Mitochondrial health is essential for maintaining cardiac function, and mitophagy—the selective degradation of damaged mitochondria—is central to maintenance of mitochondrial quality. In this review, we focus on the role of mitophagy in atherosclerotic disease, exploring both canonical and noncanonical pathways. We aim to highlight how proper regulation of mitophagy supports cardiac health, while imbalances in this process can contribute to the onset and progression of cardiovascular conditions. In addition, we examine the cardioprotective potential of mitophagy in the context of disease and discuss its close relationship with mitochondrial dynamics, particularly as they relate to both macrovascular and microvascular dysfunction. Finally, we identify current gaps in knowledge and outline key questions that remain for the field to address, with the goal of guiding future research in this critical area of cardiovascular biology.

Accurate protein quantification at low concentrations and small volumes is critical for advancing small‐scale research, such as microvasculature studies. However, existing microscale protocols often require ≥5 μL of sample or highly concentrated lysates, limiting their applicability in contexts with scarce material. To overcome these limitations, we developed the Nano‐Extraction BCA‐Optimized Workflow (NEBOW), a novel method requiring only 2 μL of sample and capable of detecting protein concentrations as low as 0.01 mg/mL. Optimized for the NanoDrop™ One UV–Vis Spectrophotometer, this workflow demonstrated significantly enhanced sensitivity and reproducibility compared to the standard BCA assay. Paired t tests ( p < 0.01) and TOST equivalence testing (15% margin) confirmed key differences, with the NEBOW method producing steeper standard curves and more consistent results at low concentrations. Bland–Altman analysis showed that standard BCA tends to overestimate protein levels, while the NEBOW method maintained accuracy across a range of low‐input samples. Western blot validation supported the improved performance of the new workflow. This approach offers a reliable, cost‐effective solution for protein quantification when sample availability is limited, without sacrificing accuracy or sample integrity.

Journal Article Accepted manuscript Angiotensin 1-7 and a TERT activator individually restore vasodilatory capacity within the microcirculation previous SARS-CoV-2 infection Get access Yoshinori Nishijima, Yoshinori Nishijima Department of Medicine, Medical College of Wisconsin, Milwaukee, WisconsinCardiovascular Center, Medical College of Wisconsin, Milwaukee, Wisconsin https://orcid.org/0000-0002-5547-4508 Search for other works by this author on: Oxford Academic PubMed Google Scholar Shelby N Hader, Shelby N Hader Department of Medicine, Medical College of Wisconsin, Milwaukee, WisconsinCardiovascular Center, Medical College of Wisconsin, Milwaukee, Wisconsin https://orcid.org/0000-0002-6564-091X Search for other works by this author on: Oxford Academic PubMed Google Scholar Erin C Birch, Erin C Birch Department of Medicine, Medical College of Wisconsin, Milwaukee, WisconsinCardiovascular Center, Medical College of Wisconsin, Milwaukee, Wisconsin Search for other works by this author on: Oxford Academic PubMed Google Scholar Yiliang Chen, Yiliang Chen Department of Medicine, Medical College of Wisconsin, Milwaukee, WisconsinVersiti Blood Research Institute, Milwaukee, Wisconsin Search for other works by this author on: Oxford Academic PubMed Google Scholar Michael E Widlansky, Michael E Widlansky Department of Medicine, Medical College of Wisconsin, Milwaukee, WisconsinCardiovascular Center, Medical College of Wisconsin, Milwaukee, WisconsinDepartment of Pharmacology, Medical College of Wisconsin, Milwaukee, Wisconsin Search for other works by this author on: Oxford Academic PubMed Google Scholar Andreas M Beyer Andreas M Beyer Department of Medicine, Medical College of Wisconsin, Milwaukee, WisconsinCardiovascular Center, Medical College of Wisconsin, Milwaukee, WisconsinDepartment of Physiology, Medical College of Wisconsin, Milwaukee, Wisconsin Correspondence: Andreas. M. Beyer, 8701 Watertown Plank Rd., Medical College of Wisconsin, Milwaukee, WI 53226 (E-mail: [email protected]) https://orcid.org/0000-0001-9129-7492 Search for other works by this author on: Oxford Academic PubMed Google Scholar Cardiovascular Research, cvaf020, https://doi.org/10.1093/cvr/cvaf020 Published: 13 February 2025 Article history Received: 19 September 2024 Revision received: 10 December 2024 Accepted: 20 December 2024 Published: 13 February 2025

BACKGROUND:Cerebral hypo-perfusion during hemodialysis (HD) may contribute to cerebral ischemic lesions and atrophy in HD patients. Vascular disease and stiffness can impair cerebrovascular reactivity (CVR) in HD patients, placing them at higher risk for cerebral hypo-perfusion during the hemodynamic stress of HD. We evaluated the relationship between CVR and change in cerebral perfusion during HD. METHODS:In a cohort of in-center HD patients, we used hypercapnia to induce a change in cerebral blood flow velocity measured with transcranial Doppler to assess CVR. We used continuous cerebral oximetry during HD to measure a change in cerebral oxygen saturation (ScO2), calculating overall decline and the largest drop as markers of cerebral perfusion. We used multiple linear regression to assess the relationship between CVR and the ScO2-associated endpoints. FINDINGS:We measured CVR in 42 HD patients and of those, 41 had the ScO2 measurements completed. The mean age was 58.5 (11.0) years, and most were male (90.5%, N = 38) with diabetes (59.5%, N = 25) and hypertension (87.5%, N = 36). The average CVR was 2.7 (1.6)%/mmHg. The average overall decline in ScO2 during HD was 2.2 (2.5)% and the average largest drop in ScO2 was 5.9 (2.8)%. CVR was negatively associated with both the largest drop in ScO2 (β = -0.67 95% CI [-1.20, -0.15], p = 0.01) and the overall decline in ScO2 (β = -0.62 95% CI [-1.09, -0.15], p = 0.01). Vascular disease was a risk factor for lower CVR (β = -1.21, 95% CI [-2.16, -0.26] p = 0.01). CONCLUSIONS:A lower CVR increases the risk for cerebral hypo-perfusion during HD. Impaired CVR may be an important part of the pathophysiology of ischemic brain injury and cognitive impairment in HD patients.

Electrocardiography (ECG) is a cornerstone of cardiac diagnostics, detecting cardiac pathologies ranging from arrhythmias to myocardial infarction. To enhance diagnostic accuracy and efficiency, deep learning models have been developed that now match or surpass human performance in ECG interpretation. However, their opaque reasoning hinders clinical trust and regulatory approval. This challenge is particularly acute for ECG signals because, unlike structured feature data, they are sequential, variable, and noise-prone, making interpretability both more difficult and more essential for clinical adoption. This review systematically evaluates ECG-specific explainable AI techniques using PRISMA guidelines. We screened 380 records across six databases and included 45 peer-reviewed studies examining diverse explainability methods including perturbation-based, gradient-based, intrinsically interpretable, sequence-aware, and counterfactual approaches. Our analysis reveals that perturbation-based techniques designed for structured data prove suboptimal for ECG signals because they treat features as independent rather than temporally dependent. In contrast, methods that transparently reveal model attention to physiologically meaningful ECG intervals such as the P wave, QRS complex, and ST segment demonstrate superior performance across localization accuracy, fidelity, and robustness metrics. While explainable AI in ECG interpretation has advanced substantially, it remains fragmented and insufficiently validated for clinical deployment. The most promising methods reveal what the model attends to in relation to known physiologic features, yet significant challenges persist in stability, computational efficiency, and regulatory readiness. Accelerating clinical translation requires open, multi-institutional benchmarks with cardiologist-annotated explanations and clinician-in-the-loop validation studies that can strengthen trust and support meaningful integration of explainable AI into patient care.

BackgroundAtherosclerotic cardiovascular disease (ASCVD) is a chronic inflammatory disease that leads to adverse events such as myocardial infarctions and stroke. Gut microbiome modulation is a promising target to reduce chronic inflammation and improve outcomes for patients with coronary artery disease (CAD). Risk profile patterns of CAD patients who target gut health with probiotics could provide insight into how gut modulation improves CAD clinical biomarkers. This study aims to evaluate the association between probiotic use and clinical markers of known atherosclerotic risk factors, in patients with CAD.MethodsWe conducted a cross-sectional large-database study using National Health and Nutrition Examination Survey (NHANES) data from years 1999–2020. The cohort included adults with at least a diagnosis of coronary heart disease, angina, and heart attack or two of the following: diabetes, high blood pressure, and high cholesterol. Analyses of clinical biomarkers compared probiotic to non-probiotic groups, between probiotic type groups, and between probiotic supplement strains.ResultsOur cohort included 14,992 survey responders. After weighting, this sample represented 46,217,980 US adults. There were 4,062,022 adults exposed to probiotics, 763,288 to probiotic supplements and 3,179,008 to probiotic foods. Probiotic exposure was associated with lower A1c (p < 0.001), lower triglycerides (p < 0.001), lower ASCVD risk score (p = 0.01) and higher HDL-C (p < 0.001). Probiotic supplement exposure was associated with lower LDL-C (p = 0.003) and total cholesterol (p = 0.047).ConclusionOur study reinforces the beneficial association between probiotic ingestion and cardiovascular health in patients with existing atherosclerotic disease. Further studies to better determine potential mechanistic connections between the gut microbiota on cardiovascular risk factors is warranted.

Arterioles are small blood vessels located just upstream of capillaries in nearly all tissues. Despite the broad and essential role of arterioles in physiology and disease, current knowledge of the functional genomics of arterioles is largely absent. Here, we report extensive maps of chromatin interactions, single-cell expression, and other molecular features in human arterioles and uncover mechanisms linking human genetic variants to gene expression in vascular cells and the development of hypertension. Compared to large arteries, arterioles exhibited a higher proportion of pericytes which were enriched for blood pressure (BP)-associated genes. BP-associated single nucleotide polymorphisms (SNPs) were enriched in chromatin interaction regions in arterioles. We linked BP-associated noncoding SNP rs1882961 to gene expression through long-range chromatin contacts and revealed remarkable effects of a 4-bp noncoding genomic segment on hypertension in vivo. We anticipate that our data and findings will advance the study of the numerous diseases involving arterioles.

Adverse changes in the gut microbiome with aging are an emerging mediator of arterial dysfunction, which contributes to cardiovascular disease (CVD) development. We investigated the therapeutic potential of enhancing the bioavailability of gut-derived short-chain fatty acids (SCFAs; produced from dietary fiber) for improving age-related arterial dysfunction. We performed gut microbial whole-genome sequencing in young (3 months) versus old (24 months) male C57BL/6N mice to explore changes in bacterial taxonomic abundance and functional pathways with aging and relations to arterial function. We then supplemented young and old mice with the SCFA acetate in drinking water versus controls and versus a high-fiber diet for 8-10 weeks to test the effects of these interventions on vascular function and explore potential mechanisms. Of the various differences in the gut microbiomes of old mice, lower SCFA-producing capacity (taxonomic abundance and functional pathways) stood out as a key feature related to worse arterial function after adjusting for age. Acetate supplementation and a high-fiber diet reversed ~30% of the age-related increase in aortic pulse wave velocity (stiffness) and fully restored carotid artery endothelium-dependent dilation (endothelial function) to young levels. Acetate and a high-fiber diet reduced age-related increases in systemic inflammation. We also found that improvements in endothelial function were likely mediated by suppressed early growth response-1 signaling using innovative siRNA-based knockdown in isolated arteries. There were no effects of the interventions in young mice. Acetate supplementation was comparably effective for ameliorating arterial dysfunction with aging as a high-fiber diet and thus shows promise for reducing CVD risk in older adults.

The circulating milieu, bioactive molecules in the bloodstream, is altered with aging and interfaces constantly with the vasculature. This anatomic juxtaposition suggests that circulating factors may actively modulate arterial function. Here, we developed a novel, translational experimental model that allows for direct interrogation of the influence of the circulating milieu on age-related arterial dysfunction (aortic stiffening and endothelial dysfunction). To do so, we exposed young and old mouse arteries to serum from young and old mice and young and midlife/older (ML/O) adult humans. We found that old mouse and ML/O adult human, but not young, serum stiffened young mouse aortic rings, assessed via elastic modulus (mouse and human serum, P = 0.003 vs. young serum control), and impaired carotid artery endothelial function, assessed by endothelium-dependent dilation (EDD) (mouse serum, P < 0.001; human serum, P = 0.006 vs. young serum control). Furthermore, young mouse and human, but not old, serum reduced aortic elastic modulus (mouse serum, P = 0.009; human serum, P < 0.001 vs. old/MLO serum control) and improved EDD (mouse and human serum, P = 0.015 vs. old/MLO serum control) in old arteries. In human serum-exposed arteries, in vivo arterial function assessed in the human donors correlated with circulating milieu-modulated arterial function in young mouse arteries (aortic stiffness, r = 0.634, P = 0.005; endothelial function, r = 0.609, P = 0.004) and old mouse arteries (aortic stiffness, r = 0.664, P = 0.001; endothelial function, r = 0.637, P = 0.003). This study establishes novel experimental approaches for directly assessing the effects of the circulating milieu on arterial function and implicates changes in the circulating milieu as a mechanism of in vivo arterial aging.

GWAS has identified >1,000 blood pressure (BP)-associated sentinel SNPs, most of which (>90%) are intronic and intergenic. How they regulate BP-relevant genes remains an open question. We hypothesized many SNPs would reside within cis -regulatory elements specific to nephron segments critical to sodium resorption such as proximal tubule (PT) and thick ascending limb (TAL). To address this, we developed chromatin state (ATAC-seq) and transcriptome (RNA-seq) maps from manually dissected human PT and medullary TAL (mTAL) segments. We identified 216,168 and 179,467 open regions in PT and mTAL, respectively, with the majority (70%) being unique to one tissue. Interestingly, peaks shared between the two segments resided more frequently in promoters, whereas unique peaks were more often found in distal introns or intergenic regions, suggesting they lie within enhancers ( Figure 1A ). Both chromatin state and transcriptomic data demonstrated a correlation between open chromatin peaks within gene promoters and mRNA levels (r=0.11; p<4.6e-16). This correlation was weaker at open regions distal to the promoter, illustrating the difficulty of mapping distal open regions to the gene(s) they regulate. We hypothesized these open regions would be enriched for BP-relevant SNPs. We curated 1,071 bp-associated SNPs and found 87 and 67 in mTAL and PT, open regions respectively, many of which are near BP-relevant genes. This enrichment was significant based on 10 4 randomizations (pValue 10 -4 ) ( Figure 1B ). We conclude that these novel tissue-specific chromatin maps can be used to identify potential mechanisms by which SNPs influence gene expression and BP. Additional studies will be needed to demonstrate both causality and molecular mechanisms.

Background and Research Question: We have previously shown that mitochondrial fission protein 1 (Fis1) is over-expressed in endothelial cells obtained from humans with diabetes. However, whether Fis1 is mechanistically linked to impaired microvascular endothelial function, either health or cardiometabolic disease, remains unknown. Methods: Fresh resistance arterioles from 73 healthy individuals and 24 individuals with type 2 diabetes (T2DM) by gluteal adipose tissue biopsy or discarded surgical tissue. Endothelium-dependent vasoreactivity was tested with increasing doses of acetylcholine (10 -10 to 10 -5 M) following two different molecular manipulations: (1) intraluminal lentiviral transfection for endothelial-specific Fis 1voverexpression vs. control or (2) intraluminal Fis1 siRNA transfection vs. scrambled control. We developed a small peptide Fis1 inhibitor, pep213, and tested if pep213 improved endothelial function in vessels from T2DM patients and healthy individuals exposed to high glucose (HG, 33mM). The impact of molecular manipulations on mitochondrial superoxide (mtO 2 ●- ) function were performed using mitoNeoD. Result: Fis1 knockdown significantly improved vasodilation to Ach in an endothelial nitric oxide synthase (eNOS)-dependent manner (N=6, P<0.0001) in T2DM vessels and healthy vessels exposed to high glucose (HG=33 mM, N=5, P<0.0001), but had no effect Ach vasodilation in healthy vessels exposed to normal glucose concentrations (NG=90 mg/dL., N=3, P=NS). Fis1 overexpression impaired Ach vasodilation in an eNOS-dependent manner in healthy vessels (N=5, P<0.0001) compared to control transfection. NONOate and papaverine controls for these studies showed effects on smooth muscle reactivity. Intraluminal pep213 (1 µM for 1 hour), reversed impaired eNOS-dependent vasodilation in T2DM vessels (N=5, P<0.001), healthy vessels exposed to HG (N=5, P<0.001), and healthy vessels over-expressing Fis1 (N=5, P<0.05), while a scrambled version of pep213 without ability to bind Fis1 had no effects in any of these scenarios. Pep213 had no effect on vasodilation to NONOate. Pep213 reduced mtO 2 ●- in T2DM and healthy vessels over-expressing Fis1 (N=5, P<0.001). Conclusion: Fis1 mechanistically regulates eNOS-dependent vasoreactivity in human vessels, potentially through reducing mtO 2 ●- levels. Fis1 over-expression may be particularly important in T2DM, and pharmacological inhibition of Fis1 can reverse endothelial dysfunction in T2DM human vessels.

Endothelial dysfunction represents a measurable and early manifestation of vascular disease. Emerging evidence suggests cardiovascular risk remains elevated after COVID-19 infection for at least 12 months, regardless of cardiovascular disease status prior to infection. We review the relationship between the severity of endothelial dysfunction and the severity of acute COVID-19 illness, the degree of impairment following recovery in both those with and without postacute sequalae SARS-CoV-2 infection, and current therapeutic efforts targeting endothelial function in patients following COVID-19 infection. We identify gaps in the literature to highlight specific areas where clinical research efforts hold promise for progress in understanding the connections between endothelial function, COVID-19, and clinical outcomes that will lead to beneficial therapeutics.

Many non-coding SNPs identified in Genome-Wide Association Studies (GWAS) likely affect BP-related gene expression through epigenetic mechanisms. This study analyzes the comparative genomic and epigenomic landscapes in human and rat kidney tissues, including kidney proximal tubule (PT) and medullary thick ascending limb (mTAL), focusing on the major challenge of identifying conserved regulatory elements in intergenic regions. The purpose of our study is to identify BP gene regulatory elements in intergenic regions that are conserved from human to rat for in vivo validation and mechanistic studies. Our ultimate goal is to generate high-resolution, genome-wide epigenomic maps of key BP-relevant tissues, using ATAC-seq, Hi-C, CUT&Tag, DNA methylation profiling and RNA-seq. We are creating visualization hubs for human and rat, integrating our data within the UCSC Genome Browser environment. UCSC contains extensive data for humans but very little for rats, particularly for the updated genome assemblies. To address this gap, we integrated our results with data from the Rat Genome Database (RGD). To date, we harmonized ATAC-seq data from human and rat PT and mTAL and displayed these data in RGD’s JBrowse2 genome browser for unique comparative genome views. Based on our initial analysis of data density and intensity, we are developing a novel pattern recognition algorithm to identify epigenomic marks that are conserved across rat and human genomes. Existing algorithms predominantly focus on the conservation of coding genes, gene families, and genomic sequences. Here, average normalized peak intensities for ATAC-seq data across tissues were subject to canonical correlation between human and rat to identify conserved epigenomic patterns within the BP GWAS loci. Here we present an example of a human BP locus in the intergenic region between NPR3 and TARS1 on chromosome 5 which has a region of conserved synteny with rat chromosome 2. Comparative analysis and visualization of this region show significant correlation between ATAC-seq peaks from rat and human PT and mTAL at this locus, with predicted conserved regulatory elements. These visualizations can uncover novel biological insights and identify potential regulatory targets for intervention. This work underscores the importance of advanced visualization techniques in understanding hypertension pathogenesis, facilitating cross-species comparisons, and enhancing the utility of genomic databases.

Resistance arteries, or arterioles, are key determinants of the total peripheral vascular resistance, which, in turn, is a key determinant of arterial blood pressure (BP). However, the amount of protein available from one isolated human arteriole may be less than 5 μg, making proteomic analysis challenging. In addition, obtaining human arterioles requires manual dissection of unfrozen clinical specimens. This limits its feasibility, especially for powerful multi-center clinical studies in which clinical specimens need to be shipped overnight to a research lab for arteriole isolation. We performed a study to address low input, test overnight tissue storage, and develop a reference human arteriolar proteomic profile. We found that, in tandem mass tag proteomic analysis, the use of a booster channel consisting of endothelial and vascular smooth muscle cells (1:5 ratio) increased the number of proteins detected in a human arteriole segment with FDR < 0.01 from 1,051 to more than 3,000. We collected adipose tissues from three human subjects and isolated two arterioles from each fresh aliquot of the tissue or after 24h of cold storage of unfrozen aliquots in MACS tissue solution. The correlation coefficient of proteomic profile was similar (p=0.6) between replicate arterioles isolated freshly, following the cold storage, or before and after the cold storage. We built a human arteriolar proteomic profile consisting of 3,836 proteins based on the analysis of 12 arteriole samples from the three subjects. The average arteriolar protein had a mean copy number of 2.76х10 6 per cell using the histone proteomic ruler, which is based on the fact that the mass of DNA per cell is approximately equal to the protein mass of histones. Transgelin was the most abundant protein detected. We curated a set of BP-relevant human genes, which encode 1,945 proteins. Of these BP-relevant proteins, 476 (12.5%) were detected in the arteriolar proteome, which was a significant overrepresentation (p<0.05, Chi squared test). These findings demonstrate that proteomic analysis is feasible with arterioles isolated from human adipose tissue following cold overnight storage and provide a reference human arteriolar proteome profile highly valuable for studies of arteriole-related traits.

Abstract Introduction. Breast cancer (BC) is the most frequently diagnosed cancer with over 4 million survivors living in the United States. Recent treatment (Tx) advances such as in targeted (e.g. anti-HER2), systemic (e.g. anthracyclines), or combined therapies have supported the increased survival rates but continue to present adverse effects. Cardiotoxicity, which may be induced during or after Tx, is the second highest cause of mortality in BC survivors and also negatively impacts quality of life (QoL). Black/African American (B/AA) patients have higher rates of this effect along with a higher incidence of aggressive BC types. Limited research has targeted vascular function as an area of focus in the cardio-oncology field. Methods. The Discovery and Elimination of Cardio-Oncology Disparities Toward Equity in the Heartland Center is examining how anti-cancer treatments impact vascular function, the potential mitigating effects of exercise and how socioenvironmental factors moderate these effects. The Take Charge during Treatment randomized trial within this Center is testing the feasibility and acceptability of an exercise during treatment program for B/AA and non-Hispanic White (NHW) women newly diagnosed with BC. The study also examines and compares the effects of exercise on VO2 peak and QoL in participants. The 16-week intervention promotes adherence to the American College of Sports Medicine exercise guidelines for cancer survivors and is guided by a certified exercise trainer. To evaluate the intervention, patient's complete surveys including the FACT-B for QoL, a VO2 peak assessment, physical assessments, and anthropometry measurements at baseline (before BC Tx), after the 16-week program, and at a 12-month timepoint. Socioenvironmental factors are examined using participant addresses. Results. To date, 1330 patients were assessed for eligibility, 60 consented, and 49 enrolled and randomized. Herein, we present data on 45 of the 49 (35 NHW, 10 B/AA). The mean age of participants is 52 years (SD=12), most were diagnosed with Stage I or II BC, and they have varied education and income levels. Baseline (pre-treatment) differences are noted by racial group for QoL (Total score – 88 (SD=9) for NHW vs 74 (SD=29) for B/AA; Physical function – 25 (SD=3) for NHW vs 22 (SD=5) for B/AA; Functional – 21 (SD=4) for NHW vs 18 (SD=8) for B/AA), VO2 peak (22 (SD=5) for NHW vs 17 (SD=6) for B/AA), and obesity (NHW – 30%, B/AA – 70%). Cognitive function, pain interference, social isolation and perceived stress were similar across groups. B/AA women were more likely to report living in a highly redlined area (50%) compared to NHW women (3%), with lower percent tree canopy (13% vs 24%) and lower perceived social cohesion (15 vs 19). Reports of discrimination and resilience were similar across groups. Conclusions. Preliminary baseline results suggest that B/AA women enter BC Tx more vulnerable to developing QoL challenges and cardiotoxicity. Neighborhood environmental factors will be further examined to understand the association with adverse outcomes. Citation Format: Estefania Alonso, Melinda Stolley, Kirsten Beyer, Alison Kriegel, Shane Phillips, Matt Durand, Michael E. Widlansky, Amanda L. Kong, Kent Hoskins, Andreas Beyer. Take Charge during Treatment: A randomized exercise trial for breast cancer patients [abstract]. In: Proceedings of the 17th AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2024 Sep 21-24; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2024;33(9 Suppl):Abstract nr C032.

We have previously shown that endothelial function in the human placental microcirculation is impacted by excessive oxidative stress and TLR9 may be part of this mechanism. Therefore, to evaluate whether TLR9 knockdown restores vascular function in preeclamptic microvessels. Placental microvessels (< 150μM diameter) were dissected from the maternal side of control or preeclamptic placentas and then incubated with media and scrambled siRNA or TLR9-specific siRNA (50nM) for 12-16 hours. Vessels were cannulated on glass micropipettes, pressurized to 60mmHg, and then pre-constricted with 0.1-2.0 nM endothelin-1. A pressure gradient was applied to induce flow and vessel inner diameters were measured by videomicroscopy. Flow-mediated dilation (FMD) was calculated as a % of maximum diameter and expressed as mean ± SEM. Results were analyzed by two-way ANOVA with Dunnett’s multiple comparisons. The maximum dilation values were evaluated by a Kruskal-Wallis test with Dunn’s multiple comparative test or t-test as appropriate. Preeclamptic microvessels demonstrated impaired endothelium-dependent vasodilation compared to healthy controls (Fig. A, p=0.0018). Vasodilator capacity in healthy controls was not significantly impacted by siRNA treatment (Fig. B, p=0.4429). TLR9 siRNA restored impaired endothelium-dependent vasodilation to flow compared to both untreated and scrambled siRNA-transfected vessels from patients with preeclampsia (Fig. C, p=0.0082). Suppression of TLR9 expression restores endothelium-dependent vasodilation to flow in preeclamptic vessels and merits further evaluation.