Hypoxia-inducible factor 1 alpha is required for the tumourigenic and aggressive phenotype associated with Rab25 expression in ovarian cancer

The small GTPase Rab25 has been functionally linked to tumour progression and aggressiveness in ovarian cancer and promotes invasion in three-dimensional environments. This type of migration has been shown to require the expression of the hypoxia-inducible factor 1 alpha (HIF-1α). In this report we demonstrate that Rab25 regulates HIF-1α protein expression in an oxygen independent manner in a panel of cancer cell lines. Regulation of HIF-1α protein expression by Rab25 did not require transcriptional upregulation, but was dependent on de novo protein synthesis through the Erbb2/ERK1/2 and p70S6K/mTOR pathways. Rab25 expression induced HIF-1 transcriptional activity, increased cisplatin resistance, and conferred intraperitoneal growth to the A2780 cell line in immunocompromised mice. Targeting HIF1 activity by silencing HIF-1β re-sensitised cells to cisplatin in vitro and reduced tumour formation of A2780-Rab25 expressing cells in vivo in a mouse ovarian peritoneal carcinomatosis model. Similar effects on cisplatin resistance in vitro and intraperitoneal tumourigenesis in vivo were obtained after HIF1b knockdown in the ovarian cancer cell line SKOV3, which expresses endogenous Rab25 and HIF-1α at atmospheric oxygen concentrations. Our results suggest that Rab25 tumourigenic potential and chemoresistance relies on HIF1 activity in aggressive and metastatic ovarian cancer. Targeting HIF-1 activity may potentially be effective either alone or in combination with standard chemotherapy for aggressive metastatic ovarian cancer

The role of ethnicity in the increased susceptibility to non-alcoholic fatty liver disease in people of Indian origin

Introduction Non-alcoholic fatty liver disease (NAFLD) is fast becoming a global health concern. It is closely associated with obesity, diabetes and the metabolic syndrome, has a global pooled prevalence of 25% and is the leading cause of chronic liver disease in Europe and the United States. People of Indian ethnicity are at increased risk of diabetes and metabolic complications at a lower body mass index (BMI) than Caucasians. This predisposition for NAFLD is further compounded by Westernisation of Asian culture resulting in increased intake of sugar-rich, energy-dense foods and decreased levels of physical activity. Despite this well documented propensity to metabolic disease (including NAFLD), there is a lack of knowledge about the true disease prevalence in India. The studies that have been published are biased towards urban, tertiary centres, where access to healthcare and appropriate diagnostic tools is more readily available. In addition to this, there is conflicting data about the role of changing lifestyle habits on NAFLD risk – particularly in relation to diet. Reduced brown adipose tissue (BAT) activity has also been linked to obesity and diabetes, however there have been no prospective studies done to examine whether reduced BAT activity could also contribute to the increased NAFLD risk within this ethnic group. Aims The aim of this thesis is therefore to fill the above gaps in knowledge, to estimate accurately the NAFLD prevalence within a large Indian population, to identify the impact of different NAFLD risk factors within this population and compare NAFLD risk profile of native Indians with their UK migrant counterparts. It will also investigate the impact of BAT activity on NAFLD risk and understand the ethnic differences in BAT activity. Results Through population-level sampling of a large Southern Indian district, NAFLD prevalence (as diagnosed by ultrasound) was shown to be 49.8%. Risk factors for NAFLD within India were the same as those seen worldwide, namely male gender (adjusted OR 2.29 1.86-2.83, p<0.001), obesity (adjusted OR 2.81 2.015-3.68, p<0.001) – in particular central obesity – and components of the metabolic syndrome (diabetes adjusted OR 1.76 1.40-2.21, p<0.001). Dietary fat intake was also independently associated with NAFLD within India (adjusted OR 1.02 1.00-1.03, p=0.019). These risk factors appear unchanged by migration to the UK. Although there were no significant differences in dietary habits following UK migration, there appeared to be an element of dietary acculturation with decreased consumption of carbohydrate within the UK-migrant Indian cohort. This may result in changes to NAFLD phenotype over time. BAT activity was lower in native Indians (ΔTrel 0.36°C) compared to UK Caucasians (ΔTrel 0.50°C, p=0.010) and UK South Asian migrants (ΔTrel 0.57°C, p<0.001). This difference was however due to environment, as there was no difference in BAT activity between control groups of different ethnicity living in the same country. BAT activity does not directly influence risk of NAFLD (adjusted OR 0.47 0.07-3.20, p=0.444). Any differences in BAT activity were related to increasing BMI, which is itself a risk factor for NAFLD. Conclusion The prevalence of NAFLD in India is significantly higher than current national and global estimates. There is a commonality of risk between India and the rest of the world – namely obesity, diabetes and a diet high in fat. Within India, increased consumption of saturated fat in the form of edible oils and meat appears to impact NAFLD risk additionally. Whilst BAT activity is lower in Indians, it is not a direct cause for the increased NAFLD risk in this ethnic group. This work highlights the need for NAFLD screening for people of Indian ethnicity, both in India and in migrant populations. Further research needs to focus on education and interventional strategies to reduce prevalence of obesity and diabetes, which may be achieved through dietary manipulation

The role of ethnicity in the increased susceptibility to non-alcoholic fatty liver disease in people of Indian origin

Introduction Non-alcoholic fatty liver disease (NAFLD) is fast becoming a global health concern. It is closely associated with obesity, diabetes and the metabolic syndrome, has a global pooled prevalence of 25% and is the leading cause of chronic liver disease in Europe and the United States. People of Indian ethnicity are at increased risk of diabetes and metabolic complications at a lower body mass index (BMI) than Caucasians. This predisposition for NAFLD is further compounded by Westernisation of Asian culture resulting in increased intake of sugar-rich, energy-dense foods and decreased levels of physical activity. Despite this well documented propensity to metabolic disease (including NAFLD), there is a lack of knowledge about the true disease prevalence in India. The studies that have been published are biased towards urban, tertiary centres, where access to healthcare and appropriate diagnostic tools is more readily available. In addition to this, there is conflicting data about the role of changing lifestyle habits on NAFLD risk – particularly in relation to diet. Reduced brown adipose tissue (BAT) activity has also been linked to obesity and diabetes, however there have been no prospective studies done to examine whether reduced BAT activity could also contribute to the increased NAFLD risk within this ethnic group. Aims The aim of this thesis is therefore to fill the above gaps in knowledge, to estimate accurately the NAFLD prevalence within a large Indian population, to identify the impact of different NAFLD risk factors within this population and compare NAFLD risk profile of native Indians with their UK migrant counterparts. It will also investigate the impact of BAT activity on NAFLD risk and understand the ethnic differences in BAT activity. Results Through population-level sampling of a large Southern Indian district, NAFLD prevalence (as diagnosed by ultrasound) was shown to be 49.8%. Risk factors for NAFLD within India were the same as those seen worldwide, namely male gender (adjusted OR 2.29 1.86-2.83, p<0.001), obesity (adjusted OR 2.81 2.015-3.68, p<0.001) – in particular central obesity – and components of the metabolic syndrome (diabetes adjusted OR 1.76 1.40-2.21, p<0.001). Dietary fat intake was also independently associated with NAFLD within India (adjusted OR 1.02 1.00-1.03, p=0.019). These risk factors appear unchanged by migration to the UK. Although there were no significant differences in dietary habits following UK migration, there appeared to be an element of dietary acculturation with decreased consumption of carbohydrate within the UK-migrant Indian cohort. This may result in changes to NAFLD phenotype over time. BAT activity was lower in native Indians (ΔTrel 0.36°C) compared to UK Caucasians (ΔTrel 0.50°C, p=0.010) and UK South Asian migrants (ΔTrel 0.57°C, p<0.001). This difference was however due to environment, as there was no difference in BAT activity between control groups of different ethnicity living in the same country. BAT activity does not directly influence risk of NAFLD (adjusted OR 0.47 0.07-3.20, p=0.444). Any differences in BAT activity were related to increasing BMI, which is itself a risk factor for NAFLD. Conclusion The prevalence of NAFLD in India is significantly higher than current national and global estimates. There is a commonality of risk between India and the rest of the world – namely obesity, diabetes and a diet high in fat. Within India, increased consumption of saturated fat in the form of edible oils and meat appears to impact NAFLD risk additionally. Whilst BAT activity is lower in Indians, it is not a direct cause for the increased NAFLD risk in this ethnic group. This work highlights the need for NAFLD screening for people of Indian ethnicity, both in India and in migrant populations. Further research needs to focus on education and interventional strategies to reduce prevalence of obesity and diabetes, which may be achieved through dietary manipulation

L-carnitine supplementation in non-alcoholic fatty liver disease: A systematic review and meta-analysis

BACKGROUNDNon-alcoholic fatty liver disease (NAFLD) dominates the landscape of modern hepatology. Affecting 25% of the general population, there is critical unmet need to identify broadly available, safe and cost-effective treatments. Cumulative evidence in animal and human models suggests that intrahepatic and skeletal muscle fatty acid oxidation is impaired in NAFLD, such that lipid accretion is not matched by efficient utilisation. L-carnitine is a crucial mediator of fatty acid metabolism in vivo, promoting mitochondrial lipid β-oxidation and enhancing tissue metabolic flexibility. These physiological properties have generated research interest in L-carnitine as a potentially effective adjunctive therapy in NAFLD.AIMTo systematically review randomised trials reporting effects of dietary L-carnitine supplementation on liver biochemistry, liver fat and insulin sensitivity in NAFLD.METHODSSearch strategies, eligibility criteria and analytic methods were specified a priori (PROSPERO reference: CRD42018107063). Ovid MEDLINE, Ovid EMBASE, PubMed, Web of Science and the Cochrane Library were searched from their inception until April 2019. Outcome measures included serum concentrations of alanine and aspartate aminotransferase (ALT and AST), liver fat and insulin sensitivity assessed by the homeostasis model of insulin resistance (HOMA-IR). A random effects meta-analysis was performed for, ALT, AST and HOMA-IR measures separately. Between-study heterogeneity was measured using I2 statistics.RESULTSFive eligible randomised trials were included in the qualitative and quantitative synthesis (n = 338). All of the 5 included trials assessed the effect of L-carnitine on serum ALT, identified from Italy, South Korea and Iran. Weighted mean difference (WMD) for ALT between L-carnitine and control groups after intervention was -25.34 IU/L [95%CI: -41.74-(-8.94); P = 0.002]. WMD for AST between L-carnitine and control groups was -13.68 IU/L (95%CI: -28.26-0.89; P = 0.066). In three studies (n = 204), HOMA-IR was evaluated. WMD for HOMA-IR between L-carnitine and control groups was -0.74 units [95%CI: -1.02-(-0.46); P < 0.001]. Two studies using validated outcome measures reported a significant reduction in liver fat in L-carnitine vs control groups post-intervention (P < 0.001).CONCLUSIONPooled results indicate that L-carnitine supplementation attenuates ALT, liver fat and insulin resistance in NAFLD cohorts, confirming a beneficial effect of L-carnitine for a highly prevalent condition with a growing economic burden

Shear susceptibility of human mesenchymal stem cells increases with generation number: Implications for stem cell therapy scale-up and manufacturing

The ability of human mesenchymal stem cells (hMSCs) to withstand shear forces during processing is still debated as there have been reports of hMSCs being damaged during manufacturing. “Shear susceptibility” of suspended hMSCs (harvested from T-flasks) was investigated using a contractional flow device “torture chamber”.1 Surprisingly, hMSCs were found not to be any more shear susceptible than vero cells (commonly used for vaccine production) provided they are not passaged extensively. (figure 1) Therefore, the number of hMSC doublings before harvesting is limited, which presents a challenge for stem cell manufacturing and scale-up. In order to develop a scale up protocol for hMSCs, we first used HEK293T cells seeded on microcarriers. ANSYS FLUENT was used to model gentle agitation of cells seeded with microcarriers in a “100mm” culture dish to determine the lowest suitable agitation speed. Cells were seeded along with microcarriers in 10mL of media spinner flask with no agitation for 24 hours followed by orbital agitation at 35rpm. HEK293T cells scaled-up using orbital agitation were found to attach and spread to fresh microcarriers more efficiently than cells seeded into an impeller-mixed spinner flask. Transfer from “loaded” microcarriers to fresh microcarriers was found to occur via “contact transfer” or “bridging” between carriers. Hence, orbital agitation is thought to promote this transfer mechanism. For anchorage-dependent hMSCs, attachment efficiency to microcarriers upon seeding plays a significant role in cell production given the apparent passage limitations. Therefore, we expect that when the orbital agitation protocol is used for scale-up of hMSCs, significantly more hMSCs will survive the seeding/attachment process and transfer between microcarriers will be more efficient than in traditional spinner flask microcarrier culture. Please click Additional Files below to see the full abstract

Health related quality of life in individuals at high risk of chronic liver disease: Impact of a community diagnostic pathway

Objectives: There is a lack of understanding of health related quality of life (HRQoL) in chronic liver disease (CLD). With the rising prevalence of alcohol and obesity driven CLD, and the increasing ability to screen for fibrosis, it is important to understand the impact of the diagnostic process for patients.Study design: Prospective cohort studyMethods: A cohort study conducted utilising the Nottingham Adult Liver Disease Stratification Pathway, UK. All patients referred as high risk for CLD (due to metabolic, alcohol or abnormal liver enzymes) completed the EQ-5D before diagnosis and at three and 12 months after. HRQoL was investigated by domain, CLD severity (transient elastography) and temporally.Results: 493 patients participated with 300 (60.9%) completing at least one follow-up HRQoL assessment.Pre-diagnosis the median (IQR) utility index was 0.75 (0.61-0.85) and visual analogue scale was 75/100 (60-90). The median utility index was significantly lower amongst those with advanced liver disease compared to those without at all time points (baseline 0.68 vs 0.77, three-months 0.65 vs 0.79, 12-months 0.69 vs 0.84, all

Outcomes of elective induction of labour compared with expectant management: population based study

Objective To determine neonatal outcomes (perinatal mortality and special care unit admission) and maternal outcomes (mode of delivery, delivery complications) of elective induction of labour compared with expectant management

Consumer representation in financial services: report into consumer representation in the payments sector

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How do pilot and feasibility studies inform randomised placebo-controlled trials in surgery? : A systematic review

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