OBJECTIVE: To explore the mechanism of antibiotic delivery system targeting bacterial biofilm with linezolid (LZD) based on ε-poly- L-lysine (ε-PLL) and cyclodextrin (CD) (ε-PLL-CD-LZD), aiming to enhance antibiotic bioavailability, effectively penetrate and disrupt biofilm structures, and thereby improve the treatment of bone and joint infections. METHODS: ε-PLL-CD-LZD was synthesized via chemical methods. The grafting rate of CD was characterized using nuclear magnetic resonance. In vitro biocompatibility was evaluated through live/dead cell staining after co-culturing with mouse embryonic osteoblast precursor cells (MC3T3-E1), human umbilical vein endothelial cells, and mouse embryonic fibroblast cells (3T3-L1). The biofilm-enrichment capacity of ε-PLL-CD-LZD was assessed using Staphylococcus aureus biofilms through enrichment studies. Its biofilm eradication efficacy was investigated via minimum inhibitory concentration (MIC) determination, scanning electron microscopy, and live/dead bacterial staining. A bone and joint infection model in male Sprague-Dawley rats was established to validate the antibacterial effects of ε-PLL-CD-LZD. RESULTS: In ε-PLL-CD-LZD, the average grafting rate of CD reached 9.88%. The cell viability exceeded 90% after co-culturing with three types cells. The strong biofilm enrichment capability was observed with a MIC of 2 mg/L. Scanning electron microscopy observations revealed the effective disruption of biofilm structure, indicating potent biofilm eradication capacity. In vivo rat experiments demonstrated that ε-PLL-CD-LZD significantly reduced bacterial load and infection positivity rate at the lesion site ( P<0.05). CONCLUSION The ε-PLL-CD antibiotic delivery system provides a treatment strategy for bone and joint infections with high clinical translational significance. By effectively enhancing antibiotic bioavailability, penetrating, and disrupting biofilms, it demonstrated significant anti-infection effects in animal models.
Biofilms/drug effects* ; Animals ; Anti-Bacterial Agents/pharmacology* ; Polylysine/chemistry* ; Cyclodextrins/administration & dosage* ; Humans ; Linezolid/pharmacology* ; Staphylococcus aureus/physiology* ; Rats, Sprague-Dawley ; Mice ; Rats ; Male ; Drug Delivery Systems ; Staphylococcal Infections/drug therapy* ; Microbial Sensitivity Tests ; Human Umbilical Vein Endothelial Cells ; Osteoblasts/cytology*

In most types of erectile dysfunction, particularly in advanced stages, typical pathological features observed are reduced parenchymal cells coupled with increased tissue fibrosis. However, the current treatment methods have shown limited success in reversing these pathologic changes. Recent research has revealed that changes in autophagy levels, along with alterations in apoptosis and fibrosis-related proteins, are linked to the progression of erectile dysfunction, suggesting a significant association. Autophagy, known to significantly affect cell fate and tissue fibrosis, is currently being explored as a potential treatment modality for erectile dysfunction. However, these present studies are still in their nascent stage, and there are limited experimental data available. This review analyzes erectile dysfunction from a pathological perspective. It provides an in-depth overview of how autophagy is involved in the apoptotic processes of smooth muscle and endothelial cells and its role in the fibrotic processes occurring in the cavernosum. This study aimed to develop a theoretical framework for the potential effectiveness of autophagy in preventing and treating erectile dysfunction, thus encouraging further investigation among researchers in this area.
Male ; Humans ; Autophagy/physiology* ; Apoptosis/physiology* ; Erectile Dysfunction/physiopathology* ; Fibrosis ; Penis/pathology* ; Animals ; Endothelial Cells/pathology* ; Myocytes, Smooth Muscle/pathology*

OBJECTIVES: To explore the role of berberine (BBR) in ameliorating coronary endothelial cell injury in Kawasaki disease (KD) by regulating the complement and coagulation cascade. METHODS: Human coronary artery endothelial cells (HCAEC) were divided into a healthy control group, a KD group, and a BBR treatment group (n=3 for each group). The healthy control group and KD group were supplemented with 15% serum from healthy children and KD patients, respectively, while the BBR treatment group received 15% serum from KD patients followed by the addition of 20 mmol/L BBR. Differential protein expression was analyzed and identified using isobaric tags for relative and absolute quantitation technology and liquid chromatography-tandem mass spectrometry, followed by GO functional enrichment analysis and KEGG signaling pathway enrichment analysis of the differential proteins. Western blot was used to detect differential protein expression. RESULTS: A total of 518 differential proteins were identified between the KD group and the healthy control group (300 upregulated proteins and 218 downregulated proteins). A total of 422 differential proteins were identified between the BBR treatment group and the KD group (221 upregulated proteins and 201 downregulated proteins). Bioinformatics analysis showed that compared to the healthy control group, the differential proteins in the KD group were enriched in the complement and coagulation cascade and ribosome biogenesis in eukaryotes. Compared to the KD group, the differential proteins in the BBR treatment group were also enriched in the complement and coagulation cascade and ribosome biogenesis in eukaryotes. Western blot results indicated that compared to the healthy control group, the expression of complement C1q subcomponent subunit C (C1QC), kininogen-1 (KNG1), complement C1s subcomponent (C1S), and C4b-binding protein alpha chain (C4BPA) was increased in the KD group (P<0.05). Compared to the KD group, the expression of KNG1, C1S, C1QC, and C4BPA was decreased in the BBR treatment group (P<0.05). CONCLUSIONS The complement and coagulation cascade may be involved in the regulation of BBR treatment for coronary injury in KD, and C1QC, KNG1, C1S, and C4BPA may serve as biomarkers for this treatment.
Mucocutaneous Lymph Node Syndrome/blood* ; Humans ; Endothelial Cells/pathology* ; Complement System Proteins/physiology* ; Coronary Vessels/drug effects* ; Male ; Blood Coagulation/drug effects* ; Berberine/therapeutic use* ; Female ; Child, Preschool ; Infant

OBJECTIVES: To investigate the role and mechanism of fibroblast growth factor (FGF) 19 in inflammation-induced injury of vascular endothelial cells caused by high glucose (HG). METHODS: Human umbilical vein endothelial cells (HUVECs) were randomly divided into four groups: control, HG, FGF19, and HG+FGF19 (n=3 each). The effect of different concentrations of glucose and/or FGF19 on HUVEC viability was assessed using the CCK8 assay. Flow cytometry was utilized to examine the impact of FGF19 on HUVEC apoptosis. Levels of interleukin-6 (IL-6), inducible nitric oxide synthase (iNOS), total superoxide dismutase (T-SOD), and malondialdehyde (MDA) were measured by ELISA. Real-time quantitative PCR and Western blotting were used to determine the mRNA and protein expression levels of vascular endothelial growth factor (VEGF), nuclear factor erythroid 2 related factor 2 (Nrf2), and heme oxygenase-1 (HO-1). Cells were further divided into control, siRNA-Nrf2 (siNrf2), HG, HG+FGF19, HG+FGF19+negative control, and HG+FGF19+siNrf2 groups (n=3 each) to observe the effect of FGF19 on oxidative stress injury in HUVECs induced by high glucose after silencing the Nrf2 gene. RESULTS: Compared to the control group, the HG group exhibited increased apoptosis rate, increased IL-6, iNOS and MDA levels, and increased VEGF mRNA and protein expression, along with decreased T-SOD activity and decreased mRNA and protein expression of Nrf2 and HO-1 (P<0.05). Compared to the HG group, the HG+FGF19 group showed reduced apoptosis rate, decreased IL-6, iNOS and MDA levels, and decreased VEGF mRNA and protein expression, with increased T-SOD activity and increased Nrf2 and HO-1 mRNA and protein expression (P<0.05). Compared to the HG+FGF19+negative control group, the HG+FGF19+siNrf2 group had decreased T-SOD activity and increased MDA levels (P<0.05). CONCLUSIONS FGF19 can alleviate inflammation-induced injury in vascular endothelial cells caused by HG, potentially through the Nrf2/HO-1 signaling pathway.
Humans ; NF-E2-Related Factor 2/genetics* ; Signal Transduction ; Human Umbilical Vein Endothelial Cells/drug effects* ; Fibroblast Growth Factors/pharmacology* ; Heme Oxygenase-1/physiology* ; Apoptosis/drug effects* ; Glucose ; Inflammation ; Interleukin-6/analysis* ; Vascular Endothelial Growth Factor A/genetics* ; Nitric Oxide Synthase Type II/analysis* ; Cells, Cultured

Meniscus injuries are widespread and the available treatments do not offer enough healing potential. Here, we provide critical support for using pigs as a biological model for meniscal degeneration and the development of cutting-edge therapies in orthopedics. We present a single-cell transcriptome atlas of the meniscus, consisting of cell clusters corresponding to four major cell types: chondrocytes, endothelial cells, smooth muscle cells, and immune cells. Five distinct chondrocyte subclusters (CH0‒CH4) were annotated, of which only one was widespread in both the red and white zones, indicating a major difference in the cellular makeup of the zones. Subclusters distinct to the white zone appear responsible for cartilage-specific matrix deposition and protection against adverse microenvironmental factors, while those in the red zone exhibit characteristics of mesenchymal stem cells and are more likely to proliferate and migrate. Additionally, they induce remodeling actions in other chondrocyte subclusters and promote the proliferation and maturation of endothelial cells, inducing healing and vascularization processes. Considering that they have substantial remodeling capabilities, these subclusters should be of great interest for tissue engineering studies. We also show that the cellular makeup of the pig meniscus is comparable to that of humans, which supports the use of pigs as a model in orthopedic therapy development.
Animals ; Swine ; Chondrocytes/physiology* ; Single-Cell Analysis ; Meniscus/blood supply* ; Endothelial Cells/cytology* ; Transcriptome ; Mesenchymal Stem Cells/cytology* ; Neovascularization, Physiologic ; Cell Proliferation

The angiogenic response is essential for the repair of ischemic brain tissue. Integrin α6 (Itga6) expression has been shown to increase under hypoxic conditions and is expressed exclusively in vascular structures; however, its role in post-ischemic angiogenesis remains poorly understood. In this study, we demonstrate that mice with endothelial cell-specific knockout of Itga6 exhibit reduced neovascularization, reduced pericyte coverage on microvessels, and accelerated breakdown of microvascular integrity in the peri-infarct area. In vitro, endothelial cells with ITGA6 knockdown display reduced proliferation, migration, and tube-formation. Mechanistically, we demonstrated that ITGA6 regulates post-stroke angiogenesis through the PI3K/Akt-eNOS-VEGFA axis. Importantly, the specific overexpression of Itga6 in endothelial cells significantly enhanced neovascularization and enhanced the integrity of microvessels, leading to improved functional recovery. Our results suggest that endothelial cell Itga6 plays a crucial role in key steps of post-stroke angiogenesis, and may represent a promising therapeutic target for promoting recovery after stroke.
Animals ; Nitric Oxide Synthase Type III/metabolism* ; Mice ; Proto-Oncogene Proteins c-akt/metabolism* ; Integrin alpha6/genetics* ; Endothelial Cells/metabolism* ; Phosphatidylinositol 3-Kinases/metabolism* ; Stroke/pathology* ; Vascular Remodeling/physiology* ; Vascular Endothelial Growth Factor A/metabolism* ; Mice, Knockout ; Signal Transduction/physiology* ; Mice, Inbred C57BL ; Male ; Neovascularization, Physiologic/physiology*

Hypoxia-inducible factor (HIF-1α), a core transcription factor responding to changes in cellular oxygen levels, is closely associated with a wide range of physiological and pathological conditions. However, its differential impacts on vascular cell types and molecular programs modulating human vascular homeostasis and regeneration remain largely elusive. Here, we applied CRISPR/Cas9-mediated gene editing of human embryonic stem cells and directed differentiation to generate HIF-1α-deficient human vascular cells including vascular endothelial cells, vascular smooth muscle cells, and mesenchymal stem cells (MSCs), as a platform for discovering cell type-specific hypoxia-induced response mechanisms. Through comparative molecular profiling across cell types under normoxic and hypoxic conditions, we provide insight into the indispensable role of HIF-1α in the promotion of ischemic vascular regeneration. We found human MSCs to be the vascular cell type most susceptible to HIF-1α deficiency, and that transcriptional inactivation of ANKZF1, an effector of HIF-1α, impaired pro-angiogenic processes. Altogether, our findings deepen the understanding of HIF-1α in human angiogenesis and support further explorations of novel therapeutic strategies of vascular regeneration against ischemic damage.
Humans ; Vascular Endothelial Growth Factor A/metabolism* ; Endothelial Cells/metabolism* ; Transcription Factors/metabolism* ; Gene Expression Regulation ; Hypoxia/metabolism* ; Cell Hypoxia/physiology*

A complex pathophysiological mechanism is involved in brain injury following cerebral infarction. The neurovascular unit (NVU) is a complex multi-cellular structure consisting of neurons, endothelial cells, pericyte, astrocyte, microglia and extracellular matrix, etc. The dyshomeostasis of NVU directly participates in the regulation of inflammatory immune process. The components of NVU promote inflammatory overreaction and synergize with the overactivation of autonomic nervous system to initiate stroke-induced immunodepression (SIID). SIID can alleviate the damage caused by inflammation, however, it also makes stroke patients more susceptible to infection, leading to systemic damage. This article reviews the mechanism of SIID and the roles of NVU in SIID, to provide a perspective for reperfusion, prognosis and immunomodulatory therapy of cerebral infarction.
Humans ; Endothelial Cells ; Stroke ; Neurons/physiology* ; Immunosuppression Therapy/adverse effects* ; Cerebral Infarction

OBJECTIVES: To explore the effect of hydrogel loaded with exosomes from Wharton's Jelly-derived mesenchymal stem cell (WJMSC) on wound healing. METHODS: Exosomes were extracted from WJMSC, and the morphology and size of WJMSC-derived exosomes (WEX) were analyzed by transmission electron microscopy and nanoparticle size analyzer, respectively. The surface markers CD9, CD81, and Calnexin of WEX were detected by Western blotting. Exosome-loaded alginate hydrogel (WEX-gel) was prepared; its morphology was studied by scanning electron microscope, and its rheological behavior was examined by a rheometer. The in vitro drug release performance of WEX-gel was investigated by BCA method. RAW264.7 cells were treated with alginate hydrogel, WEX and WEX-gel, respectively; and the expression of CD86 and CD206 in macrophages was detected by flow cytometry. A full-thickness skin wound model was established in mice; the model mice were randomly divided into blank control group, WEX control group and WEX-gel group, and PBS, WEX and WEX-gel were applied to the wound area of mice, respectively. On day 3, the skin tissue of mice was excised, and the antibacterial effect of WEX hydrogel was evaluated by plate counting. On day 15, the mice were euthanized and the percentage of residual wounds was calculated. The histological changes of the skin wound were observed after hematoxylin and eosin (HE) and Masson stainings. The expression of CD86, CD206, CD31 and vascular endothelial growth factor (VEGF) in the skin wound tissue was detected by immunohistochemistry. RESULTS: Exosomes were successfully extracted from WJMSC. WEX-gel presented a regular three-dimensional network structure, good rheology and controlled drug release performance. WEX-gel promoted the polarization of RAW264.7 cells from the M1 phenotype to M2 phenotype in vitro. The residual wound percentage in blank control group, WEX control group and WEX-gel group were (27.5±3.4)%, (15.3±1.2)% and (7.6±1.1)%, respectively (P<0.05). The antibacterial property of WEX-gel is better than that of WEX (P<0.05). The dermis thickness, the number of new hair follicles, and the rate of collagen deposition in the WEX-gel group were significantly higher than those in the other two groups (all P<0.05). The expression of CD206, CD31 and VEGF in skin wound tissue was higher and the expression of CD86 was lower in WEX-gel group than those in other two groups (all P<0.05). CONCLUSIONS WEX-gel can significantly promote wound healing in mice by regulating the polarization of macrophages.
Mice ; Animals ; Vascular Endothelial Growth Factor A ; Wharton Jelly ; Exosomes ; Hydrogels ; Wound Healing/physiology* ; Mesenchymal Stem Cells ; Anti-Bacterial Agents ; Alginates

Mesenchymal stem cell (MSC)-derived exosomes (Exos) were reported to a prospective candidate in accelerating diabetic wound healing due to their pro-angiogenic effect. MSCs pretreated with chemistry or biology factors were reported to advance the biological activities of MSC-derived exosomes. Hence, this study was designed to explore whether exosomes derived from human umbilical cord MSCs (hucMSCs) preconditioned with Nocardia rubra cell wall skeleton (Nr-CWS) exhibited superior proangiogenic effect on diabetic wound repair and its underlying molecular mechanisms. The results showed that Nr-CWS-Exos facilitated the proliferation, migration and tube formation of endothelial cells in vitro. In vivo, Nr-CWS-Exos exerted great effect on advancing wound healing by facilitating the angiogenesis of wound tissues compared with Exos. Furthermore, the expression of circIARS1 increased after HUVECs were treated with Nr-CWS-Exos. CircIARS1 promoted the pro-angiogenic effects of Nr-CWS-Exos on endothelial cellsvia the miR-4782-5p/VEGFA axis. Taken together, those data reveal that exosomes derived from Nr-CWS-pretreated MSCs might serve as an underlying strategy for diabetic wound treatment through advancing the biological function of endothelial cells via the circIARS1/miR-4782-5p/VEGFA axis.
Humans ; Endothelial Cells/metabolism* ; Exosomes/metabolism* ; Cell Wall Skeleton/metabolism* ; Neovascularization, Physiologic ; Wound Healing/physiology* ; MicroRNAs/metabolism* ; Diabetes Mellitus ; Vascular Endothelial Growth Factor A/metabolism*