Murine Disease Models Research Articles

Probenecid slows disease progression in a murine model of autosomal dominant polycystic kidney disease.

Development of autosomal dominant polycystic kidney disease (ADPKD) involves renal epithelial cell abnormalities. Cystic fluid contains a high level of ATP that, among other effects, leads to a reduced reabsorption of electrolytes in cyst-lining cells, and thus results in cystic fluid accumulation. Earlier, we demonstrated that Pkd1RC/RC mice, a hypomorphic model of ADPKD, exhibit increased expression of pannexin-1, a membrane channel capable of ATP release. In the current study, we found that human ADPKD cystic epithelia have higher pannexin-1 abundance than normal collecting ducts. We hypothesized that inhibition of pannexin-1 function with probenecid can be used to attenuate ADPKD development. Renal function in male and female Pkd1RC/RC and control mice was monitored between 9 and 20 months of age. To test the therapeutic effects of probenecid (a uricosuric agent and a pannexin-1 blocker), osmotic minipumps were implanted in male and female Pkd1RC/RC mice, and probenecid or vehicle was administered for 42 days until 1 year of age. Probenecid treatment improved glomerular filtration rates and slowed renal cyst formation in male mice (as shown in histopathology). The mechanistic effects of probenecid on sodium reabsorption and fluid transport were tested on polarized mpkCCDcl4 cells subjected to short-circuit current measurements, and in 3D cysts grown in Matrigel. In the mpkCCDcl4 epithelial cell line, probenecid elicited higher ENaC currents and attenuated in vitro cyst formation, indicating lower sodium and less fluid retention in the cysts. Our studies open new avenues of research into targeting pannexin-1 in ADPKD pathology.

Epigallocatechin gallate alleviates high-fat diet-induced hepatic lipotoxicity by targeting mitochondrial ROS-mediated ferroptosis.

Background: Non-alcoholic fatty liver disease (NAFLD) is a chronic advanced liver disease that is highly related to metabolic disorders and induced by a high-fat diet (HFD). Recently, epigallocatechin gallate (EGCG) has been regarded as a protective bioactive polyphenol in green tea that has the ability to protect against non-alcoholic fatty liver disease, but the molecular mechanism remains poorly deciphered. Ferroptosis plays a vital role in the progression of non-alcoholic fatty liver disease, but experimental evidence of ferroptosis inhibition by epigallocatechin gallate is limited. Hence, our study aimed to investigate the effect and mechanisms of epigallocatechin gallate on hepatic ferroptosis to mitigate hepatic injury in high-fat diet-fed mice. Methods: Fifty male C57BL/6 mice were fed either a standard chow diet (SCD), a high-fat diet, or a high-fat diet and administered epigallocatechin gallate or ferrostatin-1 (a ferroptosis-specific inhibitor) for 12weeks. Liver injury, lipid accumulation, hepatic steatosis, oxidative stress, iron overload, and ferroptosis marker proteins were examined. In vitro, steatotic L-02 cells were used to explore the underlying mechanism. Results: In our research, we found that epigallocatechin gallate notably alleviated liver injury and lipid accumulation, oxidative stress, hepatic steatosis, decreased iron overload and inhibited ferroptosis in a high-fat diet-induced murine model of non-alcoholic fatty liver disease. In vitro experiments, using ferrostatin-1 and a mitochondrial reactive oxygen species (MtROS) scavenger (Mito-TEMPO), we found that epigallocatechin gallate remarkably alleviated oxidative stress and inhibited ferroptosis by reducing the level of mitochondrial reactive oxygen species in steatotic L-02 cells. Conclusion: Taken together, our results revealed that epigallocatechin gallate may exert protective effects on hepatic lipotoxicity by inhibiting mitochondrial reactive oxygen species-mediated hepatic ferroptosis. Findings from our study provide new insight into prevention and treatment strategies for non-alcoholic fatty liver disease pathological processes.

Efficacy of platelet-inspired hemostatic nanoparticles on bleeding in von Willebrand disease murine models

AbstractThe lack of innovation in von Willebrand disease (VWD) originates from many factors including the complexity and heterogeneity of the disease but also from a lack of recognition of the impact of the bleeding symptoms experienced by patients with VWD. Recently, a few research initiatives aiming to move past replacement therapies using plasma-derived or recombinant von Willebrand factor (VWF) concentrates have started to emerge. Here, we report an original approach using synthetic platelet (SP) nanoparticles for the treatment of VWD type 2B (VWD-2B) and severe VWD (type 3 VWD). SP are liposomal nanoparticles decorated with peptides enabling them to concomitantly bind to collagen, VWF, and activated platelets. In vitro, using various microfluidic assays, we show the efficacy of SPs to improve thrombus formation in VWF-deficient condition (with human platelets) or using blood from mice with VWD-2B and deficient VWF (VWF-KO, ie, type 3 VWD). In vivo, using a tail-clip assay, SP treatment reduced blood loss by 35% in mice with VWD-2B and 68% in mice with VWF-KO. Additional studies using nanoparticles decorated with various combinations of peptides demonstrated that the collagen-binding peptide, although not sufficient by itself, was crucial for SP efficacy in VWD-2B; whereas all 3 peptides appeared necessary for mice with VWF-KO. Clot imaging by immunofluorescence and scanning electron microscopy revealed that SP treatment of mice with VWF-KO led to a strong clot, similar to those obtained in wild-type mice. Altogether, our results show that SP could represent an attractive therapeutic alternative for VWD, especially considering their long half-life and stability.

The Safety of Adiponectin Receptor Agonist AdipoRon in a Rabbit Model of Arthrofibrosis.

AdipoRon is an adiponectin receptor 1, 2 (ADIPOR1 and ADIPOR2) agonist with numerous reported physiological benefits in murine models of human disease, including a proposed reduction in fibrosis. However, AdipoRon has never been investigated in rabbits, which provide a robust model for orthopedic conditions. We examined the safety of intravenous (IV) AdipoRon in New Zealand White (NZW) female rabbits surgically stressed by a procedure that mimics human arthrofibrosis. Fifteen female NZW rabbits were prospectively studied using increasing AdipoRon doses based on established literature. AdipoRon was dissolved in dimethyl sulfoxide (DMSO), diluted in normal saline, and administered IV preoperatively and for 5 subsequent days postoperatively. The primary outcome was overall toxicity to rabbits, whereas secondary outcomes were change in rabbit weights and hemodynamics and defining acid-base characteristics of the drug formulation. Two rabbits expired during preoperative drug administration at 25 mg/kg. Remaining rabbits received preoperative doses of DMSO (vehicle), 2.5, 5, or 10 mg/kg of AdipoRon without complications. On postoperative day 1, one rabbit sustained a tonic-clonic seizure after their second dose of 10 mg/kg AdipoRon. The remaining 12 rabbits (4 in each group) received six serial doses of vehicle, 2.5, or 5 mg/kg of AdipoRon without adverse effects. All formulations of AdipoRon were within safe physiological pH ranges (4-5). We are the first to report the use of IV AdipoRon in a surgically stressed rabbit model of orthopedic disease. AdipoRon doses of 5 mg/kg or less appear to be well-tolerated in female NZW rabbits. Impact statement We provided the first in vivo toxicity assessment and dose optimization of a new antifibrotic experimental medication, AdipoRon, in a surgically stressed rabbit model of knee arthrofibrosis.

Metabolic correlates to critical speed in murine models of sickle cell disease.

Introduction: Exercise intolerance is a common clinical manifestation in patients with sickle cell disease (SCD), though the mechanisms are incompletely understood. Methods: Here we leverage a murine mouse model of sickle cell disease, the Berkeley mouse, to characterize response to exercise via determination of critical speed (CS), a functional measurement of mouse running speed upon exerting to exhaustion. Results: Upon observing a wide distribution in critical speed phenotypes, we systematically determined metabolic aberrations in plasma and organs-including heart, kidney, liver, lung, and spleen-from mice ranked based on critical speed performances (top vs. bottom 25%). Results indicated clear signatures of systemic and organ-specific alterations in carboxylic acids, sphingosine 1-phosphate and acylcarnitine metabolism. Metabolites in these pathways showed significant correlations with critical speed across all matrices. Findings from murine models were thus further validated in 433 sickle cell disease patients (SS genotype). Metabolomics analyses of plasma from 281 subjects in this cohort (with HbA < 10% to decrease confounding effects of recent transfusion events) were used to identify metabolic correlates to sub-maximal exercise test performances, as measure by 6min walking test in this clinical cohort. Results confirmed strong correlation between test performances and dysregulated levels of circulating carboxylic acids (especially succinate) and sphingosine 1-phosphate. Discussion: We identified novel circulating metabolic markers of exercise intolerance in mouse models of sickle cell disease and sickle cell patients.

The Murine Oral Metatranscriptome Reveals Microbial and Host Signatures of Periodontal Disease

Periodontal disease is accompanied by alterations to cellular profiles and biological activities of both the subgingival microbiome and host tissues. Although significant progress has been made in describing the molecular basis of the homeostatic balance of host–commensal microbe interactions in health compared to the destructive imbalance in disease, particularly with respect to immune and inflammatory systems, few studies have attempted a comprehensive analysis in diverse host models. Here, we describe the development and application of a metatranscriptomic approach to analysis of host–microbe gene transcription in a murine periodontal disease model, based on oral gavage infection using Porphyromonas gingivalis in C57BL6/J mice. We generated 24 metatranscriptomic libraries from individual mouse oral swabs, representing health and disease. On average, 76% ± 11.7% reads in each sample belonged to the murine host genome and the remainder to the microbes. We found 3,468 (2.4% of the total) murine host transcripts differentially expressed between health and disease, of which 76% were overexpressed in periodontitis. Predictably, there were prominent alterations to genes and pathways linked with the host immune compartment in disease—the CD40 signaling pathway being the top enriched biological process in this data set. However, in addition, we observed significant alterations to other biological processes in disease, particularly cellular/metabolic processes and biological regulation. The number of differentially expressed microbial genes particularly indicated shifts in carbon metabolism pathways in disease with potential consequences for metabolic end-product formation. Together, these metatranscriptome data reveal marked changes between the gene expression patterns in both the murine host and microbiota, which may represent signatures of health and disease, providing the basis for future functional studies of prokaryotic and eukaryotic cellular responses in periodontal disease. In addition, the noninvasive protocol developed in this study will enable further longitudinal and interventionist studies of host–microbe gene expression networks.

A4 PREDNISOLONE, A GLUCOCORTICOID WIDELY USED FOR TREATMENT OF IBD, ENHANCES A HUMAN INTERLEUKIN-4-ACTIVATED MACROPHAGE PHENOTYPE

Abstract Background With cellular immunotherapy, the individuals’ medication could ablate (or enhance) any therapeutic benefit of the transferred cells. Murine and human macrophages activated with IL-4 (i.e., M(IL4)) improve wound healing and reduce the severity of disease in murine models of colitis. Advancing the position that autologous M(IL4) could be a novel approach to IBD, a critical question arises: will concurrent medication impact the M(IL4)s anti-colitic effect? To address this, we tested if prednisolone, a synthetic, anti-inflammatory glucocorticoid used to induce remission in IBD flares,impacts human M(IL4) phenotype and function. Purpose To determine if prednisolone suppresses or enhances a human M(IL4) phenotype as defined by canonical marker molecules and wound healing and anti-colitic activities. Method Macrophages were differentiated from the blood monocytes of healthy volunteers using M-CSF (7 days) and treated with GMP-grade IL-4 (10 ng/mL, 48h) ± a 24h treatment with prednisolone (1μg/mL). Subsequently, conditioned medium was collected for TGFb measurement by ELISA and for use in a T84 epithelial cell in vitro wound healing assay. Retrieved M(IL4) and M(IL4,pred.) were characterized by mRNA expression of CD206 (mannose receptor), RAMP1 (CGRP receptor), and CD14 (LPS co-receptor). One million murine bone marrow-derived M(IL4) or M(IL4,pred.) were injected into BALB/c mice 48h prior to intra-rectal DNBS (3mg), and colitis was assessed 72h-post DNBS. Result(s) Human M(IL4)s displayed increased mRNA expression of CD206 and RAMP1, and reduced CD14 compared to M(0), with the CD206 and RAMP1 being further increased by prednisolone treatment. M(IL4,pred.) produced more TGF-β than M(IL4) upon LPS stimulation [363 ± 30 vs. 241 ± 24 pg/ml, n= 4, p&amp;lt;0.05], which would predict an enhanced wound healing capacity. Stimulated M(IL4,pred.) produced more IL-10 than M(IL4). Furthermore, murine M(IL4,pred.) retained an anti-colitic capacity comparable to M(IL4) as determined by disease activity score in the DNBS model. Conclusion(s) Human M(IL4)s subsequently exposed to the potent immunomodulatory glucocorticoid, prednisolone show increased expression of phenotypic markers and increased output of TGFb and IL-10. Crucially M(IL4,pred.) retained an anti-colic effect in the murine DNBS model of colitis. Interpreting these data, we suggest that the anti-colitic effect of M(IL4) immunotherapy would not be adversely offset by the individuals concomitant use of steroids. Our preliminary findings support pursuing M(IL4) transfers as a novel approach to the management of IBD. Please acknowledge all funding agencies by checking the applicable boxes below Other Please indicate your source of funding; Helmsley Charitable Trust Disclosure of Interest None Declared

Analysis of structural variation among inbred mouse strains

Background‘Long read’ sequencing methods have been used to identify previously uncharacterized structural variants that cause human genetic diseases. Therefore, we investigated whether long read sequencing could facilitate genetic analysis of murine models for human diseases.ResultsThe genomes of six inbred strains (BTBR T + Itpr3tf/J, 129Sv1/J, C57BL/6/J, Balb/c/J, A/J, SJL/J) were analyzed using long read sequencing. Our results revealed that (i) Structural variants are very abundant within the genome of inbred strains (4.8 per gene) and (ii) that we cannot accurately infer whether structural variants are present using conventional short read genomic sequence data, even when nearby SNP alleles are known. The advantage of having a more complete map was demonstrated by analyzing the genomic sequence of BTBR mice. Based upon this analysis, knockin mice were generated and used to characterize a BTBR-unique 8-bp deletion within Draxin that contributes to the BTBR neuroanatomic abnormalities, which resemble human autism spectrum disorder.ConclusionA more complete map of the pattern of genetic variation among inbred strains, which is produced by long read genomic sequencing of the genomes of additional inbred strains, could facilitate genetic discovery when murine models of human diseases are analyzed.

Profiling the oxylipidome in aged mice after chronic ethanol feeding: Identifying lipid metabolites as drivers of hepatocyte stress.

The global population of people over the age of 65 is increasing and expected to reach 1.5 billion by 2050. While aging is associated with a number of chronic illnesses including dementia, the underlying contribution of alcohol misuse in the elderly is understudied. Long-term chronic alcohol misuse can lead to alcohol-associated liver disease, consisting of a spectrum of pathologies, including steatosis and cirrhosis; liver disease can be rapidly accelerated by non-resolving inflammation. Despite this knowledge, the mechanistic underpinnings of dysregulated host immunity and accelerated liver disease progression in the aged by alcohol is unknown. Alcohol misuse in the elderly is on the rise and aging is associated with progressive increases in pro-inflammatory cytokine production. The goals of the current study are to characterize bioactive lipid mediators of inflammation by making use of a murine model of ethanol-induced liver disease in 3-month-old and 20-month-old mice by quantitatively profiling selected oxylipins in liver, brain and plasma. Following chronic ethanol exposure, liver injury, steatosis, and senescence markers were robustly increased in aged mice compared to young adult mice. Expression of proinflammatory cytokines and lipid metabolizing enzymes were increased in liver by both age and ethanol feeding. Lipoxygenase-derived lipid metabolites 9- and 13-hydroxy-octadecadienoic acid and 15-hydroxyeicosatetraenoic acid were increased in liver and plasma in ethanol-fed aged mice and positively correlated with liver injury. In plasma, 9,10-dihydroxy-octadecenoic acid/epoxy-octadecenoic acid plasma ratios correlated with liver injury in ethanol-fed aged mice. Finally, 15-hydroxyeicosatetraenoic acid and 9,10-dihydroxy-octadecenoic acid positively correlated between liver and plasma. Importantly, leukotriene E4, 9,10-dihydroxy-octadecenoic acid and 15-hydroxyeicosatetraenoic acid increased lipid accumulation and ER stress in cultured AML12 hepatocytes. These data highlight the complexity of lipid metabolite networks but identify key mediators that may be used for diagnostic and prognostic markers in early stages of alcohol-related liver disease in patients of all ages.

Vertical sleeve gastrectomy associates with airway hyperresponsiveness in a murine model of allergic airway disease and obesity.

Asthma is a chronic airway inflammatory disease marked by airway inflammation, remodeling and hyperresponsiveness to allergens. Allergic asthma is normally well controlled through the use of beta-2-adrenergic agonists and inhaled corticosteroids; however, a subset of patients with comorbid obesity experience resistance to currently available therapeutics. Patients with asthma and comorbid obesity are also at a greater risk for severe disease, contributing to increased risk of hospitalization. Bariatric surgery improves asthma control and airway hyperresponsiveness in patients with asthma and comorbid obesity, however, the underlying mechanisms for these improvements remain to be elucidated. We hypothesized that vertical sleeve gastrectomy (VSG), a model of metabolic surgery in mice, would improve glucose tolerance and airway inflammation, resistance, and fibrosis induced by chronic allergen challenge and obesity. Male C57BL/6J mice were fed a high fat diet (HFD) for 13 weeks with intermittent house dust mite (HDM) allergen administration to induce allergic asthma, or saline as control. At week 11, a subset of mice underwent VSG or Sham surgery with one week recovery. A separate group of mice did not undergo surgery. Mice were then challenged with HDM or saline along with concurrent HFD feeding for 1-1.5 weeks before measurement of lung mechanics and harvesting of tissues, both of which occurred 24 hours after the final HDM challenge. Systemic and pulmonary cytokine profiles, lung histology and gene expression were analyzed. High fat diet contributed to increased body weight, serum leptin levels and development of glucose intolerance for both HDM and saline treatment groups. When compared to saline-treated mice, HDM-challenged mice exhibited greater weight gain. VSG improved glucose tolerance in both saline and HDM-challenged mice. HDM-challenged VSG mice exhibited an increase in airway hyperresponsiveness to methacholine when compared to the non-surgery group. The data presented here indicate increased airway hyperresponsiveness in allergic mice undergoing bariatric surgery.

Insulin-like growth factor 1 reduces coronary atherosclerosis in pigs with familial hypercholesterolemia.

Although murine models of coronary atherosclerotic disease have been used extensively to determine mechanisms, limited new therapeutic options have emerged. Pigs with familial hypercholesterolemia (FH pigs) develop complex coronary atheromas that are almost identical to human lesions. We reported previously that insulin-like growth factor 1 (IGF-1) reduced aortic atherosclerosis and promoted features of stable plaque in a murine model. We administered human recombinant IGF-1 or saline (control) in atherosclerotic FH pigs for 6 months. IGF-1 decreased relative coronary atheroma in vivo (intravascular ultrasound) and reduced lesion cross-sectional area (postmortem histology). IGF-1 increased plaque's fibrous cap thickness, and reduced necrotic core, macrophage content, and cell apoptosis, consistent with promotion of a stable plaque phenotype. IGF-1 reduced circulating triglycerides, markers of systemic oxidative stress, and CXCL12 chemokine levels. We used spatial transcriptomics (ST) to identify global transcriptome changes in advanced plaque compartments and to obtain mechanistic insights into IGF-1 effects. ST analysis showed that IGF-1 suppressed FOS/FOSB factors and gene expression of MMP9 and CXCL14 in plaque macrophages, suggesting possible involvement of these molecules in IGF-1's effect on atherosclerosis. Thus, IGF-1 reduced coronary plaque burden and promoted features of stable plaque in a pig model, providing support for consideration of clinical trials.

Functional TFEB activation characterizes multiple models of renal cystic disease and loss of polycystin-1.

Polycystic kidney disease is a disorder of renal epithelial growth and differentiation. Transcription factor EB (TFEB), a master regulator of lysosome biogenesis and function, was studied for a potential role in this disorder. Nuclear translocation and functional responses to TFEB activation were studied in three murine models of renal cystic disease, including knockouts of folliculin, folliculin interacting proteins 1 and 2, and polycystin-1 (Pkd1) as well as in mouse embryonic fibroblasts lacking Pkd1 and three-dimensional cultures of Madin-Darby canine kidney cells. Nuclear translocation of Tfeb characterized cystic but not noncystic renal tubular epithelia in all three murine models as both an early and sustained response to cyst formation. Epithelia expressed elevated levels of Tfeb-dependent gene products, including cathepsin B and glycoprotein nonmetastatic melanoma protein B. Nuclear Tfeb translocation was observed in mouse embryonic fibroblasts lacking Pkd1 but not wild-type fibroblasts. Pkd1 knockout fibroblasts were characterized by increased Tfeb-dependent transcripts, lysosomal biogenesis and repositioning, and increased autophagy. The growth of Madin-Darby canine kidney cell cysts was markedly increased following exposure to the TFEB agonist compound C1, and nuclear Tfeb translocation was observed in response to both forskolin and compound C1 treatment. Nuclear TFEB also characterized cystic epithelia but not noncystic tubular epithelia in human patients with autosomal dominant polycystic kidney disease. Noncanonical activation of TFEB is characteristic of cystic epithelia in multiple models of renal cystic disease including those associated with loss of Pkd1. Nuclear TFEB translocation is functionally active in these models and may be a component of a general pathway contributing to cystogenesis and growth.NEW & NOTEWORTHY Changes in epithelial cell metabolism are important in renal cyst development. The role of TFEB, a transcriptional regulator of lysosomal function, was explored in several models of renal cystic disease and human ADPKD tissue sections. Nuclear TFEB translocation was uniformly observed in cystic epithelia in each model of renal cystic disease examined. TFEB translocation was functionally active and associated with lysosomal biogenesis and perinuclear repositioning, increased TFEB-associated protein expression, and activation of autophagic flux. Compound C1, a TFEB agonist, promoted cyst growth in 3-D cultures of MDCK cells. Nuclear TFEB translocation is an underappreciated signaling pathway for cystogenesis that may represent a new paradigm for cystic kidney disease.

Daily Brief Heat Therapy Reduces Seizures in A350V IQSEC2 Mice and Is Associated with Correction of AMPA Receptor-Mediated Synaptic Dysfunction.

Purposeful induction of fever for healing, including the treatment of epilepsy, was used over 2000 years ago by Hippocrates. More recently, fever has been demonstrated to rescue behavioral abnormalities in children with autism. However, the mechanism of fever benefit has remained elusive due in large part to the lack of appropriate human disease models recapitulating the fever effect. Pathological mutations in the IQSEC2 gene are frequently seen in children presenting with intellectual disability, autism and epilepsy. We recently described a murine A350V IQSEC2 disease model, which recapitulates important aspects of the human A350V IQSEC2 disease phenotype and the favorable response to a prolonged and sustained rise in body core temperature in a child with the mutation. Our goal has been to use this system to understand the mechanism of fever benefit and then develop drugs that can mimic this effect and reduce IQSEC2-associated morbidity. In this study, we first demonstrate a reduction in seizures in the mouse model following brief periods of heat therapy, similar to what was observed in a child with the mutation. We then show that brief heat therapy is associated with the correction of synaptic dysfunction in neuronal cultures of A350V mice, likely mediated by Arf6-GTP.

CS27109, A Selective Thyroid Hormone Receptor-β Agonist Alleviates Metabolic-Associated Fatty Liver Disease in Murine Models.

Thyroid hormone receptor-β (THR-β) agonists play crucial roles in dyslipidemia and metabolic associated fatty liver disease (MAFLD). We developed a novel oral and liver-targeted THR-β agonist, CS27109, and evaluated its efficacy in the treatment of metabolic disorders. We evaluated in vitro and in vivo efficacy and/or safety of CS27109 along with MGL3196 (a phase III THR-β agonist). CS27109 showed pronounced activity and selectivity to THR-β and favorable PK properties, which was equivalent to MGL3196. In the hamster model, animals treated with a high dose of CS27109 showed equivalent reductions in serum TC and LDL-c with groups treated with MGL3196. In the rat model, CS27109 and MGL3196 reduced serum ALT, TC, TG, LDL-c, liver weight ratio, and liver steatosis. CS27109 simultaneously decreased liver TG and TC, and MGL3196 additionally reduced AST. In the mouse model, CS27109 dose-dependently reduced serum AST, ALT, liver inflammation, and NAS score, and also downregulated TC, LDL-c, liver steatosis, and fibrosis, but not in a dose-dependent manner. MGL3196 revealed an equivalent effect with CS27109 in that model. CS27109 also exhibited tolerable toxicity to the heart. CS27109 shows comparative in vitro and in vivo efficacy with MGL3196, suggesting its potential therapeutic application in the treatment of MAFLD such as dyslipidemia and steatohepatitis.

Periodontal disease is associated with increased gut colonization of pathogenic Haemophilusparainfluenzae in patients with Crohn's disease.

Intestinal colonization of the oral bacterium Haemophilus parainfluenzae has been associated with Crohn's disease (CD) severity and progression. This study examines the role of periodontal disease (PD) as a modifier for colonization of H. parainfluenzae in patients with CD and explores the mechanisms behind H. parainfluenzae-mediated intestinal inflammation. Fifty subjects with and without CD were evaluated for the presence of PD, and their oral and fecal microbiomes were characterized. PD is associated with increased levels of H. parainfluenzae strains in subjects with CD. Oral inoculation of H. parainfluenzae elicits strain-dependent intestinal inflammation in murine models of inflammatory bowel disease, which is associated with increased intestinal interferon-γ (IFN-γ)+ CD4+ Tcells and disruption of the host hypusination pathway. In summary, this study establishes a strain-specific pathogenic role of H. parainfluenzae in intestinal inflammation and highlights the potential effect of PD on intestinal colonization by pathogenic H. parainfluenzae strains in patients with CD.

Involvement of the CXCR4/CXCL12 axis in skeletal muscle wasting in a murine model of chronic obstructive pulmonary disease (COPD)

Involvement of the CXCR4/CXCL12 axis in skeletal muscle wasting in a murine model of chronic obstructive pulmonary disease (COPD)

Lack of Cyclin E1 in hepatocytes aggravates ethanol-induced liver injury and hepatic steatosis in experimental murine model of acute and chronic alcohol-associated liver disease

BackgroundCyclin E1 is the regulatory subunit of cyclin-dependent kinase 2 (Cdk2) and one of the central players in cell cycle progression. We recently showed its crucial role for initiation of liver fibrosis and hepatocarcinogenesis. In the present study, we investigated the role of Cyclin E1 in the development of alcohol-associated liver disease (ALD). MethodsMice with constitutive (E1−/−), hepatocyte-specific (Cyclin E1Δhepa), or intestinal-epithelial-cell-specific (Cyclin E1ΔIEC) inactivation of Cyclin E1 and corresponding wild type littermate controls (WT) were administered either a Lieber-DeCarli ethanol diet (LDE) for 3 weeks or acute ethanol binges (6 g/kg) through oral gavage. Serum parameters of liver functionality were measured; hepatic tissues were collected for biochemical and histological analyses. ResultsThe administration of acute EtOH binge and chronic LDE diet to E1−/− mice enhanced hepatic steatosis, worsened liver damage and triggered body weight loss. Similarly, in the acute EtOH binge model, Cyclin E1Δhepa mice revealed a significantly worsened liver phenotype. In contrast, inactivation of Cyclin E1 only in intestinal epithelial cell (IECs)did not lead to any significant changes in comparison to WT mice after acute EtOH challenge. Remarkably, both acute and chronic EtOH administration in E1−/− animals resulted in increased levels of ADH and decreased expression of ALDH1/2. The additional application of a pan-Cdk inhibitor (S-CR8) further promoted liver damage in EtOH-treated WT mice. ConclusionOur data point to a novel unexpected role of Cyclin E1 in hepatocytes for alcohol metabolism, which seems to be independent of the canonical Cyclin E1/Cdk2 function as a cell cycle regulator.

Infusion of Some but Not All Types of Human Perinatal Stromal Cells Prevent Organ Fibrosis in a Humanized Graft versus Host Disease Murine Model

Allogeneic transplant rejection represents a medical complication that leads to high morbidity and mortality rates. There are no treatments to effectively prevent fibrosis; however, there is great interest in evaluating the use of perinatal mesenchymal stromal cells (MSCs) and other MSCs to prevent fibrosis associated with chronic rejection. In this study, we isolated human perinatal stromal cells (PSCs) from amnion (AM-PSC), placental villi (PV-PSC), and umbilical cord (UC-PSC) tissues, demonstrating the phenotypic characteristics of MSCs as well as a >70% expression of the immunomodulatory markers CD273 and CD210. The administration of a single dose (250,000 cells) of each type of PSC in a humanized graft versus host disease (hGvHD) NSG® murine model delayed the progression of the disease as displayed by weight loss and GvHD scores ranging at various levels without affecting the hCD3+ population. However, only PV-PSCs demonstrated an increased survival rate of 50% at the end of the study. Furthermore, a histopathological evaluation showed that only PV-PSC cells could reduce human CD45+ cell infiltration and the fibrosis of the lungs and liver. These findings indicate that not all PSCs have similar therapeutic potential, and that PV-PSC as a cell therapeutic may have an advantage for targeting fibrosis related to allograft rejection.

The helminth derived peptide FhHDM-1 redirects macrophage metabolism towards glutaminolysis to regulate the pro-inflammatory response.

We have previously identified an immune modulating peptide, termed FhHDM-1, within the secretions of the liver fluke, Fasciola hepatica, which is sufficiently potent to prevent the progression of type 1 diabetes and multiple sclerosis in murine models of disease. Here, we have determined that the FhHDM-1 peptide regulates inflammation by reprogramming macrophage metabolism. Specifically, FhHDM-1 switched macrophage metabolism to a dependence on oxidative phosphorylation fuelled by fatty acids and supported by the induction of glutaminolysis. The catabolism of glutamine also resulted in an accumulation of alpha ketoglutarate (α-KG). These changes in metabolic activity were associated with a concomitant reduction in glycolytic flux, and the subsequent decrease in TNF and IL-6 production at the protein level. Interestingly, FhHDM-1 treated macrophages did not express the characteristic genes of an M2 phenotype, thereby indicating the specific regulation of inflammation, as opposed to the induction of an anti-inflammatory phenotype per se. Use of an inactive derivative of FhHDM-1, which did not modulate macrophage responses, revealed that the regulation of immune responses was dependent on the ability of FhHDM-1 to modulate lysosomal pH. These results identify a novel functional association between the lysosome and mitochondrial metabolism in macrophages, and further highlight the significant therapeutic potential of FhHDM-1 to prevent inflammation.

The mesenchymal compartment in myelodysplastic syndrome: Its role in the pathogenesis of the disorder and its therapeutic targeting.

Myelodysplastic syndromes include a broad spectrum of malignant myeloid disorders that are characterized by dysplastic ineffective hematopoiesis, reduced peripheral blood cells counts and a high risk of progression to acute myeloid leukemia. The disease arises primarily because of accumulating chromosomal, genetic and epigenetic changes as well as immune-mediated alterations of the hematopoietic stem cells (HSCs). However, mounting evidence suggests that aberrations within the bone marrow microenvironment critically contribute to myelodysplastic syndrome (MDS) initiation and evolution by providing permissive cues that enable the abnormal HSCs to grow and eventually establish and propagate the disease. Mesenchymal stromal cells (MSCs) are crucial elements of the bone marrow microenvironment that play a key role in the regulation of HSCs by providing appropriate signals via soluble factors and cell contact interactions. Given their hematopoiesis supporting capacity, it has been reasonable to investigate MSCs' potential involvement in MDS. This review discusses this issue by summarizing existing findings obtained by in vitro studies and murine disease models of MDS. Furthermore, the theoretical background of targeting the BM-MSCs in MDS is outlined and available therapeutic modalities are described.