http://cmotifs.tchlab.org.
BackgroundLiver regeneration is inhibited by chronic ethanol consumption and this impaired repair response may contribute to the risk for alcoholic liver disease. We developed and applied a novel data analysis approach to assess the effect of chronic ethanol intake in the mechanisms responsible for liver regeneration. We performed a time series transcriptomic profiling study of the regeneration response after 2/3rd partial hepatectomy (PHx) in ethanol-fed and isocaloric control rats.ResultsWe developed a novel data analysis approach focusing on comparative pattern counts (COMPACT) to exhaustively identify the dominant and subtle differential expression patterns. Approximately 6500 genes were differentially regulated in Ethanol or Control groups within 24 h after PHx. Adaptation to chronic ethanol intake significantly altered the immediate early gene expression patterns and nearly completely abrogated the cell cycle induction in hepatocytes post PHx. The patterns highlighted by COMPACT analysis contained several non-parenchymal cell specific markers indicating their aberrant transcriptional response as a novel mechanism through which chronic ethanol intake deregulates the integrated liver tissue response.ConclusionsOur novel comparative pattern analysis revealed new insights into ethanol-mediated molecular changes in non-parenchymal liver cells as a possible contribution to the defective liver regeneration phenotype. The results revealed for the first time an ethanol-induced shift of hepatic stellate cells from a pro-regenerative phenotype to that of an anti-regenerative state after PHx. Our results can form the basis for novel interventions targeting the non-parenchymal cells in normalizing the dysfunctional repair response process in alcoholic liver disease. Our approach is illustrated online at http://compact.jefferson.edu.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-2492-x) contains supplementary material, which is available to authorized users.
Highlights Spatially-tracked single neuron transcriptomics map of intrinsic cardiac ganglia Distinct composition of intrinsic cardiac neurons compared to the central neurons Correlation of cholinergic and catecholaminergic markers suggesting neuroplasticity Adaptive and dynamic system driven by putative paracrine neuromodulatory networks
Following partial hepatectomy, a coordinated series of molecular events occurs to regulate hepatocyte entry into the cell cycle to recover lost mass. In rats during the first 6 h following resection, hepatocytes are primed by a tightly controlled cytokine response to prepare hepatocytes to begin replication. Although it appears to be a critical element driving regeneration, the cytokine response to resection has not yet been fully characterized. Specifically, the role of one of the key response elements to cytokine signaling (NF-κB) remains incompletely characterized. In this study, we present a novel, genome-wide, pattern-based analysis characterizing NF-κB binding during the priming phase of liver regeneration. We interrogated the dynamic regulation of priming by NF-κB through categorizing NF-κB binding in different temporal profiles: immediate sustained response, early transient response, and delayed response to partial hepatectomy. We then identified functional regulation of NF-κB binding by relating the temporal response profile to differential gene expression. We found that NF-κB bound genes govern negative regulation of cell growth and inflammatory response immediately following hepatectomy. NF-κB also transiently regulates genes responsible for lipid biosynthesis and transport as well as induction of apoptosis following hepatectomy. By the end of the priming phase, NF-κB regulation of genes involved in inflammatory response, negative regulation of cell death, and extracellular structure organization became prominent. These results suggest that NF-κB regulates target genes through binding and unbinding in immediate, transient, and delayed patterns. Such dynamic switch-like patterns of NF-κB binding may govern different functional transitions that drive the onset of regeneration.
NF-κB is a major inflammatory response mediator in the liver, playing a key role in the pathogenesis of alcoholic liver injury. We investigated zonal as well as liver cell type-specific distribution of NF-κB activation across the liver acinus following adaptation to chronic ethanol intake and 70% partial hepatectomy (PHx). We employed immunofluorescence staining, digital image analysis and statistical distributional analysis to quantify subcellular localization of NF-κB in hepatocytes and hepatic stellate cells (HSCs). We detected significant spatial heterogeneity of NF-κB expression and cellular localization between cytoplasm and nucleus across liver tissue. Our main aims involved investigating the zonal bias in NF-κB localization and determining to what extent chronic ethanol intake affects this zonal bias with in hepatocytes at baseline and post-PHx. Hepatocytes in the periportal area showed higher NF-κB expression than in the pericentral region in the carbohydrate-fed controls, but not in the ethanol group. However, the distribution of NF-κB nuclear localization in hepatocytes was shifted towards higher levels in pericentral region than in periportal area, across all treatment conditions. Chronic ethanol intake shifted the NF-κB distribution towards higher nuclear fraction in hepatocytes as compared to the pair-fed control group. Ethanol also stimulated higher NF-κB expression in a subpopulation of HSCs. In the control group, PHx elicited a shift towards higher NF-κB nuclear fraction in hepatocytes. However, this distribution remained unchanged in the ethanol group post-PHx. HSCs showed a lower NF-κB expression following PHx in both ethanol and control groups. We conclude that adaptation to chronic ethanol intake attenuates the liver zonal variation in NF-κB expression and limits the PHx-induced NF-κB activation in hepatocytes, but does not alter the NF-κB expression changes in HSCs in response to PHx. Our findings provide new insights as to how ethanol treatment may affect cell-type specific processes regulated by NF-κB activation in liver cells.
Activation and deactivation of hepatic stellate cells (HSCs) is an important mechanism contributing to both healthy liver function and development of liver diseases, which relies on the interplay between numerous signaling pathways. There is accumulating evidence for the regulatory role of microRNAs that are downstream from these pathways in HSC activation. However, the relative contribution of these pathways and interacting microRNA regulators to the activation process is unknown. We pursued a computational modeling approach to explore the timing and regulatory balances that are critical to HSC activation and quiescence. We developed an integrated model incorporating three signaling pathways with crosstalk (NF-κB, STAT3 and TGF-β) and two microRNAs (miR-146a, miR-21) that are differentially regulated by these pathways. Simulations demonstrated that TGF-β-mediated regulation of microRNAs is critical to drive the HSC phenotypic switch from quiescence (miR-146a high miR-21 low) to an activated state (miR-146a low miR-21 high). We found that the relative timing between peak NF-κB and STAT3 activation plays a key role driving the initial dynamics of miR-146a. We observed re-quiescence from the activated
Building on the observation that chromatin compaction can be locally modulated by activity, we propose a model of in vivo chromatin as an active polymer and study its large scale conformations. In particular, we study an active mechanochemical model of chromosomal folding based on the interplay among polymer elasticity, confinement, topological constraints, and fluctuating active stresses arising from the ATP-dependent action of a variety of chromatin-associated protein machines and chromatin-remodeling proteins and their stochastic turnover. We find that activity drives the chromatin to a nonequilibrium steady state; the statistics of conformations in this nonequilibrium steady state are consistent with recent measurements on intrachromosomal contact probabilities and chromosomal compaction. The contact exponents at steady state show a systematic variation with changes in the nature of activity and the rates of turnover. The steady state configuration of the active chromatin in two dimensions resembles a space-filling Peano curve, which might have implications for the optimization of genome information storage.
Chronic ethanol intake impairs liver regeneration through a system-wide alteration in the regulatory networks driving the response to injury. Our study focused on the initial phase of response to 2/3rd partial hepatectomy (PHx) to investigate how adaptation to chronic ethanol intake affects the genome-wide binding profiles of the transcription factors C/EBP-β and C/EBP-α. These factors participate in complementary and often opposing functions for maintaining cellular differentiation, regulating metabolism, and governing cell growth during liver regeneration. We analyzed ChIP-seq data with a comparative pattern count (COMPACT) analysis, which exhaustively enumerates temporal patterns of discretized binding profiles to identify dominant as well as subtle patterns that may not be apparent from conventional clustering analyses. We found that adaptation to chronic ethanol intake significantly alters the genome-wide binding profile of C/EBP-β and C/EBP-α before and following PHx. A subset of these ethanol-induced changes include C/EBP-β binding to promoters of genes involved in the profibrogenic transforming growth factor-β pathway, and both C/EBP-β and C/EBP-α binding to promoters of genes involved in the cell cycle, apoptosis, homeostasis, and metabolic processes. The shift in C/EBP binding loci, coupled with an ethanol-induced increase in C/EBP-β binding at 6 h post-resection, indicates that ethanol adaptation may change both the amount and nature of C/EBP binding postresection. Taken together, our results suggest that chronic ethanol consumption leads to a spatially and temporally reorganized activity at many genomic loci, resulting in a shift in the dynamic balance and coordination of cellular processes underlying regenerative response.
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