CRISPR-edited hepatic organoids as drug screening platform for non-alcoholic fatty liver disease
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PROVIDER: S-EPMC10432308 | biostudies-literature | 2023 Jul
REPOSITORIES: biostudies-literature
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Project description:We report the generation of human ESC-derived, expandable hepatic organoids (hEHOs) using our newly established method with wholly defined (serum-free, feeder free) media. The hEHOs stably maintain phenotypic features of bipotential liver stem/progenitor cells that can differentiate into functional hepatocytes or cholangiocytes. The hEHOs can expand for 20 passages enabling large scale expansion to cell numbers requisite for industry or clinical programs. The cells from hEHOs display remarkable repopulation capacity in injured livers of FRG mice following transplantation, and they differentiate in vivo into mature hepatocytes. If implanted into the epididymal fat pads of immune-deficient mice, they do not generate non-hepatic lineages and have no tendency to form teratomas. We further develop a derivative model by incorporating human fetal liver mesenchymal cells (hFLMCs) into the hEHOs, referred to as hFLMC/hEHO, which can model alcoholic liver disease-associated pathophysiologic changes, including oxidative stress generation, steatosis, inflammatory mediators release and fibrosis, under ethanol treatment. Our work demonstrates that the hEHOs have considerable potential to be a novel, ex vivo pathophysiological model for studying alcoholic liver disease as well as a promising cellular source for treating human liver diseases.
| S-EPMC6951343 | biostudies-literature
Project description:Non-alcoholic fatty liver disease (NAFLD) is currently the world's most common liver disease, estimated to affect up to one-fourth of the population. Hallmarked by hepatic steatosis, NAFLD is associated with a multitude of detrimental effects and increased mortality. This narrative review investigates the molecular mechanisms of hepatic steatosis in NAFLD, focusing on the four major pathways contributing to lipid homeostasis in the liver. Hepatic steatosis is a consequence of lipid acquisition exceeding lipid disposal, i.e., the uptake of fatty acids and de novo lipogenesis surpassing fatty acid oxidation and export. In NAFLD, hepatic uptake and de novo lipogenesis are increased, while a compensatory enhancement of fatty acid oxidation is insufficient in normalizing lipid levels and may even promote cellular damage and disease progression by inducing oxidative stress, especially with compromised mitochondrial function and increased oxidation in peroxisomes and cytochromes. While lipid export initially increases, it plateaus and may even decrease with disease progression, sustaining the accumulation of lipids. Fueled by lipo-apoptosis, hepatic steatosis leads to systemic metabolic disarray that adversely affects multiple organs, placing abnormal lipid metabolism associated with NAFLD in close relation to many of the current life-style-related diseases.
| S-EPMC6105174 | biostudies-literature
Project description:AimsMetformin is first-line treatment of type 2 diabetes mellitus and reduces cardiovascular events in patients with insulin resistance and type 2 diabetes. Target tissue for metformin action is thought to be the liver, where metformin distribution depends on facilitated transport by polyspecific transmembrane organic cation transporters (OCTs). Non-alcoholic fatty liver disease (NAFLD) is the most common liver disease in the western world with strong associations to insulin resistance and the metabolic syndrome, but whether NAFLD affects metformin biodistribution to the liver is not known. In this study, the primary aim was to investigate in vivo hepatic uptake of metformin dynamically in humans with variable degrees of liver affection. As a secondary aim, we wished to correlate hepatic metformin distribution with OCT gene transcription determined in diagnostic liver biopsies.MethodsEighteen patients with biopsy-proven NAFLD were investigated using 11C-metformin PET/CT technique. Gene transcripts of OCTs were determined by real-time polymerase chain reaction (PCR).ResultsWe observed similar hepatic volume of distribution of metformin between patients with simple steatosis and non-alcoholic steatohepatitis (NASH) (Vd 2.38 ± 0.56 vs. 2.10 ± 0.39, P = 0.3). There was no association between hepatic exposure to metformin and the degree of inflammation or fibrosis, and no clear correlation between metformin distribution and OCT gene transcription.ConclusionMetformin is distributed to the liver in patients with NAFLD and the distribution is not impaired by inflammation or fibrosis. The findings imply that metformin action in liver in patients with NAFLD may be preserved.
| S-EPMC6624391 | biostudies-literature
Project description:Three-dimensional (3D) cell culture technologies, such as organoids, are physiologically relevant models for basic and clinical applications. Automated microfluidics offers advantages in high-throughput and precision analysis of cells but is not yet compatible with organoids. Here, we present an automated, high-throughput, microfluidic 3D organoid culture and analysis system to facilitate preclinical research and personalized therapies. Our system provides combinatorial and dynamic drug treatments to hundreds of cultures and enables real-time analysis of organoids. We validate our system by performing individual, combinatorial, and sequential drug screens on human-derived pancreatic tumor organoids. We observe significant differences in the response of individual patient-based organoids to drug treatments and find that temporally-modified drug treatments can be more effective than constant-dose monotherapy or combination therapy in vitro. This integrated platform advances organoids models to screen and mirror real patient treatment courses with potential to facilitate treatment decisions for personalized therapy.
| S-EPMC7573629 | biostudies-literature
Project description:Nonalcoholic fatty liver disease (NAFLD) is considered the next major health epidemic with an estimated 25% worldwide prevalence. No drugs have yet been approved and NAFLD remains a major unmet need. Here, we identify MCJ (Methylation-Controlled J protein) as a target for non-alcoholic steatohepatitis (NASH), an advanced phase of NAFLD. MCJ is an endogenous negative regulator of the respiratory chain Complex I that acts to restrain mitochondrial respiration. We show that therapeutic targeting of MCJ in the liver with nanoparticle- and GalNAc-formulated siRNA efficiently reduces liver lipid accumulation and fibrosis in multiple NASH mouse models. Decreasing MCJ expression enhances the capacity of hepatocytes to mediate β-oxidation of fatty acids and minimizes lipid accumulation, which results in reduced hepatocyte damage and fibrosis. Moreover, MCJ levels in the liver of NAFLD patients are elevated relative to healthy subjects. Thus, inhibition of MCJ emerges as an alternative approach to treat NAFLD.
| S-EPMC7334216 | biostudies-literature
Project description:Telomere length has been linked to the prevalence and progression of metabolic disease. However, clinical implications of telomere length in biopsy-proven non-alcoholic fatty liver disease (NAFLD) patients remain unclear. Therefore, this study aimed to investigate the association of telomere length with the histological severity of NAFLD. The cross-sectional data derived from the prospectively enrolled Boramae NAFLD registry (n = 91) were analyzed. The liver tissues and clinical information were obtained from both NAFLD patients and non-NAFLD subjects. Binary logistic regression was performed to identify the independent association between telomere length and the histological severity of NAFLD. A total of 83 subjects with or without biopsy-proven NAFLD were included for analysis: non-NAFLD in 23 (27.7%), non-alcoholic fatty liver in 15 (18.1%), and non-alcoholic steatohepatitis (NASH) in 45 (54.2%). Telomere length measured from liver tissues showed a strong negative correlation (p < 0.001) with age, regardless of NAFLD status. Therefore, telomere length was corrected for age. Age-adjusted telomere length than decreased gradually with an increasing severity of fibrosis in patients with NAFLD (p < 0.028). In multivariate analysis, age-adjusted telomere length (odds ratio [OR] 0.59; 95% CI 0.37-0.92; p = 0.019) and high-density lipoprotein cholesterol (OR 0.94; 95% CI 0.80-0.99; p = 0.039) were independently associated with significant fibrosis. The age-adjusted telomere length tends to decrease along with the fibrosis stage of NAFLD. In particular, among the histological components of NAFLD, fibrosis severity seems to be related to telomere length in the liver.
| S-EPMC8429461 | biostudies-literature
Project description:ObjectiveIncreased hepatic expression of dipeptidyl peptidase 4 (DPP4) is associated with non-alcoholic fatty liver disease (NAFLD). Whether this is causative for the development of NAFLD is not yet clarified. Here we investigate the effect of hepatic DPP4 overexpression on the development of liver steatosis in a mouse model of diet-induced obesity.MethodsPlasma DPP4 activity of subjects with or without NAFLD was analyzed. Wild-type (WT) and liver-specific Dpp4 transgenic mice (Dpp4-Liv-Tg) were fed a high-fat diet and characterized for body weight, body composition, hepatic fat content and insulin sensitivity. In vitro experiments on HepG2 cells and primary mouse hepatocytes were conducted to validate cell autonomous effects of DPP4 on lipid storage and insulin sensitivity.ResultsSubjects suffering from insulin resistance and NAFLD show an increased plasma DPP4 activity when compared to healthy controls. Analysis of Dpp4-Liv-Tg mice revealed elevated systemic DPP4 activity and diminished active GLP-1 levels. They furthermore show increased body weight, fat mass, adipose tissue inflammation, hepatic steatosis, liver damage and hypercholesterolemia. These effects were accompanied by increased expression of PPARγ and CD36 as well as severe insulin resistance in the liver. In agreement, treatment of HepG2 cells and primary hepatocytes with physiological concentrations of DPP4 resulted in impaired insulin sensitivity independent of lipid content.ConclusionsOur results give evidence that elevated expression of DPP4 in the liver promotes NAFLD and insulin resistance. This is linked to reduced levels of active GLP-1, but also to auto- and paracrine effects of DPP4 on hepatic insulin signaling.
| S-EPMC5641684 | biostudies-literature
Project description:Despite considerable efforts in modeling liver disease in vitro, it remains difficult to recapitulate the pathogenesis of the advanced phases of non-alcoholic fatty liver disease (NAFLD) with inflammation and fibrosis. Here, a liver-on-a-chip platform with bioengineered multicellular liver microtissues is developed, composed of four major types of liver cells (hepatocytes, endothelial cells, Kupffer cells, and stellate cells) to implement a human hepatic fibrosis model driven by NAFLD: i) lipid accumulation in hepatocytes (steatosis), ii) neovascularization by endothelial cells, iii) inflammation by activated Kupffer cells (steatohepatitis), and iv) extracellular matrix deposition by activated stellate cells (fibrosis). In this model, the presence of stellate cells in the liver-on-a-chip model with fat supplementation showed elevated inflammatory responses and fibrosis marker up-regulation. Compared to transforming growth factor-beta-induced hepatic fibrosis models, this model includes the native pathological and chronological steps of NAFLD which shows i) higher fibrotic phenotypes, ii) increased expression of fibrosis markers, and iii) efficient drug transport and metabolism. Taken together, the proposed platform will enable a better understanding of the mechanisms underlying fibrosis progression in NAFLD as well as the identification of new drugs for the different stages of NAFLD.
| S-EPMC8035291 | biostudies-literature
Project description:Upper tract urothelial carcinomas (UTUCs) are rare entities that are usually diagnosed at advanced stages. Research on UTUC pathobiology and clinical management has been hampered by the lack of models accurately reflecting disease nature and diversity. In this study, a modified organoid culture system is used to generate a library of 25 patient-derived UTUC organoid lines retaining the histological architectures, marker gene expressions, genomic landscapes, and gene expression profiles of their parental tumors. The study demonstrates that the responses of UTUC organoids to anticancer drugs can be identified and the model supports the exploration of novel treatment strategies. This work proposes a modified protocol for generating patient-derived UTUC organoid lines that may help elucidate UTUC pathophysiology and assess the responses of these diseases to various drug therapies in personalized medicine.
| S-EPMC8811809 | biostudies-literature
Project description:Background and aimsBrain metastases are prevalent in the late stages of malignant melanoma. Multimodal therapy remains challenging. Patient-derived organoids (PDOs) represent a valuable pre-clinical model, faithfully recapitulating key aspects of the original tumor, including the heterogeneity and the mutational status. This study aimed to establish PDOs from melanoma brain metastases (MBM-PDOs) and to test the feasibility of using them as a model for in vitro targeted-therapy drug testing.MethodsSurgical resection samples from eight patients with melanoma brain metastases were used to establish MBM-PDOs. The samples were enzymatically dissociated followed by seeding into low-attachment plates to generate floating organoids. The MBM-PDOs were characterized genetically, histologically, and immunohistologically and compared with the parental tissue. The MBM-PDO cultures were exposed to dabrafenib (BRAF inhibitor) and trametinib (MEK inhibitor) followed by a cell viability assessment.ResultsSeven out of eight cases were successfully cultivated, maintaining the histological, immunohistological phenotype, and the mutational status of the parental tumors. Five out of seven cases harbored BRAF V600E mutations and were responsive to BRAF and MEK inhibitors in vitro. Two out of seven cases were BRAF wild type: one case harboring an NRAS mutation and the other harboring a KIT mutation, and both were resistant to BRAF and MEK inhibitor therapy.ConclusionsWe successfully established PDOs from melanoma brain metastases surgical specimens, which exhibited a consistent histological and mutational profile with the parental tissue. Using FDA-approved BRAF and MEK inhibitors, our data demonstrate the feasibility of employing MBM-PDOs for targeted-therapy in vitro testing.
| S-EPMC11360789 | biostudies-literature