Search Results (240)

Background: Postmenopausal osteoporosis is associated with cellular senescence and the accumulation of the senescence-associated secretory phenotype (SASP). While mesenchymal stem cell (MSC)-derived exosomes show tissue repair potential, the efficacy and mechanisms of MSC-derived apoptotic vesicles (apoVs) remain unclear. This study compared MSC-apoVs and exosomes in postmenopausal osteoporosis and investigated the underlying epigenetic mechanisms. Methods: Therapeutic efficacy was evaluated in an ovariectomized (OVX) mouse model and senescent human bone marrow mesenchymal stem cells (hBMMSCs). Small RNA sequencing identified differential microRNA (miRNA) cargos between vesicle types. SASP-related cytokine expression (IL-6, TNF-α, MCP-1) and pathway activation were assessed by RT-qPCR, ELISA, and Western blot. Results: MSC-apoV treatment attenuated bone loss in OVX mice and reduced SASP expression in senescent hBMMSCs to a greater extent than exosomes. Small RNA sequencing revealed that apoVs were enriched with a specific miRNA cluster, including hsa-let-7b-5p, hsa-miR-92a-3p, and hsa-miR-98-5p. Bioinformatic analyses identified BRAF and CRKL as downstream targets of this miRNA cluster, supported by reduced protein levels after apoV treatment. Subsequent molecular assays showed that apoV treatment inhibited the phosphorylation of both the MAPK (p38 and JNK) and NF-κB (p65) signaling pathways, which correlated with reduced local inflammation in the bone marrow microenvironment and preserved osteogenic differentiation capacity. Conclusions: MSC-apoVs attenuate postmenopausal osteoporosis more effectively than exosomes. This enhanced efficacy is associated with the delivery of an enriched miRNA cluster that inhibits MAPK and NF-κB signaling, together with suppression of BRAF and CRKL protein expression. ApoVs may represent a cell-free therapeutic strategy for age-related bone loss. Full article

Background and Objectives: Treatment-resistant depression (TRD) is a major clinical challenge in the management of major depressive disorder (MDD). While pharmacogenetics has been suggested to be clinically useful in guiding antidepressant treatment, few studies have explored if and how pharmacogenes can be involved in TRD pathophysiology and its clinical outcomes. Material amd Methods: We explored the role of differences in metabolizer phenotypes, gene expression levels, and microRNAs of three key pharmacogenes (CYP2D6, CYP2C19, CYP2B6) in TRD pathophysiology and antidepressant response in a cohort of 300 patients with MDD from the PROMPT consortium. Results: CYP2D6 phenotype distribution did not differ significantly between TRD and non-TRD groups, but mRNA expression was significantly upregulated in TRD. Hsa-miR-26b-5p, a microRNA predicted to regulate CYP2D6, was significantly downregulated in TRD. For CYP2C19, intermediate metabolizers (IMs) were underrepresented in TRD versus non-TRD (IMs vs. normal metabolizers (NMs): χ² = 6.07, p = 0.019). microRNA hsa-let-7d-5p and hsa-miR-27a-3p, predicted to regulate CYP2C19, were significantly downregulated in TRD. No significant differences were found for CYP2B6. Conclusions: This study contributes valuable insights to the PROMPT project on how pharmacokinetic gene variants and their expression and regulatory mechanisms may influence antidepressant response and resistance in MDD. Full article

Exposure to high-altitude hypoxia leads to complex physiological and molecular adaptations, particularly in skeletal muscle. MicroRNAs (miRNAs), including muscle-enriched (myomiRNAs) and hypoxia-responsive (hypoxamiRNAs), play critical roles in regulating these responses. We investigated miRNA expression changes in the skeletal muscle of healthy, non-smoking Italian adults (mean age 36.7 ± 12.4 years) participating in the Himalayan expedition “Lobuche Peak—Pyramid Exploration & Physiology” conducted in the Sagaramāthā (Mount Everest) National Park, Nepal. The peak overnight stay altitude was ≈5000 m at the Pyramid International Laboratory—Observatory. Muscle biopsies were taken before and after the expedition from Vastus lateralis, at one-third of the distance from the upper margin of the rotula to the anterior superior iliac spine. Small RNA sequencing was used to profile differentially expressed miRNAs. Several miRNAs were differentially expressed (exploratory analysis), suggesting potential involvement in hypoxia-related adaptation. These encompass both canonical myomiRNAs (e.g., miR-206, miR-486-5p) and hypoxamiRNAs (e.g., miR-378a-5p, miR-199a-3p, let-7b-5p). In enrichment analysis, we found several connections between miRNAs and pathways that may play a role in physiological regeneration or differentiation in muscle cells. Among functions, focal adhesion (p-value = 0.001), regulation of actin cytoskeleton (p-value = 0.026), Rap-1 (p-value = 0.007), cAMP (p-value = 0.017), MAPK (p-value = 0.019), and Hippo (p-value = <0.001) signaling pathways were predicted to be the most targeted. These findings provide preliminary insights into physiological adaptation, requiring confirmation in larger and controlled cohorts. Full article

Let $\epsilon >0$ and $\mathcal{TB}(\mathcal{X}\times \mathcal{X})$ be the Banach algebra of all $2\times 2$ bounded upper triangular operator matrices on a separable Hilbert space $\mathcal{X}\times \mathcal{X}$. In this paper, we first establish the spectrum equalities for special cases of upper triangular operator matrices—diagonal block operator matrix $M_0=\left({\displaystyle \genfrac{}{}{0pt}{}{A0}{0B}}\right)$. We obtain that ${\widehat{\Sigma }}_{bi,\epsilon }\left(M_0\right)={\Sigma }_{bi,\epsilon }\left(A\right)\cup {\Sigma }_{bi,\epsilon }\left(B\right)$, $i\in \{1,2,4\}$, where ${\Sigma }_{bi,\epsilon }(·)$ and ${\widehat{\Sigma }}_{bi,\epsilon }(·)$ denote the noncommutative pseudo-upper (resp. lower) semi-Browder essential spectrum, noncommutative pseudo-Browder essential spectrum, sub-noncommutative pseudo-upper (resp. lower) semi-Browder essential spectrum, and sub-noncommutative pseudo-Browder essential spectrum. Secondly, based on Cao and Bai’s works, we study the perturbation of the sub-noncommutative pseudo-Browder essential spectrum ${\widehat{\Sigma }}_{b4,\epsilon }(·)$ of a 2 × 2 bounded upper triangular operator matrix $M_C=\left({\displaystyle \genfrac{}{}{0pt}{}{AC}{0B}}\right)$ on a separable Hilbert space. We obtain that ${\displaystyle \bigcap _{C\in \mathcal{B}\left(\mathcal{X}\right)}{\widehat{\Sigma }}_{b4,\epsilon }\left(M_C\right)={\Sigma }_{b1,\epsilon }\left(A\right)\cup {\Sigma }_{b2,\epsilon }\left(B\right)\cup \Delta }$, where $\Delta =\{\lambda \in \mathbb{C}:$ there exist $P_i\in \mathcal{B}\left(\mathcal{X}\right)$ with $\parallel P_i\parallel <\epsilon ,i\in \{1,2\},$ such that $\alpha (A+P_1-\lambda I)+\alpha (B+P_2-\lambda I)\ne \beta (A+P_1-\lambda I)+\beta (B+P_2-\lambda I)\}$. Finally, we obtain ${\Sigma }_{bi,\epsilon }\left(A\right)\cup {\Sigma }_{bi,\epsilon }\left(B\right)={\widehat{\Sigma }}_{bi,\epsilon }\left(M_C\right)\cup W,i\in \{1,2,4\}$, where W is the union of certain holes in $({\Sigma }_{bi,\epsilon }\left(A\right)\cup {\Sigma }_{bi,\epsilon }\left(B\right))\backslash {\widehat{\Sigma }}_{bi,\epsilon }\left(M_C\right)$. Full article

Background: Cardiovascular diseases remain important causes of morbidity and potential premature mortality in children. Although clinical imaging and electrophysiologic testing have advanced, early, minimally invasive biomarkers that can both detect myocardial injury and help differentiate among overlapping pediatric phenotypes are still limited. Circulating microRNAs (miRNAs; miRs) are becoming attractive biomarker candidates because many are abundant in the heart, actively released into the circulation, and remarkably stable in plasma. The study aimed to assess the expression of miR-1-3p, miR-let-7b-5p, miR-21-5p, and miR-26b-5p in children with cardiovascular disease. Methods: Children aged 10–18 years with cardiac arrhythmias, myocarditis, or cardio-myopathies were recruited. The control group consisted of healthy age- and sex-matched children. For each participant, peripheral venous blood was collected for plasma isolation and miRNA profiling. The expression of miR-1-3p, miR-let-7b-5p, miR-21-5p, miR-26b-5p, and UniSp6 molecules was analyzed using the comparative cycle threshold delta Ct (ΔCt) method. A p-value ≤ 0.05 was considered statistically significant. Results: miR-26b-5p was significantly downregulated in patients with cardiac disease compared with healthy controls. miR-21-5p and miR-26b-5p were downregulated in patients with ventricular arrhythmia. Moreover, miR-26b-5p was downregulated in arrhythmia in general. We found no significant difference in the expression of miR-1-3p, miR-let-7b-5p, miR-21b-5p, and miR-26b-5p between patients with and without myocarditis, as well as with and without hypertrophic cardiomyopathy. Conclusions: miR-26b-5p may distinguish young patients with cardiovascular disease and those with arrhythmias from healthy individuals. miR-21-5p and miR-26b-5p may also be seen as potential biomarkers of ventricular arrhythmia. Further studies involving a larger sample size are required to obtain sufficient data and validate these findings. Full article

Urinary microRNAs (miRNAs) are promising noninvasive biomarkers for cancer detection, but their clinical utility is reduced by inconsistent normalization strategies, reducing reproducibility and comparability across studies. In this study, we assessed the stability of miR-20a as an endogenous normalizer for urinary miRNA profiling in clear cell renal cell carcinoma (ccRCC) while standardizing the pre-analytical phase using a urine stabilizing solution. Ninety-nine urine samples were analyzed: 47 from healthy individuals, 30 from ccRCC patients pre-surgery, and 22 post-operative patients. Six candidate miRNAs—miR-20a, miR-15b, miR-16, miR-15a, miR-210-3p, and miR-let-7b—were quantified via RT-qPCR. Stability analysis with RefFinder, integrating multiple algorithms (geNorm, normFinder, BestKeeper, and ΔCt methods), identified miR-20a as the most stable among the six candidates. Raw Ct values of miR-20a were normally distributed (Shapiro–Wilk test, p > 0.05), with no significant intergroup differences (one-way ANOVA, F(2.96) = 2.324, p = 0.103) and minimal intragroup variability (CV% 4.98–6.38). MiR-20a expression remained stable across different tumor staging, grading, and urine storage durations. These findings confirm miR-20a as a robust endogenous normalizer for urinary miRNA analyses and support the feasibility of developing reproducible urinary liquid biopsy workflows for ccRCC, even in settings where immediate sample processing is not feasible. Full article

Triple-negative breast cancer (TNBC) is an aggressive subtype lacking targeted therapies and characterized by pronounced heterogeneity and widespread dysregulation of microRNAs (miRNAs) that influence epithelial-to-mesenchymal transition (EMT) and metastasis. Tumor-derived extracellular vesicles (tEVs) further contribute to TNBC progression by transporting oncogenic cargo that can enhance pro-inflammatory signaling. The synthetic SMRwt peptide has been suggested to modulate oncogenic pathways; however, its effects on EV miRNA composition and inflammatory transcript profiles in TNBC remain unclear. Here, we investigated whether SMRwt alters tEV-associated miRNAs and cytokine transcript signatures relevant to EMT and inflammasome-linked pathways. Extracellular vesicles were isolated from SMR-treated and untreated MDA-MB-231 cells, followed by nanoparticle tracking analysis and small RNA sequencing. SMRwt treatment enriched 11 tumor-suppressive miRNAs (including Let-7a-5p, Let-7b-5p, miR-24-3p, miR-26b-5p, miR-92a-3p, miR-93-5p, and miR-496) previously associated with the regulation of proliferation, EMT, migration, and metastasis. We also observed modest, non-significant decreases (1.01–1.27-fold) in oncogenic miR-1200, miR-374a-5p, and miR-937-3p, which have been implicated in the progression of breast, lung, and bone malignancies. Complementary transcriptomic profiling using the NanoString nCounter Breast Cancer 360 Gene Expression Panel (NanoString Technologies, Inc., Seattle, CA, USA) demonstrated reduced expression of inflammasome-associated cytokines in TNBC cells relative to non-tumorigenic controls, including a log₂ fold change of −1.15 for IL 1β (MDA-MB-231 vs. MCF10A). These transcript-level changes suggest potential modulation. Additionally, SMRwt suppresses ASC-mediated caspase-1 activation and reduces IL-1β secretion, thereby inhibiting NLRP3 inflammasome signaling. Therefore, we infer that SMRwt simultaneously restores tumor-suppressive miRNA networks and suppresses inflammasome-driven inflammation, supporting its potential as a dual-target therapeutic strategy for TNBC. Full article

Staphylococcus aureus (S. aureus) is the most prevalent pathogen associated with subclinical mastitis, which significantly impacts dairy farming worldwide. Fluctuations in reproductive hormones, such as bovine prolactin (bPRL) and 17β-estradiol (E2), are known to compromise the innate immune response (IIR) of the mammary gland (MG). In this study, we evaluated the effects of bPRL and E2 on the effector response of primary bovine macrophages, isolated from lactating Holstein cows, challenged with S. aureus. We demonstrated that physiological concentrations of bPRL (5 ng/mL) and E2 (50 pg/mL) induced differential changes in the expression of pro-inflammatory (TNF-α, IL-6, and IL-1β) and anti-inflammatory (IL-10) cytokines, chemokines (IL-8), antimicrobial peptides (BNBD10 and S100A7), and miRNAs (miR-451, miR-155, miR-7863, miR-146a, miR-21a, Let-7a-5p, miR-30b, and miR-23a) in S. aureus-challenged macrophages. Moreover, these hormones promoted global histone H3 acetylation and the epigenetic H3K9ac mark without affecting H3K9me2 levels. Hormonal treatment also modulated histone deacetylase (HDAC) activity. Furthermore, hormonal treatment altered macrophage chemotaxis and phagocytosis. In conclusion, bPRL and E2 modulate the effector functions of bovine macrophages during S. aureus infection. This process could be associated with the regulation of histone H3 modifications, such as H3K9ac, in IIR-related genes. Full article

Post-infectious Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) is a chronic disease with unresolved pathophysiology and limited diagnostic options. Extracellular vesicles (EVs) carry disease-specific protein and miRNA signatures and may enable improved disease profiling. We aimed to identify novel protein and miRNA markers as potential biomarkers in plasma EVs from female ME/CFS patients, including post-COVID-19 ME/CFS and post-infectious ME/CFS of other origins, compared with healthy controls. EVs were isolated from plasma by size-exclusion chromatography and characterized for number, size, morphology, and surface marker expression. Flow cytometry showed that small EVs strongly expressed tetraspanins, with only minor differences between ME/CFS patients and healthy donors. Proteomic profiling of EVs from ME/CFS patients identified altered cargo proteins, including hemoglobin subunit alpha and insulin-like growth factor-binding protein acid labile subunit compared with healthy controls (n ≤ 10/cohort). Small RNA sequencing followed by qPCR revealed significant downregulation of hsa-let-7b-5p in EVs from post-COVID-19 ME/CFS patients (n = 12) versus healthy controls (n = 15). Reduced hsa-let-7b-5p expression correlated with impaired physical functioning and increased fatigue, pain, and immune activation. These findings indicate that EV cargo differences, particularly hemoglobin subunit alpha and insulin-like growth factor-binding protein acid labile subunit, as well as hsa-let-7b-5p, represent promising candidates for ME/CFS diagnosis and patient stratification. Full article

Apigenin, a naturally occurring flavonoid with low toxicity, exhibits anticancer activity, yet its effects on microRNAs (miRNAs) and downstream gene networks in esophageal squamous cell carcinoma (ESCC) remain unclear. Here, we evaluated apigenin’s antitumor effects in TE-1 and Eca-109 cells, assessing proliferation, apoptosis, colony formation, and invasion. Differentially expressed miRNAs were identified via small RNA sequencing, and candidate target genes were predicted, annotated using GO and KEGG analyses, and validated by qRT-PCR, revealing miRNA-mediated regulatory mechanisms underlying apigenin’s inhibitory effects in ESCC. Apigenin markedly suppressed cell proliferation, clonogenic growth, wound closure, and invasive capacity, while promoting apoptosis in a dose-dependent manner. In TE-1 cells, apigenin upregulated hsa-let-7c-3p, hsa-miR-374c-3p, hsa-miR-3177-3p hsa-miR-4454, and hsa-miR-4728-3p, while downregulating hsa-miR-573, hsa-miR-548az-5p, hsa-miR-33b-5p, hsa-miR-4479, and hsa-miR-3198. Correspondingly, tumor-associated target genes including ALDH3A2, SEMA3F, MAP4K5, and TRIP13 were upregulated, whereas PIK3IP1, AGO2, MMP2, and RALBP1 were suppressed. In Eca-109 cells, apigenin altered the expression of distinct miRNAs, including the upregulation of hsa-miR-891-5p, hsa-miR-3170, hsa-miR-4421, and hsa-miR-675-5p and the downregulation of hsa-miR-153, hsa-miR-3188, and hsa-miR-4435, thereby modulating key oncogenic targets such as MAPK1, SALL4, and COX15. Functional enrichment analyses indicated that apigenin-regulated genes are involved in multiple cancer-related pathways across cytoplasmic and nuclear compartments. Overall, these results suggest that apigenin suppresses ESCC progression via coordinated miRNA–mRNA regulation, highlighting its potential as a therapeutic agent. Full article

Let $(X,\mathfrak{A},P)$ and $(Y,\mathfrak{B},Q)$ be two probability spaces, R be their skew product on the product $\sigma$-algebra $\mathfrak{A}\otimes \mathfrak{B}$ and $\{({\mathfrak{A}}_y,S_y):y\in Y\}$ be a Q-disintegration of R. Then, let $\mathfrak{A}\divideontimes \mathfrak{B}$ be the $\sigma$-algebra generated $\mathfrak{A}\otimes \mathfrak{B}$ and by the family $\mathcal{M}:=\{E\subset X\times Y:\exists N\in {\mathfrak{B}}_0\forall y\notin N\widehat{S_y}\left(E^y\right)=0\}$ and $R_{\divideontimes }$ be the extension of R such that $\mathcal{M}$ becomes the family of $R_{*}$-zero sets ($\widehat{S_y}$ is the completion of $S_y$ and ${\mathfrak{B}}_0=\{B\in \mathfrak{B}:Q\left(B\right)=0\}$). We prove that there exists a lifting $\pi$ on ${\mathcal{L}}^{\infty }\left(R_{\divideontimes }\right)$ and liftings ${\sigma }_y$ on ${\mathcal{L}}^{\infty }\left(\widehat{S_y}\right)$, $y\in Y$, such that sections of $\pi$ determined by Y are lifting invariant (in particular, the sections are measurable), i.e., ${\left[\pi \left(f\right)\right]}^y={\sigma }_y\left({\left[\pi \left(f\right)\right]}^y\right) \mathrm{for} \mathrm{every} y\in Y \mathrm{and} \mathrm{every} f\in {\mathcal{L}}^{\infty }\left(R_{\divideontimes }\right).$ In general, if $\pi$ is an arbitrary lifting on the product, then some sections of $\pi \left(f\right)$ may be even nonmeasurable. The main novelty of my paper lies in expanding the domain of the measure in the product to $\mathfrak{A}\divideontimes \mathfrak{B}$ and constructing on such a much larger abstract space the suitable lifting. Such expansions used to be made only in case of topological spaces, where product of marginal Borel sets was replaced by the Borel subsets of the product space. However, several topological technics are then applied, not approachable in the abstract case. The main theorem is a generalization of earlier lifting results, where either separability of $\mathfrak{A}$ in the Frechet–Nikodým pseudometric was assumed or $R\ll P\times Q$. In case of a separable P and in the case when $R\ll P\times Q$, a characterization of stochastic processes possessing an equivalent measurable version is presented. The theorem is a strong generalization of earlier results (see the introduction) where it was proved only that the lifting modification of a measurable stochastic process (via the lifting constructed there) is again measurable. Full article

The recognition of microRNAs as components of animal-source foods (ASFs) with epigenetic characteristics and regulation has spurred research in an interesting direction, particularly in understanding their microRNAs (miRNAs) fraction. Thus, a constant supply of them through food intake, with equally conserved targets, may facilitate their accumulation in tissues rich in their targets. Here, we consider the potentially dominant miRNAs in animal-source foods (ASFs) documented in the literature, identified through a frequency-weighted ordinal recurrence approach. let-7d-5p, miR-101-3p, and miR-133b consistently showed dominant rankings in a product-specific manner in lean meat. In meat fat, let-7i-5p, miR-30c-5p, and miR-23a-3p were highly ranked. Among various types of meat offal, miR-145-5p, miR-92-5p, and miR-24-3p emerged as the predominant miRNAs. Similarly, in dairy products, miR-200a-3p, miR-200c-3p, miR-223-3p, miR-25-3p, miR-29a-3p, and miR-29b-3p were recurrently dominant, whereas miR-17-5p, miR-184, miR-30e-5p, and miR-92b-3p showed a comparable prevalence in seafood. Even though bioinformatic approaches suggest miRNAs from raw ASFs showed major enrichment of processes and pathways culminating in epithelial barrier integrity modulation, such putative functions tend to be equally enriched by predicted targets of the miRNAs in processed products. Product-specific highly ranked miRNAs from food categories stipulate possible preferential enrichment in contexts of cell–cell adhesion, cytoskeletal dynamics, and inflammatory control by meat (lean, fat, offal), immune homeostasis by dairy, and neural signalling by seafood, providing hypotheses for future functional studies. However, a limited understanding of their stability during gastrointestinal transit may present a more immediate limitation to their potential translational applicability. Full article

Let ${\mathcal{M}}_{\alpha }$ be the bilinear fractional maximal operator. In this paper, we prove that the commutators ${\mathcal{M}}_{\alpha ,b}^i$ in the i-th entry $(i=1,2)$ and the bilinear iterated commutators ${\mathcal{M}}_{\alpha ,\overrightarrow{b}}$ of ${\mathcal{M}}_{\alpha }$ are bounded operators from product weighted Morrey spaces $L^{p_1,\frac{\kappa p_1}{q_1}}\left(w_1^{p_1},w_1^{q_1}\right)\times L^{p_2,\frac{\kappa p_2}{q_2}}\left(w_2^{p_2},w_2^{q_2}\right)$ to weighted Morrey spaces $L^{q,\kappa }\left(v_{\overrightarrow{w}}^q\right)$, provided that $b\in \mathrm{BMO}\left({\mathbb{R}}^{\mathrm{n}}\right)$ and $\overrightarrow{b}=(b_1,b_2)\in \mathrm{BMO}\left({\mathbb{R}}^{\mathrm{n}}\right)\times \mathrm{BMO}\left({\mathbb{R}}^{\mathrm{n}}\right)$. Furthermore, by using the techniques of function decompositions and the Fréchet–Kolmogorov theorem on weighted Morrey spaces, the compactness of ${\mathcal{M}}_{\alpha ,b}^i(i=1,2)$ and ${\mathcal{M}}_{\alpha ,\overrightarrow{b}}$ are also established whenever $b\in \mathrm{CMO}\left({\mathbb{R}}^{\mathrm{n}}\right)$ and $\overrightarrow{b}=(b_1,b_2)\in \mathrm{CMO}\left({\mathbb{R}}^{\mathrm{n}}\right)\times \mathrm{CMO}\left({\mathbb{R}}^{\mathrm{n}}\right)$, where $\mathrm{CMO}\left({\mathbb{R}}^{\mathrm{n}}\right)$ denotes the closure of $C_c^{\infty }\left({\mathbb{R}}^n\right)$ in the $\mathrm{BMO}\left({\mathbb{R}}^{\mathrm{n}}\right)$ topology. Full article

Outer membrane vesicles (OMVs) are tractable, cell-free microbial outputs that can shape innate immune programs. Here, we compared OMVs from the probiotic Escherichia coli Nissle 1917 (EcN) and the commensal strain ECOR12 in a paired within-donor model of human monocyte-derived dendritic cells (Mo-DCs) (N = 20). In the core integrated arm, Mo-DCs were exposed to iDC control, EcN OMVs, or ECOR12 OMVs (10 µg/mL, 24 h) and profiled for maturation markers (CD14, CD83, CD209), cytokines (IL-6, TNF-α, IL-10), and a targeted miRNA panel (miR-155-5p, let-7i-3p, miR-146b-5p, miR-29a-5p). Both OMV types promoted maturation (increased CD83 and reduced CD14), but generated distinct cytokine–miRNA configurations, with ECOR12 tending toward an IL-10–high profile and EcN toward higher IL-6/TNF-α tendencies. Multivariate integration separated conditions into reproducible, strain-specific immune fingerprints, supporting the key take-home that probiotic versus commensal E. coli OMVs imprint distinguishable coordinated response states in human DCs. In an extended phenotyping arm, ECOR63 OMVs were evaluated by ELISA and flow cytometry only and were not included in miRNA profiling or integrated PCA due to unavailable matched miRNA measurements. Full article

Background/Objectives: In severe peripheral artery disease (PAD) with limb ischemia, hypercholesterolemia (HC) is associated with impaired neovascularization. Extracellular vesicles (EVs) are present within ischemic muscles, and they contain microRNAs (miRs) involved in several biological functions, including angiogenesis and neovascularization. Methods: We used a mouse model of PAD and compared the response to hindlimb ischemia in hypercholesterolemic ApoE^−/− vs. normocholesterolemic mice. Next-generation sequencing (NGS) was used to perform full miR expression profiling in ischemic skeletal muscles and in EVs of varying sizes—large EVs (lEVs) and small EVs (sEVs)—within these muscles. Results: We identified several miRs with potential pro-angiogenic effects (angiomiRs) that are reduced by HC in lEVs (Let-7b-5p, miR-151-3p, Let-7c-5p) or sEVs (miR-21a-5p, miR-196b-5p, miR-340-5p). As proof of principle, we showed that the overexpression of Let-7b-5p in lEVs, or miR-21a-5p in sEVs, can significantly increase the angiogenic capacity of these EVs in vitro. HC also impaired the enrichment of specific angiomiRs in lEVs (miR-100-5p), sEVs (miR-142a-3p), or in both lEVs and sEVs (miR-146b-5p). In silico approaches, including the prediction of miR targets, pathway unions, and gene unions, identified the resulting predictive effects of HC-modulated miRs in EVs on processes with key roles in the modulation of angiogenesis and neovascularization, such as the regulation of the actin cytoskeleton and focal adhesion and the HIF-1, MAPK, AMPK, and PI3K-Akt signaling pathways. Conclusions: Our results constitute an important first step towards the identification of specific miRs that could be targeted to improve EV angiogenic function in hypercholesterolemic conditions and reduce tissue ischemia in patients with severe PAD. Full article