Gut microbiome: A possible common therapeutic target for treatment of atherosclerosis and cancer.

Human gut microbiota is a dynamic and variable system that can change over time and in response to different diets and treatments. There is currently no doubt that gut microbiota can provide interesting therapeutic opportunities, since it can metabolize biologically active molecules, drugs, and their precursors, and control their bioavailability. Moreover, it can produce both beneficial and dangerous metabolites that influence host's health. In this review, we summarize the current knowledge on the involvement of gut microbiota in two chronic human pathologies that represent the greatest challenges of modern medicine: atherosclerosis and cancer. Interesting parallels are observed between the mechanisms and possible treatment approaches of these pathologies. Some of the common effects of therapeutic agents targeting both pathologies, such as anti-inflammatory activity, are partially mediated by the gut microbiota. We will discuss the effects of common drugs (metformin, statins and aspirin) and various nutraceuticals on gut microbiota and outline the pathways of microbial involvement in mediating the pleiotropic beneficial effects of these agents in atherosclerosis and cancer.

Loss of swiss cheese in Neurons Contributes to Neurodegeneration with Mitochondria Abnormalities, Reactive Oxygen Species Acceleration and Accumulation of Lipid Droplets in Drosophila Brain.

Various neurodegenerative disorders are associated with human NTE/PNPLA6 dysfunction. Mechanisms of neuropathogenesis in these diseases are far from clearly elucidated. Hereditary spastic paraplegia belongs to a type of neurodegeneration associated with NTE/PNLPLA6 and is implicated in neuron death. In this study, we used Drosophila melanogaster to investigate the consequences of neuronal knockdown of swiss cheese (sws)-the evolutionarily conserved ortholog of human NTE/PNPLA6-in vivo. Adult flies with the knockdown show longevity decline, locomotor and memory deficits, severe neurodegeneration progression in the brain, reactive oxygen species level acceleration, mitochondria abnormalities and lipid droplet accumulation. Our results suggest that SWS/NTE/PNPLA6 dysfunction in neurons induces oxidative stress and lipid metabolism alterations, involving mitochondria dynamics and lipid droplet turnover in neurodegeneration pathogenesis. We propose that there is a complex mechanism in neurological diseases such as hereditary spastic paraplegia, which includes a stress reaction, engaging mitochondria, lipid droplets and endoplasmic reticulum interplay.

Genetics of Arterial-Wall-Specific Mechanisms in Atherosclerosis: Focus on Mitochondrial Mutations.

PURPOSE OF REVIEW: Mutations in both nuclear and mitochondrial genes are associated with the development of atherosclerotic lesions in arteries and may provide a partial explanation to the focal nature of lesion distribution in the arterial wall. This review is aimed to discuss the genetic aspects of atherogenesis with a special focus on possible pro-atherogenic variants (mutations) of the nuclear and mitochondrial genomes that may be implicated in atherosclerosis development and progression. RECENT FINDINGS: Mutations in the nuclear genes generally do not cause a phenotype restricted to a specific vascular wall cell and manifest themselves mostly at the organism level. Such mutations can act as important contributors to changes in lipid metabolism and modulate other risk factors of atherosclerosis. By contrast, mitochondrial DNA (mtDNA) mutations occurring locally in the arterial wall cells or in circulating immune cells may play a site-specific role in atherogenesis. The mosaic distribution of heteroplasmic mtDNA mutations in the arterial wall tissue may explain, at least to some extent, the locality and focality of atherosclerotic lesions distribution. The genetic mechanisms of atherogenesis include alterations of both nuclear and mitochondrial genomes. Altered lipid metabolism and inflammatory response of resident arterial wall and circulating immune cells may be related to mtDNA damage and defective mitophagy, which hinders clearance of dysfunctional mitochondria. Mutations of mtDNA can have mosaic distribution and locally affect functionality of endothelial and subendothelial intimal cells in the arterial wall contributing to atherosclerotic lesion development.

Thermal effects of monopolar electrosurgery detected by real-time infrared thermography: an experimental appendectomy study.

BACKGROUND: Monopolar energy (ME) is routinely used in appendectomy. This study aimed to investigate the degree of lateral thermal spread generated by ME and to evaluate the thermal injury sustained by the close-lying tissues. METHODS: Appendectomy with a monopolar Maryland dissector was performed in 8 rabbits (at 30 and 60 W power settings). A high-resolution infrared camera was used to record tissue heating during the intervention. After autopsy macroscopic changes were evaluated and tissue samples were subjected to myeloperoxidase (MPO) assay and histological examination. RESULTS: No significant differences in the extent of thermal spread, MPO activity and histological signs of inflammation were observed between groups. Regardless of the power settings, the heat spread exceeded 2 cm laterally along the mesoappendix when application time exceeded 3 s. The spread of heat through tubular structures in both groups caused a significant temperature rise in the nearby intestinal loop, resulting in perforation (n = 3) and necrosis (n = 1). CONCLUSIONS: Application time is critical in thermal spread during appendectomy aided by ME. Tubular anatomic structures can enhance thermal injury on distant tissues. The observed effects of ME bear clinical relevance that need further investigation.

Noninvasive Testing for Diagnosis of Stable Coronary Artery Disease in the Elderly.

Efficient diagnostic approaches to detect coronary artery disease (CAD) in elderly patients are necessary to ensure optimal and timely treatment. The population of suspected CAD patients older than 70 years is especially vulnerable and constantly growing. Finding the optimal diagnostic approach is challenging due to certain features of this population, such as high prevalence of comorbidities, existing contraindications to exercise tests or cognitive decline, which hinders correct assessment of the patient's situation. Moreover, some symptoms of CAD can have variable significance in the elderly compared to younger adult groups. In this review, we present current recommendations of the United States (US) and European cardiologists' associations and discuss their applicability for diagnostics in the elderly population. Exercise electrocardiogram (ECG) and exercise stress echocardiography (SE) tests are not feasible for a substantial proportion of elderly patients. Coronary computed tomography angiography (CTA) appears to be an attractive alternative for such patients, but is not universally applicable; for instance, it is problematic in patients with significant calcification of the vessels. Moreover, more studies are needed to compare the results delivered by CTA to those of other diagnostic methods. Future efforts should be focused on comparative studies to better understand the limits and advantages of different diagnostic methods and their combinations. It is possible that some of the currently used diagnostic criteria could be improved to better accommodate the needs of the elderly population.

Cystinosin deficiency causes podocyte damage and loss associated with increased cell motility.

The involvement of the glomerulus in the pathogenesis of cystinosis, caused by loss-of-function mutations in cystinosin (CTNS, 17p13), is a matter of controversy. Although patients with cystinosis demonstrate glomerular lesions and high-molecular-weight proteinuria starting from an early age, a mouse model of cystinosis develops only signs of proximal tubular dysfunction. Here we studied podocyte damage in patients with cystinosis by analyzing urinary podocyte excretion and by in vitro studies of podocytes deficient in cystinosin. Urine from patients with cystinosis presented a significantly higher amount of podocytes compared with controls. In culture, cystinotic podocytes accumulated cystine compatible with cystinosin deficiency. The expression of podocyte specific genes CD2AP, podocalyxin, and synaptopodin and of the WT1 protein was evident in all cell lines. Conditionally immortalized podocyte lines of 2 patients with different CTNS mutations had altered cytoskeleton, impaired cell adhesion sites, and increased individual cell motility. Moreover, these cells showed enhanced phosphorylation of both Akt1 and Akt2 (isoforms of protein kinase B). Inhibition of Akt by a specific inhibitor (Akti inhibitor 1/2) resulted in normalization of the hypermotile phenotype. Thus, our study extends the list of genetic disorders causing podocyte damage and provides the evidence of altered cell signaling cascades resulting in impaired cell adhesion and enhanced cell motility in cystinosis.

Altered mTOR signalling in nephropathic cystinosis.

Lysosomes play a central role in regulating autophagy via activation of mammalian target of rapamycin complex 1 (mTORC1). We examined mTORC1 signalling in the lysosomal storage disease nephropathic cystinosis (MIM 219800), in which accumulation of autophagy markers has been previously demonstrated. Cystinosis is caused by mutations in the lysosomal cystine transporter cystinosin and initially affects kidney proximal tubules causing renal Fanconi syndrome, followed by a gradual development of end-stage renal disease and extrarenal complications. Using proximal tubular kidney cells obtained from healthy donors and from cystinotic patients, we demonstrate that cystinosin deficiency is associated with a perturbed mTORC1 signalling, delayed reactivation of mTORC1 after starvation and abnormal lysosomal retention of mTOR during starvation. These effects could not be reversed by treatment with cystine-depleting drug cysteamine. Altered mTORC1 signalling can contribute to the development of proximal tubular dysfunction in cystinosis and points to new possibilities in therapeutic intervention through modulation of mTORC-dependent signalling cascades.

The Role of Endoplasmic Reticulum Stress and Unfolded Protein Response in Atherosclerosis.

Pathogenesis of atherosclerosis is a complex process involving several metabolic and signalling pathways. Accumulating evidence demonstrates that endoplasmic reticulum stress and associated apoptosis can be induced in the pathological conditions of atherosclerotic lesions and contribute to the disease progression. Notably, they may play a role in the development of vulnerable plaques that induce thrombosis and are therefore especially dangerous. Endoplasmic reticulum stress response is regulated by several signaling mechanisms that involve protein kinases and transcription factors. Some of these molecules can be regarded as potential therapeutic targets to improve treatment of atherosclerosis. In this review we will discuss the role of endoplasmic reticulum stress and apoptosis in atherosclerosis development in different cell types and summarize the current knowledge on potential therapeutic agents targeting molecules regulating these pathways and their possible use for anti-atherosclerotic therapy.

Study of the activated macrophage transcriptome.

Transcriptome analysis is a powerful modern tool to study possible alterations of gene expression associated with human diseases. It turns out to be especially promising for evaluation of gene expression changes in immunopathology, as immune cells have flexible gene expression patterns that can be switched in response to infection, inflammatory stimuli and exposure to various cytokines. In particular, macrophage polarization towards pro-inflammatory (M1) and anti-inflammatory (M2) phenotypes can be successfully studied using the modern transcriptome analysis approaches. The two mostly used techniques for transcriptome analysis are microarray and next generation sequencing. In this review we will provide an overview of known gene expression changes associated with immunopathology and discuss the advantage and limitations of different methods of transcriptome analysis.

Phenomenon of individual difference in human monocyte activation.

Macrophages play an important role in the pathogenesis of atherosclerosis, including the early pre-clinical stages of the disease development. We have explored the possibility that the disease onset could be associated with altered monocyte/macrophage response to activating pro- and anti-inflammatory stimuli. We evaluated the susceptibility of circulating monocytes from healthy individuals and patients with asymptomatic carotid atherosclerosis to M1 and M2 activation. The obtained data indicated the existence of a remarkable individual difference in susceptibility to activation among monocytes isolated from the blood of different subjects, regardless of the presence or absence of atherosclerosis. The identified differences in susceptibility to activation between monocytes may explain the individual peculiarities of the immune response in different subjects.

Structure-transfection activity relationships in a series of novel cationic lipids with heterocyclic head-groups.

Cationic liposomes are promising candidates for the delivery of various therapeutic nucleic acids. Here, we report a convenient synthesis of carbamate-type cationic lipids with various hydrophobic domains (tetradecanol, dialkylglycerol, cholesterol) and positively charged head-groups (pyridinium, N-methylimidazolium, N-methylmorpholinium) and data on the structure-transfection activity relationships. It was found that single-chain lipids possess high surface activity, which correlates with high cytotoxicity due to their ability to disrupt the cellular membrane by combined hydrophobic and electrostatic interactions. Liposomes containing these lipids also display high cytotoxicity with respect to all cell lines. Irrespective of chemical structures, all cationic lipids form liposomes with similar sizes and surface potentials. The characteristics of complexes composed of cationic liposomes and nucleic acids depend mostly on the type of nucleic acid and P/N ratios. In the case of oligodeoxyribonucleotide delivery, the transfection activity depends on the type of cationic head-group regardless of the type of hydrophobic domain: all types of cationic liposomes mediate efficient oligonucleotide transfer into 80-90% of the eukaryotic cells, and liposomes based on lipids with N-methylmorpholinium cationic head-group display the highest transfection activity. In the case of plasmid DNA and siRNA, the type of hydrophobic domain determines the transfection activity: liposomes composed of cholesterol-based lipids were the most efficient in DNA transfer, while liposomes containing glycerol-based lipids exhibited reasonable activity in siRNA delivery under serum-free conditions.

Drosophila Lysophospholipase Gene swiss cheese Is Required for Survival and Reproduction.

Drosophila melanogaster is one of the most famous insects in biological research. It is widely used to analyse functions of different genes. The phosphatidylcholine lysophospholipase gene swiss cheese was initially shown to be important in the fruit fly nervous system. However, the role of this gene in non-nervous cell types has not been elucidated yet, and the evolutional explanation for the conservation of its function remains elusive. In this study, we analyse expression pattern and some aspects of the role of the swiss cheese gene in the fitness of Drosophila melanogaster. We describe the spatiotemporal expression of swiss cheese throughout the fly development and analyse the survival and productivity of swiss cheese mutants. We found swiss cheese to be expressed in salivary glands, midgut, Malpighian tubes, adipocytes, and male reproductive system. Dysfunction of swiss cheese results in severe pupae and imago lethality and decline of fertility, which is impressive in males. The latter is accompanied with abnormalities of male locomotor activity and courtship behaviour, accumulation of lipid droplets in testis cyst cells and decrease in spermatozoa motility. These results suggest that normal swiss cheese is important for Drosophila melanogaster fitness due to its necessity for both specimen survival and their reproductive success.

Morpho-Functional Consequences of Swiss Cheese Knockdown in Glia of Drosophila melanogaster.

Glia are crucial for the normal development and functioning of the nervous system in many animals. Insects are widely used for studies of glia genetics and physiology. Drosophila melanogaster surface glia (perineurial and subperineurial) form a blood-brain barrier in the central nervous system and blood-nerve barrier in the peripheral nervous system. Under the subperineurial glia layer, in the cortical region of the central nervous system, cortex glia encapsulate neuronal cell bodies, whilst in the peripheral nervous system, wrapping glia ensheath axons of peripheral nerves. Here, we show that the expression of the evolutionarily conserved swiss cheese gene is important in several types of glia. swiss cheese knockdown in subperineurial glia leads to morphological abnormalities of these cells. We found that the number of subperineurial glia nuclei is reduced under swiss cheese knockdown, possibly due to apoptosis. In addition, the downregulation of swiss cheese in wrapping glia causes a loss of its integrity. We reveal transcriptome changes under swiss cheese knockdown in subperineurial glia and in cortex + wrapping glia and show that the downregulation of swiss cheese in these types of glia provokes reactive oxygen species acceleration. These results are accompanied by a decline in animal mobility measured by the negative geotaxis performance assay.

Possible Role of Mitochondrial DNA Mutations in Chronification of Inflammation: Focus on Atherosclerosis.

Chronification of inflammation is the process that lies at the basis of several human diseases that make up to 80% of morbidity and mortality worldwide. It can also explain a great deal of processes related to aging. Atherosclerosis is an example of the most important chronic inflammatory pathology in terms of public health impact. Atherogenesis is based on the inflammatory response of the innate immunity arising locally or focally. The main trigger for this response appears to be modified low-density lipoprotein (LDL), although other factors may also play a role. With the quick resolution of inflammation, atherosclerotic changes in the arterial wall do not occur. However, a violation of the innate immunity response can lead to chronification of local inflammation and, as a result, to atherosclerotic lesion formation. In this review, we discuss possible mechanisms of the impaired immune response with a special focus on mitochondrial dysfunction. Some mitochondrial dysfunctions may be due to mutations in mitochondrial DNA. Several mitochondrial DNA mutations leading to defective mitophagy have been identified. The regulatory role of mitophagy in the immune response has been shown in recent studies. We suggest that defective mitophagy promoted by mutations in mitochondrial DNA can cause innate immunity disorders leading to chronification of inflammation.

NADPH Oxidases and Their Role in Atherosclerosis.

The current view on atherosclerosis positions it as a multifactorial disorder that results from the interplay between lipid metabolism disturbances and inflammatory processes. Oxidative stress is proven to be one of the initiating factors in atherosclerosis development, being implicated both in the inflammatory response and in atherogenic modifications of lipoproteins that facilitate lipid accumulation in the arterial wall. The hallmark of oxidative stress is the elevated level of reactive oxygen species (ROS). Correspondingly, the activity of major ROS-generating enzymes, including nicotinamide adenine dinucleotide phosphate (NADPH) oxidases, xanthine oxidases, and cyclooxygenases, is an important element in atherosclerosis development. In particular, the role of NADPH oxidases in atherosclerosis development has become a subject of intensive research. Aberrant activity of NADPH oxidases was shown to be associated with cardiovascular disease in humans. With regard to atherosclerosis, several important pathological components of the disease development, including endothelial dysfunction, inflammation, and vascular remodeling, involve aberrations in NADPH oxidases functioning. In humans, NADPH oxidases are represented by four isoforms expressed in vascular tissues, where they serve as the main source of ROS during atherogenesis. Moreover, recent studies have demonstrated their impact on vascular remodeling processes. Interestingly, one of the NADPH oxidase isoforms, NOX4, was shown to have an atheroprotective effect. Despite the growing evidence of the crucial involvement of NADPH oxidases in atherosclerosis pathogenesis, the available data still remains controversial. In this narrative review, we summarize the current knowledge of the role of NADPH oxidases in atherosclerosis and outline the future directions of research.

Endoplasmic Reticulum Stress in Macrophages: The Vicious Circle of Lipid Accumulation and Pro-Inflammatory Response.

The endoplasmic reticulum (ER) stress is an important event in the pathogenesis of different human disorders, including atherosclerosis. ER stress leads to disturbance of cellular homeostasis, apoptosis, and in the case of macrophages, to foam cell formation and pro-inflammatory cytokines production. In atherosclerosis, several cell types can be affected by ER stress, including endothelial cells, vascular smooth muscular cells, and macrophages. Modified low-density lipoproteins (LDL) and cytokines, in turn, can provoke ER stress through different processes. The signaling cascades involved in ER stress initiation are complex and linked to other cellular processes, such as lysosomal biogenesis and functioning, autophagy, mitochondrial homeostasis, and energy production. In this review, we discuss the underlying mechanisms of ER stress formation and the interplay of lipid accumulation and pro-inflammatory response. We will specifically focus on macrophages, which are the key players in maintaining chronic inflammatory milieu in atherosclerotic lesions, and also a major source of lipid-accumulating foam cells.

The Role of Mitochondria in Cardiovascular Diseases.

The role of mitochondria in cardiovascular diseases is receiving ever growing attention. As a central player in the regulation of cellular metabolism and a powerful controller of cellular fate, mitochondria appear to comprise an interesting potential therapeutic target. With the development of DNA sequencing methods, mutations in mitochondrial DNA (mtDNA) became a subject of intensive study, since many directly lead to mitochondrial dysfunction, oxidative stress, deficient energy production and, as a result, cell dysfunction and death. Many mtDNA mutations were found to be associated with chronic human diseases, including cardiovascular disorders. In particular, 17 mtDNA mutations were reported to be associated with ischemic heart disease in humans. In this review, we discuss the involvement of mitochondrial dysfunction in the pathogenesis of atherosclerosis and describe the mtDNA mutations identified so far that are associated with atherosclerosis and its risk factors.

Mitochondrion as a Selective Target for the Treatment of Atherosclerosis: Role of Mitochondrial DNA Mutations and Defective Mitophagy in the Pathogenesis of Atherosclerosis and Chronic Inflammation.

BACKGROUND: Atherosclerosis is a chronic inflammatory condition that affects different arteries in the human body and often leads to severe neurological complications, such as stroke and its sequelae. Affected blood vessels develop atherosclerotic lesions in the form of focal thickening of the intimal layer, so called atherosclerotic plaques. OBJECTIVES: Despite the high priority of atherosclerosis research for global health and the numerous preclinical and clinical studies conducted, currently, there is no effective pharmacological treatment that directly impacts atherosclerotic plaques. Many knowledge gaps exist in our understanding of the mechanisms of plaque formation. In this review, we discuss the role of mitochondria in different cell types involved in atherogenesis and provide information about mtDNA mutations associated with the disease. RESULTS: Mitochondria of blood and arterial wall cells appear to be one of the important factors in disease initiation and development. Significant experimental evidence connects oxidative stress associated with mitochondrial dysfunction and vascular disease. Moreover, mitochondrial DNA (mtDNA) deletions and mutations are being considered as potential disease markers. Further study of mtDNA damage and associated dysfunction may open new perspectives for atherosclerosis treatment. CONCLUSION: Mitochondria can be considered as important disease-modifying factors in several chronic pathologies. Deletions and mutations of mtDNA may be used as potential disease markers. Mitochondria-targeting antioxidant therapies appear to be promising for the development of treatment of atherosclerosis and other diseases associated with oxidative stress and chronic inflammation.

The difference between cellulolytic 'culturomes' and microbiomes inhabiting two contrasting soil types.

High-throughput 16S rRNA sequencing was performed to compare the microbiomes inhabiting two contrasting soil types-sod-podzolic soil and chernozem-and the corresponding culturome communities of potentially cellulolytic bacteria cultured on standard Hutchinson media. For each soil type, soil-specific microorganisms have been identified: for sod-podzolic soil-Acidothermus, Devosia, Phenylobacterium and Tumebacillus, and for chernozem soil-Sphingomonas, Bacillus and Blastococcus. The dynamics of differences between soil types for bulk soil samples and culturomes varied depending on the taxonomic level of the corresponding phylotypes. At high taxonomic levels, the number of common taxa between soil types increased more slowly for bulk soil than for culturome. Differences between soil-specific phylotypes were detected in bulk soil at a low taxonomic level (genus, species). A total of 13 phylotypes were represented both in soil and in culturome. No relationship was shown between the abundance of these phylotypes in soil and culturome.

Optical Mapping-Validated Machine Learning Improves Atrial Fibrillation Driver Detection by Multi-Electrode Mapping.

BACKGROUND: Atrial fibrillation (AF) can be maintained by localized intramural reentrant drivers. However, AF driver detection by clinical surface-only multielectrode mapping (MEM) has relied on subjective interpretation of activation maps. We hypothesized that application of machine learning to electrogram frequency spectra may accurately automate driver detection by MEM and add some objectivity to the interpretation of MEM findings. METHODS: Temporally and spatially stable single AF drivers were mapped simultaneously in explanted human atria (n=11) by subsurface near-infrared optical mapping (NIOM; 0.3 mm2 resolution) and 64-electrode MEM (higher density or lower density with 3 and 9 mm2 resolution, respectively). Unipolar MEM and NIOM recordings were processed by Fourier transform analysis into 28 407 total Fourier spectra. Thirty-five features for machine learning were extracted from each Fourier spectrum. RESULTS: Targeted driver ablation and NIOM activation maps efficiently defined the center and periphery of AF driver preferential tracks and provided validated annotations for driver versus nondriver electrodes in MEM arrays. Compared with analysis of single electrogram frequency features, averaging the features from each of the 8 neighboring electrodes, significantly improved classification of AF driver electrograms. The classification metrics increased when less strict annotation, including driver periphery electrodes, were added to driver center annotation. Notably, f1-score for the binary classification of higher-density catheter data set was significantly higher than that of lower-density catheter (0.81±0.02 versus 0.66±0.04, P<0.05). The trained algorithm correctly highlighted 86% of driver regions with higher density but only 80% with lower-density MEM arrays (81% for lower-density+higher-density arrays together). CONCLUSIONS: The machine learning model pretrained on Fourier spectrum features allows efficient classification of electrograms recordings as AF driver or nondriver compared with the NIOM gold-standard. Future application of NIOM-validated machine learning approach may improve the accuracy of AF driver detection for targeted ablation treatment in patients.