Nicotinamide mononucleotide and melatonin counteract myocardial ischemia-reperfusion injury by activating SIRT3/FOXO1 and reducing apoptosis in aged male rats

Molecular Biology Reports - It has been documented that aging increases the risk of cardiovascular disease including myocardial ischemia/reperfusion (IR) injury and acute myocardial infarction. In...     

Two Weeks of Metformin Treatment Enhances Mitochondrial Respiration in Skeletal Muscle of AMPK Kinase Dead but Not Wild Type Mice

Metformin is used as an anti-diabetic drug. Metformin ameliorates insulin resistance by improving insulin sensitivity in liver and skeletal muscle. Reduced mitochondrial content has been reported in type 2 diabetic muscles and it may contribute to decreased insulin sensitivity characteristic for diabetic muscles. The molecular mechanism behind the effect of metformin is not fully clarified but inhibition of complex I in the mitochondria and also activation of the 5′AMP activated protein kinase (AMPK) has been reported in muscle. Furthermore, both AMPK activation and metformin treatment have been associated with stimulation of mitochondrial function and biogenesis. However, a causal relationship in skeletal muscle has not been investigated. We hypothesized that potential effects of in vivo metformin treatment on mitochondrial function and protein expressions in skeletal muscle are dependent upon AMPK signaling. We investigated this by two weeks of oral metformin treatment of muscle specific kinase dead α₂ (KD) AMPK mice and wild type (WT) littermates. We measured mitochondrial respiration and protein activity and expressions of key enzymes involved in mitochondrial carbohydrate and fat metabolism and oxidative phosphorylation. Mitochondrial respiration, HAD and CS activity, PDH and complex I-V and cytochrome c protein expression were all reduced in AMPK KD compared to WT tibialis anterior muscles. Surprisingly, metformin treatment only enhanced respiration in AMPK KD mice and thereby rescued the respiration defect compared to the WT mice. Metformin did not influence protein activities or expressions in either WT or AMPK KD mice.We conclude that two weeks of in vivo metformin treatment enhances mitochondrial respiration in the mitochondrial deficient AMPK KD but not WT mice. The improvement seems to be unrelated to AMPK, and does not involve changes in key mitochondrial proteins.     

Differential Plasma MicroRNA Profiles in HBeAg Positive and HBeAg Negative Children with Chronic Hepatitis B

Background and Aim

Children chronically infected with hepatitis B virus (HBV) are at high risk of progressive liver disease. However, no treatment is available that is consistently effective in curing chronic hepatitis B (CHB) in children. Improved understanding of the natural course of disease is warranted. Identification of specific microRNA (miRNA) profiles in children chronically infected with HBV may provide insight into the pathogenesis of CHB and lead to advances in the management of children with CHB.

Patients and Methods

MiRNA PCR panels were employed to screen plasma levels of 739 miRNAs in pooled samples from HBeAg positive, HBeAg negative, and healthy children. The three groups’ plasma miRNA profiles were compared, and aberrantly expressed miRNAs were identified. The identified miRNAs were then validated. Individual RT-qPCRs were performed on plasma from 34 HBeAg positive, 26 HBeAg negative, and 60 healthy children.

Results

A panel of 16 plasma miRNAs were identified as aberrantly expressed in HBeAg positive and HBeAg negative children (p<0.001). Levels of all of the miRNAs were upregulated in HBeAg positive children compared with in HBeAg negative children. A positive correlation was furthermore found between plasma levels of the identified miRNAs and HBV DNA (p<0.001).

Conclusion

We are the first to investigate the plasma miRNA profile of children chronically infected with HBV. Our data indicates the existence of a relationship between abundance of circulating miRNAs and immunological stages in the natural course of disease. Certain miRNAs may contribute to the establishment and maintenance of CHB in children. Further studies are warranted to advance understanding of miRNAs in the pathogenesis of CHB, hopefully leading to the identification of future therapeutic targets.     

Environmental Records from Great Barrier Reef Corals: Inshore versus Offshore Drivers

The biogenic structures of stationary organisms can be effective recorders of environmental fluctuations. These proxy records of environmental change are preserved as geochemical signals in the carbonate skeletons of scleractinian corals and are useful for reconstructions of temporal and spatial fluctuations in the physical and chemical environments of coral reef ecosystems, including The Great Barrier Reef (GBR). We compared multi-year monitoring of water temperature and dissolved elements with analyses of chemical proxies recorded in Porites coral skeletons to identify the divergent mechanisms driving environmental variation at inshore versus offshore reefs. At inshore reefs, water Ba/Ca increased with the onset of monsoonal rains each year, indicating a dominant control of flooding on inshore ambient chemistry. Inshore multi-decadal records of coral Ba/Ca were also highly periodic in response to flood-driven pulses of terrigenous material. In contrast, an offshore reef at the edge of the continental shelf was subject to annual upwelling of waters that were presumed to be richer in Ba during summer months. Regular pulses of deep cold water were delivered to the reef as indicated by in situ temperature loggers and coral Ba/Ca. Our results indicate that although much of the GBR is subject to periodic environmental fluctuations, the mechanisms driving variation depend on proximity to the coast. Inshore reefs are primarily influenced by variable freshwater delivery and terrigenous erosion of catchments, while offshore reefs are dominated by seasonal and inter-annual variations in oceanographic conditions that influence the propensity for upwelling. The careful choice of sites can help distinguish between the various factors that promote Ba uptake in corals and therefore increase the utility of corals as monitors of spatial and temporal variation in environmental conditions.     

A GDF5 Point Mutation Strikes Twice - Causing BDA1 and SYNS2

Growth and Differentiation Factor 5 (GDF5) is a secreted growth factor that belongs to the Bone Morphogenetic Protein (BMP) family and plays a pivotal role during limb development. GDF5 is a susceptibility gene for osteoarthritis (OA) and mutations in GDF5 are associated with a wide variety of skeletal malformations ranging from complex syndromes such as acromesomelic chondrodysplasias to isolated forms of brachydactylies or multiple synostoses syndrome 2 (SYNS2). Here, we report on a family with an autosomal dominant inherited combination of SYNS2 and additional brachydactyly type A1 (BDA1) caused by a single point mutation in GDF5 (p.W414R). Functional studies, including chondrogenesis assays with primary mesenchymal cells, luciferase reporter gene assays and Surface Plasmon Resonance analysis, of the GDF5^W414R variant in comparison to other GDF5 mutations associated with isolated BDA1 (p.R399C) or SYNS2 (p.E491K) revealed a dual pathomechanism characterized by a gain- and loss-of-function at the same time. On the one hand insensitivity to the main GDF5 antagonist NOGGIN (NOG) leads to a GDF5 gain of function and subsequent SYNS2 phenotype. Whereas on the other hand, a reduced signaling activity, specifically via the BMP receptor type IA (BMPR1A), is likely responsible for the BDA1 phenotype. These results demonstrate that one mutation in the overlapping interface of antagonist and receptor binding site in GDF5 can lead to a GDF5 variant with pathophysiological relevance for both, BDA1 and SYNS2 development. Consequently, our study assembles another part of the molecular puzzle of how loss and gain of function mutations in GDF5 affect bone development in hands and feet resulting in specific types of brachydactyly and SYNS2. These novel insights into the biology of GDF5 might also provide further clues on the pathophysiology of OA.     

Influence of Erythropoietin on Cognitive Performance during Experimental Hypoglycemia in Patients with Type 1 Diabetes Mellitus: A Randomized Cross-Over Trial

Introduction

The incidence of severe hypoglycemia in type 1 diabetes has not decreased over the past decades. New treatment modalities minimizing the risk of hypoglycemic episodes and attenuating hypoglycemic cognitive dysfunction are needed. We studied if treatment with the neuroprotective hormone erythropoietin (EPO) enhances cognitive function during hypoglycemia.

Materials and Methods

Eleven patients with type 1 diabetes, hypoglycemia unawareness and recurrent severe hypoglycemia completed the study. In a double-blind, randomized, balanced, cross-over study using clamped hypoglycemia they were treated with 40,000 IU of EPO or placebo administered intravenously six days before the two experiments. Cognitive function (primary endpoint), hypoglycemic symptoms, and counter-regulatory hormonal response were recorded.

Results

Compared with placebo, EPO treatment was associated with a significant reduction in errors in the most complex reaction time task (−4.7 (−8.1 to −1.3), p = 0.01) and a less reaction time prolongation (−66 (−117 to −16) msec, p = 0.02). EPO treatment did not change performance in other measures of cognition. Hypoglycemic symptoms, EEG-changes, and counter-regulatory hormone concentrations did not differ between EPO and placebo treatment.

Conclusion

In patients with type 1 diabetes and hypoglycemia unawareness, treatment with EPO is associated with a beneficial effect on cognitive function in a complex reaction time task assessing sustained attention/working memory. Hypoglycemic symptoms and hormonal responses were not changed by EPO treatment.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00615368","term_id":"NCT00615368"}}NCT00615368     

An Optimized Histochemical Method to Assess Skeletal Muscle Glycogen and Lipid Stores Reveals Two Metabolically Distinct Populations of Type I Muscle Fibers

Skeletal muscle energy metabolism has been a research focus of physiologists for more than a century. Yet, how the use of intramuscular carbohydrate and lipid energy stores are coordinated during different types of exercise remains a subject of debate. Controversy arises from contradicting data from numerous studies, which used different methodological approaches. Here we review the “pros and cons” of previously used histochemical methods and describe an optimized method to ensure the preservation and specificity of detection of both intramyocellular carbohydrate and lipid stores. For optimal preservation of muscle energy stores, air drying cryosections or cycles of freezing-thawing need to be avoided. Furthermore, optimization of the imaging settings in order to specifically image intracellular lipid droplets stained with oil red O or Bodipy-493/503 is shown. When co-staining lipid droplets with associated proteins, Bodipy-493/503 should be the dye of choice, since oil red O creates precipitates on the lipid droplets blocking the light. In order to increase the specificity of glycogen stain, an antibody against glycogen is used. The resulting method reveals the existence of two metabolically distinct myosin heavy chain I expressing fibers: I-1 fibers have a smaller crossectional area, a higher density of lipid droplets, and a tendency to lower glycogen content compared to I-2 fibers. Type I-2 fibers have similar lipid content than IIA. Exhaustive exercise lead to glycogen depletion in type IIA and IIX fibers, a reduction in lipid droplets density in both type I-1 and I-2 fibers, and a decrease in the size of lipid droplets exclusively in type I-1 fibers.     

Hepatitis B Surface Antigen Quantity Positively Correlates with Plasma Levels of microRNAs Differentially Expressed in Immunological Phases of Chronic Hepatitis B in Children

Background and Aim

Children with chronic hepatitis B (CHB) are at high risk of progressive liver disease. It is suggested that a newly-identified panel of 16 microRNAs is important in the pathogenesis of CHB in children. Subviral hepatitis B surface antigen (HBsAg) particles are produced in large excess over infectious virions. Interestingly, circulating HBsAg particles have been shown to carry microRNAs. A thorough characterisation of the identified microRNAs and HBsAg over time in plasma from children with CHB may provide useful information about the natural course of childhood CHB.

Patients and Methods

A cohort of 42 children with CHB was followed over time. Three to five blood samples were obtained from each child at minimum intervals of half a year; in total 180 blood samples. Plasma levels of the 16 microRNAs previously identified were analysed by quantitative real-time polymerase-chain-reaction. Plasma HBsAg was quantified using ARCHITECT® HBsAg assay.

Results

The presence of 14/16 plasma microRNAs in children with CHB was confirmed. All 14 microRNAs were significantly differentially expressed in different immunological phases of the disease. MicroRNA plasma levels were highest in immune-tolerant children, lower in immune-active children, and reached the lowest values in immune-inactive children, p<0.001. Plasma levels of four microRNAs decreased significantly over time in immune-tolerant and immune-active children whereas the microRNA plasma levels were stable in immune-inactive children, p<0.004. HBsAg quantity was positively correlated with plasma levels of 11/14 microRNAs, p<0.004.

Conclusion

This is the first study to characterise plasma microRNAs and HBsAg over time in children with CHB. Our data suggest that plasma levels of selected microRNAs and HBsAg are inversely correlated with immunological control of CHB in children. Further studies are, however, needed to advance the understanding of microRNAs and HBsAg in the pathogenesis of CHB in children.     

Human Cytomegalovirus Infection of M1 and M2 Macrophages Triggers Inflammation and Autologous T-Cell Proliferation

Macrophages (Mϕ) are first targets during human cytomegalovirus (HCMV) infection and are thought to be crucial for viral persistence and dissemination. However, since Mϕ are also a first line of defense and key modulators of the immune response, these cells are at the crossroad between protection and viral pathogenesis. To date, the Mϕ-specific contribution to the immune response against HCMV is still poorly understood. In view of the opposite roles of M1 and M2 Mϕ during initiation and resolution of the immune response, we characterized the effects of HCMV infection on classically activated M1 Mϕ and alternatively activated M2 Mϕ. Although HCMV susceptibility was higher in M2 Mϕ, HCMV established a productive and persistent infection in both types of Mϕ. Upon HCMV encounter, both types of Mϕ acquired similar features of classical activation and secreted high levels of proinflammatory cytokines and chemokines. As a functional consequence, conditioned media obtained from HCMV-infected M1 and M2 Mϕ potently activated freshly isolated monocytes. Finally, compared to HCMV-infected monocyte-derived dendritic cells, infected M1 and M2 Mϕ were more efficient in stimulating proliferation of autologous T cells from HCMV-seropositive donors at early times (24 h) postinfection, while the Mϕ immunostimulatory properties were reduced, but not abrogated, at later times (72 h postinfection). In summary, our findings indicate that Mϕ preserve proper antigen presentation capacity upon HCMV infection while enhancing inflammation, thus suggesting that Mϕ play a role in the maintenance of the large HCMV-specific T-cell repertoire in seropositive individuals.