Investigation the effects of vitreous humor on proliferation and dedifferentiation of differentiated NTERA2 cells

Mammals have a limited capacity to regenerate their tissues and organs. One of the mechanisms associated with natural regeneration is dedifferentiation. Several small molecules such as vitamin C and growth factors could improve reprogramming efficiency. In this study, the NTERA2-D1 (NT2) cells were induced towards differentiation (NT2-RA) with 10-5 M retinoic acid (RA) for three days and then subjected to various amounts of vitreous humor (VH). Results show that the growth rate of these cells was reduced, while this rate was partly restored upon treatment with VH (NT2-RA-VH). Cell cycle analysis with PI method also showed that the numbers of cells at the S phase of the cell cycle in these cells were increased. The levels of SSEA3 and TRA-1-81 antigens in NT2-RA were dropped but they increased in NT2- RA-VH to a level similar to the NT2 cells. The level of SSEA1 had an opposite pattern. Expression of OCT4 gene dropped after RA treatment, but it was recovered in NT2-RA-VH cells. In conclusion, we suggest VH as a potent mixture for improving the cellular reprogramming leading to dedifferentiation.

SYNCRIP Modulates the Epithelial-Mesenchymal Transition in Hepatocytes and HCC Cells

Heterogeneous nuclear ribonucleoproteins (hnRNPs) control gene expression by acting at multiple levels and are often deregulated in epithelial tumors; however, their roles in the fine regulation of cellular reprogramming, specifically in epithelial–mesenchymal transition (EMT), remain largely unknown. Here, we focused on the hnRNP-Q (also known as SYNCRIP), showing by molecular analysis that in hepatocytes it acts as a “mesenchymal” gene, being induced by TGFβ and modulating the EMT. SYNCRIP silencing limits the induction of the mesenchymal program and maintains the epithelial phenotype. Notably, in HCC invasive cells, SYNCRIP knockdown induces a mesenchymal–epithelial transition (MET), negatively regulating their mesenchymal phenotype and significantly impairing their migratory capacity. In exploring possible molecular mechanisms underlying these observations, we identified a set of miRNAs (i.e., miR-181-a1-3p, miR-181-b1-3p, miR-122-5p, miR-200a-5p, and miR-let7g-5p), previously shown to exert pro- or anti-EMT activities, significantly impacted by SYNCRIP interference during EMT/MET dynamics and gathered insights, suggesting the possible involvement of this RNA binding protein in their transcriptional regulation.

CellRank for directed single-cell fate mapping

Computational trajectory inference enables the reconstruction of cell state dynamics from single-cell RNA sequencing experiments. However, trajectory inference requires that the direction of a biological process is known, largely limiting its application to differentiating systems in normal development. Here, we present CellRank (https://cellrank.org) for single-cell fate mapping in diverse scenarios, including regeneration, reprogramming and disease, for which direction is unknown. Our approach combines the robustness of trajectory inference with directional information from RNA velocity, taking into account the gradual and stochastic nature of cellular fate decisions, as well as uncertainty in velocity vectors. On pancreas development data, CellRank automatically detects initial, intermediate and terminal populations, predicts fate potentials and visualizes continuous gene expression trends along individual lineages. Applied to lineage-traced cellular reprogramming data, predicted fate probabilities correctly recover reprogramming outcomes. CellRank also predicts a new dedifferentiation trajectory during postinjury lung regeneration, including previously unknown intermediate cell states, which we confirm experimentally.

Hepatocyte generation in liver homeostasis, repair, and regeneration

The liver has remarkable capability to regenerate, employing mechanism to ensure the stable liver-to-bodyweight ratio for body homeostasis. The source of this regenerative capacity has received great attention over the past decade yet still remained controversial currently. Deciphering the sources for hepatocytes provides the basis for understanding tissue regeneration and repair, and also illustrates new potential therapeutic targets for treating liver diseases. In this review, we describe recent advances in genetic lineage tracing studies over liver stem cells, hepatocyte proliferation, and cell lineage conversions or cellular reprogramming. This review will also evaluate the technical strengths and limitations of methods used for studies on hepatocyte generation and cell fate plasticity in liver homeostasis, repair and regeneration.

NETISCE: A Network-Based Tool for Cell Fate Reprogramming

The search for effective therapeutic targets in fields like regenerative medicine and cancer research has generated interest in cell fate reprogramming. This cellular reprogramming paradigm can drive cells to a desired target state from any initial state. However, methods for identifying reprogramming targets remain limited for biological systems that lack large sets of experimental data or a dynamical characterization. We present NETISCE, a novel computational tool for identifying cell fate reprogramming targets in static networks. NETISCE identifies reprogramming targets through the innovative use of control theory within a dynamical systems framework. Through validations in studies of cell fate reprogramming from developmental, stem cell, and cancer biology, we show that NETISCE can predict previously identified cell fate reprogramming targets and identify potentially novel combinations of targets. NETISCE extends cell fate reprogramming studies to larger-scale biological networks without the need for full model parameterization and can be implemented by experimental and computational biologists to identify parts of a biological system that are relevant for the desired reprogramming task.

Extracellular matrix guidance of autophagy: a mechanism regulating cancer growth

The extracellular matrix (ECM) exists as a dynamic network of biophysical and biochemical factors that maintain tissue homeostasis. Given its sensitivity to changes in the intra- and extracellular space, the plasticity of the ECM can be pathological in driving disease through aberrant matrix remodelling. In particular, cancer uses the matrix for its proliferation, angiogenesis, cellular reprogramming and metastatic spread. An emerging field of matrix biology focuses on proteoglycans that regulate autophagy, an intracellular process that plays both critical and contextual roles in cancer. Here, we review the most prominent autophagic modulators from the matrix and the current understanding of the cellular pathways and signalling cascades that mechanistically drive their autophagic function. We then critically assess how their autophagic functions influence tumorigenesis, emphasizing the complexities and stage-dependent nature of this relationship in cancer. We highlight novel emerging data on immunoglobulin-containing and proline-rich receptor-1, heparanase and thrombospondin 1 in autophagy and cancer. Finally, we further discuss the pro- and anti-autophagic modulators originating from the ECM, as well as how these proteoglycans and other matrix constituents specifically influence cancer progression.

Transcriptomic-Based Quantification of the Epithelial-Hybrid-Mesenchymal Spectrum Across Biological Contexts

Epithelial-mesenchymal plasticity (EMP) underlies embryonic development, wound healing, and cancer metastasis and fibrosis. Cancer cells exhibiting EMP often have more aggressive behavior, characterized by drug resistance, and tumor-initiating and immuno-evasive traits. Thus, the EMP status of cancer cells can be a critical indicator of patient prognosis. Here, we compare three distinct transcriptomic-based metrics—each derived using a different gene list and algorithm—that quantify the EMP spectrum. Our results for over 80 cancer-related RNA-seq datasets reveal a high degree of concordance among these metrics in quantifying the extent of EMP. Moreover, each metric, despite being trained on cancer expression profiles, recapitulates the expected changes in EMP scores for non-cancer contexts such as lung fibrosis and cellular reprogramming into induced pluripotent stem cells. Thus, we offer a scoring platform to quantify the extent of EMP in vitro and in vivo for diverse biological applications including cancer.

Directly Reprogrammed Human Neurons to Understand Age-Related Energy Metabolism Impairment and Mitochondrial Dysfunction in Healthy Aging and Neurodegeneration

Brain aging is characterized by several molecular and cellular changes grouped as the hallmarks or pillars of aging, including organelle dysfunction, metabolic and nutrition-sensor changes, stem cell attrition, and macromolecular damages. Separately and collectively, these features degrade the most critical neuronal function: transmission of information in the brain. It is widely accepted that aging is the leading risk factor contributing to the onset of the most prevalent pathological conditions that affect brain functions, such as Alzheimer’s, Parkinson’s, and Huntington’s disease. One of the limitations in understanding the molecular mechanisms involved in those diseases is the lack of an appropriate cellular model that recapitulates the “aged” context in human neurons. The advent of the cellular reprogramming of somatic cells, i.e., dermal fibroblasts, to obtain directly induced neurons (iNs) and induced pluripotent stem cell- (iPSC-) derived neurons is technical sound advances that could open the avenues to understand better the contribution of aging toward neurodegeneration. In this review, we will summarize the commonalities and singularities of these two approaches for the study of brain aging, with an emphasis on the role of mitochondrial dysfunction and redox biology. We will address the evidence showing that iNs retain age-related features in contrast to iPSC-derived neurons that lose the aging signatures during the reprogramming to pluripotency, rendering iNs a powerful strategy to deepen our knowledge of the processes driving normal cellular function decline and neurodegeneration in a human adult model. We will finally discuss the potential utilization of these novel technologies to understand the differential contribution of genetic and epigenetic factors toward neuronal aging, to identify and develop new drugs and therapeutic strategies.

The Profile of Expression of SG2NAs Differs Between Cancer Types and it is Involved in the Reprogramming of Tumour Cell Proteome.

Striatin and SG2NA are scaffold proteins that from signalling complexes called STRIPAK. It has been associated with cancer and other diseases. Our earlier studies have shown that SG2NA forms a complex with the cancer associate protein DJ-1 and signalling kinase Akt, promoting cancer cell survival. In the present study, we used bioinformatics analyses to confirm the existence of two isoforms of human SG2NA i.e., 78 and 87 kDas. In addition, several smaller isoforms like 35 kDa were also seen in western blot analyses of human cell lysates. The expression of these isoforms varies between different human cancer cell lines. Also, the protein level does not corroborate with its transcript level, suggesting a complex regulation of its expression. In breast tumour tissues, the expression of the 35 and 78 kDa isoforms was higher as compared to the adjacent normal tissues, while the 87 kDa isoform was detected in the breast tumour tissues only. With the progression of stages of breast cancer, the expression of 78 kDa isoform decreased, while 87 kDa became undetectable. In coimmunoprecipitation assay, the profile of SG2NA interactome in breast tumor vis-à-vis adjacent normal breast tissues shows hundreds of common proteins, while some proteins specifically interacted in breast tumour tissue only. We conclude that SG2NA is involve in diverse cellular pathways and have roles in cellular reprogramming during tumorigenesis.