Therapeutic targeting of injuries that require transient restoration of proteins by mRNA delivery is an attractive approach that, until recently, has remained poorly explored. In this study, we ...examined the therapeutic utility of mRNA delivery for liver fibrosis and cirrhosis. Specifically, we aimed to demonstrate the therapeutic efficacy of human hepatocyte nuclear factor alpha (HNF4A) mRNA in mouse models of fibrosis and cirrhosis.
We investigated restoration of hepatocyte functions by HNF4A mRNA transfection in vitro, and analyzed the attenuation of liver fibrosis and cirrhosis in multiple mouse models, by delivering hepatocyte-targeted biodegradable lipid nanoparticles (LNPs) encapsulating HNF4A mRNA. To identify potential mechanisms of action, we performed microarray-based gene expression profiling, single-cell RNA sequencing, and chromatin immunoprecipitation. We used primary liver cells and human liver buds for additional functional validation.
Expression of HNF4A mRNA led to restoration of the metabolic activity of fibrotic primary murine and human hepatocytes in vitro. Repeated in vivo delivery of LNP-encapsulated HNF4A mRNA induced a robust inhibition of fibrogenesis in 4 independent mouse models of hepatotoxin- and cholestasis-induced liver fibrosis. Mechanistically, we discovered that paraoxonase 1 is a direct target of HNF4A and it contributes to HNF4A-mediated attenuation of liver fibrosis via modulation of liver macrophages and hepatic stellate cells.
Collectively, our findings provide the first direct preclinical evidence of the applicability of HNF4A mRNA therapeutics for the treatment of fibrosis in the liver.
Liver fibrosis and cirrhosis remain unmet medical needs and contribute to high mortality worldwide. Herein, we take advantage of a promising therapeutic approach to treat liver fibrosis and cirrhosis. We demonstrate that restoration of a key gene, HNF4A, via mRNA encapsulated in lipid nanoparticles decreased injury in multiple mouse models of fibrosis and cirrhosis. Our study provides proof-of-concept that mRNA therapy is a promising strategy for reversing liver fibrosis and cirrhosis.
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•Restoration of HNF4A via mRNA delivery improves functions of fibrotic primary hepatocytes from both mice and humans.•The mRNA encapsulated in lipid nanoparticles can be delivered into hepatocytes of the fibrotic liver.•Lipid nanoparticle-mediated HNF4A mRNA delivery ameliorates fibrosis and cirrhosis in different chronic liver injury models.•Paraoxonase 1, a therapeutic target of HNF4A, contributes to anti-fibrotic effects of HNF4A.•HNF4A mRNA delivery affects macrophage infiltration and polarization as well as hepatic stellate cell activation.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Pre‐B‐cell leukemia homeobox (PBX) transcription factors are known to regulate organogenesis, but their molecular targets and function in midbrain dopaminergic neurons (mDAn) as well as their role in ...neurodegenerative diseases are unknown. Here, we show that PBX1 controls a novel transcriptional network required for mDAn specification and survival, which is sufficient to generate mDAn from human stem cells. Mechanistically, PBX1 plays a dual role in transcription by directly repressing or activating genes, such as Onecut2 to inhibit lateral fates during embryogenesis, Pitx3 to promote mDAn development, and Nfe2l1 to protect from oxidative stress. Notably, PBX1 and NFE2L1 levels are severely reduced in dopaminergic neurons of the substantia nigra of Parkinson's disease (PD) patients and decreased NFE2L1 levels increases damage by oxidative stress in human midbrain cells. Thus, our results reveal novel roles for PBX1 and its transcriptional network in mDAn development and PD, opening the door for new therapeutic interventions.
Synopsis
PBX1 controls a novel transcriptional network required for midbrain dopaminergic specification and survival. PBX1 and its target NFE2L1 protect dopaminergic neurons from oxidative stress, and their levels are severely reduced in the substantia nigra of Parkinson's disease patients.
Pbx1 is expressed in a subpopulation of dopaminergic neuroblasts and controls the specification and survival of midbrain dopaminergic neurons.
PBX1 plays a dual role in transcription by directly repressing or activating genes, such as Onecut2 to inhibit lateral fates during embryogenesis, Pitx3 to promote mDAn development, and Nfe2l1 to protect from oxidative stress.
PBX1 and NFE2L1 protect dopaminergic neurons from oxidative stress.
Nuclear PBX1 and NFE2L1 levels are severely reduced in midbrain dopaminergic neurons of Parkinson's disease patients.
The homeobox factor PBX1 directly represses or activates key genes involved in specification, survival, and oxidative stress protection of midbrain dopaminergic neurons.
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FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
•Honeybees are less likely to transfer pollen to oilseed rape than solitary bees.•Though less common solitary bees may contribute significantly to pollination.•The importance of solitary bees is ...particularly high away from honeybee hives.•Management for wild pollinators is crucial where honeybee hives are infrequent.
In Europe, oilseed rape is the principal crop used in the production of edible and renewable fuel oil products. Insect pollinators, in particular bees, have been shown to have a positive effect on the seed set of this crop. We undertook experiments looking at behavioural differences between honeybees, bumblebees and solitary bees visiting oilseed rape flowers, and related this to landscape scale responses in visitation rates. We found that behavioural differences between honeybees, bumblebees and solitary bees alter the likelihood of pollen transfer from their bodies to the plant stigma. Solitary bees and bumblebees tend to have greater rates of stigmal contact than honeybees. The interactions between the likelihood of free pollen on bodies and the probability of stigmal contact suggest that only 34.0% of visitations by honeybees were likely to result in pollen transfer to the stigma, relative to 35.1% for the bumblebees and 71.3% for solitary bees. Visitation rates were higher for honeybees in high quality landscapes with relatively large areas of alternative foraging habitat. Visitation rates of honeybees were also more frequent in the vicinity of managed hives. For solitary bees and bumblebees visitation rates did not respond to landscape structure, although more species of solitary bees were found in landscapes with a high cover of semi-natural grassland. While honeybees may be less efficient in pollen transfer per unit visit, where they numerically outweigh other types of bees in a crop (e.g. around managed hives) this may not be important. For this reason the relative ease with which hives can be moved across landscape means that honeybees are perhaps the most suitable taxa for use as a pro-active mitigation measure against pollinator deficits. However, the greater efficiency of solitary bees compensates for the effort required to implement longer term management (i.e. the establishment of flower rich field margins and open soil nesting sites) to support their populations.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
A beam-normal single-spin asymmetry generated in the scattering of transversely polarized electrons from unpolarized nucleons is an observable related to the imaginary part of the two-photon exchange ...process. We report a 2% precision measurement of the beam-normal single-spin asymmetry in elastic electron-proton scattering with a mean scattering angle of θlab=7.9° and a mean energy of 1.149 GeV. The asymmetry result is Bn=−5.194±0.067(stat)±0.082 (syst) ppm. This is the most precise measurement of this quantity available to date and therefore provides a stringent test of two-photon exchange models at far-forward scattering angles (θlab→0) where they should be most reliable.
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CMK, CTK, FMFMET, IJS, NUK, PNG, UL, UM
Background
Lower activation of higher threshold (type-II) motor units (MUs) has been suggested in children compared with adults. We examined child–adult differences in discrete MU activation of the ...flexor carpi radialis (FCR).
Methods
Fifteen boys (10.2 ± 1.4 years), and 17 men (25.0 ± 2.7 years) completed 2 laboratory sessions. Following a habituation session, maximal voluntary isometric wrist flexion torque (MVIC) was determined before completing trapezoidal isometric contractions at 70%MVIC. Surface electromyography was captured by Delsys Trigno Galileo sensors and decomposed into individual MU action potential trains. Recruitment threshold (RT), and MU firing rates (MUFR) were calculated.
Results
MVIC was significantly greater in men (10.19 ± 1.92 Nm) than in boys (4.33 ± 1.47 Nm) (
p
< 0.05), but not statistically different after accounting for differences in body size. Mean MUFR was not different between boys (17.41 ± 7.83 pps) and men (17.47 ± 7.64 pps). However, the MUFR–RT slope was significantly (
p
< 0.05) steeper (more negative) in boys, reflecting a progressively greater decrease in MUFR with increasing RT. Additionally, boys recruited more of their MUs early in the ramped contraction.
Conclusion
Compared with men, boys tended to recruit their MUs earlier and at a lower percentage of MVIC. This difference in MU recruitment may explain the greater decrease in MUFR with increasing RT in boys compared with men. Overall, these findings suggest an age-related difference in the neural strategy used to develop moderate–high torque in wrist flexors, where boys recruit more of their MUs earlier in the force gradation process, possibly resulting in a narrower recruitment range.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Cardiac surgery induces many physiologic changes including major inflammatory and sympathetic nervous system responses. Here, we conducted a single-centre pilot study to generate hypotheses on the ...potential immune impact of adding high spinal anaesthesia to general anaesthesia during cardiac surgery in adults. We hypothesized that this strategy, previously shown to blunt the sympathetic response and improve pain management, could reduce the undesirable systemic inflammatory responses caused by cardiac surgery.
This prospective randomized unblinded pilot study was conducted on 14 patients undergoing cardiac surgery for coronary artery bypass grafting and/or aortic valve replacement secondary to severe aortic stenosis. The primary outcome measures examined longitudinally were serum pro-inflammatory (IL-6, IL-1b, CCL2), anti-inflammatory (IL-10, TNF-RII, IL-1Ra), acute phase protein (CRP, PTX3) and cardiovascular risk (sST2) biomarkers.
The kinetics of pro- and anti-inflammatory biomarker was determined following surgery. All pro-inflammatory and acute phase reactant biomarker responses induced by surgical stress were indistinguishable in intensity and duration between control groups and those who also received high spinal anaesthesia. Conversely, IL-10 levels were markedly elevated in both intensity and duration in the group receiving high spinal anesthesia (p = 0.005).
This hypothesis generating pilot study suggests that high spinal anesthesia can alter the net inflammatory response that results from cardiac surgery. In appropriately selected populations, this may add incremental benefit by dampening the net systemic inflammatory response during the week following surgery. Larger population studies, powered to assess immune, physiologic and clinical outcomes in both acute and longer term settings, will be required to better assess potential benefits of incorporating high spinal anesthesia.
ClinicalTrials.gov NCT00348920.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
In the era of organ machine perfusion, experimental models to optimize reconditioning of (marginal) liver grafts are needed. Although the relevance of cytokine signatures in liver transplantation has ...been analyzed previously, the significance of molecular monitoring during normothermic machine perfusion (NMP) remains elusive. Therefore, we developed a porcine model of cold ischemic liver graft injury after prolonged static cold storage (SCS) and subsequent NMP: Livers obtained from ten minipigs underwent NMP for 6 h directly after procurement (control group) or after 20 h of SCS. Grafts after prolonged SCS showed significantly elevated AST, ALT, GLDH and GGT perfusate concentrations, and reduced lactate clearance. Bile analyses revealed reduced bile production, reduced bicarbonate and elevated glucose concentrations after prolonged SCS. Cytokine analyses of graft perfusate simultaneously demonstrated an increase of pro-inflammatory cytokines such as Interleukin-1α, Interleukin-2, and particularly Interleukin-18. The latter was the only significantly elevated cytokine compared to controls, peaking as early as 2 h after reperfusion (11,012 ng/ml vs. 1,493 ng/ml;
p
= 0.029). Also, concentrations of High-Mobility-Group-Protein B1 were significantly elevated after 2 h of reperfusion (706.00 ng/ml vs. 148.20 ng/ml;
p
< 0.001) and showed positive correlations with AST (
r
2
= 0.846) and GLDH (
r
2
= 0.918) levels. Molecular analyses during reconditioning of liver grafts provide insights into the degree of inflammation and cell damage and could thereby facilitate future interventions during NMP reducing acute and chronic graft injury.
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