Molecular angiogenic events of a two-herb wound healing formula involving MAPK and Akt signaling pathways in human vascular endothelial cells

<p>The process of wound healing consists of the following phases: inflammation, proliferation, remodeling. Non-steroidal antiinflammatory drugs may be important in this process, especially in a stage called angiogenesis. For this reason, it was decided to investigate the effect of selective COX-2 (cyclooxygenase 2) inhibitor (NS-398) on the proliferation of endothelial cells and their ability to secrete bFGF (fibroblast growth factor) for vascular endothelial cells (HMEC-1). For determination of the secretion of bFGF in a cell line HMEC-1 immunosorbent ELISA assays were used. In turn, the cell proliferation assay was performed using the MTT method. Using MTT method, it was found that NS-398 at 10 μM did not affect cell viability. Whereas selective COX-2 inhibitor at 100 μM decreased cell viability in a statistically significant manner and inhibited the proliferative effect of 100 μg/mL LPS at concentrations of 10 and 100 μM. In the further step, application of NS-398 (10 and 100 μM) with LPS (100 μg/mL; inflammatory environment) reduced the secretion of bFGF in a statistically significant manner. The investigations showed that NS-398 has an antiangiogenic effect which is based on reducing the proliferation of vascular endothelial cells and inhibiting the secretion of bFGF- factor responsible for angiogenesis during wound healing.</p>

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Rat epidermal stem cells promote the angiogenesis of full-thickness wound

10.21203/rs.3.rs-28767/v1 ◽

2020 ◽

Author(s):

Shaobin Huang ◽

Zhicheng Hu ◽

Peng Wang ◽

Yi zhang ◽

Xiaoling Cao ◽

...

Keyword(s):

Stem Cells ◽

Wound Healing ◽

Endothelial Cells ◽

Cell Suspension ◽

Vascular Endothelial Cells ◽

Control Group ◽

Vascular Endothelial ◽

Epidermal Stem Cells ◽

Full Thickness ◽

Full Thickness Wound

Abstract Background: Full-thickness wounds are a serious problem which badly affects patients’ life quality and also become the difficult problem for clinicians. Stem cells have great prospects in the treatment of wounds. Our previous experiments proved that autologous basal cell suspension can promote wound healing, and there are epidermal stem cells (ESCs) in basal cell suspension. We then conducted experiments to explore the effect of ESCs on full-thickness wound. Methods: In our study, the rat ESCs were isolated and expanded, and transfected with lentivirus to stably express EGFP. Experimental rats were randomly divided into 2 groups, in the ESCs group, the rat ESCs were sprayed on the Full-thickness wounds of the rats, while in control group, sprayed the PBS on the wound. Wound healing and neovascularization were then evaluated. Colonization, division and differentiation of ESCs on the wound were discovered by immunofluorescence.Results: The result suggested that rat ESCs can colonize, divide and proliferate in the wound. What’s more, the rat ESCs around blood vessels can differentiate into vascular endothelial cells and form a lumen-like structure. Compared with the control group, spraying the rat ESCs on the wound bed can promote angiogenesis and accelerate wound healing. Conclusions: Our study proved that rat ESCs were safe and effective for treating full-thickness wounds, and under certain conditions, ESCs can differentiate into vascular endothelial cells to promote angiogenesis and wound healing.

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Effects of IL-1β and TNF-α on the Expression of P311 in Vascular Endothelial Cells and Wound Healing in Mice

Frontiers in Physiology ◽

10.3389/fphys.2020.545008 ◽

2020 ◽

Vol 11 ◽

Author(s):

Daijun Zhou ◽

Tengfei Liu ◽

Song Wang ◽

Weifeng He ◽

Wei Qian ◽

...

Keyword(s):

Wound Healing ◽

Endothelial Cells ◽

Cell Migration ◽

Protein Expression ◽

Vascular Endothelial Cells ◽

Control Group ◽

Vascular Endothelial ◽

Expression Levels ◽

Relative Expression ◽

Tnf Α

ObjectiveThis study aimed to define the role of interleukine-1β (IL-1β) and tumor necrosis factor-α (TNF-α) in the expression of P311 in vascular endothelial cells (VECs) and in wound healing.MethodsDAPI staining, a CCK-8 assay, cell migration assay, and an angiogenesis assay were used to assess the effects exerted by TNF-α and IL-1β at various concentrations on morphology, proliferation, migration, and angiogenesis of VECs. Western blot (WB) and reverse transcription-polymerase chain reaction (RT-PCR) models were employed to observe the effects exerted by proteins related to the nuclear factor-kappa B (NF-κB) signaling pathway and P311 mRNA expression. Bioinformatics analysis was performed on the binding sites of P311 and NF-κB. Finally, to investigate the effects of IL-1β and TNF-α on wound healing and the length of new epithelium in mice, we established a full-thickness wound defect model in mice. Immunohistochemistry was used to measure changes in P311, proliferating cell nuclear antigen (PCNA), vascular endothelial growth factor (VEGF), CD31 (platelet endothelial cell adhesion molecule-1, PECAM-1/CD31), as well as other related proteins.ResultsWhen levels of TNF-α and IL-1β were both 20 ng/ml, their effects on cell proliferation, cytoskeleton protein expression, cell migration, and angiogenesis were the greatest (P < 0.05). IL-1β and TNF-α at moderate concentrations effectively promoted P311 mRNA and p-NF-κB protein expression (P < 0.05), while p-NF-K b protein expression was decreased (P < 0.05). Luciferase assays showed that P311 expression was also relatively greater when stimulated at moderate concentrations (P < 0.05), while relative expression was significantly lower when the p-NF-K b inhibitor CAPE was added (P < 0.05). On 7-day wound healing rate comparison, the control, IL-1β, IL-1βab, TNF-α, and TNF-αab groups were 18, 37, 35, 39, and 36%, respectively, while control group + P311 siRNA was 31% (P < 0.05). New epithelial length, granulation tissue thickness, and number of blood vessels trends were also the same. In the control group, P311 showed lower relative expression levels than the others (P < 0.05). P311 relative expression levels trended as follows: control group > IL-1βab > IL-1β > TNF-αab > TNF-α (P < 0.05).ConclusionWhen IL-1β and TNF-α concentrations are moderate, they effectively promote the proliferation, expression, migration, and angiogenesis of VECs, possibly by promoting the expression of the NF-K b pathway and thereby promoting the expression of P311. In vitro experiments on mice suggest that P311 effectively promotes wound healing, and its mechanism may be closely related to PCNA, CD31, and VEGF.

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Activated Platelets Induce Tissue Factor Expression on Human Umbilical Vein Endothelial Cells by Ligation of CD40

Thrombosis and Haemostasis ◽

10.1055/s-0037-1615402 ◽

1998 ◽

Vol 80 (12) ◽

pp. 1008-1041 ◽

Cited By ~ 137

Author(s):

Matthias Kalbas ◽

Antje Willuweit ◽

Volker Henn ◽

Richard Kroczek ◽

Gert Müller-Berghaus ◽

...

Keyword(s):

Wound Healing ◽

Endothelial Cells ◽

Tissue Factor ◽

Vascular Endothelial Cells ◽

Tissue Injury ◽

Pyrrolidine Dithiocarbamate ◽

Type I ◽

Dependent Manner ◽

Vascular Endothelial ◽

Activated Platelets

SummaryCD40 is a type I member of the tumour necrosis factor (TNF) receptor superfamily of proteins, and is present on a wide variety of cells including vascular endothelial cells. Ligation of this receptor on endothelial cells is known to increase expression of inflammatory adhesion molecules. We have recently demonstrated that platelets express the ligand of CD40 (CD154) within seconds of exposure to agonist, and interact with endothelial cells to participate directly in the induction of an inflammatory response. Here we show that activated platelets induce tissue factor (TF) expression on endothelial cells in a CD40/CD154-dependent manner, and that the magnitude of this response can equal that induced by TNFα. Moreover, CD40 ligation on endothelial cells downregulates the expression of thrombomodulin. We also show that CD40-mediated TF expression is less sensitive to inhibition with the oxidative radical scavenger pyrrolidine dithiocarbamate than is that mediated by TNFα, indicating that CD40 has a distinct signalling pathway. Tissue factor is a cell membrane protein which functions as the main trigger of the extrinsic pathway of blood coagulation, and its expression on endothelial cells is implicated in wound healing and angiogenesis. Since platelets are among the first cells involved in haemostasis following tissue injury, our data showing that ligation of CD40 by CD154 induces a procoagulant phenotype on vascular endothelial cells suggests that platelets may play an important role in the induction of wound healing.

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