Comparative analysis of calcified soft tissues revealed shared deregulated pathways

IntroductionCalcification of soft tissues is a common age-related pathology that primarily occurs within vascular tissue. The mechanisms underlying pathological calcification in humans and tissue specificity of the process is still poorly understood. Previous studies examined calcified tissues on one to one basis, thus preventing comparison of deregulated pathways across tissues.PurposeThis study aimed to establish common and tissue-specific changes associated with calcification in aorta, artery tibial, coronary artery and pituitary gland in subjects from the Genotype-Tissue Expression (GTEx) dataset using its RNA sequencing and histological data.MethodsWe used publicly available data from the GTEx database https://gtexportal.org/home/aboutGTEx. All GTEx tissue samples were derived by the GTEx consorcium from deceased donors, with age from 20 to 79, both men and women. GTEx study authorization was obtained via next-of-kin consent for the collection and banking of de-identified tissue samples for scientific research. Hematoxylin and eosin (H&E) staining of arteries were manually graded based on the presence of calcification on a scale from zero to four, where zero designates absence of calcification and four designates severe calcification. Samples with fat contamination and mislabeled tissues were excluded, which left 430 aorta, 595 artery tibial, 124 coronary artery, and 283 pituitary samples for downstream gene expression analysis. Transcript levels of protein-coding genes were associated with calcification grade using sex, age bracket and cause of death as covariates, and tested for pathway enrichment using gene set enrichment analysis.ResultsWe identified calcification deposits in 28 (6.5%) aortas, 121 (20%), artery tibials, 54 (43%), coronary arteries, and 24 (8%) pituitary glands of GTEx subjects. We observed an age-dependent increase in incidence of calcification in all vascular tissues, but not in pituitary. Subjects with calcification in the artery tibial were significantly more likely to have calcification in the coronary artery (OR = 2.56, p = 6.3e-07). Markers of calcification previously established in preclinical and in vitro studies, e.g., BMP2 and RUNX2, were deregulated in the calcified tibial and coronary arteries, confirming the relevance of these genes to human pathology. Differentially expressed genes associated with calcification poorly overlapped across tissues suggesting tissue-specific nuances in mechanisms of calcification. Nevertheless, calcified arteries unanimously down-regulated pathways of intracellular transport and up-regulated inflammatory pathways suggesting these as universal targets for pathological calcification. In particular, PD-1 and PD-L1 genes were up-regulated in calcified tissues but not in the blood of the same subjects, suggesting that localized inflammation contributes to pathological calcification.ConclusionPathological calcification is a prevalent disease of aging that shares little changes in expression in individual genes across tissues. However, our analysis suggests that it potentially can be targeted by alleviating local inflammation of soft tissues

Identification of differentially expressed micro- and mRNA clusters in blood of patients with coronary artery disease: intersection with calcification pathways and implications in cardio-vascular diseases

<p>Coronary artery disease (CAD) is one of the main and common types of cardio-vascular diseases, which occurs as a result of atherosclerotic plaque formation in blood vessels during ageing. Increased levels of calcium deposition were detected in blood vessels of patients with CAD and linked to early CAD formation. Here, we hypothesized that these two processes are linked and possess common molecular basis, which is realized through regulation of target genes involved in calcification and CAD.</p> <p>To validate our hypothesis, we used a bioinformatics approach, in which micro- and mRNA data of blood samples from 12 patients with CAD were analyzed. Several high quality data sets were extracted from GEO database, analyzed by commercially available program TR GeneGo Metacore, statistically verified by J-express algorithms and analyzed using statistical methods implemented in R.</p> <p>Analysis of GEO data revealed patterns of differentially expressed microRNAs in patients with CAD. We have shown the involvement of the differentially expressed microRNAs (hsa-miR-154; hsa-miR-451; hsa-miR-17) in signalling pathways associated with vascular calcification. Additionally, we obtained data of mRNA expression, showing several clusters of genes, which are involved in vascular calcification pathways in patients with CAD and developed an interaction map of differentially expressed genes in the blood of CAD patients and genes associated with vascular calcification.</p> <p>We conclude that patients with CAD have several differentially expressed gene markers, which may have different impact on calcification development. Data provided may have a positive outcome for cardio-vascular diagnosis strategy and personalized medicine.</p

An Alternative Nested Reading Frame May Participate in the Stress-Dependent Expression of a Plant Gene

Although plants as sessile organisms are affected by a variety of stressors in the field, the stress factors for the above-ground and underground parts of the plant and their gene expression profiles are not the same. Here, we investigated NbKPILP, a gene encoding a new member of the ubiquitous, pathogenesis-related Kunitz peptidase inhibitor (KPI)-like protein family, that we discovered in the genome of Nicotiana benthamiana and other representatives of the Solanaceae family. The NbKPILP gene encodes a protein that has all the structural elements characteristic of KPI but in contrast to the proven A. thaliana KPI (AtKPI), it does not inhibit serine peptidases. Unlike roots, NbKPILP mRNA and its corresponding protein were not detected in intact leaves, but abiotic and biotic stressors drastically affected NbKPILP mRNA accumulation. In search of the causes of suppressed NbKPILP mRNA accumulation in leaves, we found that the NbKPILP gene is “matryoshka,” containing an alternative nested reading frame (ANRF) encoding a 53-amino acid (aa) polypeptide (53aa-ANRF) which has an amphipathic helix (AH). We confirmed ANRF expression experimentally. A vector containing a GFP-encoding sequence was inserted into the NbKPILP gene in frame with 53aa-ANRF, resulting in a 53aa-GFP fused protein that localized in the membrane fraction of cells. Using the 5′-RACE approach, we have shown that the expression of ANRF was not explained by the existence of a cryptic promoter within the NbKPILP gene but was controlled by the maternal NbKPILP mRNA. We found that insertion of mutations destroying the 53aa-ANRF AH resulted in more than a two-fold increase of the NbKPILP mRNA level. The NbKPILP gene represents the first example of ANRF functioning as a repressor of a maternal gene in an intact plant. We proposed a model where the stress influencing the translation initiation promotes the accumulation of NbKPILP and its mRNA in leaves

The Intergenic Interplay between Aldose 1-Epimerase-Like Protein and Pectin Methylesterase in Abiotic and Biotic Stress Control

The mechanical damage that often precedes the penetration of a leaf by a pathogen promotes the activation of pectin methylesterase (PME); the activation of PME leads to the emission of methanol, resulting in a “priming” effect on intact leaves, which is accompanied by an increased sensitivity to Tobacco mosaic virus (TMV) and resistance to bacteria. In this study, we revealed that mRNA levels of the methanol-inducible gene encoding Nicotiana benthamiana aldose 1-epimerase-like protein (NbAELP) in the leaves of intact plants are very low compared with roots. However, stress and pathogen attack increased the accumulation of the NbAELP mRNA in the leaves. Using transiently transformed plants, we obtained data to support the mechanism underlying AELP/PME-related negative feedback The insertion of the NbAELP promoter sequence (proNbAELP) into the N. benthamiana genome resulted in the co-suppression of the natural NbAELP gene expression, accompanied by a reduction in the NbAELP mRNA content and increased PME synthesis. Knockdown of NbAELP resulted in high activity of PME in the cell wall and a decrease in the leaf glucose level, creating unfavorable conditions for Agrobacterium tumefaciens reproduction in injected leaves. Our results showed that NbAELP is capable of binding the TMV movement protein (MPTMV) in vitro and is likely to affect the cellular nucleocytoplasmic transport, which may explain the sensitivity of NbAELP knockdown plants to TMV. Although NbAELP was primarily detected in the cell wall, the influence of this protein on cellular PME mRNA levels might be associated with reduced transcriptional activity of the PME gene in the nucleus. To confirm this hypothesis, we isolated the N. tabacum PME gene promoter (proNtPME) and showed the inhibition of proNtPME-directed GFP and GUS expression in leaves when co-agroinjected with the NbAELP-encoding plasmid. We hypothesized that plant wounding and/or pathogen attack lead to PME activation and increased methanol emission, followed by increased NbAELP expression, which results in reversion of PME mRNA level and methanol emission to levels found in the intact plant

The Antioxidant Cofactor Alpha-Lipoic Acid May Control Endogenous Formaldehyde Metabolism in Mammals

The healthy human body contains small amounts of metabolic formaldehyde (FA) that mainly results from methanol oxidation by pectin methylesterase, which is active in a vegetable diet and in the gastrointestinal microbiome. With age, the ability to maintain a low level of FA decreases, which increases the risk of Alzheimer's disease and dementia. It has been shown that 1,2-dithiolane-3-pentanoic acid or alpha lipoic acid (ALA), a naturally occurring dithiol and antioxidant cofactor of mitochondrial α-ketoacid dehydrogenases, increases glutathione (GSH) content and FA metabolism by mitochondrial aldehyde dehydrogenase 2 (ALDH2) thus manifests a therapeutic potential beyond its antioxidant property. We suggested that ALA can contribute to a decrease in the FA content of mammals by acting on ALDH2 expression. To test this assumption, we administered ALA in mice in order to examine the effect on FA metabolism and collected blood samples for the measurement of FA. Our data revealed that ALA efficiently eliminated FA in mice. Without affecting the specific activity of FA-metabolizing enzymes (ADH1, ALDH2, and ADH5), ALA increased the GSH content in the brain and up-regulated the expression of the FA-metabolizing ALDH2 gene in the brain, particularly in the hippocampus, but did not impact its expression in the liver in vivo or in rat liver isolated from the rest of the body. After ALA administration in mice and in accordance with the increased content of brain ALDH2 mRNA, we detected increased ALDH2 activity in brain homogenates. We hypothesized that the beneficial effects of ALA on patients with Alzheimer's disease may be associated with accelerated ALDH2-mediated FA detoxification and clearance

Genetic and phenotypic analysis of the causal relationship between aging and COVID-19

Background: Epidemiological studies revealed that the elderly and those with comorbidities are most affected by COVID-19, but it is important to investigate shared genetic mechanisms between COVID-19 risk and aging. Methods: We conducted a multi-instrument Mendelian Randomization analysis of multiple lifespan-related traits and COVID-19. Aging clock models were applied to the subjects with different COVID-19 conditions in the UK-Biobank cohort. We performed a bivariate genomic scan for age-related COVID-19 and Mendelian Randomization analysis of 389 immune cell traits to investigate their effect on lifespan and COVID-19 risk. Results: We show that the genetic variation that supports longer life is significantly associated with the lower risk of COVID-19 infection and hospitalization. The odds ratio is 0.31 (P = 9.7 × 10-6) and 0.46 (P = 3.3 × 10-4), respectively, per additional 10 years of life. We detect an association between biological age acceleration and future incidence and severity of COVID-19 infection. Genetic profiling of age-related COVID-19 infection indicates key contributions of Notch signaling and immune system development. We reveal a negative correlation between the effects of immune cell traits on lifespan and COVID-19 risk. We find that lower B-cell CD19 levels are indicative of an increased risk of COVID-19 and decreased life expectancy, which is further validated by COVID-19 clinical data. Conclusions: Our analysis suggests that the factors that accelerate aging lead to an increased COVID-19 risk and point to the importance of Notch signaling and B cells in both. Interventions that target these factors to reduce biological age may reduce the risk of COVID-19

Downregulation of mitochondrial metabolism is a driver for fast skeletal muscle loss during mouse aging

Abstract Skeletal muscle aging is characterized by the loss of muscle mass, strength and function, mainly attributed to the atrophy of glycolytic fibers. Underlying mechanisms driving the skeletal muscle functional impairment are yet to be elucidated. To unbiasedly uncover its molecular mechanisms, we recurred to gene expression and metabolite profiling in a glycolytic muscle, Extensor digitorum longus (EDL), from young and aged C57BL/6JRj mice. Employing multi-omics approaches we found that the main age-related changes are connected to mitochondria, exhibiting a downregulation in mitochondrial processes. Consistent is the altered mitochondrial morphology. We further compared our mouse EDL aging signature with human data from the GTEx database, reinforcing the idea that our model may recapitulate muscle loss in humans. We are able to show that age-related mitochondrial downregulation is likely to be detrimental, as gene expression signatures from commonly used lifespan extending interventions displayed the opposite direction compared to our EDL aging signature

Endogenous Methanol Regulates Mammalian Gene Activity

<div><p>We recently showed that methanol emitted by wounded plants might function as a signaling molecule for plant-to-plant and plant-to-animal communications. In mammals, methanol is considered a poison because the enzyme alcohol dehydrogenase (ADH) converts methanol into toxic formaldehyde. However, the detection of methanol in the blood and exhaled air of healthy volunteers suggests that methanol may be a chemical with specific functions rather than a metabolic waste product. Using a genome-wide analysis of the mouse brain, we demonstrated that an increase in blood methanol concentration led to a change in the accumulation of mRNAs from genes primarily involved in detoxification processes and regulation of the alcohol/aldehyde dehydrogenases gene cluster. To test the role of ADH in the maintenance of low methanol concentration in the plasma, we used the specific ADH inhibitor 4-methylpyrazole (4-MP) and showed that intraperitoneal administration of 4-MP resulted in a significant increase in the plasma methanol, ethanol and formaldehyde concentrations. Removal of the intestine significantly decreased the rate of methanol addition to the plasma and suggested that the gut flora may be involved in the endogenous production of methanol. ADH in the liver was identified as the main enzyme for metabolizing methanol because an increase in the methanol and ethanol contents in the liver homogenate was observed after 4-MP administration into the portal vein. Liver mRNA quantification showed changes in the accumulation of mRNAs from genes involved in cell signalling and detoxification processes. We hypothesized that endogenous methanol acts as a regulator of homeostasis by controlling the mRNA synthesis.</p></div