disease modeling
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Although most same-stage non-alcoholic fatty liver disease (NAFLD) patients exhibit similar histologic sequelae, the underlying mechanisms appear to be highly heterogeneous. Therefore, it was recently proposed to redefine NAFLD to metabolic dysfunction-associated fatty liver disease (MAFLD) in which other known causes of liver disease such as alcohol consumption or viral hepatitis do not need to be excluded. Revised nomenclature envisions speeding up and facilitating anti-MAFLD drug development by means of patient stratification whereby each subgroup would benefit from distinct pharmacological interventions. As human-based in vitro research fulfils an irrefutable step in drug development, action should be taken as well in this stadium of the translational path. Indeed, most established in vitro NAFLD models rely on short-term exposure to fatty acids and use lipid accumulation as a phenotypic benchmark. This general approach to a seemingly ambiguous disease such as NAFLD therefore no longer seems applicable. Human-based in vitro models that accurately reflect distinct disease subgroups of MAFLD should thus be adopted in early preclinical disease modeling and drug testing. In this review article, we outline considerations for setting up translational in vitro experiments in the MAFLD era and allude to potential strategies to implement MAFLD heterogeneity into an in vitro setting so as to better align early drug development with future clinical trial designs.

There is an urgent need to understand how SARS-CoV-2 infects the airway epithelium and in a subset of individuals leads to severe illness or death. Induced pluripotent stem cells (iPSCs) provide a near limitless supply of human cells that can be differentiated into cell types of interest, including airway epithelium, for disease modeling. We present a human iPSC-derived airway epithelial platform, composed of the major airway epithelial cell types, that is permissive to SARS-CoV-2 infection. Subsets of iPSC-airway cells express the SARS-CoV-2 entry factors ACE2 and TMPRSS2. Multiciliated cells are the primary initial target of SARS-CoV-2 infection. Upon infection with SARS-CoV-2, iPSC-airway cells generate robust interferon and inflammatory responses and treatment with remdesivir or camostat methylate causes a decrease in viral propagation and entry, respectively. In conclusion, iPSC-derived airway cells provide a physiologically relevant in vitro model system to interrogate the pathogenesis of, and develop treatment strategies for, COVID-19 pneumonia.

Around one third of patients with mitochondrial disorders develop a kind of cardiomyopathy. In these cases, severity is quite variable ranging from asymptomatic status to severe manifestations including heart failure, arrhythmias, and sudden cardiac death. ATP is primarily generated in the mitochondrial respiratory chain via oxidative phosphorylation by utilizing fatty acids and carbohydrates. Genes in both the nuclear and the mitochondrial DNA encode components of this metabolic route and, although mutations in these genes are extremely rare, the risk to develop cardiac symptoms is significantly higher in this patient cohort. Additionally, infants with cardiovascular compromise in mitochondrial deficiency display a worse late survival compared to patients without cardiac symptoms. At this point, the mechanisms behind cardiac disease progression related to mitochondrial gene mutations are poorly understood and current therapies are unable to substantially restore the cardiac performance and to reduce the disease burden. Therefore, new strategies are needed to uncover the pathophysiological mechanisms and to identify new therapeutic options for mitochondrial cardiomyopathies. Here, human induced pluripotent stem cell (iPSC) technology has emerged to provide a suitable patient-specific model system by recapitulating major characteristics of the disease in vitro, as well as to offer a powerful platform for pre-clinical drug development and for the testing of novel therapeutic options. In the present review, we summarize recent advances in iPSC-based disease modeling of mitochondrial cardiomyopathies and explore the patho-mechanistic insights as well as new therapeutic approaches that were uncovered with this experimental platform. Further, we discuss the challenges and limitations of this technology and provide an overview of the latest techniques to promote metabolic and functional maturation of iPSC-derived cardiomyocytes that might be necessary for modeling of mitochondrial disorders.

Abstract Background Spinal cord motor neurons (MNs) from human iPS cells (iPSCs) have wide applications in disease modeling and therapeutic development for amyotrophic lateral sclerosis (ALS) and other MN-associated neurodegenerative diseases. We need highly efficient MN differentiation strategies for generating iPSC-derived disease models that closely recapitulate the genetic and phenotypic complexity of ALS. An important application of these models is to understand molecular mechanisms of action of FDA-approved ALS drugs that only show modest clinical efficacy. Novel mechanistic insights will help us design optimal therapeutic strategies together with predictive biomarkers to achieve better efficacy. Methods We induce efficient MN differentiation from iPSCs in 4 days using synthetic mRNAs coding two transcription factors (Ngn2 and Olig2) with phosphosite modification. These MNs after extensive characterization were applied in electrophysiological and neurotoxicity assays as well as transcriptomic analysis, to study the neuroprotective effect and molecular mechanisms of edaravone, an FDA-approved drug for ALS, for improving its clinical efficacy. Results We generate highly pure and functional mRNA-induced MNs (miMNs) from control and ALS iPSCs, as well as embryonic stem cells. Edaravone alleviates H2O2-induced neurotoxicity and electrophysiological dysfunction in miMNs, demonstrating its neuroprotective effect that was also found in the glutamate-induced miMN neurotoxicity model. Guided by the transcriptomic analysis, we show a previously unrecognized effect of edaravone to induce the GDNF receptor RET and the GDNF/RET neurotrophic signaling in vitro and in vivo, suggesting a clinically translatable strategy to activate this key neuroprotective signaling. Notably, edaravone can replace required neurotrophic factors (BDNF and GDNF) to support long-term miMN survival and maturation, further supporting the neurotrophic function of edaravone-activated signaling. Furthermore, we show that edaravone and GDNF combined treatment more effectively protects miMNs from H2O2-induced neurotoxicity than single treatment, suggesting a potential combination strategy for ALS treatment. Conclusions This study provides methodology to facilitate iPSC differentiation and disease modeling. Our discoveries will facilitate the development of optimal edaravone-based therapies for ALS and potentially other neurodegenerative diseases. Graphical abstract

AbstractHuman pluripotent stem cells (hPSCs) have great potential for disease modeling, drug discovery, and regenerative medicine as they can differentiate into many different functional cell types via directed differentiation. However, the application of disease modeling is limited due to a time-consuming and labor-intensive process of introducing known pathogenic mutations into hPSCs. Base editing is a newly developed technology that enables the facile introduction of point mutations into specific loci within the genome of living cells without unwanted genome injured. We describe an optimized stepwise protocol to introduce disease-specific mutations of long QT syndrome (LQTs) into hPSCs. We highlight technical issues, especially those associated with introducing a point mutation to obtain isogenic hPSCs without inserting any resistance cassette and reproducible cardiomyocyte differentiation. Based on the protocol, we succeeded in getting hPSCs carrying LQTs pathogenic mutation with excellent efficiency (31.7% of heterozygous clones, 9.1% of homozygous clones) in less than 20 days. In addition, we also provide protocols to analyze electrophysiological of hPSC-derived cardiomyocytes using multi-electrode arrays. This protocol is also applicable to introduce other disease-specific mutations into hPSCs. Graphical abstract

Infectious disease transmission is a nonlinear process with complex, sometimes unintuitive dynamics. Modeling can transform information about a disease process and its parameters into quantitative projections that help decision makers compare public health response options. However, modelers face methodologic challenges, data challenges, and communication challenges, which are exacerbated under the time constraints of a public health emergency. We review methods, applications, challenges and opportunities for real-time infectious disease modeling during public health emergencies, with examples drawn from the two deadliest pandemics in recent history: HIV/AIDS and coronavirus disease 2019 (COVID-19). Expected final online publication date for the Annual Review of Public Health, Volume 43 is April 2022. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

Transdifferentiation is the process of converting terminally differentiated cells to another cell type. Being less time-consuming and free from tumorigenesis, it is a promising alternative to directed differentiation, which provides cell sources for tissue regeneration therapy and disease modeling. In the past decades, transdifferentiation was found to happen within or across the cell lineages, being induced by overexpression of key transcription factors, chemical cocktail treatments, etc. Implementing next-generation biotechnologies, such as genome editing tools and scRNA-seq, improves current protocols and has the potential to facilitate discovery in new pathways of transdifferentiation, which will accelerate its application in clinical use.

AbstractStudying the etiology of human neurodevelopmental diseases has long been a challenging task due to the brain’s complexity and its limited accessibility. Human pluripotent stem cells (hPSCs)-derived brain organoids are capable of recapitulating various features and functionalities of the human brain, allowing the investigation of intricate pathogenesis of developmental abnormalities. Over the past years, brain organoids have facilitated identifying disease-associated phenotypes and underlying mechanisms for human neurodevelopmental diseases. Integrating with more cutting-edge technologies, particularly gene editing, brain organoids further empower human disease modeling. Here, we review the latest progress in modeling human neurodevelopmental disorders with brain organoids.

Abstract Background Evolution in the in vitro cell culture from conventional 2D to 3D technique has been a significant accomplishment. The 3D culture models have provided a close and better insight into the physiological study of the human body. The increasing demand for organs like liver, kidney, and pancreas for transplantation, rapid anti-cancer drug screening, and the limitations associated with the use of animal models have attracted the interest of researchers to explore 3D organ culture. Main body Natural, synthetic, and hybrid material-based hydrogels are being used as scaffolds in 3D culture and provide 'close-to-in vivo’ structures. Organoids: the stem cell-derived small size 3D culture systems are now favored due to their ability to mimic the in-vivo conditions of organ or tissue and this characteristic has made it eligible for a variety of clinical applications, drug discovery and regenerative medicine are a few of the many areas of application. The use of animal models for clinical applications has been a long-time ethical and biological challenge to get accurate outcomes. 3D bioprinting has resolved the issue of vascularization in organoid culture to a great extent by its layer-by-layer construction approach. The 3D bioprinted organoids have a popular application in personalized disease modeling and rapid drug development and therapeutics. Short conclusions This review paper, focuses on discussing the novel organoid culture approach, its advantages and limitations, and potential applications in a variety of life science areas namely cancer research, cell therapy, tissue engineering, and personalized medicine and drug discovery. Graphical Abstract

Macrophage colony-stimulating factor receptor (M-CSFR/CSF1R) signaling is crucial for the differentiation, proliferation, and survival of myeloid cells. The CSF1R pathway is a promising therapeutic target in many human diseases, including neurological disorders or cancer. Zebrafish are commonly used for human disease modeling and preclinical therapeutic screening. Therefore, it is necessary to understand the proper function of cytokine signaling in zebrafish to reliably model human-related diseases. Here, we investigate the roles of zebrafish Csf1rs and their ligands - Csf1a, Csf1b and Il34, in embryonic and adult myelopoiesis. The proliferative effect of exogenous Csf1a on embryonic macrophages is connected to both receptors, Csf1ra and Csf1rb, however there is no evident effect of Csf1b in zebrafish embryonic myelopoiesis. Furthermore, we uncover an unknown role of Csf1rb in zebrafish granulopoiesis. Deregulation of Csf1rb signaling leads to failure in myeloid differentiation resulting in neutropenia throughout the whole lifespan. Surprisingly, Il34 signaling through Csf1rb seems to be of high importance as both csf1rbΔ4bp and il34Δ5bp deficient zebrafish larvae lack granulocytes. Our single-cell RNA sequencing analysis of adult whole kidney marrow (WKM) hematopoietic cells suggests that csf1rb is expressed mainly by blood and myeloid progenitors and that the expression of csf1ra and csf1rb is non-overlapping. We point out differentially expressed genes important in hematopoietic cell differentiation and immune response in selected WKM populations. Our findings could improve the understanding of myeloid cell function and lead to the further study of CSF1R pathway deregulation in disease, mostly in cancerogenesis.