NIHRDataInsights/HRCSData huggingface dataset:
Title: Reprogramming adult human hepatocytes into liver progenitors with unlimited self-renewal, efficient differentiation, and transplantation capacities
Funder: Medical Research Council

Abstract: In 2017, the Ochiya group reprogrammed rat mature hepatocytes by the use of 3 small molecules, and generated liver progenitors with unlimited self-renewal and efficient differentiation capacities, called chemically induced liver progenitors (CLiPs). While several groups, including ours, have applied the same or similar culture conditions to human hepatocytes, so far only limited success was obtained. Human CLiPs (hCLiPs) derived from the adult human hepatocytes did not proliferate over 1 month. hCLiPs from the fetal/neonatal hepatocytes had an unlimited proliferation capacity, but lost their differentiation capacity within 2 weeks. In this project, 1) we aim to generate hCLiPs with unlimited proliferation and efficient differentiation capacities using doxycycline (Dox)-inducible shRNA and transcription factors (TF) expression based on the existing data sets. In this candidate approach, we have already found that down-regulation of a hepatocellular carcinoma (HCC) suppressor was sufficient to overcome the proliferation block in adult hCLiPs, and a cocktail of 6 TFs up-regulated ALBUMIN (ALB) expression in late passage adult hCLiPs. High ALB expression correlates with a CLiP state with a good differentiation capacity in rat, mouse and human. In parallel, 2) we will look for pathways essential for the maintenance of CLiP proliferation/self-renewal and differentiation capacities using CRISPR/Cas9-mediated genome-wide knockout screens with early passage mouse CLiPs and genetically engineered human CLiPs generated in 1). Finally, 3) we will apply the compounds that can activate the essential pathways identified in 2) to establish a culture condition that supports self-renewal of the fully functional hCLiPs upon removal of Dox using the cell lines established in 1). This work will not only provide an unlimited source of fully functional hepatocytes for toxicology tests and drug screens, but also pave a way to generate non-genetically modified hCLiPs for cell therapies.


Research activity codes: 5.2 CELLULAR AND GENE THERAPIES
Health categories: ORAL AND GASTROINTESTINAL
Title: NOVEL TOOLS FOR EVALUATING INTESTINAL DYSFUNCTION IN CHILDREN AND ADULTS WITH MALNUTRITION DISORDERS
Funder: Medical Research Council

Abstract: Severe acute malnutrition (SAM) is a medical emergency, especially when there are infectious or metabolic complications, and under these circumstances mortality can reach 30% during a 2-3 week hospital admission. Clinical trials of novel interventions suffer from the drawback of inadequate endpoints which do not predict mortality or non-response well, and which reflect pathophysiology only poorly. In addition, those tests which are available are expensive and cumbersome to administer. Here, we propose five work packages, all designed to develop new tools for measuring elements of gut functional capacity which are common disturbances in many nutritional disorders. Stable isotope techniques will be used to analyse digestion and absorption, derangements of which have been neglected for decades. New laser-based sensing will be used to define increased permeability at point of care, using oral fluorescent tracers. Measurements of dendritic cell/monocyte responses to translocated pathogen associated molecular patterns will enable features to be defined for future development. The composition of the microbiome will be quantified using DNA sequencing, and its functionality assessed using metabolomics. Finally, all of these will be brought to bear on children with SAM in Zambia, Zimbabwe and Uganda, who will be followed up during and after their critical illness to determine which tools best map the clinical course. This 5-year study brings together a unique consortium with diverse skill sets to address a singularly important question in global health, but the data generated may be of wider application for people anywhere with intestinal and nutritional problems.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS;2.2 FACTORS RELATING TO PHYSICAL ENVIRONMENT
Health categories: ORAL AND GASTROINTESTINAL
Title: Effects of alpha1-antitrypsin polymerisation on organelle structure and fluidity in hepatocytes
Funder: Medical Research Council

Abstract: How mutant alpha-1-antitrypsin (A1AT) impairs liver function is unclear. Many mutants of A1AT accumulate in the endoplasmic reticulum (ER) causing it to fragment. This leads to increased sensitivity to ER-stress, caused when ER protein folding is impaired. ER-stress elicits an unfolded protein response (UPR), which is the liver can cause inflammation and cell death. We discovered that A1AT polymers form a matrix within the ER through which ER proteins can diffuse. Formation of this matrix changes the biophysical properties of the ER including microviscosity and crowding. We will elucidate the mechanisms of A1AT-induced ER dysfunction by focusing on the relationship between A1AT polymers and molecular motion within the ER. Aim 1: Determine the mechanism linking A1AT accumulation and the UPR in hepatocytes. FRAP and point FCS will report A1AT and chaperone mobility in iPSC-derived hepatocytes like cells (HLCs). The chaperones responsible for driving A1AT polymers into an immobile form will be identified. Manipulate ER-shaping proteins will reveal the relationship between ER shape and ER viscosity and crowding. How these parameters alter ER stress signalling will be determined. Aim 2: Determine how A1AT polymers affect chaperone mobility within intact reticular ER. Single particle tracking will report A1AT and chaperone mobility in reticular ER of HLCs. We will determine how mutant A1AT alters the movement of large complexes, including chaperones, in morphologically normal ER in HLCs. Aim 3: Modify ER luminal A1AT mobility for therapeutic gain. We will manipulate ER structure (ER shaping proteins), ER stress signalling (proteostasis enhancer drugs) and A1AT polymerisation status (polymerisation inhibitors) in an effort to ameliorate the inflammatory and fibrogenic signals emanating from Z-A1AT HLCs. These experiments will reveals the mechanisms linking A1AT polymers to altered ER structure, viscosity and crowding, and to inflammation and fibrosis.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: ORAL AND GASTROINTESTINAL
Title: Structural and cellular basis of alpha-1-antitrypsin (AT) deficiency and the serpinopathies
Funder: Medical Research Council

Abstract: We described the polymerisation of mutants that underlie antitrypsin deficiency and developed new paradigms for the liver and lung disease associated with this condition. We have also described the polymerisation of mutants of antichymotrypsin that have been associated with COPD and mutants of neuroserpin that cause a novel dementia that we called familial encephalopathy with neuroserpin inclusion bodies (FENIB). We grouped these diseases with others that result from polymerisation of mutants of the serpins as the serpinopathies. We have used biochemistry, biophysical analysis, crystallography, NMR, cell and induced pluripotent stem cells, monoclonal antibodies and Drosophila and C. elegans models of disease to dissect the structural basis of polymer formation and the cellular consequences. We are in the late stage of developing small molecules that block polymerisation with BioMarin. However, the current molecules also block inhibitory activity against neutrophil elastase. The specific aims are to: (i) determine the linkage in pathological polymers isolated from the livers of Z homozygotes at near-atomic resolution by cryo-electron microscopy; (ii) define the structure of the pathological polymers caused by shutter domain mutants of antitrypsin that cause liver disease and neuroserpin that cause FENIB (these are likely different from those caused by the Z mutation); (iii) use NMR to characterize intermediates on serpin polymerisation pathways and fingerprint and define the structure of the pathological polymer; (iv) use cutting-edge tomographic approaches to determine the morphology of Z antitrypsin polymers in situ within the cellular environment and (v) determine the use of pathological polymers as a biomarker and diagnostic tool for antitrypsin deficiency. This work will allow us to explore new diagnostic and therapeutic strategies for antitrypsin deficiency and the serpinopathies. The work is enabled by the platform facilities and technical support at UCL.


Research activity codes: 4.1 DISCOVERY AND PRECLINICAL TESTING OF MARKERS AND TECHNOLOGIES;4.2 EVALUATION OF MARKERS AND TECHNOLOGIES
Health categories: ORAL AND GASTROINTESTINAL
Title: SOLID: Determining the optimum biomarker strategies for the detection of advanced liver disease at the primary-secondary care interface
Funder: Medical Research Council

Abstract: Aim: develop a primary care pathway as a biomarker evaluation platform to identify to individuals with advanced liver fibrosis Patients: Patients with obesity, T2DM or harmful alcohol consumption attending annual reviews will be recruited. Searches in 2 GP practices indicate 2600 of 20,000 patients meet eligibility criteria. Test biomarkers: FIB-4 is inexpensive with good negative predictive values and proven utility in 2-step fibrosis pathway with TE in NAFLD. ELF and PRO-C3 have good diagnostic accuracy alone or in combination with FIB-4 so we believe evaluating these biomarkers is warranted. Transient elastography will be our standard to stage fibrosis because it is non-invasive and validated against liver biopsy. Although TE is imperfect, it is unethical to conduct liver biopsies in a case finding study. Liver stiffness (fibrosis) and controlled attenuation parameter (steatosis) will be measured using the M or XL probe. A valid scan will be defined as 10 valid elastography measurements with an IQR/median ratio <0.3. Only patients with valid TE readings will be used to assess biomarker performance. Our study has been powered to allow 5% failure and 10% non-attendance for TE rate. Primary endpoint is correct diagnosis of advanced fibrosis (TE >10 kPa) using a 2-step pathway (FIB-4 and ELF or PRO-C3). In NAFLD, specificity for advanced fibrosis is 75% (71% sens) at 9.7 kPa and 90% at 14.1 kPa. >10 kPa has been chosen to maintain sensitivity. We recognise that there is a false positive rate for this cut-off so we will assess the final fibrosis stage after review in liver clinic with all test results including liver biopsy if performed. Given all 3 biomarkers have high sensitivity for advanced fibrosis, only patients with 1 or more elevated biomarker will have TE performed to rationalise resource. Novel pathways will be developed using a per-protocol analysis and modelled to develop the most effective pathway on an 'intention to diagnose' basis.


Research activity codes: 4.4 POPULATION SCREENING
Health categories: ORAL AND GASTROINTESTINAL
Title: Hepatocytes Encapsulated with mesenchymal stromal cells in alginate microbeads for the treatment of acute Liver failure in Paediatric patients (HELP)
Funder: Medical Research Council

Abstract: Acute liver failure (ALF) in children and adults is a rare disease with high mortality without liver transplantation (LTx), as other therapies like extracorporeal liver assist devices have failed to demonstrate efficacy. LTx is life saving but limited by the scarcity of donor organs. In addition, life-long immunosuppression needs make liver transplant itself another disease. Success of auxiliary liver transplantation, demonstrating more than 70% native liver recovery, suggests that few weeks of support with minimally invasive hepatocytes microbeads could provide similar outcome to auxiliary liver transplant in children with ALF of diverse aetiologies. We have proven the feasibility and safety of the transplantation of human hepatocyte microbeads (HMB001) for the treatment of ALF in children, and the partial success of this approach, supported by a NIHR grant. In the last few years, further supported by the NIHR, we have developed HMB002, made of primary human hepatocytes and support cells encapsulated in a novel alginate. HMB002 showed greater function and longevity than HMB001: after 14 days, in vitro albumin production was 60x higher in HMB002 than in HMB001 and the A1AT production 32x higher. In vivo data showed a similar pattern, leading us to believe that HMB002 would be more potent in providing effective liver support to bridge the patients to recovery without liver transplantation, or bridge them to liver transplantation. The i4i NIHR grant allowed us to develop HMB002 as a GMP (ATIMP) clinical product. Our clinical trial application was awarded MHRA approval in September 2020. The aim of this project is to undertake a phase I/II clinical trial to test HMB002 in children with ALF to assess its safety/efficacy. Success of this trial would open a road to commercialisation with consideration of extended use to treat adults with ALF and bridge patients with acute-on-chronic liver failure to liver transplantation.


Research activity codes: 6.2 CELLULAR AND GENE THERAPIES
Health categories: ORAL AND GASTROINTESTINAL
Title: Structural Studies into E. coli SslE and its role in Diarrhoeal Diseases
Funder: Medical Research Council

Abstract: The gastrointestinal tract is populated by a complex community of microorganisms, the gut microbiota, which have a major role in maintaining a healthy immune system. E. coli is a gram-negative bacterium that colonizes the human intestine and is generally beneficial; however, virulent strains are the cause of enteric diseases, urinary tract infections, sepsis and meningitis. The epithelial cells that line the gastrointestinal tract are covered by a layer of secreted mucus which acts as a barrier to separate epithelial cells and adjacent host tissue from the residing commensal microbiota but also invading pathogens. Mucins are a family of large glycosylated proteins and the primary constituents of mucus. As such, a major pathogenic mechanism of successful enteric pathogens is the targeting and degradation of these mucin structures. A broad range of E. coli pathotypes export the protein SslE, which is essential for intestinal colonization and has a dual role in promoting mucosal clearance in the host intestine and promoting biofilm maturation. However, we lack a full structural and mechanistic understanding of how SslE interacts with mucins. We have identified a unique feature of SslE where it can form polymers that stabilize biofilm structures and our preliminary studies have provided a partial Cryo-EM structure of the monomeric SslE core region which hints at a novel mode of interaction with mucins. Using the commensal E. coli W and enterotoxigenic H10407 strains as model systems, we now plan to (i) elucidate the atomic structure of intact SslE, (ii) determine how SslE recognizes and processes mucins and (iii) establish the role of SslE/mucin interactions during colonization. These studies will provide insight into new mechanisms related to E. coli pathogenesis and may have implications for the development of new vaccines and strategies to reduce bacterial virulence.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING
Health categories: ORAL AND GASTROINTESTINAL
Title: Improving tolerance for FODMAPs using modified celluloses: defining the role of gelation in reducing gas production in vitro and in vivo
Funder: Medical Research Council

Abstract: Low FODMAP (fermentable, oligo-di-mono-saccharides and polyhydric alcohols) diets have revolutionised management of irritable bowel syndrome (IBS). FODMAPs cause abdominal discomfort, bloating and flatulence by their rapid colonic fermentation which produces gas and distends the colon. Low FODMAP diets, although widely adopted, are expensive, inconvenient and may have negative effects on health by excluding high fibre foods (HFF) that protect against obesity-related diseases and colon cancer. This project aims to use modified celluloses to trap FODMAPs in a gel, thereby limiting bacterial access and modifying fermentation to produce less gas. We have shown this in IBS patients by co-administered psyllium, a viscous and gel forming fibre, with the FODMAP, inulin. Initial experiments in healthy volunteers (HVs) using breath hydrogen response to inulin as the endpoint, will co-administer modified celluloses to define the relative importance of gelation versus viscosity and the optimum cellulose for subsequent use. A model of colonic fermentation using stool samples from the HVs will measure gas and short chain fatty acid (SCFA) production to a) characterise the response to increased viscosity and gelation b) define the variation in SCFA production associated with the wide range of gas production observed in vivo. Subsequent HV studies will use MRI and the pH-sensitive SmartPill to measure the effect of cellulose on key parameters (flow rate of inulin into colon, colon volume, pH). The effect of these changes will be measured in the model colon, validated by comparison of in vitro and in vivo results. They will also inform a clinical trial of 3 week of cellulose treatment in IBS patients with endpoints including MRI colonic gas volumes, symptoms, microbiota and SCFAs. This new understanding of the relationship between fermentation and symptoms could lead to inexpensive treatments for IBS and functional foods to improve population tolerance of beneficial HFFs.


Research activity codes: 2.2 FACTORS RELATING TO PHYSICAL ENVIRONMENT
Health categories: ORAL AND GASTROINTESTINAL
Title: Optical Confirmation of Nasogastric Tube Placement with Early Photon Imaging
Funder: Medical Research Council

Abstract: Placement of enteral feeding tubes (nasogastric tubes/NGTs) is a standard medical procedure, yet misplacement is a serious issue (e.g. food entering the lungs, with consequent death and disability from pulmonary complications). Current practice relies heavily on ionising X-ray radiation for medical device localisation, causing significant disruption due to the dual requirements to protect staff and waiting for the availability of, or transport to, X-ray equipment. Current X-ray based techniques for NGT localisation often yield ambiguous and difficult-to-interpret results, resulting in (at best) delays to feeding whilst the procedure is repeated or (at worst) inappropriate commencement of feeding and subsequent severe harm to the patient. We have developed a compact bedside system capable of providing a practitioner with real-time guidance/feedback/visualisation when placing NGTs. This device utilises an imaging implementation of the technique known as time-correlated single-photon counting (TCSPC). When a point source of light is placed inside the body, a tiny fraction of the light will emerge from the body with a near line-of-sight / direct path. These initial photons can be differentiated from later-arriving photons (which have been subject to scattering by tissue) by a time resolved single-photon sensitive camera positioned outside of the patient. The direct path photons hold precise information about the location of the point source inside the tissue. In combination with a custom NGT/insert that controllably emits light at distinct points along the tube, we can therefore detect the complete path of inserted medical devices. The path is visually projected back onto the patient, in real-time, during placement or reconfirmation. This device has the potential to improve clinical workflows, increase safety and shorten time to feeding (linked with improved clinical outcomes). We now aim to clinically translate and evaluate our device in a first-in-human study.


Research activity codes: 6.3 MEDICAL DEVICES
Health categories: ORAL AND GASTROINTESTINAL
Title: MICA: Tissue ecology in IBD-development and pathophysiological function
Funder: Medical Research Council

Abstract: IBD is a complex multifactorial disease involving maladaptation of the host microbe interface and ensuing relapsing and remitting chronic inflammation of the gastrointestinal tract. Encompassing Crohn's disease (CD), ulcerative colitis (UC) and IBD unclassified (IBDu), IBD is extremely heterogeneous in disease phenotype, behaviour, location and response to therapy limiting our ability to direct new biological therapies to those most likely to benefit. In recent work we mapped distinct cellular and molecular pathways to discreet intestinal tissue niches termed pathotypes that underlie treatment non-response in IBD. Building on these results we will employ a multi-disciplinary approach, iterating between mouse models and study of human IBD tissue to provide mechanistic insight into pathotype development and function. Bulk and single cell sequencing alongside spatial transcriptomics and proteomics on human IBD tissue will be used to further characterise established pathotypes, as well as to identify new ones related to disease outcome measures. Mechanistic analysis will be performed using disease positioned mouse models that reproduce tissular pathotypes to identify upstream microbial drivers and sensing pathways triggering pathotype development, including the contribution of inflammasome and autophagy pathways to epithelial cell responses. In parallel, we will assess the contribution of cytokine networks and fibroblasts to the formation of pathotype-specific inflammatory tissue niches. Finally, pathologic processes identified in mouse models will be validated functionally in ex vivo patient tissue assays, and assessed for therapeutic utility. By generating new understanding of the diversity of tissular processes that underlie IBD heterogeneity, we aim to provide a biological evidence-based strategy for the therapeutic management of IBD.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: ORAL AND GASTROINTESTINAL
Title: Hepatic Progenitor Cell response to ischemia reperfusion injury; biological pathways involved in post-transplant biliary injury and repair
Funder: Medical Research Council

Abstract: Background: A current hypothesis states that ischemia reperfusion injury (IRI) results in damage to the Hepatic Progenitor Cell (HPC) population (a subset of cholangiocytes within the biliary tree), which leads to insufficient regeneration of the injured bile ducts and development of biliary complications (BC) post orthotopic liver transplantation (OLT). Preliminary data: Human HPCs (hHPCs) display a change in phenotype when subjected to hypoxia. Genes associated with Primary Cilia (PC), organelles exclusively present in cholangiocytes in the liver, display a significantly reduced expression following hypoxia. Length of biliary PC has been assessed in human livers declined for OLT. Length of PC decreases with prolonged warm and cold ischemia. These results have been confirmed in a murine model of liver procurement and cold storage. Hypothesis and Aims: IRI decreases the hHPCs` capacity to repair biliary injury via a mechanism involving PC. This study aims at modelling the development of BCs to understand their pathology and identify therapeutic targets. Methods: hHPCs have been extracted from livers declined for OLT, cultured in vitro and characterized. hHPCs will be used in functional IRI studies in vitro (hypoxia chamber, hydrogen peroxide) and analysed including RNA-sequencing. Findings will be confirmed ex vivo in human samples using liver protocol biopsies from transplanted grafts. Findings will be correlated with the clinical outcome. Functional studies will be performed. First in vitro by modifying the hHPCs using CrispR technology and then in vivo by transplantation of unmodified and modified hHPCs into a murine model of biliary disease. Outcomes: To enhance the understanding of the biochemical processes involved in biliary IRI and regeneration. Secondly, to identify therapeutic targets to treat/ prevent BCs to help redefine graft utilisation by improving graft quality and thus expand the clinical acceptance criteria for liver grafts.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS;5.2 CELLULAR AND GENE THERAPIES
Health categories: ORAL AND GASTROINTESTINAL