin, nuclear relocalisation of CDC5L and cytoplasmic displacement of SFRS2 in the more advanced stages of colorectal cancer. PHB showed increased cytoplasmic staining in Dukes’ stage A and B cancers. These results were backed up by appropriate scoring systems. Our results concerning these proteins in colorectal cancer are novel and agree with several studies by other research groups describing their roles in other cancers or cancer models. Due to the relatively late detection of lesions in humans, the early changes which were observed in animals might have been missed in humans. NAP1L1, RPL6, SFRS2 and NCL showed early overexpression during colorectal tumourigenesis. These proteins therefore have the potential as screening biomarkers. Due to their role in regulating cellular proliferation, NAP1L1, RPL6 and CDC5L are potential therapeutic targets for colorectal cancer. PHB is also a potential marker for Dukes’ stage A and B cancers. Mechanistically, the interplay between SFRS2 and CDC5L during colorectal tumourigenesis is a promising case to follow. Based on this study and other studies conducted in our group, the above proteins are promising screening, predictive or prognostic markers as well as potential therapeutic targets for colorectal cancer.

Non-alcoholic fatty liver disease (NAFLD) is characterised by the accumulation of lipid in liver. Liver is the principal organ involved in drug biotransformation. The central hypothesis of this project is that alterations in the liver environment due to lipid accumulation aggravate sensitivity of hepatocytes to toxicity of some commonly prescribed drugs (paracetamol, alcohol, phenobarbital, cisplatin or doxorubicin). The aim of this study was to investigate the effect of lipid overloading (steatosis) on drug cytotoxicity in the liver model Huh7 cell line. Hepatic steatosis was induced in Huh7 cells by exposing cells to 300 μM FFA mixture or 1 mM oleic acid. Impact of steatosis on drug toxicity was examined by co-treating the cells with FFA and paracetamol, alcohol, phenobarbital, cisplatin or doxorubicin. Cell viability MTT assay showed that neither 300 μM FFA mixture nor 1 mM OA caused significant reduction in cell viability after 24 h incubation. However, a significant reduction seen after incubation for 48 h, therefore for the subsequent experiments the time frame chosen was 24 h. The results showed that 300 μM FFA mixture did not induce a significant amount of intracellular lipid, whereas 1 mM OA induced significant amount of intracellular lipid. The subsequent experiments were carried out to test the hypothesis that lipid-loaded Huh7 cells are more sensitive to drug toxicity. Co-treatment with FFA and either paracetamol, ethanol or doxorubicin resulted in further reduction in cell viability. Phenobarbital resulted in enhanced cell viability, and no significant changes in cell viability observed after co-treatment with cisplatin. To investigate mode of cell death, caspase3/7 activity was measured as mediator of apoptosis. Generally, an advance apoptosis was observed in steatotic cells treated with the different drugs. Reactive oxygen species were measured as a possible trigger of apoptosis. A significant amount of ROS were generated by FFAs, and generally, drug treatment induced a higher significant amount of ROS in the steatotic Huh7 cells. To elucidate the specific changes observed upon treatment of steatotic cells with hepatotoxic drugs, a single FFA dose (300 μM FFA mixture) and a single drug (doxorubicin) were selected to identify major alterations in gene expression. The microarray data confirmed alterations in oxidative stress-related genes. Such changes were functionally relevant as confirmed by cellular assay and In-Cell Western blot. In conclusion, the hypothesised effect of steatosis on drug toxicity has been confirmed and steatosis may enhance drug toxicity through changing cellular oxidative state and activating the apoptotic pathway.

The prevalence of overweight and obesity continues to rise substantially across the world. It is the leading preventable cause of death worldwide and is associated with a large number of comorbidities that present a perpetual burden on healthcare costs. Much of the recent work to understand and address the problem of obesity has focused on the role of gastrointestinal hormones on the regulation of appetite, satiety, and food intake, and how interventions such as physical activity and exercise can affect the secretion of these hormones. However, the gastrointestinal system and the role of gastric emptying are often overlooked. The aim of this thesis was to enhance understanding of the physiology and regulation of gastric emptying and its interactions with carbohydrates. This will help in the development of novel non-pharmacological dietary interventions or foods that can modulate appetite and energy intake. A series of studies on human volunteers are presented in this thesis. Firstly, the gastric emptying rate of different 6% simple sugar solutions (water control, fructose, glucose, sucrose, 50:50 fructose and glucose) and gut hormone responses of circulating acylated ghrelin, active glucagon like peptide-1 (GLP-1), glucose dependent insulinotropic polypeptide (GIP) and insulin were investigated. Hepatic metabolism and function in response to the different simple sugar solutions were also examined. The time of maximal gastric emptying rate (Tlag) differed significantly between between sucrose and glucose solutions. Differences in insulin and GIP responses between fructose containing solutions and glucose only solutions were also seen. No differences in hepatic metabolism measures or function were observed following the intake of 36 g of the various test sugars. However, lactate production was significantly greater for fructose containing solutions. Following on from these results, the effect of increased dietary fructose intake on gastric emptying rate of glucose and fructose was investigated. Three days supplementation with 120g/d fructose resulted in acceleration of gastric emptying rate of a fructose but not a glucose solution. No significant differences in the circulating concentration of gastrointestinal hormones, but subtle differences in responses over time were suggested which may explain the specific monosaccharide adaptations of gastric emptying. Further work is required to confirm this and to investigate the longevity and reversibility of the gastrointestinal adaptation and the mechanism involved. Lastly, several tagging single nucleotide polymorphisms (SNP) of the GLP-1 receptor gene were associated with gastric emptying rate. Further work is required on the regions identified to pinpoint the exact SNP or SNPs responsible.

Liver Fluke (Fasciola hepatica and F. gigantica) infection causes fasciolosis or Liver Rot disease, which is estimated to cause losses to the agricultural sector in the order of US$10 billion per year and is growing in significance as a neglected tropical disease that infects 17 million people worldwide. Following the accidental ingestion of metacercariae, infective stage liver fluke excyst, penetrate the duodenum of the host and migrate through the liver towards the bile ducts. During the process of invasion, liver fluke excrete/secretes multiple proteolytic enzymes that are key virulence proteins that operate at the host-parasite interface. Prominent amongst the virulence proteins released by liver fluke are cathepsin 8 proteases; these proteases not only facilitate excystment and invasion of the duodenal wall but also take part in feeding, digestion, and immune evasion. Until recently only 10 clades of cathepsin B proteases were reported, however our integrated PCR and bioinformatic-based approaches have uncovered new data on liver fluke cathepsin B proteases. Based on our findings, we propose a new clade-based classification system for cathepsin 8 proteases of the family Fasciolidae. Expression analyses identified a total of 22 cathepsin B proteases across the key life stages (metacercariae, newly excysted juvenile [NEJ] and adult) of three species, Fasciola hepatica, Fasciola gigantica and Fascioloides magna. End point PCR analyses and real-time PCR based approaches were used to interrogate the relative expression of all the cathepsin B clades identified. The results \showed that almost all the cathespin 8 clades are activated during the dormant metacercarial stage. Cathepsin B2 is highly expressed in metacercaria and NEJs but is weakly expressed in adult worms. The latter has been widely studied as a key protease in the early infection stages and has been touted as a potential control target for fluke. We also identified inconsistencies in the naming of cathepsin Bs and, based on key structrual features, we propose that F.hepatica cathepsin B1 is renamed as cathepsin B5; its homologue, FgCB5, was found to be expressed in F. gigantica. The putative cathepsins B5 and B8 (Robinson et. al., 2009) were found to be part of cathepsins 87 and 86, respectively and have been redesignated accordingly. A homologue of cathepsin 87 (FmCB7) was also discovered in F. magna and it was more like FgCB7 than FhCB7. Some cathepsin 8 proteases from fluke have been proposed to be inactive, e.g. F. hepatica cathepsin 84 (FhCB4) because it possess possesses an active site serine residue rather than a cysteine. In contrast, F. gigantica cathepsin 84 does possess a cysteine in the active site. Interestingly, our PCR expression profiling data indicated that FhCB4 is expressed in both the infective life stages of the parasite and in the adult worms. To investigate this further we set out to elucidate the activity of FhCB4 in following heterologous expression in Pichia pastoris along with a mutant enzyme in which the active site cysteine was present (FhCB4S29C). Unfortunately, none of the proteases showed functional activity against the f1uorogenic substrates Z-Arg-Arg-AMC, Z-Phe-Arg-AMC and Z-Leu-Arg-AMC, as the mature peptide failed to get auto-processed from the propeptide. The structural analysis of FhC84, FgCB4 and FhC82 using PyMOL software has shown negligible differences in the catalytic triad, questioning the proposal that it is an inactive protease.

This thesis presents the design, development and efficacy of antimicrobial nanotherapies, to overcome the challenges faced by conventional antimicrobial delivery for the treatment of Pseudomonas aeruginosa respiratory infections in cystic fibrosis (CF). In the CF lungs, mucus and biofilms pre.sent complex barriers to the optimal delivery of antimicrobial therapy, leading to low antimicrobial exposure to resident bacteria and inadequate eradication. Respiratory disease, associated with chronic respiratory infection, is ultimately the main cause of morbidity and mortality in CF patients. There is a need to develop improved antimicrobial and anti-biofilm therapies in this patient group. Firstly, alginate/chitosan polymeric nanoparticle (NP) delivery vehicles for the antimicrobial tobramycin were formulated. Tobramycin NPs demonstrated antimicrobial efficacy against laboratory and clinically isolated strains of P. aeruginosa in vitro and in vivo against infected Galleria mellonella. Next, functionalisation of the mucolytic DNase to tobramycin NPs was investigated with the aim to improve penetration of the mucus and biofilm barriers, to increase the accessibility of tobramycin to the difficult-to-reach sites of infection. Functional activity of both drugs was established against DNA, P. aeruginosa strains and in CF sputum. DNase functionalisation was found to improve the sputum penetration of NPs and these DNase tobramycin NPs possessed anti microbia! activity in sputum samples from CF patients. Finally, the antimicrobial activity of the developed nanotherapies against P. aeruginosa biofilms was studied. DNase tobramycin NPs have the potential to target both the biofilm matrix and bacterial cells through the combined DNA degradation and bactericidal actions of DNase and tobramycin. Against established P. aeruginosa biofilms, both tobramycin NPs and DNase tobramycin NPs were found to be effective antimicrobial agents. The developed antimicrobial nanotherapies represent a promising therapeutic strategy for improving tobramycin delivery to mucus-embedded, biofilm-associated respiratory infections in CF.

Cystic Fibrosis (CF) is a genetic disorder primarily characterised by chronic infection and inflammation within the lungs. Serine proteases, in particular trypsin-Ilke serine proteases are associated with dehydration of the airways through proteolytiC activation of the Epithelial Sodium Channel (ENaC), leading to a reduction in mucociliary clearance and colonisation of the airways. The development of novel inhibitors and activity-based probes (ABPs) targeting these proteases was hypothesised as potential therapeutic agents and research tools to allow further insights into the disease mechanism. Novel decarboxylated substrate mimetics were designed and synthesised as potential reversible inhibitors of these Channel Activating Proteases (CAPs). This led to the development of a 4-amidinobenzylamine based compound which exhibited selective inhibition of matriptase and trypsin whilst displaying limited effects on other CAPs. An additional approach undertaken was the screen of a fragment library, previously identified through virtual screening as potential inhibitors of prostasin. This approach identified a 2,4-diamino-1,3,5-triazine analogue as a potential lead compound for the development of novel prostasin inhibitors. In the development of novel chemical tools for the detection and characterisation of novel CAPs, a library of broad-spectrum probes featuring a diphenyl ɑ-aminoalkylphosphonate moiety was synthesised. These compounds displayed potent irreversible inhibition with limited cross-reactivity and enabled successful detection of serine proteases utilising 'click-chemistry' techniques. In addition to the application of diphenyl ɑ-aminoalkylphosphonate ABPs, a novel class of irreversible protease inhibitor utilising N-alkyl glycine residues was developed. Consequently, a range of irreversible serine protease inhibitors and activity-based probes were synthesised displaying both broad-spectrum and more specific inhibitory properties. Promisingly, such compounds have displayed similar or improved inhibitory potency in comparison to their diphenyl ɑ-aminoalkylphosphonate counterparts. These compounds have also been applied successfully within Activity-Based ELlSA systems for the quantification of active serine protease. In conclusion, the development of these novel ABPs will aid future research into the proteolytic activation of ENaC and furthermore will allow insight into protease involvement within other disease states and may enable the quantification of protease biomarkers for use within both research and the clinic.

This thesis showed that there is overlap between the PXR and a related nuclear receptor termed the constitutive androstane receptor (CAR). The murine pyruvate dehydrogenase complex E2 (PDC-E2) was cloned, expressed, purified and used as antigen for ELISA experiments. Next, three annual studies were carried out in SJL/J mice, by sensitising them with a PDC-E2 derived peptide (p163), in order to check if these mice develop cholangiopathy. Then, two animal studies were carried out in order to check the effect of pregnenolone-16alpha-carbonitrile (PCN) in PXR-/- and PXR +/+(C57BL6 background) mice chronically treated with carbon tetrachloride (CCI4). The result was that PCN did not inhibit the hepatotoxicity of CCI4. Furthermore, a mouse line was derived that could be susceptible to the development of cholangiopathy (SJL/J- PXR-/- ) and wild type (SJL/J- PXR+/+) control. Immunisation of SJL/J- PXR-/- and SJL/J-PXR+/+ with p163 peptide increased the portal tract inflammation equally in mice over CFA control. Treating these mice with lithocholic acid (LCA) produced a higher level of portal tract inflammation in knockout mice than in wild type. Moreover, twelve Single Nucleotide Polymorphisms (SNPs), on human PXR gene, were studied in relation to lipid profile in Glasgow population (P-population). Interestingly, two of them (PXR3 and PXR8) had strong association with lipoprotein variations especially in the male population. Three SNPs (PXR1, PXR2 and PXR8) on human PXR gene were studied between PBC samples and control groups and the results showed that these three genetic variations did not particularly predispose to the disease. Finally one polymorphism (PXR11) on human PXR gene was studied between hyperlipidemia and control group but again our results showed that this specific SNP did not predispose to hyperlipidemia.