Atherosclerotic Plaque Samples Research Articles

Microflora of atherosclerotic plaques and blood of patients with atherosclerosis

Background. Over the past decades, the role of the microbial factor in the development of atherosclerosis has been discussed. To date, data on the composition of the microflora of atherosclerotic plaques have been accumulated, but in these studies the blood microflora in atherosclerosis was not studied.
 Aim. To determine the contamination frequency of biopsy samples of atherosclerotic plaques and peripheral blood with cultured microorganisms in patients with atherosclerosis of the brachiocephalic arteries.
 Material and methods. A cross-sectional study in 35 patients with atherosclerosis of the carotid arteries; the average age of the patients was 58.2±8.3 years, was performed. Blood and atherosclerotic plaque samples from 23 men (mean age 58.6±10.3 years) and 12 women (mean age 57.3±14.3 years) were examined. Anaerobic flasks for blood cultivation were incubated at 35 °C for up to 6 months, from which they were regularly inoculated onto solid nutrient media to obtain culture growth. Cultures of tissues of atherosclerotic plaques of the carotid arteries in test tubes with thioglycollate medium were thermostated until visible growth of the culture was obtained; observation was carried out for up to 2 months. Identification of crops was carried out using commercial kits. For statistical analysis of the results obtained, the equality of the average values in two samples was checked using the Student’s statistical reliability test. Correlation analysis of the tightness of the connection was carried out using the Spearman rank correlation method.
 Results. The frequency of detection of microorganisms in samples of atherosclerotic plaques was 48.6% (17/35), including Cutibacterium acnes — in 31.4% (11/35), the genus Staphylococcus — in 22.9% (8/35), associations of these microorganisms — in 5.7% (2/35) of samples. C. acnes was cultured in the blood of 11.4% (4/35) of patients. In 5.7% (2/35) of patients, cultures of C. acnes bacteria were isolated from both atherosclerotic plaque biopsies and blood. Bacterial cultures exhibited slow growth on nutrient media. Correlation analysis of the closeness of the connection according to the Chaddock scale demonstrated the presence of a high and very high closeness of connection between the amount of high-density lipoproteins and the daily growth of C. acnes in atherosclerotic plaques (p=0.812) and blood (rxy=–0.969), the content of leukocytes showed a very high closeness of connection with days of growth of C. acnes in the blood (rxy=–0.957); Student's t-test revealed a statistically significant relationship with the fact of detection of C. acnes cultures in atherosclerotic plaques (p=0.013).
 Conclusion. Bacterial cultures isolated from blood samples and atherosclerotic plaques of patients with atherosclerosis had slow growth on nutrient media, and the period of their growth or the fact of detection correlated with the number of leukocytes and the level of high-density lipoproteins in the blood of patients.

Circadian gene CLOCK accelerates atherosclerosis by promoting endothelial autophagy

ABSTRACT Atherosclerosis (AS) is a chronic inflammatory disease which gives rise to life-threatening complications like ischemic stroke. Rupture of carotid atherosclerotic plaque is the main cause of ischemic stroke. Emerging evidence has demonstrated that disturbed circadian rhythms could accelerate the progression of atherosclerosis by regulating endothelial function. Moreover, our previous study implicated the circadian gene circadian locomotor output cycles kaput (CLOCK) in the pathogenesis of unstable plaques. In this study, we explored the underlying mechanism that CLOCK mediates endothelial cell autophagy involved in the progression of AS. Circadian and autophagy gene expression was analyzed in the GSE41571 dataset and human carotid atherosclerotic plaque samples. Then we used ox-LDL to treat HUVECs, and analyzed CLOCK and autophagy gene in endothelial cells. Besides that, we comprehensively analyzed in vivo experiments to explore the function of CLOCK in autophagy and atherosclerosis using different staining including HE, MT and IF staining. In the dataset and patient samples, CLOCK expression and autophagy were decreased in the unstable plaque group compared with the stable group. Decreased Beclin1, ATG5, LC3, and CLOCK were also observed in HUVECs under oxidative stress condition which also enhances cell proliferation. In vivo, we also found decreasing level of CLOCK, Beclin1, LC3 and ATG5 in ApoE−/− mice compared with WT mice. Silencing of CLOCK in ApoE−/− mice may further aggravate atherosclerosis including decreased cap thickness and collagens. Our findings implicated that downregulation CLOCK would impair endothelial cell autophagy and accelerate atherosclerotic plaque, which provides a novel strategy for treatment of progression in AS.

Exploring the shared molecular mechanism of microvascular and macrovascular complications in diabetes: Seeking the hub of circulatory system injury.

Microvascular complications, such as diabetic retinopathy (DR) and diabetic nephropathy (DN), and macrovascular complications, referring to atherosclerosis (AS), are the main complications of diabetes. Blindness or fatal microvascular diseases are considered to be identified earlier than fatal macrovascular complications. Exploring the intrinsic relationship between microvascular and macrovascular complications and the hub of pathogenesis is of vital importance for prolonging the life span of patients with diabetes and improving the quality of life. The expression profiles of GSE28829, GSE30529, GSE146615 and GSE134998 were downloaded from the Gene Expression Omnibus database, which contained 29 atherosclerotic plaque samples, including 16 AS samples and 13 normal controls; 22 renal glomeruli and tubules samples from diabetes nephropathy including 12 DN samples and 10 normal controls; 73 lymphoblastoid cell line samples, including 52 DR samples and 21 normal controls. The microarray datasets were consolidated and DEGs were acquired and further analyzed by bioinformatics techniques including GSEA analysis, GO-KEGG functional clustering by R (version 4.0.5), PPI analysis by Cytoscape (version 3.8.2) and String database, miRNA analysis by Diana database, and hub genes analysis by Metascape database. The drug sensitivity of characteristic DEGs was analyzed. A total of 3709, 4185 and 8086 DEGs were recognized in AS, DN, DR, respectively, with 1820, 1666, 888 upregulated and 1889, 2519, 7198 downregulated. GO and KEGG pathway analyses of DEGs and GSEA analysis of common differential genes demonstrated that these significant sites focused primarily on inflammation-oxidative stress and immune regulation pathways. PPI networks show the connection and regulation on top-250 significant sites of AS, DN, DR. MiRNA analysis explored the non-coding RNA upstream regulation network and significant pathway in AS, DN, DR. The joint analysis of multiple diseases shows the common influenced pathways of AS, DN, DR and explored the interaction between top-1000 DEGs at the same time. In the microvascular and macrovascular complications of diabetes, immune-mediated inflammatory response, chronic inflammation caused by endothelial cell activation and oxidative stress are the three links linking atherosclerosis, diabetes retinopathy and diabetes nephropathy together. Our study has clarified the intrinsic relationship and common tissue damage mechanism of microcirculation and circulatory system complications in diabetes, and explored the mechanism center of these two vascular complications. It has far-reaching clinical and social value for reducing the incidence of fatal events and early controlling the progress of disabling and fatal circulatory complications in diabetes.

Integrated investigation of DNA methylation, gene expression and immune cell population revealed immune cell infiltration associated with atherosclerotic plaque formation

BackgroundThe clinical consequences of atherosclerosis are significant source of morbidity and mortality throughout the world, while the molecular mechanisms of the pathogenesis of atherosclerosis are largely unknown.MethodsIn this study, we integrated the DNA methylation and gene expression data in atherosclerotic plaque samples to decipher the underlying association between epigenetic and transcriptional regulation. Immune cell classification was performed on the basis of the expression pattern of detected genes. Finally, we selected ten genes with dysregulated methylation and expression levels for RT-qPCR validation.ResultsGlobal DNA methylation profile showed obvious changes between normal aortic and atherosclerotic lesion tissues. We found that differentially methylated genes (DMGs) and differentially expressed genes (DEGs) were highly associated with atherosclerosis by being enriched in atherosclerotic plaque formation-related pathways, including cell adhesion and extracellular matrix organization. Immune cell fraction analysis revealed that a large number of immune cells, especially macrophages, activated mast cells, NK cells, and Tfh cells, were specifically enriched in the plaque. DEGs associated with immune cell fraction change showed that they were mainly related to the level of macrophages, monocytes, resting NK cells, activated CD4 memory T cells, and gamma delta T cells. These genes were highly enriched in multiple pathways of atherosclerotic plaque formation, including blood vessel remodeling, collagen fiber organization, cell adhesion, collagen catalogic process, extractable matrix assembly, and platelet activation. We also validated the expression alteration of ten genes associated with infiltrating immune cells in atherosclerosis.ConclusionsIn conclusion, these findings provide new evidence for understanding the mechanisms of atherosclerotic plaque formation, and provide a new and valuable research direction based on immune cell infiltration.

Assessment of periodontal disease severity in patients with cardiovascular disease: A cross-sectional study

Background: Periodontitis is a biofilm-induced chronic inflammatory disease that affects the tooth-supporting structures. Transient bacteremia during periodontal infection may lead to direct bacterial invasion of endothelial cells. Aim: To observe if the patient's periodontal status has an influence on the severity of the cardiac disease. Materials and Methods: Subgingival and atherosclerotic plaques were extracted from patients with coronary artery disease in this cross-sectional analytical study. Based on the nested polymerase chain reaction results, 44 patients were age- and gender-matched and divided into two groups based on the presence or absence of periodontal pathogens belonging to the red complex in the atherosclerotic plaque samples. Version 20.0 of IBM SPSS Statistics for the Windows was used to analyze the data. The Pearson Chi-square test was done to investigate the association between periodontal status and pathogen detection, as well as determine the relationship between periodontal severity and cardiac severity. P < 0.05 was considered to be statistically significant. Results: Statistically insignificant association was observed between periodontal severity and the detection of the periodontal pathogens Porphyromonas gingivalis and Tannerella forsythia in the subgingival and atherosclerotic plaques in Group A; however, a significant association was observed for the pathogen Treponema denticola in the subgingival plaque for both Groups A (P = 0.039) and B (P = 0.005). No significant association was observed between the periodontal severity and cardiac severity for Groups A (P = 0.40) and B (P = 0.277). A weak positive but statistically insignificant (P = 0.097) correlation was observed for overall periodontal and cardiac severity. Conclusion: A higher percentage of patients had generalized chronic periodontitis in the group that tested positive for the pathogens in the atherosclerotic plaques, indicating a possible influence of periodontal status on cardiac outcomes.

Chlamydia pneumoniae is Prevalent in Symptomatic Coronary Atherosclerotic Plaque Samples Obtained From Directional Coronary Atherectomy, but its Quantity is Not Associated With Plaque Instability: An Immunohistochemical and Molecular Study.

Aim:To clarify whether there is any association between the extent of Chlamydia pneumoniae (C. pneumoniae) infection and plaque instability or post–directional coronary atherectomy (DCA) restenosis, we determined the frequency of C. pneumoniae infection and its localization in symptomatic coronary atherosclerotic plaques using specimens obtained from DCA.Methods and results:Immunohistochemistry (IHC) and real-time polymerase chain reaction (RT-PCR) revealed the existence of C. pneumoniae in all 50 specimens of coronary atherosclerotic plaques obtained by DCA. C. pneumoniae–positive cell ratio determined with IHC or copy numbers of C. pneumoniae DNA detected by RT-PCR did not differ significantly between patients with stable angina pectoris and those with acute coronary syndrome (IHC: 16.4 ± 7.6% vs 18.0 ± 7.1%, P = .42; RT-PCR: no. of cases with high copy numbers 12/25 vs 10/25, P = .78), or between patients with subsequent post-DCA restenosis and those without (IHC: 17.1 ± 8.0% vs 18.0 ± 7.4%, P = .74; RT-PCR: 5/12 vs 10/21, P = 1.00).Conclusions:C. pneumoniae was highly prevalent in coronary atherosclerotic plaques of patients who underwent DCA. However, the extent of C. pneumoniae infection in coronary atherosclerotic plaques was not associated with plaque instability or post-DCA restenosis.

Comprehensive analysis of dysregulated genes associated with atherosclerotic plaque destabilization.

Atherosclerotic plaque destabilization is a dominating cause of acute cardiovascular events such as myocardial infarction and stroke. This study aims to identify genetic biomarkers related to atherosclerotic plaque destabilization using bioinformatics. Three transcriptome datasets of human carotid atherosclerotic plaque samples were downloaded from ArrayExpress and Gene Expression Omnibus databases, including E-MATB-2055, E-TABM-190, and GSE120521. With Robust Rank Aggregation analysis, we documented 46 differentially expressed genes between stable and unstable/ruptured plaques. Functional enrichment analysis using DAVID tool demonstrated that these genes were mainly related to biological functions such as extracellular matrix disassembly, collagen catabolic process, response to mechanical stimulus, and PPAR signaling pathway. A protein-protein interaction network for the differentially expressed genes was constructed, and eight pivotal genes (ITGAM, MMP9, PLAUR, CCR1, CD163, CD36, ADAM8, and IL1RN) were obtained from the network with a connective degree > 5. The expression patterns of these hub differentially expressed genes could be verified in atherosclerotic plaque samples with intraplaque hemorrhage. Using gene set variation analysis, the eight genes were integrated to generate an atherosclerotic plaque destabilization score, which showed a high performance in not only discriminating individuals with myocardial infarction from those with stable coronary illness, but also in predicting future acute cardiovascular events in atherosclerotic patients. In conclusion, the findings of this study will enhance our knowledge on the pathological mechanisms involved in atherosclerotic plaque destabilization, and provide potential gene biomarkers for risk stratification of patients with atherosclerotic cardiovascular disease.

Streptococcus mutans in atherosclerotic plaque: Molecular and immunohistochemical evaluations.

To verify the presence of Streptococcus mutans (S.mutans) in atherosclerotic plaque (AP) using techniques with different sensitivities, correlating with histological changes in plaque and immunoexpression of inflammatory markers. Thirteen AP samples were subjected to real-time polymerase chain reaction (qRT-PCR), histopathological analyses, histochemical analysis by Giemsa staining (GS), and immunohistochemical analysis for S.mutans, IL-1β, and TNF-α (streptavidin-biotin-peroxidase method). Ten necropsy samples of healthy vessels were used as controls. All AP samples showed histopathological characteristics of severe atherosclerosis and were positive for S.mutans (100.0%) in qRT-PCR and immunohistochemical analyses. GS showed that Streptococcus sp. colonized the lipid-rich core regions and fibrous tissue, while the control group was negative for Streptococcus sp. IL-1β and TNF-α were expressed in 100% and 92.3% of the AP tested, respectively. The control samples were positive for S.mutans in qRT-PCR analysis, but negative for S.mutans, IL-1β, and TNF-α in immunohistochemical analyses. The detection of S.mutans in AP and the visualization of Streptococcus sp. suggested a possible association between S.mutans and atherosclerosis. The results obtained from the control samples suggested the presence of DNA fragments or innocuous bacteria that were not associated with tissue alteration. However, future studies are necessary to provide more information.

Material Decomposition in Low-Energy Micro-CT Using a Dual-Threshold Photon Counting X-Ray Detector

Material decomposition in computed tomography is a method for differentiation and quantification of materials in a sample and it utilizes the energy dependence of the linear attenuation coefficient. In this study, a post-image reconstruction material decomposition method is constructed for a low-energy micro-CT setup using a photon counting x-ray detector. The low photon energy range (4–11 keV) allows for K-edge contrast separation of naturally occurring materials in organic tissue without the need of additional contrast agents. The decomposition method was verified using a phantom and its capability to decompose biomedical samples was evaluated with paraffin embedded human atherosclerotic plaques. Commonly, the necessary dual energy data for material decomposition is obtained by manipulating the emitted x-ray spectrum from the source. With the photon counting detector, this data was obtained by acquiring two energy window images on each side of the K-edge of one material in the sample. The samples were decomposed into three materials based on attenuation values in manually selected regions. The method shows a successful decomposition of the verification phantom and a distinct distribution of iron, calcium and paraffin in the atherosclerotic plaque samples. Though the decompositions are affected by beam hardening and ring artifacts, the method shows potential for spectral evaluation of biomedical samples.

Loss Of Insulin‐like Growth Factor 1 Receptor In Vascular Smooth Muscle Cells Promotes Increased Mirna‐221 And ‐222 In Type 2 Diabetes

Type 2 diabetes mellitus is associated with a 2 to 4 times higher risk of a heart attack or stroke. This increased risk of heart attacks and strokes results from an accelerated development of atherosclerotic plaques. During atherosclerotic plaque development, vascular smooth muscle cells (VSMC) proliferate and migrate leading to thickening of the intima of the arterial walls. This process is accelerated in diabetic patients by increase in two miRNAs, miR-221 and -222. These miRNAs are sensitive to changes in insulin receptor (IR) signaling. Under normal conditions, most of IR subunits are bound to insulin-like growth factor-1 receptor (IGFR) subunits forming heterodimers, which largely function as IGFR homodimers. A loss of IGFR thus promotes increased IR homodimers and an increase in insulin signaling. Our objective is to evaluate the effect of loss of IGFR on miR-221 and -222 expression in VSMCs. We hypothesize that the loss of IGFR coupled with physiological insulin stimulation promotes increased expression of miR-221/-222, which will in turn promote increased VSMC proliferation and migration. IGFR expression was measured in human atherosclerotic plaque samples from diabetic patients and nondiabetic patients. The effects of insulin stimulation coupled with the absence of IGFR was modeled with VSMCs isolated from an VSMC-specific IGFR knockout mouse. Our data demonstrates that arteries from diabetic subjects exhibit a 5.5-fold loss of the insulin-like growth factor receptor (IGFR-1). Murine vascular smooth muscle cells lacking IGFR exhibit a dose-dependent increase in miR-221/222 in response to physiological insulin stimulation. We conclude that loss of IGFR, likely increasing IR holoreceptors, alters insulin signaling to promote increased miR-221/222 in response to physiological insulin. These data provide support for the loss of IGFR as an initiator of the increase in intimal thickening seen in type 2 diabetes associated with miR-221/222.

Monocyte-Chemoattractant Protein-1 Levels in Human Atherosclerotic Lesions Associate With Plaque Vulnerability.

[Figure: see text].

Robust quantitative assessment of collagen fibers with picrosirius red stain and linearly polarized light as demonstrated on atherosclerotic plaque samples.

Collagen is an important component in maintaining structural integrity and functionality of tissues and is modulated in various biological processes. Its visualization and possible quantification using histopathological stains can be important for understanding disease progression or therapeutic response. Visualization of collagen fiber with the histological stain picrosirius red (PSR) is enhanced with polarized light and quantitative analysis is possible using circular polarizers. However, linear polarizers are more commonly available and easier to optically align. The objective of the present study is to demonstrate a novel image acquisition technique and analysis method using linearly polarized light. The proposed imaging technique is based on image acquisition at multiple slide rotation angles, which are co-registered to form a composite image used for quantitative analysis by pixel intensity or pixel counting. The technique was demonstrated on multiple human coronary samples with varying histopathologies and developed specifically to analyze cap collagen in atherosclerotic plaque. Pixel counting image analysis was found to be reproducible across serial tissue sections and across different users and sufficiently sensitive to detect differences in cap structural integrity that are likely relevant to prediction of rupture risk. The benefit of slide rotation angle under linear polarization to acquire images represents a feasible and practical implementation for expanding the general utility of PSR for quantitative analysis.

Molecular analysis shows the presence of periodontal bacterial DNA in atherosclerotic plaques from patients with coronary artery disease

Links between periodontitis and atherosclerosis can be predicted based on inflammatory mechanisms initiated by bacteria associated with periodontal lesions, which then influence the initiation or propagation of the atherosclerotic lesion. This study aimed to detect the presence of three periodontal pathogens, in atheromatous plaques of patients with coronary artery disease. Subgingival and atherosclerotic plaque samples were obtained from 80 patients scheduled for CABG or angioplasty. A nested PCR was done for the detection of the pathogens in the plaque samples. Porphyromonas gingivalis, Tanarella forsythia, and Treponema denticola were detected in 10%, 12.5%, and 1.3% of the atherosclerotic plaque samples respectively. It was also observed that patients whose atherosclerotic plaques tested positive for one or more of the pathogens had chronic periodontitis.

Quantitative Detection of Periodontopathogenic Bacteria in Atherosclerotic Plaques from Coronary Arteries by Real-Time PCR

Objectives:Epidemiologic studies have suggested periodontitis as a risk factor for Coronary Artery Diseas (CAD). Detection of periopathogens in atheromatous plaque provides some evidence for the causal relationship between these two conditions. The aim of this study was to determine the presence and quantity of periopathogens in coronary atherosclerotic plaques in patients undergoing Coronary Artery Bypass Graft (CABG) surgery.Methods:20 patients who were candidates for endarterectomy were enrolled in this study for the periodontal examination. Subgingival and coronary atherosclerotic plaque samples were then collected. Thereafter, quantitative detection ofAggregatibacter actinomycetemcomitans (A.a), Porphyromonas gingivali (P.g), and all bacteria detected by Real-Time PCR (RT-PCR) were measured. The correlation analysis was also used to evaluate the relationship between quantities of periopathogens in atherosclerotic and subgingival plaque samples.Results:A.awas detected in 13 patients (65%) with subgingival plaques and 4 patients (20%) with atherosclerotic plaques. In addition,P.gwas found in 15 patients (75%) with subgingival and 10 patients (50%) with atherosclerotic plaques.A.arepresented means of 2.7% and 10.04% of detected bacteria in both atherosclerotic and subgingival plaque samples, respectively. The mean of quantity ofP.gwas 10.85% and 12.87% of the detected bacteria obtained from atherosclerotic and subginigival samples, respectively. Correlation analysis showed a significant correlation between the quantities ofA.ain the atherosclerotic and subgingival plaques, but such a significant relationship was not found forP.g.Conclusion:This study confirmed the detection ofA.aandP.gin atheromatous plaque. The quantitative data suggested that periopathogens comprise a significant proportion of atherosclerotic plaque microbiome, which may consequently contribute to the development of CAD.

Analysis of atherosclerosis by inductively coupled plasma optical emission spectroscopy and scanning acoustic microscopy

Calcium deposition within the atherosclerotic plaques is the precursor of cardiovascular complications. Therefore, determination of levels of minerals and trace elements in blood plays an important role in assigning the stage of atherosclerosis. In this study, determination of mineral and trace element levels in atherosclerotic patients is aimed. Mineral and trace element levels within serum samples of 12 atherosclerotic patients were evaluated by Inductively Coupled Plasma Optical Emission Spectrometry (ICP-OES) and phosphorous (P), iron (Fe), magnesium (Mg) and calcium (Ca) levels were examined. Human carotid atherosclerotic plaque samples were previously screened by Scanning Acoustic Microscopy (SAM) and sound speed maps of the plaques showed higher sound speed values in the calcified regions, when compared to collagen-rich regions, indicating accumulation of calcium. Element analysis also showed increased Ca levels within serum samples. Therefore, it can be concluded that Ca deposition can be examined by ICP-OES and SAM, indicating that these techniques are confirmatory and may be combined to characterize atherosclerosis in the future.

Smoking is Associated to DNA Methylation in Atherosclerotic Carotid Lesions.

Tobacco smoking is a major risk factor for atherosclerotic disease and has been associated with DNA methylation (DNAm) changes in blood cells. However, whether smoking influences DNAm in the diseased vascular wall is unknown but may prove crucial in understanding the pathophysiology of atherosclerosis. In this study, we associated current tobacco smoking to epigenome-wide DNAm in atherosclerotic plaques from patients undergoing carotid endarterectomy. DNAm at commonly methylated sites (cytosine-guanine nucleotide pairs separated by a phospho-group [CpGs]) was assessed in atherosclerotic plaque samples and peripheral blood samples from 485 carotid endarterectomy patients. We tested the association of current tobacco smoking with DNAm corrected for age and sex. To control for bias and inflation because of cellular heterogeneity, we applied a Bayesian method to estimate an empirical null distribution as implemented by the R package bacon. Replication of the smoking-associated methylated CpGs in atherosclerotic plaques was executed in the second sample of 190 carotid endarterectomy patients, and results were meta-analyzed using a fixed-effects model. Tobacco smoking was significantly associated to differential DNAm in atherosclerotic lesions of 4 CpGs (false discovery rate <0.05) mapped to 2 different genes ( AHRR, ITPK1) and 17 CpGs mapped to 8 genes and RNAs in blood. The strongest associations were found for CpGs mapped to the gene AHRR, a repressor of the aryl hydrocarbon receptor transcription factor involved in xenobiotic detoxification. One of these methylated CpGs were found to be regulated by local genetic variation. The risk factor tobacco smoking associates with DNAm at multiple loci in carotid atherosclerotic lesions. These observations support further investigation of the relationship between risk factors and epigenetic regulation in atherosclerotic disease.

Periopathogens in atherosclerotic plaques of patients with both cardiovascular disease and chronic periodontitis.

BACKGROUNDAtherosclerosis and periodontitis are both chronic inflammatory diseases. Although a strong relationship between the two has already been established, the underlying mechanism is unknown. The present study was conducted aiming to detect the deoxyribonucleic acid (DNA) of Aggregatibacter actinomycetemcomitans (A.a), Campylobacter rectus (C.r), and Porphyromonas gingivalis (P.g) in subgingival and atherosclerotic plaques of patients with both chronic periodontitis and cardiovascular disease (CVD).METHODSIn this cross sectional study, patients with coronary artery disease (CAD) and moderate to severe periodontitis which were scheduled for coronary artery bypass grafting (CABG) were enrolled in the study. The subgingival plaques were collected before surgery. All samples were examined for the detection of selected periopathogens using polymerase chain reaction (PCR).RESULTSThe subgingival and atherosclerotic plaque samples of 23 patients were examined. The DNA of P.g, A.a, and C.r were found to be positive in 43.47%, 43.47%, and 78.26% of subgingival plaques, and 13.04%, 17.39%, and 8.69% of atherosclerotic plaques, respectively. In all cases, the bacterial species found in atherosclerotic plaques were also found in the subgingival plaques of the same patient.CONCLUSIONThis study demonstrated the presence of periopathogens in atherosclerotic plaques of patients with chronic periodontitis. More studies are required to ascertain the exact role of these periopathogens in atherosclerotic plaque formation.

Relationship between CHOP/GADD153 and unstable human carotid atherosclerotic plaque

Background and aims: The signaling protein C/EBP homologous protein (CHOP) and corresponding growth-arrest-and-DNA-damage-inducible gene 153 (GADD153) is associated with endoplasmic reticulum stress (ERS), which can lead to apoptosis. Our study aims to elucidate the role of CHOP/GADD153 in unstable atherosclerotic (AS) plaque formation isolated from confounding factors such as diabetes mellitus, primary hyperlipidemia, autoimmune deficiencies/abnormalities, essential hypertension, chronic heart failure, chronic kidney disease, and smoking.Material and methods: We collected carotid artery tissue samples from patients aged 50–80 years-old who received carotid endarterectomies (CEA) at our institution. We obtained fresh AS plaque samples during CEA and preserved the specimens immediately in the operating room with liquid nitrogen. Samples were categorized as stable or unstable AS plaques according to a six-stage histologic classification. CHOP/GADD153 expression was then examined with immunohistochemistry and reverse transcription polymerase chain reaction (RT-PCR).Results: A total of 32 patients met our inclusion and exclusion criteria, with 24 (75.0%) classified as unstable lesions. The mean optical density ratio normalized to GAPDH for CHOP/GADD153 in stable and unstable groups was 0.357 ± 0.025 and 0.490 ± 0.027, respectively (p < .05). Positive immunostaining of CHOP/GADD153 was found in macrophages and smooth muscle cells of unstable AS plaques with a mean integrated optical density of 0.63 ± 0.03, compared to 0.17 ± 0.05 in the stable group (p < .05).Conclusions: In conclusion, we were able to show significant elevation of CHOP/GADD153 in unstable plaques independent of other confounding factors that induce ERS.

Identification of Key Pathways and Genes in Advanced Coronary Atherosclerosis Using Bioinformatics Analysis.

Background Coronary artery atherosclerosis is a chronic inflammatory disease. This study aimed to identify the key changes of gene expression between early and advanced carotid atherosclerotic plaque in human. Methods Gene expression dataset GSE28829 was downloaded from Gene Expression Omnibus (GEO), including 16 advanced and 13 early stage atherosclerotic plaque samples from human carotid. Differentially expressed genes (DEGs) were analyzed. Results 42,450 genes were obtained from the dataset. Top 100 up- and downregulated DEGs were listed. Functional enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) identification were performed. The result of functional and pathway enrichment analysis indicted that the immune system process played a critical role in the progression of carotid atherosclerotic plaque. Protein-protein interaction (PPI) networks were performed either. Top 10 hub genes were identified from PPI network and top 6 modules were inferred. These genes were mainly involved in chemokine signaling pathway, cell cycle, B cell receptor signaling pathway, focal adhesion, and regulation of actin cytoskeleton. Conclusion The present study indicated that analysis of DEGs would make a deeper understanding of the molecular mechanisms of atherosclerosis development and they might be used as molecular targets and diagnostic biomarkers for the treatment of atherosclerosis.

Linkages between oral commensal bacteria and atherosclerotic plaques in coronary artery disease patients.

Coronary artery disease is an inflammatory disorder characterized by narrowing of coronary arteries due to atherosclerotic plaque formation. To date, the accumulated epidemiological evidence supports an association between oral bacterial diseases and coronary artery disease, but has failed to prove a causal link between the two. Due to the recent surge in microbial identification and analyses techniques, a number of bacteria have been independently found in atherosclerotic plaque samples from coronary artery disease patients. In this study, we present meta-analysis from published studies that have independently investigated the presence of bacteria within atherosclerotic plaque samples in coronary artery disease patients. Data were collated from 63 studies covering 1791 patients spread over a decade. Our analysis confirms the presence of 23 oral commensal bacteria, either individually or in co-existence, within atherosclerotic plaques in patients undergoing carotid endarterectomy, catheter-based atherectomy, or similar procedures. Of these 23 bacteria, 5 (Campylobacter rectus, Porphyromonas gingivalis, Porphyromonas endodontalis, Prevotella intermedia, Prevotella nigrescens) are unique to coronary plaques, while the other 18 are additionally present in non-cardiac organs, and associate with over 30 non-cardiac disorders. We have cataloged the wide spectrum of proteins secreted by above atherosclerotic plaque-associated bacteria, and discuss their possible roles during microbial migration via the bloodstream. We also highlight the prevalence of specific poly-microbial communities within atherosclerotic plaques. This work provides a resource whose immediate implication is the necessity to systematically catalog landscapes of atherosclerotic plaque-associated oral commensal bacteria in human patient populations.