Recent Advances in the Modification and Improvement of Bioprosthetic Heart Valves.

Valvular heart disease (VHD) has become a burden and a growing public health problem in humans, causing significant morbidity and mortality worldwide. An increasing number of patients with severe VHD need to undergo heart valve replacement surgery, and artificial heart valves are in high demand. However, allogeneic valves from donors are lacking and cannot meet clinical practice needs. A mechanical heart valve can activate the coagulation pathway after contact with blood after implantation in the cardiovascular system, leading to thrombosis. Therefore, bioprosthetic heart valves (BHVs) are still a promising way to solve this problem. However, there are still challenges in the use of BHVs. For example, their longevity is still unsatisfactory due to the defects, such as thrombosis, structural valve degeneration, calcification, insufficient re-endothelialization, and the inflammatory response. Therefore, strategies and methods are needed to effectively improve the biocompatibility and longevity of BHVs. This review describes the recent research advances in BHVs and strategies to improve their biocompatibility and longevity.

Detoxification and Activating Blood Circulation Decoction Promotes Reendothelialization of Damaged Blood Vessels via VEGF Signaling Pathway Activation by miRNA-126.

The occurrence of in-stent restenosis (ISR) poses a significant challenge for percutaneous coronary intervention (PCI). Thus, the promotion of vascular reendothelialization is essential to inhibit endothelial proliferation. In this study, we clarified the mechanism by which Detoxification and Activating Blood Circulation Decoction (DABCD) promotes vascular reendothelialization to avoid ISR by miRNA-126-mediated modulation of the vascular endothelial growth factor (VEGF) signaling pathway. A rat model of post-PCI restenosis was established by balloon injury. The injured aortic segment was collected 14 and 28 d after model establishment. Our findings indicate that on the 14th and 28th days following balloon injury, DABCD reduced intimal hyperplasia and inflammation and promoted vascular reendothelialization. Additionally, DABCD markedly increased nitric oxide (NO) expression and significantly decreased ET-1 production in rat serum. DABCD also increased the mRNA level of endothelial nitric oxide synthase (eNOS) and the protein expression of VEGF, p-Akt, and p-extracellular signal-regulated kinase (ERK)1/2 in vascular tissue. Unexpectedly, the expression of miR-126a-5p mRNA was significantly lower in the aortic tissue of balloon-injured rats than in the aortic tissue of control rats, and higher miR-126a-5p levels were observed in the DABCD groups. The results of this study indicated that the vascular reendothelialization effect of DABCD on arterial intimal injury is associated with the inhibition of neointimal formation and the enhancement of vascular endothelial activity. More specifically, the effects of DABCD were mediated, at least in part, through miR-126-mediated VEGF signaling pathway activation.

Reendothelialization of Acellular Adipose Flaps under Mimetic Physiological Dynamic Conditions.

The extensive soft-tissue defects resulting from trauma and tumors pose a prevalent challenge in clinical practice, characterized by a high incidence rate. Autologous tissue flap transplantation, considered the gold standard for treatment, is associated with various drawbacks, including the sacrifice of donor sources, postoperative complications, and limitations in surgical techniques, thereby impeding its widespread applicability. The emergence of tissue-engineered skin flaps, notably the acellular adipose flap (AAF), offers potential alternative solutions. However, a critical concern confronting large-scale tissue-engineered skin flaps currently revolves around the reendothelialization of internal vascular networks. In our study, we have developed an AAF utilizing perfusion decellularization, demonstrating excellent physical properties. Cytocompatibility experiments have confirmed its cellular safety, and cell adhesion experiments have revealed spatial specificity in facilitating endothelial cells adhesion within the adipose flap scaffold. Using a novel mimetic physiological fluid shear stress setting, endothelial cells were dynamically inoculated and cultured within the acellular vascular network of the pedicled AAF in our research. Histological and gene expression analyses have shown that the mimetic physiological fluid dynamic model significantly enhanced the reendothelialization of the AAF. This innovative platform of acellular adipose biomaterials combined with hydrodynamics may offer valuable insights for the design and manufacturing of 3D vascularized tissue constructs, which can be applied to the repair of extensive soft-tissue defects.

Liver tissue engineering using decellularized scaffolds: Current progress, challenges, and opportunities.

Liver transplantation represents the only definitive treatment for patients with end-stage liver disease. However, the shortage of liver donors provokes a dramatic gap between available grafts and patients on the waiting list. Whole liver bioengineering, an emerging field of tissue engineering, holds great potential to overcome this gap. This approach involves two main steps; the first is liver decellularization and the second is recellularization. Liver decellularization aims to remove cellular and nuclear materials from the organ, leaving behind extracellular matrices containing different structural proteins and growth factors while retaining both the vascular and biliary networks. Recellularization involves repopulating the decellularized liver with appropriate cells, theoretically from the recipient patient, to reconstruct the parenchyma, vascular tree, and biliary network. The aim of this review is to identify the major advances in decellularization and recellularization strategies and investigate obstacles for the clinical application of bioengineered liver, including immunogenicity of the designed liver extracellular matrices, the need for standardization of scaffold fabrication techniques, selection of suitable cell sources for parenchymal repopulation, vascular, and biliary tree reconstruction. In vivo transplantation models are also summarized for evaluating the functionality of bioengineered livers. Finally, the regulatory measures and future directions for confirming the safety and efficacy of bioengineered liver are also discussed. Addressing these challenges in whole liver bioengineering may offer new solutions to meet the demand for liver transplantation and improve patient outcomes.

Diabetic conditions promote drug coating degradation but prevent endothelial coverage after stenting.

The endothelialization of drug-eluting stents is delayed after implantation in patients with diabetes. Although numerous factors were implicated in hyperglycemia-induced endothelial dysfunction, the effects of stent drug coating degradation on endothelial dysfunction remains unclear. We hypothesized that diabetic conditions promote drugcoating degradation and enhance antiproliferative agent release, but that the rapid release of these antiproliferative agents inhibits endothelial cell proliferation leading to poor reendothelialization post-stenting. To verify this hypothesis, a dynamic hyperglycemic circulation system was introduced to measure the profile of drugcoating degradation in vitro. Flow cytometry and RNA sequencing were performed to evaluate endothelial cell proliferation. Moreover, a Type 1 diabetic rabbit model was generated and a rescue experiment conducted to evaluate the effects of rapid drugcoating elution on endothelial coverage in vivo. The main findings were as follows: 1) diabetic conditions promoted drugcoating degradation and increased antiproliferative agent release; 2) this increase in antiproliferative agent release inhibited endothelial cell proliferation and delayed endothelial coverage; and 3) strict glycemic control attenuated drugcoating degradation and promoted endothelial coverage post-stenting. This is the first study to illustrate rapid drugcoating degradation and its potential effects on endothelial recovery under diabetic conditions, highlighting the importance of strict glycemic management in patients with diabetes after drug-eluting stent implantation. STATEMENT OF SIGNIFICANCE: Diabetic conditions promote drug coating degradation and increase the release of antiproliferative agents. Rapid drug coating degradation under diabetic conditions inhibits endothelial cell proliferation and delays endothelialization. Strict glycemic control attenuates drug coating degradation and promotes endothelialization.

Nanotechnology in development of next generation of stent and related medical devices: Current and future aspects.

Coronary stents have saved millions of lives in the last three decades by treating atherosclerosis especially, by preventing plaque protrusion and subsequent aneurysms. They attenuate the vascular SMC proliferation and promote reconstruction of the endothelial bed to ensure superior revascularization. With the evolution of modern stent types, nanotechnology has become an integral part of stent technology. Nanocoating and nanosurface fabrication on metallic and polymeric stents have improved their drug loading capacity as well as other mechanical, physico-chemical, and biological properties. Nanofeatures can mimic the natural nanofeatures of vascular tissue and control drug-delivery. This review will highlight the role of nanotechnology in addressing the challenges of coronary stents and the recent advancements in the field of related medical devices. Different generations of stents carrying nanoparticle-based formulations like liposomes, lipid-polymer hybrid NPs, polymeric micelles, and dendrimers are discussed highlighting their roles in local drug delivery and anti-restenotic properties. Drug nanoparticles like Paclitaxel embedded in metal stents are discussed as a feature of first-generation drug-eluting stents. Customized precision stents ensure safe delivery of nanoparticle-mediated genes or concerted transfer of gene, drug, and/or bioactive molecules like antibodies, gene mimics via nanofabricated stents. Nanotechnology can aid such therapies for drug delivery successfully due to its easy scale-up possibilities. However, limitations of this technology such as their potential cytotoxic effects associated with nanoparticle delivery that can trigger hypersensitivity reactions have also been discussed in this review. This article is categorized under: Implantable Materials and Surgical Technologies > Nanotechnology in Tissue Repair and Replacement Therapeutic Approaches and Drug Discovery > Nanomedicine for Cardiovascular Disease Therapeutic Approaches and Drug Discovery > Emerging Technologies.

3D-Printed Stents Loaded with Panax notoginseng Saponin for Promoting Re-endothelialization and Reducing Local Inflammation in the Carotid Artery of Rabbits.

Endovascular treatment (EVT) using stents has become the primary option for severe cerebrovascular stenosis. However, considerable challenges remain to be addressed, such as in-stent restenosis (ISR) and late thrombosis. Many modified stents have been developed to inhibit the hyperproliferation of vascular smooth muscle cells (SMCs) and protect vascular endothelial cells (VECs), thereby reducing such complications. Some modified stents, such as those infused with rapamycin, have improved in preventing acute thrombosis. However, ISR and late thrombosis, which are long-term complications, remain unavoidable. Panax notoginseng saponin (PNS), a traditional Chinese medicine consisting of various compounds, is beneficial in promoting the proliferation and migration of VECs and inhibiting the proliferation of SMCs. Herein, a 3D-printed polycaprolactone (PCL) stent loaded with PNS (PNS-PCL stent) was developed based on a previous study. In vitro studies confirmed that PNS promotes the migration and proliferation of VECs, which were damaged, by increasing the expression levels of microRNA-126, p-AKT, and endothelial nitric oxide synthase. In vivo, the PNS-PCL stents maintained the patency of the carotid artery in rabbits for up to three months, outperforming the PCL stents. The PNS-PCL stents may present a new solution for the EVT of cerebrovascular atherosclerotic stenosis in the future.

Passivation protein-adhesion platform promoting stent reendothelialization using two-electron-assisted oxidation of polyphenols.

Superhydrophilic surfaces play an important role in nature. Inspired by this, scientists have designed various superhydrophilic materials that are widely used in the field of biomaterials, such as PEG molecular brushes and zwitterionic materials. However, superhydrophilic coatings with only anti-fouling properties do not satisfy the requirements for rapid reendothelialization of cardiovascular stent surfaces. Herein, a novel polyphenol superhydrophilic surface with passivated protein-adsorption properties was developed using two-electron oxidation of dopamine and polyphenols. This coating has a multiscale effects: 1) macroscopically: anti-fouling properties of superhydrophilic; 2) microscopically: protein adhesion properties of active groups (quinone-, amino-, hydroxyphenyl groups and aromatic ring). Polyphenols not only enhance the ability of coating to passivate protein-adsorption, but also make the coating have polyphenol-related biological functions. Therefore, the polyphenol and passivated protein-adsorption platform together maintain the stability of the scaffold microenvironment. This, in turn, provides favorable conditions for the growth of endothelial cells on the scaffold surface. In vivo implantation of the coated stents into the abdominal aorta resulted in uniform and dense endothelial cells covering the surface of the neointima. Moreover, new endothelial cells secreted large amounts of functional endothelial nitric oxide synthase like healthy endothelial cells. These results indicate that the polyphenol superhydrophilic coating potentially resists intra-stent restenosis and promotes surface reendothelialization. Hence, polyphenol superhydrophilic coatings with passivated protein-adsorption properties constructed by two-electron-assisted oxidation are a highly effective and versatile surface-modification strategy for implantable cardiovascular devices.

Coptisine inhibits neointimal hyperplasia through attenuating Pak1/Pak2 signaling in vascular smooth muscle cells without retardation of re-endothelialization.

BACKGROUND AND AIMS: Vascular injury-induced endothelium-denudation and profound vascular smooth muscle cells (VSMCs) proliferation and dis-regulated apoptosis lead to post-angioplasty restenosis. Coptisine (CTS), an isoquinoline alkaloid, has multiple beneficial effects on the cardiovascular system. Recent studies identified it selectively inhibits VSMCs proliferation. However, its effects on neointimal hyperplasia, re-endothelialization, and the underlying mechanisms are still unclear. METHODS: Cell viability was assayed by 3-[4,5-dimethylthiazole-2-yl]-2,5-diphenyltetrazolium bromide (MTT) and cell counting kit-8 (CCK-8). Cell proliferation and apoptosis were measured by flow cytometry and immunofluorescence of Ki67 and TUNEL. Quantitative phosphoproteomics (QPP) was employed to screen CTS-responsive phosphor-sites in the key regulators of cell proliferation and apoptosis. Neointimal hyperplasia was induced by balloon injury of rat left carotid artery (LCA). Adenoviral gene transfer was conducted in both cultured cells and LCA. Re-endothelialization was evaluated by Evan's blue staining of LCA. RESULTS: 1) CTS had strong anti-proliferative and pro-apoptotic effects in cultured rat VSMCs, with the EC50 4∼10-folds lower than that in endothelial cells (ECs). 2) Rats administered with CTS, either locally to LCA's periadventitial space or orally, demonstrated a potently inhibited balloon injury-induced neointimal hyperplasia, but had no delaying effect on re-endothelialization. 3) The QPP results revealed that the phosphorylation levels of Pak1S144/S203, Pak2S20/S197, Erk1T202/Y204, Erk2T185/Y187, and BadS136 were significantly decreased in VSMCs by CTS. 4) Adenoviral expression of phosphomimetic mutants Pak1D144/D203/Pak2D20/D197 enhanced Pak1/2 activities, stimulated the downstream pErk1T202/Y204/pErk2T185/Y187/pErk3S189/pBadS136, attenuated CTS-mediated inhibition of VSMCs proliferation and promotion of apoptosis in vitro, and potentiated neointimal hyperplasia in vivo. 5) Adenoviral expression of phosphoresistant mutants Pak1A144/A203/Pak2A20/A197 inactivated Pak1/2 and totally simulated the inhibitory effects of CTS on platelet-derived growth factor (PDGF)-stimulated VSMCs proliferation and PDGF-inhibited apoptosis in vitro and neointimal hyperplasia in vivo. 6) LCA injury significantly enhanced the endogenous phosphorylation levels of all but pBadS136. CTS markedly attenuated all the enhanced levels. CONCLUSIONS: These results indicate that CTS is a promising medicine for prevention of post-angioplasty restenosis without adverse impact on re-endothelialization. CTS-directed suppression of pPak1S144/S203/pPak2S20/S197 and the subsequent effects on downstream pErk1T202/Y204/pErk2T185/Y187/pErk3S189 and pBadS136 underline its mechanisms of inhibition of VSMCs proliferation and stimulation of apoptosis. Therefore, the phosphor-sites of Pak1S144/S203/Pak2S20/S197 constitute a potential drug-screening target for fighting neointimal hyperplasia restenosis.

C1q/Tumor Necrosis Factor-Related Protein-9 Is a Novel Vasculoprotective Cytokine That Restores High Glucose-Suppressed Endothelial Progenitor Cell Functions by Activating the Endothelial Nitric Oxide Synthase.

BACKGROUND: This study investigated whether gCTRP9 (globular C1q/tumor necrosis factor-related protein-9) could restore high-glucose (HG)-suppressed endothelial progenitor cell (EPC) functions by activating the endothelial nitric oxide synthase (eNOS). METHODS AND RESULTS: EPCs were treated with HG (25 mmol/L) and gCTRP9. Migration, adhesion, and tube formation assays were performed. Adiponectin receptor 1, adiponectin receptor 2, and N-cadherin expression and AMP-activated protein kinase, protein kinase B, and eNOS phosphorylation were measured by Western blotting. eNOS activity was determined using nitrite production measurement. In vivo reendothelialization and EPC homing assays were performed using Evans blue and immunofluorescence in mice. Treatment with gCTRP9 at physiological levels enhanced migration, adhesion, and tube formation of EPCs. gCTRP9 upregulated the phosphorylation of AMP-activated protein kinase, protein kinase B, and eNOS and increased nitrite production in a concentration-dependent manner. Exposure of EPCs to HG-attenuated EPC functions induced cellular senescence and decreased eNOS activity and nitric oxide synthesis; the effects of HG were reversed by gCTRP9. Protein kinase B knockdown inhibited eNOS phosphorylation but did not affect gCTRP9-induced AMP-activated protein kinase phosphorylation. HG impaired N-cadherin expression, but treatment with gCTRP9 restored N-cadherin expression after HG stimulation. gCTRP9 restored HG-impaired EPC functions through both adiponectin receptor 1 and N-cadherin-mediated AMP-activated protein kinase /protein kinase B/eNOS signaling. Nude mice that received EPCs treated with gCTRP9 under HG medium showed a significant enhancement of the reendothelialization capacity compared with those with EPCs incubated under HG conditions. CONCLUSIONS: CTRP9 promotes EPC migration, adhesion, and tube formation and restores these functions under HG conditions through eNOS-mediated signaling mechanisms. Therefore, CTRP9 modulation could eventually be used for vascular healing after injury.

The Supreme Biodegradable Polymer DES in Acute and Chronic Coronary Syndromes: A PIONEER III Substudy.

Background: The PIONEER III trial demonstrated noninferiority of 12-month target lesion failure (TLF) with the Supreme DES (Sinomed), a thin-strut cobalt-chromium, biodegradable polymer, sirolimus-eluting stent, compared with a durable polymer, everolimus-eluting (XIENCE/PROMUS) stent (DP-EES). The relative safety and effectiveness of the Supreme DES in patients with acute coronary syndromes (ACS) and those with chronic coronary syndromes (CCS) is not known. Methods: PIONEER III was a prospective, multicenter, international, 2:1 randomized trial stratified by clinical presentation. The primary end point was TLF at 12 months (a composite of cardiac death, target vessel myocardial infarction, or ischemia-driven target lesion revascularization). Results: A total of 1628 patients were enrolled, including 41% of patients with ACS (unstable angina and non-ST-elevation myocardial infarction) randomized to Supreme DES (n = 441) versus DP-EES (n = 232) and 59% of patients with CCS randomized to Supreme DES (n = 645) versus DP-EES (n = 310). Patients with ACS were younger, fewer presented with less diabetes, hypertension, and previous revascularization, but more were current smokers. The primary end point of TLF (6.4% vs 4.4%; P = .1), major adverse cardiac events (8.5% vs 6.5%; P = .16), and stent thrombosis (0.4% vs 0.9%; P = .25) at 12 months were similar in the ACS and CCS groups. There was no difference in TLF at 12 months between Supreme DES and DP-EES among patients with ACS (6.6% vs 6.0%; P = .89) and those with CCS (4.5% vs 4.3%; P = .83); interaction P = .51 for TLF by clinical presentation. Conclusions: Compared with the DP-EES, the Supreme DES seemed safe and effective with a similar TLF at 12 months in both patients with ACS and those with CCS.

Jiedu Huoxue Prescription Regulates NF-κB/NLRP3/Caspase-1 to Promote Vascular Reendothelialization in Rat Model of Injured Thoracic Aorta / 中国实验方剂学杂志

ObjectiveTo investigate the mechanism of Jiedu Huoxue prescription in promoting the reendothelialization of injured vessels by regulating the nuclear factor （NF）-κB/NOD-like receptor protein 3 （NLRP3）/cysteine-aspartic acid protease （Caspase）-1-mediated pyroptosis. MethodA rat model of injured thoracic aorta was established by balloon injury， and 36 rats were assigned into shame surgery， model， low-， medium-， and high-dose Jiedu Huoxue prescription， and atorvastatin calcium tablet groups. The injured aortic segment was collected 28 days after surgery. Hematoxylin-eosin （HE） staining and Evans blue staining were conducted to reveal the changes of vascular structural morphology and the reendothelialization of blood vessels， respectively. The enzyme-linked immunosorbent assay （ELISA） was employed to determine the levels of tumor necrosis factor-α （TNF-α）， intercellular adhesion molecule-1 （ICAM-1）， interleukin （IL）-1β， and nitric oxide （NO） in the serum. Western blotting was employed to determine the expression of endothelial nitric oxide synthase （eNOS）， NF-κB p65， phospho-NF-κB p65 （p-NF-κB p65）， NLRP3， and Caspase-1 in the vascular tissue. ResultThe model group showed thickened endovascular membrane， proliferation and disarrangement of smooth muscle cells of the artery wall， obvious inflammatory cell infiltration， and narrowed luminal area. Jiedu Huoxue prescription and atorvastatin calcium tablets mitigated the pathological changes of the thoracic aorta in different degrees. After balloon injury， the endothelial coverage rate of the model group decreased significantly， while Jiedu Huoxue prescription and atorvastatin calcium tablets increased the reendothelialization rate （P<0.05）. Compared with the shame surgery group， the model group showed elevated levels of TNF-α， ICAM-1， and IL-1β （P<0.01） and lowered NO level （P<0.01） in the serum. In addition， the model group presented down-regulated protein level of eNOS （P<0.01） and up-regulated phosphorylation of pyroptosis-associated proteins NLPR3， Caspase-1， and NF-κB p65 in the vascular tissue （P<0.05， P<0.01）. Compared with the model group， Jiedu Huoxue prescription and atorvastatin calcium tablets lowered TNF-α， ICAM-1， and IL-1β levels （P<0.05， P<0.01） and elevated the NO level in the serum （P<0.05， P<0.01）. Moreover， the drugs up-regulated the expression of eNOS （P<0.01） and down-regulated the expression of NLRP3， Caspase-1， and NF-κB p65 （P<0.05， P<0.01） in the vascular tissue. ConclusionJiedu Huoxue prescription can promote the reendothelialization and inhibit the intimal hyperplasia of vessels after balloon injury by regulating the NF-κB/NLRP3/Caspase-1 pathway to inhibit pyroptosis and reduce endothelial inflammatory injury.

Berberine-Promoted CXCR4 Expression Accelerates Endothelial Repair Capacity of Early Endothelial Progenitor Cells in Persons with Prehypertension / 中国结合医学杂志

<p><b>OBJECTIVE</b>To evaluate whether the berberine treatment can improve endothelial repair capacity of early endothelial progenitor cells (EPCs) from prehypertensive subjects through increasing CXC chemokine receptor 4 (CXCR4) signaling.</p><p><b>METHODS</b>EPCs were isolated from prehypertensive and healthy subjects and cultured. In vivo reendothelialization capacity of EPCs from prehypertensive patients with or without in vitro berberine treatment was examined in a nude mouse model of carotid artery injury. The protein expressions of CXCR4/Janus kinase-2 (JAK-2) signaling of in vitro EPCs were detected by Western blot analysis.</p><p><b>RESULTS</b>CXCR4 signaling and alteration in migration and adhesion functions of EPCs were evaluated. Basal CXCR4 expression was significantly reduced in EPCs from prehypertensive patients compared with normal subjects (P<0.01). Also, the phosphorylation of JAK-2 of EPCs, a CXCR4 downstream signaling, was significantly decreased (P<0.01). Berberine promoted CXCR4/JAK-2 signaling expression of in vitro EPCs (P<0.01). Transplantation of EPCs pretreated with berberine markedly accelerated in vivo reendothelialization (P<0.01). The increased in vitro function and in vivo reendothelialization capacity of EPCs were inhibited by CXCR4 neutralizing antibody or pretreatment with JAK-2 inhibitor AG490, respectively (P<0.01).</p><p><b>CONCLUSION</b>Berberinemodified EPCs via up-regulation of CXCR4 signaling contributes to enhanced endothelial repair capacity in prehypertension, indicating that berberine may be used as a novel potential primary prevention means against prehypertension-related atherosclerotic cardiovascular disease.</p>

Crocin promotes mobilization of EPCs and re-endothelialization of damaged blood vessels in mice with carotid artery injury / 中国病理生理杂志

AIM:To study the effect of crocin on the mobilization of endothelial progenitor cells (EPCs) in the peripheral blood of the mice with carotid arterial injury and its mechanism.METHODS:The carotid artery injury model of the C57BL/6 mice was established by the method of wire injury.The animals were divided into sham operation group,saline-treated model group,and low dose,medium dose and high dose (10,50 and 100 μ mol· kg-1 · L-1,respectively) of crocin treatment groups.The mobilization of the EPCs in peripheral blood of the mice with carotid artery injury was detected by flow cytometry at 3 d.The changes of vascular endothelial growth factor (VEGF),stromal-derived factor-1 (SDF-1),basic fibroblast growth factor (bFGF),epidermal growth factor (EGF) and matrix metalloproteinase-9 (MMP-9) in the peripheral blood of the mice with carotid artery injury were detected by enzyme-linked immunosorbent assay at 7 d.The vascular re-endothelialization and intimal hyperplasia of the mice with carotid artery injury were detected by Evans blue and hematoxylin-eosin staining.At the same time,real-time PCR was used to detect the mRNA expression of vascular repair factor-related receptors,vascular endothelial growth factor receotor-2 (VEGFR-2),CXC chemokine receptor-4 (CXCR4),basic fibroblast growth factor receptor (bFGFR) and epidermal growth factor receptor (EGFR),in the injured segments of carotid arteries.RESULTS:Compared with sham group,the EPCs mobilization and the content of vascular repair factors VEGF,SDF-1,bFGF,EGF and MMP-9 in peripheral blood were increased in model group (P ＜0.05).The area of vascular endothelium was decreased,while the area of intimal hyperplasia and the ratio of intimal to medial membrane area were increased significantly (P ＜ 0.05).The expression levels of VEGFR-2,CXCR4,bFGFR and EGFR were also increased in the injured segments of carotid arteries (P ＜ 0.05).Compared with model group,the EPCs mobilization and the content of vascular repair factors VEGF,SDF-1,bFGF,EGF and MMP-9 in peripheral blood were significantly increased in different concentrations of crocin-treated mice with carotid artery injury (P ＜ 0.05).The area of vascular endothelium was gradually increased,while the area of intimal hyperplasia and the ratio of intimal to medial membrane area were gradually decreased (P ＜ 0.05).The expression levels of VEGFR-2,CXCR4,bFGFR and EGFR were also gradually increased in the injured segments of cartid arteries (P ＜ 0.05).CONCLUSION:Crocin promotes the mobilization of EPCs and the re-endothelialization of damaged blood vessels in the mice with carotid artery injury,thus repairing the injured vasculature.

Crocin promotes mobilization of EPCs and re-endothelialization of damaged blood vessels in mice with carotid artery injury / 中国病理生理杂志

AIM:To study the effect of crocin on the mobilization of endothelial progenitor cells (EPCs) in the peripheral blood of the mice with carotid arterial injury and its mechanism.METHODS:The carotid artery injury model of the C57BL/6 mice was established by the method of wire injury.The animals were divided into sham operation group,saline-treated model group,and low dose,medium dose and high dose (10,50 and 100 μ mol· kg-1 · L-1,respectively) of crocin treatment groups.The mobilization of the EPCs in peripheral blood of the mice with carotid artery injury was detected by flow cytometry at 3 d.The changes of vascular endothelial growth factor (VEGF),stromal-derived factor-1 (SDF-1),basic fibroblast growth factor (bFGF),epidermal growth factor (EGF) and matrix metalloproteinase-9 (MMP-9) in the peripheral blood of the mice with carotid artery injury were detected by enzyme-linked immunosorbent assay at 7 d.The vascular re-endothelialization and intimal hyperplasia of the mice with carotid artery injury were detected by Evans blue and hematoxylin-eosin staining.At the same time,real-time PCR was used to detect the mRNA expression of vascular repair factor-related receptors,vascular endothelial growth factor receotor-2 (VEGFR-2),CXC chemokine receptor-4 (CXCR4),basic fibroblast growth factor receptor (bFGFR) and epidermal growth factor receptor (EGFR),in the injured segments of carotid arteries.RESULTS:Compared with sham group,the EPCs mobilization and the content of vascular repair factors VEGF,SDF-1,bFGF,EGF and MMP-9 in peripheral blood were increased in model group (P ＜0.05).The area of vascular endothelium was decreased,while the area of intimal hyperplasia and the ratio of intimal to medial membrane area were increased significantly (P ＜ 0.05).The expression levels of VEGFR-2,CXCR4,bFGFR and EGFR were also increased in the injured segments of carotid arteries (P ＜ 0.05).Compared with model group,the EPCs mobilization and the content of vascular repair factors VEGF,SDF-1,bFGF,EGF and MMP-9 in peripheral blood were significantly increased in different concentrations of crocin-treated mice with carotid artery injury (P ＜ 0.05).The area of vascular endothelium was gradually increased,while the area of intimal hyperplasia and the ratio of intimal to medial membrane area were gradually decreased (P ＜ 0.05).The expression levels of VEGFR-2,CXCR4,bFGFR and EGFR were also gradually increased in the injured segments of cartid arteries (P ＜ 0.05).CONCLUSION:Crocin promotes the mobilization of EPCs and the re-endothelialization of damaged blood vessels in the mice with carotid artery injury,thus repairing the injured vasculature.

Carbon Monoxide Releasing Molecule Accelerates Reendothelialization after Carotid Artery Balloon Injury in Rat / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>This study was aimed to investigate the effects of carbon monoxide releasing molecule (CORM-2), a novel carbon monoxide carrier, on the reendothelialization of carotid artery in rat endothelial denudation model.</p><p><b>METHODS</b>Male rats subjected to carotid artery balloon injury were treated with CORM-2, inactive CORM-2 (iCORM-2) or dimethyl sulfoxide (DMSO). The reendothelialization capacity was evaluated by Evans Blue dye and the immunostaining with anti-CD31 antibody. The number of circulating endothelial progenitor cells (EPCs) was detected by flow cytometry. The proliferation, migration, and adhesion of human umbilical vein endothelial cells (HUVECs) were assessed by using [3H]thymidine, Boyden chamber and human fibronectin respectively. The expressions of protein were detected by using western blot analysis.</p><p><b>RESULTS</b>CORM-2 remarkably accelerated the re-endothelialization 5 d later and inhibited neointima formation 28 d later. In addition, the number of peripheral EPCs significantly increased in CORM-2-treated rats than that in iCORM-2 or DMSO-treated rats after 5 d later. In vitro experiments, CORM-2 significantly enhanced the proliferation, migration and adhesion of HUVECs. The levels of Akt, eNOS phosphorylation, and NO generation in HUVECs were also much higher in CORM-2 treated group. Blocking of PI3K/Akt/eNOS signaling pathway markedly suppressed the enhanced migration and adhesion of HUVECs induced by CORM-2.</p><p><b>CONCLUSION</b>CORM-2 could promote endothelial repair, and inhibit neointima formation after carotid artery balloon injury, which might be associated with the function changes of HUVECs regulated by PI3K/Akt/eNOS pathway.</p>

Effects of shear stress on late endothelial progenitor cell functions / 医用生物力学

Objective To investigate the effects of shear stress on late endothelial progenitor cells (EPCs) functions in vitro and in vivo. Methods Density gradient centrifugation-isolated rat bone marrow mononuclear cells were cultured in EGM-2MV and induced into EPCs. The 3rd~4th generation of EPCs, namely late EPCs, were treated with shear stress (1.2 Pa). Then cell biological functions, such as proliferation, adhesion, migration and ability of tube formation, were assayed with EdU incorporation assay, adhesion testing, Boyden chamber assay and Matrigel, respectively. The gene expression of VEFG was analyzed by real time RT-PCR. The apoptosis and aging situation of late EPCs were assayed by FACS and senescence-associated β-galactosidase (SA-β-gal) staining. The reendothelialization capacity of late EPCs treated by shear stress was evaluated by establishing models of freshly balloon-injured carotid arteries of rats and cell transplantation in situ. Results Shear stress could increase proliferation, adhesion, migration and tube formation of late EPCs (P＜0.05), upregulate the gene expression of VEGF, inhibit EPC apoptosis and delayed EPC aging (P＜0.05). Transplantation of late EPCs treated by shear stress facilitated in vivo reendothelialization in the injured arterial segment and inhibited neointima formation. Conclusions Shear stress within the physiological range can improve the functions of late EPCs and enhance their therapeutic ability of repairing vascular endothelial injury, which provides experimental basis for the clinic application of EPCs and shear stress-mediated cell therapy.

Comparison of bare metal stent and paclitaxel-eluting stent using a novel rat aorta stent model

The purpose of our study was to create a novel rat aorta stent implantation model. Stainless steel bare metal stents (BMS) or paclitaxel-eluting stents (PES) were implanted in male Sprague-Dawley rats (BW 400 +/- 20 g). Two and four weeks after stent implantation, the aorta were collected, fixed with 2% glutaraldehyde, and cut into two segments. One segment was used for scanning electron microscopy analysis to evaluate re-endothelialization, and the other segment was used to calculate the neointimal area. At 2 weeks after stenting, the appearance of neointimal hyperplasia was less in the PES group than in the BMS group. At 4 weeks after stenting, no significant difference in neointimal hyperplasia was observed between two groups. On the other hand, the PES group showed more thrombus formation and less re-endothelialization compared to the BMS group. This study demonstrated the ability of a novel rat model of aorta stenting via a common carotid artery to measure the efficacy and safety of commercially available drug-eluting stents.

Stromal Cell-Derived factor-1 ? Involves in Mobilization of Endothelial Progenitor Cells and Reendothelialization After Artery Injury / 中华高血压杂志

Background Stromal cell-derived factor-1_?(SDF-1_?)has been demonstrated to be essential for stern cell mobilization/homing.Recent evidence indicates that SDF-1_? has been expressed in injured carotid arter- ies.Besides,high SDF-1_? plasma levels are clinically associated with stable coronary artery disease.Objective To investigate whether SDF 1 involves in mobilization of endothelial progenitor cells(EPC)and reendothelialization after vascular injury.Methods SDF-1_? was detected by RT-PCR and Western blot in carotid arteries of mice at different time points after wire-induced injury.SDF-1_? determination in peripheral blood samples and BM was per- formed by SDF-1_? enzyme-linked immunosorbent assay(ELISA)kit.EPC in peripheral blood collected at different time points after vascular injury were quantified by flow cytornetry.In subgroup,blocking SDF-1 rnonoclonal anti- body was injected,peripheral blood EPC were quantified after vascular injury and reendothelialization of injured ar- teries was determined 14 days later.Results Expression of SDF-1_? was evident at day 1,and peaked at day 3 after arterial injury.A rise in plasmatic concentration of SDF-1_? and a significant reduction of SDF-1_? in bone marrow concentration was noticed at all time points following injury.The amount of circulating EPC was increased shortly after induction of vascular injury and persisted up to 7 days(P

Effects of transplantation of peripheral blood mesenchymal stem cells with hypoxia preconditioning on postangioplasty restenosis in rabbits / 中国病理生理杂志

AIM: To investigate the function and the mechanism of transplanting bone marrow derived peripheral blood mesenchymal stem cells(PBMSCs) on restenosis after carotid balloon angioplasty in the model of carotid atherosclerosis rabbits,and to determine if the functions of PBMSCs are enhanced after hypoxia preconditioning.METHODS: Bone marrow cells were mobilized by granulocyte colony-stimulating factor(G-CSF),and PBMSCs were collected through density gradient centrifugation and adherent culture,labeled with enhancement type green fluorescent protein(EGFP) genes.All animals with carotid atherosclerosis stenosis were randomly divided into three groups: hypoxia preconditioning group(n=24,received intravenous transplantation of PBMSCs with hypoxia preconditioning),non-hypoxia preconditioning group(n=24,received normal culture of PBMSCs) and control group(n=24,only received equal-volume of culture medium).Vascular endothelial growth factor(VEGF) was determined by enzyme linked immunosorbent assay(ELISA) at 7 d,14 d and 28 d post-angioplasty,respectively.The vessel morphology,the homing of MSCs and the reendothelialization were analyzed with Weigert staining and immunohistochemistry.RESULTS: Compared to control group,the level of VEGF significantly increased in both hypoxia preconditioning group and non-hypoxia preconditioning group at all time points(P