bluuebunny/crossref_metadata_2025_split huggingface dataset:
Development of a human in vitro model of bone marrow vascular niche on a chip to study CD34+ hematopoietic stem/progenitor cells function in diabetes
Authors: Campanile M , Persico G , Casciaro F , Noto G , Sambado L , Bettinelli L , Stefani P M , Sambataro M , Giorgio M , Cerutti C , Spinetti G 
Published: November 2023

Abstract

Background/Introduction
Protective bone marrow (BM) CD34+ hematopoietic stem/progenitor cells (HSPCs) are released in the bloodstream after cardiomacrovascular (CV) tissue damage such as myocardial infarction1. Diabetes, an high risk condition to develop CV diseases, results in BM damage2-4 and detrimental pro-apoptotic and pro-oxidant shifts in BM-derived CD34+HSPCs5,6. Being able to classify and study the mechanisms of CD34+HSPC-vascular cell interaction would be a great step forward in the CV research field. However, nowadays there is no available primary human cell-based in vitro model to study the BM vascular niche.

Purpose
We aim to develop a human-based in vitro model to reproduce the BM vascular niche that allows the analysis of molecular mechanisms involved in CD34+HSPC function. This model will be relevant to identify molecular targets beyond hypoglycemic treatment and potentiate personalized medicine applications for diabetic complications.

Method
Human endothelial cells and pericytes were isolated from femoral head of patient who underwent hip replacement or from ad hoc BM aspirate by magnetic bead separation and characterized by immunofluorescence staining and flow cytometry. A culture of endothelial and pericytic cells to replicate the BM vascular niche was developed on a chip. CD34+HSPC subpopulation alteration in diabetes was preliminarily evaluated through single cell sequencing analysis (scRNAseq) on CD34+HSPC cells isolated from BM of patients with or without diabetes (N=1).

Results
We successfully set up a new method to efficiently isolate autologous BM endothelial cells and pericytes derived from patients (N=3). The cells expressing typically endothelial (CD31, ZO-1, and VE-cadherin) and pericytic (CD146, CD140b) markers were cultured in a 2D chip model. ScRNAseq reveals 15 clusters of CD34+HSPCs and important differences between the non-diabetic and diabetic samples. In particular, cluster 2 is highly represented in the non-diabetic and completely disappears in the diabetic sample, while on the contrary cluster 9 is over-represented in the diabetic compared to the non-diabetic sample. Interestingly, though the trajectory analysis cluster 2 shows the highest probability of turning into cluster 9. Through Ingenuity Canonical Pathways analysis we identified the anti-oxidant pathway as the most affected in diabetes-derived CD34+HSPCs.

Conclusions
In vitro human vascular niche model could be the gold standard to study BM cell crosstalk. In our interest, cluster 2 completely disappears in diabetes-derived CD34+HSPCs, potentially representing the CV-protective CD34+HSPC fractions more prone to be mobilized. Developing of our in vitro model would be useful to validate this hypothesis.Figure 1:scRNAseq cluster UMAPFigure 2:Endothelial cells IF staining
Circulating long noncoding RNAs and risk of ischemic stroke or death in the elderly: insights from the VITA study
Authors: Lapikova-Bryhinska T , Ministrini S , Kraler S , Puspitasari Y M , Mohammed S , Costantino S , Riederer P , Fischer P , Hinterberger M , Paneni F , Luscher T , Grunblatt E , Akhmedov A , Camici G 
Published: November 2023

Abstract

Introduction
Long non-coding RNAs (lncRNAs) represent a superfamily of non-protein coding transcripts that regulate a variety of cellular processes and functions in health and disease. While our (patho-)physiological understanding of lncRNAs constantly evolves, their potential as cardiovascular biomarkers to predict adverse outcomes in the elderly remains poorly understood.

Method
A total of 361 healthy individuals aged 75, prospectively recruited in the Vienna Transdanube Aging (VITA) cohort, were included in the present analysis. A semi biased lncRNA screening strategy was employed: "Aging", ‘’stroke", "cardiac", "inflammation", and "neurodegenerative" served as keywords. The primary endpoint was a composite outcome of death and ischemic stroke. Death was assessed over a 14-year follow-up from public registries, and ischemic stroke was evaluated over a 90-month follow-up involving magnetic resonance imaging (MRI) based assessment. Expression of lncRNAs wasassessed using qRT-PCR (TaqMan Assays) with preamplification reaction, using 18S for normalization.

Results
The initial screening retrieved 24 lncRNAs, of which 6 showed a solid amplification signal. Over a median follow-up of 14 years, the incidence of the primary endpoint was 62.6%, with 53.5% deaths, 3.3% brain infarcts, and 12.7% lacunar strokes (which were studied during the 7.5 years).
H19 predicted the incidence of the primary endpoint (HR 1.675, 95% C.I. 1.076-2.607, p= 0.023), which was mainly driven by its strong association with mortality, independent of potential confounders (adjusted HR 1.789, 95% C.I. 1.083-2.955, p= 0.023). In contrast, NKILA was found to predict the incidence of lacunar stroke beyond established risk factors (adjusted HR 0.259, 95% C.I. 0.087-0.768, p= 0.015) but was unexpectedly not associated with mortality risk.

Conclusion
In a prospective cohort of healthy elderly subjects, high levels of lncRNA H19 are associated with a high mortality risk, while low levels of lncRNA NKILA predict an increased risk of lacunar stroke.
Epigenetic regulation of PCSK9 expression by statins
Authors: Duddu S , Chakrabarti R , Sharma P , Tiwari A , Shukla P C 
Published: November 2023

Abstract

Background
Statins are lipid-lowering drugs widely used to reduce the risk of cardiovascular diseases. Although they inhibit HMG-CoA, statins are known to induce the expression of PCSK9, a key regulator of plasma LDL-cholesterol. However, the underlying epigenetic mechanisms remain unknown.

Objective
This study aimed at investigating the histone modification-mediate epigenetic regulation of PCSK9 and the underlying mechanisms during statin treatment.

Methods
Using immunoblotting, qPCR, and immunofluorescence techniques we assessed various histone modifications and expression levels of PCSK9 and other cholesterol-regulating candidates upon statin treatment. Further, 4-6 weeks old C57BL/6 mice were subjected to high-fat diet in combination with atorvastatin for 10 weeks. We then performed ChIP-qPCR to evaluate the fold enrichment of targeted histone marks at the gene promoter regions of PCSK9 and SREBP.

Results
Atorvastatin and cholesterol-treated HepG2 cells were reported to have an increased transcript and protein expression of PCSK9. Further, screening of different histone methylation and acetylation marks under atorvastatin treatment revealed significant enrichment in the levels of the activating H3K4me3 and H3K9ac marks. ChIP-qPCR analysis suggested significantly higher enrichment of H3K4me3 and H3K9ac at the gene promoter region of PCSK9 and SREBP2 (Figure 1&2). In vitro pharmacological intervention against these histone modifications not only significantly lowered their activation but also attenuated statin-induced PCSK9 expression.

Conclusion
Together, these findings suggest that statins modulate the expression of PCSK9 by regulating H3K4me3 and H3K9ac levels at the PCSK9 gene promoter. Inhibition of these activating histone modifications repressed statin-dependent PCSK9 expression. Thus, targeting chromatin remodeling may help negate the adverse effects of statin treatment, primarily of increasing PCSK9 during hypercholesterolemia.
Inhibition of farnesylation of Ras-GTPases ameliorates inflammation-induced loss of endothelial barrier integrity and leukocyte transmigration
Authors: Guenduez D , Hamm C , Aslam M 
Published: November 2023

Abstract

Background and Aims
The family of small GTPases (Rho- and Ras-GTPases) is known to regulate endothelial cytoskeleton dynamics and hence barrier integrity. The role of Rho GTPases (RhoA and Rac1) in endothelial barrier integrity is well characterized; however, little is known about the role of Ras-GTPases. The activity of small GTPases is regulated by several mechanisms, including post-translational prenylation (farnesylation or geranylgeranylation). The Ras-GTPases are preferentially post-translationally farnesylated by farnesyl transferases (FTs). The aim of the present study was to investigate whether inhibition of farnesylation of Ras-GTPases protects loss of endothelial barrier integrity induced by inflammatory mediators.

Methods and Results
Treatment of human primary endothelial cells (ECs; HUVEC and lung microvascular ECs) with pharmacological inhibitor of farnesyl transferases (FTIs), LB42708 (LB) and tipifarnib, stabilized basal endothelial barrier function (albumin-flux) and abrogated thrombin-induced hyperpermeability in a concentration- and time-dependent manner reaching maximum at a concentration of 2 mmol/L after 12 h of incubation. These concentrations of the FTIs reduced farnesylation of all the Ras isoforms but not the geranylgeranylation of Rho GTPases. Likewise, LB abrogated the TNFa-induced leucocyte transendothelial migration (Trans-well assay). Interestingly, FTIs treatment caused an enhanced translocation of VE-cadherin to cell-cell junctions (confocal microscopy) without exerting any effect on the actin cytoskeleton or endothelial contractile machinery; mainly regulated by Rho GTPases. The Ca2+-switch experiments demonstrated an inhibition of VE-cadherin internalization. These data were reproduced by lentiviral-mediated stable over-expression of the prenylation-dead mutants of Ras isoforms but not RhoA or Rac1.

Conclusion
The novel data of the present study demonstrate that FTIs protect endothelial barrier function against inflammatory mediator-induced endothelial hyperpermeability. We propose that FTIs are novel drug candidates for the treatment of edematous conditions (such as lung edema) where endothelial barrier integrity is compromised.
Kallstatin/Serpina3c improves hyperlipidemia induced by high-fat diet via regulating glycolipid metabolism in adipose tissue
Authors: Guo J , Ji Z , Ma G , Yao Y 
Published: November 2023

Abstract

Background
Hyperlipidemia is an independent risk factor for cardiovascular diseases. Studies showed that the plasma Kallstatin (KS) in obese patients was reduced, and the decreased KS in obese patients with type 2 diabetes was increased after weight loss. The mouse homologous gene of human KS is Serpina3c, and the relationship between KS/Serpina3c and hyperlipidemia and its regulatory mechanism are unclear.

Purpose
To explore whether KS/Serpina3c improves hyperlipidemia and specific mechanisms.

Methods
The serum KS level of Chinese hyperlipidemia population was detected by ELISA. Serpina3c knockout mice (Serpina3c KO) were constructed based on the strategy of Cre/loxP recombination system. The adipocyte-specific Serpina3c knockout adeno-associated virus (AAV8-shSerpina3c) and control (AAV8-shNC) were constructed, and the adipocyte-specific Serpina3c overexpression adeno-associated virus (AAV8-Serpina3c) were constructed. The 8-week-old C57BL/6J mice were injected with AAV8-shSerpina3c or AAV8-shNC in situ into the inguinal adipose tissue (iWAT). Serpina3c KO mice were injected with AAV8-Serpina3c or AAV8-shNC in situ into the iWAT. The mice model of hyperlipidemia was established by feeding high-fat diet (HFD) for 16 weeks. The fat content of mice was observed with 7.0T magnetic resonance imaging (MRI)system. The serum triglyceride, total cholesterol, LDL and HDL were detected with the kit. Serpina3c KD, Serpina3c OV or control 3T3-L1 adipocytes were stimulated by PA for RNA-seq.

Results
(1) The serum KS decreased in hyperlipidemic people in China. (2) The 7.0T MRI system displayed bigger fat in the Serpina3c KO mice and AAV8-shSerpina3c mice. (4) The serum results showed higher lipids in Serpina3c KO and AAV8-shSerpina3c mice. (5) Oil red O staining showed increased lipid deposition in liver, muscle and heart. (6) The RNA-seq results revealed that the HIF1α and glycolysis pathways of Serpina3c KD adipocyte increased, while the fatty acid metabolic pathway decreased. The results of primary adipocytes verified the increase of glycolysis and decreased β oxidation. (7) And the fatty acid uptake in liver, heart and muscle tissue of Serpina3c KO mice and AAV8-shSerpina3c mice increased, and the β oxidation decreased.

Conclusions
The expression and secretion of KS/Serpina3c decreased in HFD adipose, which promotes glycolysis and inhibits fatty acids β oxidation of adipocytes. On the one hand, the decrease of Serpina3c activates glycolysis and increases the production of glycerol 3-phosphate by activating HIF1α. On the other hand, CPT1 was inhibited by increased HIF1α, impeding the activated fatty acid (Fatty-acyl-CoA) in the cytoplasm entering the mitochondria through CPT1. Glycerol 3-phosphate and fatty acyl CoA accumulated in the cytoplasm of adipocyte synthesize triglycerides through the diglyceride pathway, aggravating hyperlipidemia induced by HFD.Schematic diagram
Dapagliflozin decreases atherosclerotic plaque instability via regulating macrophage pyroptosis
Authors: Xu L , Dai Y , Qian J , Ge J 
Published: November 2023

Abstract

Background
Vulnerable plaques are characterized by infiltration of inflammatory cells, playing a key role in the progression of acute coronary events. It’s important to clarify the inflammatory mechanism of unstable plaque formation. Several clinical trials have demonstrated that dapagliflozin could reduce major adverse cardiac events in whether diabetic or non-diabetic patients. However, the underlying cardioprotective mechanism of dapagliflozin remains unclear. This study was aimed to investigate the role of dapagliflozin in regulating macrophage pyroptosis and vulnerable plaque formation.

Methods
20 ApoE-/- mice (control) were fed with high fat diet while another 20 ApoE-/- mice were challenged with high fat diet plus dapagliflozin for 12 weeks. The extent and instability of atherosclerotic plaque was determined by oil-red staining, HE staining, immunofluorescence staining and electron microscopy. Changes in subsets of immune cells were evaluated by flow cytometry. Plasma cytokines were assessed by ELISA. Microarray analysis was applied to detect gene expressions while Western blot and real-time PCR was used to assess gene expression levels.

Results
Morphology studies revealed that dapagliflozin could inhibit plaque formation and reduce instability in ApoE-/- mice. FACS data showed that dapagliflozin could decrease CD11b+Ly6Chigh M1 macrophages differentiation and inhibit foam cells formation in ApoE-/- mice. Microarray analysis and in vitro studies exhibited that dapagliflozin could induce the down regulation of NLRP3, caspase-1, IL-1β, IL-18 and MMP-7/10/12/14 to retard macrophage pyroptosis and foam cell formation.

Conclusions
We have characterized a novel role for dapagliflozin in modulating atherosclerotic lesion development and progression. We envision that this study may provide several potential therapeutic targets for treatment of acute coronary syndromes.
ZAP (Zero-Fluoro - ALARA Procedures) EP: A UK Tertiary Centre Safety and Efficiency Evaluation
Authors: Makris N , Kotsialou Z , Elbanhawy N , Ariyaratnam J , Chalil S , Gall S , Chu G , Arujuna A 
Published: November 2023

Abstract

Background
3D mapping facilitates zero-fluoro (ZF) and as-low-as-reasonably-achievable (ALARA) procedures which allows for minimizing exposure to harmful radiation and maximizing lead-free time in the EP lab.

Objective
Safety, efficiency, radiation time and dose exposure was assessed in a consecutive unselected supraventricular tachycardias (SVT) cohort.

Methods
Retrospective and prospective analysis comparing procedure time, complications and radiation dose and time between ZAP (ZF, ALARA) and non-ALARA cases was performed between January 2021 to August 2022. Radiation exposure was expressed as dose-area product (DAP). We included AVNRTs, accessory pathway (AP) ablations and EP studies.

Results
A total of consecutive 279 patients were included, of which 116 were ZAP (40 were ZF cases) and 163 non-ALARA. Median age was 52.5 for ZAP and 47.5 for non-ALARA patients. In specific, 131 AVNRT ablations (50 ZAP, 81 non-ALARA), 45 AP ablations (23 ZAP, 22 non-ALARA) and 103 EP studies (43 ZAP, 60 non-ALARA) were included. A comparative analysis of DAP between ZAP and non-ALARA cohort in both AVNRT ablations and EP studies demonstrated statistical significance with corresponding values of 3.15±4.4 vs. 6.04±7.5 Gy.cm2 (p=0.001) and 1.94±1.8 vs. 3.78±3.9 Gy.cm2 (p=0.024) (Figure 1). Significantly shorter fluoroscopy duration was also noted 6.07±6 vs. 12.44±7.5 min (p&lt;0.001; AVNRT) and 4.23±4.2 vs. 6.16±4.1 min (p=0.01; EPS). For AP ablations there was a trend in favour of ZAP but not significant difference (DAP: 8.52±7.7 vs. 10.65±9 Gy.cm2, p=0.43; fluoroscopy duration: 12.12±8.7 vs. 15.9±8.3 min, p=0.33) (Figure 1). This might be attributed to the relatively smaller number of AP cases. Complication rates and procedure time were not different between ZAP and non-ALARA cases.

Conclusion
Lower fluoroscopy time with reduced radiation exposure is seen in ZAP cases compared to non-ALARA procedures for AVNRT ablations and EP studies. Future ICE navigation in complex cases may further reduce fluoroscopy need.Figure 1
Colchicine prevents accelerated atherosclerosis development in TET2-mutant clonal hematopoiesis
Authors: Zuriaga M A , Yu Z , Matesanz N , Truong B , Asensio-Lopez M C , Uddin M M , Nakao T , Niroula A , Zorita V , Moro R , Ebert B L , Honigberg M , Pacual-Figal D , Natarajan P , Fuster J J 
Published: November 2023

Abstract

Background
Somatic mutations in the TET2 gene that lead to clonal hematopoiesis are associated with accelerated atherosclerosis development in mice and a higher risk of atherosclerotic cardiovascular disease events in humans. Mechanistically, these observations have been linked to exacerbated vascular inflammation.

Purpose
Here, we sought to evaluate whether colchicine, a widely available and inexpensive anti-inflammatory drug, prevents the accelerated atherosclerosis associated with TET2-mutant clonal hematopoiesis.

Methods
In mice, we mimicked TET2-mutant clonal hematopoiesis using a competitive bone marrow transplantation approach in atherosclerosis prone Ldlr/- mice. We generated hematopoietic chimeras carrying initially 10% Tet2-/- hematopoietic cells, which were fed a high-cholesterol diet and treated with colchicine or placebo. Atherosclerosis development was evaluated histologically. In humans, we analyzed whole exome sequencing data and electronic medical records from 50,291 participants in a hospital-based cohort to examine the potential modifying effect of colchicine prescription on the association between clonal hematopoiesis and myocardial infarction.

Results
Colchicine prevented the development of accelerated atherosclerosis in the mouse model of TET2-mutant clonal hematopoiesis, without affecting serum cholesterol levels or the expansion of TET2-deficient hematopoietic cells. In humans, consistent with experimental findings, patients who were prescribed colchicine had attenuated associations between TET2-mutant clonal hematopoiesis and myocardial infarction (ORno colchicine [95% CI]: 1.18 [0.93-1.48]; ORcolchicine [95% CI]: 0.78 [0.45-1.35]; Pinteraction=0.03). This interaction was not observed for carriers of other clonal hematopoiesis-related mutations, such as those affecting DNMT3A or ASXL1.

Conclusions
Our results highlight the potential value of colchicine as a targeted preventive care strategy to mitigate the higher cardiovascular risk of carriers of somatic TET2 mutations in blood cells. These observations set the basis for the development of clinical trials that evaluate the efficacy of precision medicine approaches tailored to the effects of specific somatic mutations linked to clonal hematopoiesis.
The senolytic agent Dasatinib reduces fat accumulation in the heart and bone marrow of type 2 diabetic, obese mice and adipogenesis of human bone marrow pericytes
Authors: Gu Y , Alvino V V , Avolio E , Morelli M B , Thomas A C , Herman A , Miller P J , Sullivan N P T , Madeddu P 
Published: November 2023

Abstract

Background
Inflammaging, a chronic inflammatory state associated with aging and obesity, participates in the pathogenesis of cardiovascular disease and type 2 diabetes (T2D). Adipocyte accumulation in bone marrow (BM) may contribute to this phenomenon. Senolytic drugs, like Dasatinib, reportedly eliminate senescent preadipocytes. However, the effects of Dasatinib on fat deposits in the heart and BM remain unknown.

Purposes
This study aimed to identify the cellular source of adipocytes in human BM and determine whether treatment with Dasatinib reverses adiposity and improves cardiovascular function in T2D obese mice.

Methods and results
Human CD34neg CD45neg CD146pos pericytes (hBM-PCs) were sorted from BM mesenchymal stem cells (hBM-MSCs) by immunomagnetic beads. HBM-PCs showed a higher propensity to differentiate into Oil red O positive (pos) adipocytes after 21 days of stimulation in a pro-adipogenic medium (P=0.0299 vs. hBMSCs (hBM-MSCs except for hBM-PCs)). Dasatinib treatment reduced the number of viable cells (P=0.0034 vs. Vehicle (DMSO)) and the rate of adipocyte differentiation (P= 0.0179 vs. Vehicle). In a randomized blind study, Dasatinib (5 mg/kg) or vehicle (10% DMSO+90% PEG-300) was given by oral gavage to T2D, obese 21-week-old male BKS.Cg-+Leprdb/+Leprdb/OlaHsd (Db/Db) mice. Groups of animals were sacrificed at 1 or 4 weeks. Dasatinib reduced the Oil red O pos cells in the heart (P= 0.001 and 0.045) and BM (P= 0.018 and 0.0253) at both time points. Senescence marker p16INK4a expression and plasma adipokines levels were similar between the two groups at one week. With long-term treatment, the expression of senescence marker p16INK4a was significantly reduced in the murine hearts (P=0.035) and BM (P=0.068), whereas adipokines levels were similar. In vitro treatment of murine BM-MSCs with Dasatinib reduced the expression of p21Waf1/Cip1 (P= 0.030). VisualSonics 3D echocardiography showed similar diastolic and systolic function in Dasatinib- or Vehicle-treated mice: stroke volume (P= 0.494); ejection fraction (P= 0.701); fractional shorten (P= 0.807); and cardiac output (P= 0.437).

Conclusion
Pericytes are the primary source of adipocytes in human BM. The senolytic agent Dasatinib reduces pericyte commitment to adipogenesis. The senolytic agent prevented fat accumulation and cell senescence in the BM and heart of T2D mice. However, more prolonged treatment may be necessary for the reduced adiposity to translate into improvements in heart function.