Quantitative morphological analysis of Deinococcus radiodurans elucidates complex dose-dependent nucleoid condensation during recovery from ionizing radiation.

The extremophile Deinococcus radiodurans maintains a highly organized and condensed nucleoid as its default state, possibly contributing to its high tolerance to ionizing radiation (IR). Previous studies of the D. radiodurans nucleoid were limited by reliance on manual image annotation and qualitative metrics. Here, we introduce a high-throughput approach to quantify the geometric properties of cells and nucleoids using confocal microscopy, digital reconstructions of cells, and computational modeling. We utilize this novel approach to investigate the dynamic process of nucleoid condensation in response to IR stress. Our quantitative analysis reveals that at the population level, exposure to IR induced nucleoid compaction and decreased the size of D. radiodurans cells. Morphological analysis and clustering identified six distinct sub-populations across all tested experimental conditions. Results indicate that exposure to IR induced fractional redistributions of cells across sub-populations to exhibit morphologies associated with greater nucleoid condensation and decreased the abundance of sub-populations associated with cell division. Nucleoid-associated proteins (NAPs) may link nucleoid compaction and stress tolerance, but their roles in regulating compaction in D. radiodurans are unknown. Imaging of genomic mutants of known and suspected NAPs that contribute to nucleoid condensation found that deletion of nucleic acid-binding proteins, not previously described as NAPs, can remodel the nucleoid by driving condensation or decondensation in the absence of stress and that IR increased the abundance of these morphological states. Thus, our integrated analysis introduces a new methodology for studying environmental influences on bacterial nucleoids and provides an opportunity to further investigate potential regulators of nucleoid condensation.IMPORTANCEDeinococcus radiodurans, an extremophile known for its stress tolerance, constitutively maintains a highly condensed nucleoid. Qualitative studies have described nucleoid behavior under a variety of conditions. However, a lack of quantitative data regarding nucleoid organization and dynamics has limited our understanding of the regulatory mechanisms controlling nucleoid organization in D. radiodurans. Here, we introduce a quantitative approach that enables high-throughput quantitative measurements of subcellular spatial characteristics in bacterial cells. Applying this to wild-type or single-protein-deficient populations of D. radiodurans subjected to ionizing radiation, we identified significant stress-responsive changes in cell shape, nucleoid organization, and morphology. These findings highlight this methodology's adaptability and capacity for quantitatively analyzing the cellular response to stressors for screening cellular proteins involved in bacterial nucleoid organization.

Single-cell morphology encodes functional subtypes of senescence in aging human dermal fibroblasts.

Cellular senescence is an established driver of aging, exhibiting context-dependent phenotypes across multiple biological length-scales. Despite its mechanistic importance, profiling senescence within cell populations is challenging. This is in part due to the limitations of current biomarkers to robustly identify senescent cells across biological settings, and the heterogeneous, non-binary phenotypes exhibited by senescent cells. Using a panel of primary dermal fibroblasts, we combined live single-cell imaging, machine learning, multiple senescence induction conditions, and multiple protein-based senescence biomarkers to show the emergence of functional subtypes of senescence. Leveraging single-cell morphologies, we defined eleven distinct morphology clusters, with the abundance of cells in each cluster being dependent on the mode of senescence induction, the time post-induction, and the age of the donor. Of these eleven clusters, we identified three bona-fide senescence subtypes (C7, C10, C11), with C10 showing the strongest age-dependence across a cohort of fifty aging individuals. To determine the functional significance of these senescence subtypes, we profiled their responses to senotherapies, specifically focusing on Dasatinib + Quercetin (D+Q). Results indicated subtype-dependent responses, with senescent cells in C7 being most responsive to D+Q. Altogether, we provide a robust single-cell framework to identify and classify functional senescence subtypes with applications for next-generation senotherapy screens, and the potential to explain heterogeneous senescence phenotypes across biological settings based on the presence and abundance of distinct senescence subtypes.

A Two-Genome Portrayal of Mitochondrial Disorders: A Review with Clinical Presentations.

Disorders of mitochondrial function are responsible for many inherited neuromuscular and metabolic diseases. Their combination of high mortality, multi-systemic involvement, and economic burden cause devastating effects on patients and their families. Molecular diagnostic tools are becoming increasingly important in providing earlier diagnoses and guiding more precise therapeutic treatments for patients suffering from mitochondrial disorders. This review addresses fundamental molecular concepts relating to the pathogenesis of mitochondrial dysfunction and disorders. A series of short cases highlights the various clinical presentations, inheritance patterns, and pathogenic mutations in nuclear and mitochondrial genes that cause mitochondrial diseases. Graphical and tabular representations of the results are presented to guide the understanding of the important concepts related to mitochondrial molecular genetics and pathology. Emerging technology is incorporating preimplantation genetic testing for mtDNA disorders, while mitochondrial replacement shows promise in significantly decreasing the transfer of diseased mitochondrial DNA (mtDNA) to embryos. Medical professionals must maintain an in-depth understanding of the gene mutations and molecular mechanisms underlying mitochondrial disorders. Continued diagnostic advances and comprehensive management of patients with mitochondrial disorders are essential to achieve robust clinical impacts from comprehensive genomic testing. This is especially true when supported by non-genetic tests such as biochemical analysis, histochemical stains, and imaging studies. Such a multi-pronged investigation should improve the management of mitochondrial disorders by providing accurate and timely diagnoses to reduce disease burden and improve the lives of patients and their families.

Real-time measurements of ATP dynamics via ATeams in Plasmodium falciparum reveal drug-class-specific response patterns.

Malaria tropica, caused by the parasite Plasmodium falciparum (P. falciparum), remains one of the greatest public health burdens for humankind. Due to its pivotal role in parasite survival, the energy metabolism of P. falciparum is an interesting target for drug design. To this end, analysis of the central metabolite adenosine triphosphate (ATP) is of great interest. So far, only cell-disruptive or intensiometric ATP assays have been available in this system, with various drawbacks for mechanistic interpretation and partly inconsistent results. To address this, we have established fluorescent probes, based on Förster resonance energy transfer (FRET) and known as ATeam, for use in blood-stage parasites. ATeams are capable of measuring MgATP2- levels in a ratiometric manner, thereby facilitating in cellulo measurements of ATP dynamics in real-time using fluorescence microscopy and plate reader detection and overcoming many of the obstacles of established ATP analysis methods. Additionally, we established a superfolder variant of the ratiometric pH sensor pHluorin (sfpHluorin) in P. falciparum to monitor pH homeostasis and control for pH fluctuations, which may affect ATeam measurements. We characterized recombinant ATeam and sfpHluorin protein in vitro and stably integrated the sensors into the genome of the P. falciparum NF54attB cell line. Using these new tools, we found distinct sensor response patterns caused by several different drug classes. Arylamino alcohols increased and redox cyclers decreased ATP; doxycycline caused first-cycle cytosol alkalization; and 4-aminoquinolines caused aberrant proteolysis. Our results open up a completely new perspective on drugs' mode of action, with possible implications for target identification and drug development.

Substrate stiffness modulates the emergence and magnitude of senescence phenotypes in dermal fibroblasts.

Cellular senescence is a major driver of aging and disease. Here we show that substrate stiffness modulates the emergence and magnitude of senescence phenotypes after exposure to senescence inducers. Using a primary dermal fibroblast model, we show that decreased substrate stiffness accelerates senescence-associated cell-cycle arrest and regulates the expression of conventional protein-based biomarkers of senescence. We found that the expression of these senescence biomarkers, namely p21WAF1/CIP1 and p16INK4a are mechanosensitive and are in-part regulated by myosin contractility through focal adhesion kinase (FAK)-ROCK signaling. Interestingly, at the protein level senescence-induced dermal fibroblasts on soft substrates (0.5 kPa) do not express p21WAF1/CIP1 and p16INK4a at comparable levels to induced cells on stiff substrates (4GPa). However, cells express CDKN1a, CDKN2a, and IL6 at the RNA level across both stiff and soft substrates. Moreover, when cells are transferred from soft to stiff substrates, senescent cells recover an elevated expression of p21WAF1/CIP1 and p16INK4a at levels comparable to senescence cells on stiff substrates, pointing to a mechanosensitive regulation of the senescence phenotype. Together, our results indicate that the emergent senescence phenotype depends critically on the local mechanical environments of cells and that senescent cells actively respond to changing mechanical cues.

Particulate matter composition drives differential molecular and morphological responses in lung epithelial cells.

Particulate matter (PM) is a ubiquitous component of air pollution that is epidemiologically linked to human pulmonary diseases. PM chemical composition varies widely, and the development of high-throughput experimental techniques enables direct profiling of cellular effects using compositionally unique PM mixtures. Here, we show that in a human bronchial epithelial cell model, exposure to three chemically distinct PM mixtures drive unique cell viability patterns, transcriptional remodeling, and the emergence of distinct morphological subtypes. Specifically, PM mixtures modulate cell viability, DNA damage responses, and induce the remodeling of gene expression associated with cell morphology, extracellular matrix organization, and cellular motility. Profiling cellular responses showed that cell morphologies change in a PM composition-dependent manner. Finally, we observed that PM mixtures with higher cadmium content induced increased DNA damage and drove redistribution among morphological subtypes. Our results demonstrate that quantitative measurement of individual cellular morphologies provides a robust, high-throughput approach to gauge the effects of environmental stressors on biological systems and score cellular susceptibilities to pollution.

Particulate matter composition drives differential molecular and morphological responses in lung epithelial cells.

Particulate matter (PM) is a ubiquitous component of indoor and outdoor air pollution that is epidemiologically linked to many human pulmonary diseases. PM has many emission sources, making it challenging to understand the biological effects of exposure due to the high variance in chemical composition. However, the effects of compositionally unique particulate matter mixtures on cells have not been analyzed using both biophysical and biomolecular approaches. Here, we show that in a human bronchial epithelial cell model (BEAS-2B), exposure to three chemically distinct PM mixtures drives unique cell viability patterns, transcriptional remodeling, and the emergence of distinct morphological subtypes. Specifically, PM mixtures modulate cell viability and DNA damage responses and induce the remodeling of gene expression associated with cell morphology, extracellular matrix organization and structure, and cellular motility. Profiling cellular responses showed that cell morphologies change in a PM composition-dependent manner. Lastly, we observed that particulate matter mixtures with high contents of heavy metals, such as cadmium and lead, induced larger drops in viability, increased DNA damage, and drove a redistribution among morphological subtypes. Our results demonstrate that quantitative measurement of cellular morphology provides a robust approach to gauge the effects of environmental stressors on biological systems and determine cellular susceptibilities to pollution.

Thallium toxicity to temperate and tropical marine organisms: Derivation of water quality guidelines to protect marine life.

A lack of thallium (Tl) toxicity data for marine organisms has hampered the development of water quality guidelines for protecting marine life and assessing ecological hazard/risk. This study assessed the toxicity (EC10/EC50) of Tl in natural seawater (salinity 34 psu and pH 8.05) to 26 functionally diverse marine organisms (19 phyla from five trophic levels) from a variety of temperate and tropical coastal marine habitats. EC10 values ranged from 3.0 µg/L (copepod, Acartia tranteri) to 489 µg/L (cyanobacterium, Cyanobium sp.), while EC50 values ranged from 9.7 µg/L to 1550 µg/L. Thallium(I) was the dominant (86-99 %) oxidation state in test waters across the range of EC10 and EC50 values. Thallium toxicity (EC10/EC50) did not differ between temperate and tropical marine organisms. New, reliable, long-term Tl water quality guidelines were derived using species sensitivity distributions (with model-averaging) to protect marine life in Australia (e.g., 3.9 µg/L for 95 % species protection).

Changes in K+ Concentration as a Signaling Mechanism in the Apicomplexa Parasites Plasmodium and Toxoplasma.

During their life cycle, apicomplexan parasites pass through different microenvironments and encounter a range of ion concentrations. The discovery that the GPCR-like SR25 in Plasmodium falciparum is activated by a shift in potassium concentration indicates that the parasite can take advantage of its development by sensing different ionic concentrations in the external milieu. This pathway involves the activation of phospholipase C and an increase in cytosolic calcium. In the present report, we summarize the information available in the literature regarding the role of potassium ions during parasite development. A deeper understanding of the mechanisms that allow the parasite to cope with ionic potassium changes contributes to our knowledge about the cell cycle of Plasmodium spp.

Senescent stroma induces nuclear deformations in cancer cells via the inhibition of RhoA/ROCK/myosin II-based cytoskeletal tension.

The presence of senescent cells within tissues has been functionally linked to malignant transformations. Here, using tension-gauge tethers technology, particle-tracking microrheology, and quantitative microscopy, we demonstrate that senescent-associated secretory phenotype (SASP) derived from senescent fibroblasts impose nuclear lobulations and volume shrinkage on malignant cells, which stems from the loss of RhoA/ROCK/myosin II-based cortical tension. This loss in cytoskeletal tension induces decreased cellular contractility, adhesion, and increased mechanical compliance. These SASP-induced morphological changes are, in part, mediated by Lamin A/C. These findings suggest that SASP induces defective outside-in mechanotransduction from actomyosin fibers in the cytoplasm to the nuclear lamina, thereby triggering a cascade of biophysical and biomolecular changes in cells that associate with malignant transformations.

Endothelial cell-leukemia interactions remodel drug responses, uncovering T-ALL vulnerabilities.

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive and often incurable disease. To uncover therapeutic vulnerabilities, we first developed T-ALL patient-derived tumor xenografts (PDXs) and exposed PDX cells to a library of 433 clinical-stage compounds in vitro. We identified 39 broadly active drugs with antileukemia activity. Because endothelial cells (ECs) can alter drug responses in T-ALL, we developed an EC/T-ALL coculture system. We found that ECs provide protumorigenic signals and mitigate drug responses in T-ALL PDXs. Whereas ECs broadly rescued several compounds in most models, for some drugs the rescue was restricted to individual PDXs, suggesting unique crosstalk interactions and/or intrinsic tumor features. Mechanistically, cocultured T-ALL cells and ECs underwent bidirectional transcriptomic changes at the single-cell level, highlighting distinct "education signatures." These changes were linked to bidirectional regulation of multiple pathways in T-ALL cells as well as in ECs. Remarkably, in vitro EC-educated T-ALL cells transcriptionally mirrored ex vivo splenic T-ALL at single-cell resolution. Last, 5 effective drugs from the 2 drug screenings were tested in vivo and shown to effectively delay tumor growth and dissemination thus prolonging overall survival. In sum, we developed a T-ALL/EC platform that elucidated leukemia-microenvironment interactions and identified effective compounds and therapeutic vulnerabilities.

Nature-inspired delivery of mitochondria-targeted angiotensin receptor blocker.

Mitochondria are critical regulators of cellular function and survival. We have previously demonstrated that functional angiotensin receptors embedded within the inner mitochondrial membrane modulate mitochondrial energy production and free radical generation. The expression of mitochondrial angiotensin II type-1 receptors increases during aging, with a complementary decrease in angiotensin II type-2 receptor density. To address this age-associated mitochondrial dysfunction, we have developed a mitochondria-targeted delivery system to effectively transport angiotensin type-1 receptor blocker-Losartan (mtLOS) into the inner mitochondrial membrane. We engineered mtLOS to become active within the mitochondria after cleavage by mitochondrial peptidases. Our data demonstrate effective and targeted delivery of mtLOS into the mitochondria, compared to a free Losartan, or Losartan conjugated to a scrambled mitochondrial target signal peptide, with significant shifts in mitochondrial membrane potential upon mtLOS treatment. Furthermore, engineered mitochondrial-targeting modalities could open new avenues to transport nonmitochondrial proteins into the mitochondria, such as other macromolecules and therapeutic agents.

The Novel Use of Umbilical Cord Blood to Obtain Complete Blood Counts for Critical Neonatal Assessment.

BACKGROUND: Neonates undergoing clinical evaluations are often subjected to potentially painful phlebotomy for laboratory tests. The use of cord blood laboratory values for admission has been suggested as a means to decrease the risk of painful venipuncture and anemia. METHODS: Peripheral and umbilical cord blood complete blood count (CBC) results were obtained from infants who required a CBC. Results were compared using the Sysmex XN heme analyzer (Sysmex, Kobe, Japan). RESULTS:  White blood cell (WBC) and hemoglobin (HgB) values were significantly higher in peripheral samples than in cord samples. The mean cord WBC count was 14.1 × 103/mm3 versus 15.6 × 103/mm3 peripherally (p < 0.001). The mean cord HgB was 15.8 g/dL versus 16.8 g/dL peripherally (p < 0.001). Cord platelet (Plt) counts were, conversely, lower in peripheral samples than in cord samples (264.8 × 103/mm3 versus 242.3 × 103/mm3, respectively; p < 0.001). Although statistically different, the mean CBC values from both samples were within the reference ranges. Delayed cord clamping (DCC) increased peripheral versus cord HgB difference nearly threefold (0.6-1.7 g/dL; p = 0.01). CONCLUSIONS: Cord blood is an acceptable source for CBC blood sampling in newborn infants and can be used for clinical decisions. CBC laboratory values for cord blood remained within the peripheral blood reference range, with slight variability between the two samples.

HSP70 and their co-chaperones in the human malaria parasite P. falciparum and their potential as drug targets.

As part of their life-cycle, malaria parasites undergo rapid cell multiplication and division, with one parasite giving rise to over 20 new parasites within the course of 48 h. To support this, the parasite has an extremely high metabolic rate and level of protein biosynthesis. Underpinning these activities, the parasite encodes a number of chaperone/heat shock proteins, belonging to various families. Research over the past decade has revealed that these proteins are involved in a number of essential processes within the parasite, or within the infected host cell. Due to this, these proteins are now being viewed as potential targets for drug development, and we have begun to characterize their properties in more detail. In this article we summarize the current state of knowledge about one particular chaperone family, that of the HSP70, and highlight their importance, function, and potential co-chaperone interactions. This is then discussed with regard to the suitability of these proteins and interactions for drug development.

Inhibition of Integrin αVß3 Signaling Improves the Antineoplastic Effect of Bexarotene in Cutaneous T-Cell Lymphoma.

Bexarotene is a specific retinoid X receptor agonist that has been used for the treatment of cutaneous T-cell lymphoma (CTCL). Because bexarotene causes hypothyroidism, it requires the administration of levothyroxine. However, levothyroxine, in addition to its ubiquitous nuclear receptors, can activate the αVß3 integrin that is overexpressed in CTCL, potentially interfering the antineoplastic effect of bexarotene. We thus investigated the biological effect of levothyroxine in relation to bexarotene treatment. Although in isolated CTCL cells levothyroxine decreased, in an αVß3-dependent manner, the antineoplastic effect of bexarotene, levothyroxine supplementation in preclinical models was necessary to avoid suppression of lymphoma immunity. Accordingly, selective genetic and pharmacologic inhibition of integrin αVß3 improved the antineoplastic effect of bexarotene plus levothyroxine replacement while maintaining lymphoma immunity. Our results provide a mechanistic rationale for clinical testing of integrin αVß3 inhibitors as part of CTCL regimens based on bexarotene administration. TEASER: Inhibiting αVß3 integrin improves the antineoplastic effect of bexarotene while maintaining lymphoma immunity.

Deep learning identification of stiffness markers in breast cancer.

While essential to our understanding of solid tumor progression, the study of cell and tissue mechanics has yet to find traction in the clinic. Determining tissue stiffness, a mechanical property known to promote a malignant phenotype in vitro and in vivo, is not part of the standard algorithm for the diagnosis and treatment of breast cancer. Instead, clinicians routinely use mammograms to identify malignant lesions and radiographically dense breast tissue is associated with an increased risk of developing cancer. Whether breast density is related to tumor tissue stiffness, and what cellular and non-cellular components of the tumor contribute the most to its stiffness are not well understood. Through training of a deep learning network and mechanical measurements of fresh patient tissue, we create a bridge in understanding between clinical and mechanical markers. The automatic identification of cellular and extracellular features from hematoxylin and eosin (H&E)-stained slides reveals that global and local breast tissue stiffness best correlate with the percentage of straight collagen. Importantly, the percentage of dense breast tissue does not directly correlate with tissue stiffness or straight collagen content.

TLR8 is activated by 5'-methylthioinosine, a Plasmodium falciparum-derived intermediate of the purine salvage pathway.

The innate immune recognition of the malaria-causing pathogen Plasmodium falciparum (P. falciparum) is not fully explored. Here, we identify the nucleoside 5'-methylthioinosine (MTI), a Plasmodium-specific intermediate of the purine salvage pathway, as a pathogen-derived Toll-like receptor 8 (TLR8) agonist. Co-incubation of MTI with the TLR8 enhancer poly(dT) as well as synthetic or P. falciparum-derived RNA strongly increase its stimulatory activity. Of note, MTI generated from methylthioadenosine (MTA) by P. falciparum lysates activates TLR8 when MTI metabolism is inhibited by immucillin targeting the purine nucleoside phosphorylase (PfPNP). Importantly, P. falciparum-infected red blood cells incubated with MTI or cultivated with MTA and immucillin lead to TLR8-dependent interleukin-6 (IL-6) production in human monocytes. Our data demonstrate that the nucleoside MTI is a natural human TLR8 ligand with possible in vivo relevance for innate sensing of P. falciparum.

Histamine H4 Receptor Agonism Induces Antitumor Effects in Human T-Cell Lymphoma.

The discovery of the human histamine H4 receptor (H4R) has contributed to our understanding of the role of histamine in numerous physiological and pathological conditions, including tumor development and progression. The lymph nodes of patients with malignant lymphomas have shown to contain high levels of histamine, however, less is known regarding the expression and function of the H4R in T-cell lymphoma (TCL). In this work we demonstrate the expression of H4R isoforms (mRNA and protein) in three human aggressive TCL (OCI-Ly12, Karpas 299, and HuT78). Histamine and specific H4R agonists (VUF8430 and JNJ28610244) significantly reduced cell viability in a dose-dependent manner (p < 0.05). The combined treatment with the H4R antagonist (JNJ7777120, 10 µM) reversed the effects of the H4R ligands. Importantly, we screened a drug repurposing library of 433 FDA-approved compounds (1 µM) in combination with histamine (10 µM) in Hut78 cells. Histamine produced a favorable antitumor effect with 18 of these compounds, including the histone deacetylase inhibitor panobinostat. Apoptosis, proliferation, and oxidative stress studies confirmed the antitumoral effects of the combination. We conclude that the H4R is expressed in TCL, and it is involved in histamine-mediated responses.

Structure and Function of Redox-Sensitive Superfolder Green Fluorescent Protein Variant.

Aims: Genetically encoded green fluorescent protein (GFP)-based redox biosensors are widely used to monitor specific and dynamic redox processes in living cells. Over the last few years, various biosensors for a variety of applications were engineered and enhanced to match the organism and cellular environments, which should be investigated. In this context, the unicellular intraerythrocytic parasite Plasmodium, the causative agent of malaria, represents a challenge, as the small size of the organism results in weak fluorescence signals that complicate precise measurements, especially for cell compartment-specific observations. To address this, we have functionally and structurally characterized an enhanced redox biosensor superfolder roGFP2 (sfroGFP2). Results: SfroGFP2 retains roGFP2-like behavior, yet with improved fluorescence intensity (FI) in cellulo. SfroGFP2-based redox biosensors are pH insensitive in a physiological pH range and show midpoint potentials comparable with roGFP2-based redox biosensors. Using crystallography and rigidity theory, we identified the superfolding mutations as being responsible for improved structural stability of the biosensor in a redox-sensitive environment, thus explaining the improved FI in cellulo. Innovation: This work provides insight into the structure and function of GFP-based redox biosensors. It describes an improved redox biosensor (sfroGFP2) suitable for measuring oxidizing effects within small cells where applicability of other redox sensor variants is limited. Conclusion: Improved structural stability of sfroGFP2 gives rise to increased FI in cellulo. Fusion to hGrx1 (human glutaredoxin-1) provides the hitherto most suitable biosensor for measuring oxidizing effects in Plasmodium. This sensor is of major interest for studying glutathione redox changes in small cells, as well as subcellular compartments in general. Antioxid. Redox Signal. 37, 1-18.

Immune cell profile and immune-related gene expression of obese peripheral blood and liver tissue.

Obesity is associated with changes in immune cell subpopulations. However, tissue and blood obesity-responsive immune phenotypic pathways have not been contrasted. Here, the local niche immune cell population and gene expression in fatty liver is compared to peripheral blood of obese individuals. The Cibersort algorithm enumerated increased fractions of memory CD4+ T lymphocytes and reductions in natural killer and memory B cells in obese liver tissue and obese blood, with similar reductions found in nonalcoholic fatty liver disease tissue. Gene expression analysis identified inflammatory immune signatures of regulatory CD4+ T cells with inferred Th1, Th17, Th2, or Treg phenotypes that differed between liver and blood. Our study suggests that the local tissue-specific immune phenotype in the liver differs from the obese peripheral circulation, with the latter reflective of multisystemic persistent inflammation that is characteristic of obesity.