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Volume 39 Issue 1
Jan.  2023
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Article Navigation >  CHINESE JOURNAL OF BURNS AND WOUNDS  > 2023  >  39(1): 35-44
CAI Jing-ying, ZHOU Man, DAI Gui- feng, et al. Retrospective investigation of inpatients with chronic wounds[J]. Chin j Burns, 2011, 27(2): 135-138. Doi: 10.3760/cma.j.issn.1009-2587.2011.02.017
Citation: Ou ZL,Wang J,Shi R,et al.Influence of reactive oxygen species responsive self-assembled nanomicelle loaded with pyroptosis inhibitor on full-thickness skin defects in diabetic rats[J].Chin J Burns Wounds,2023,39(1):35-44.DOI: 10.3760/cma.j.cn501225-20221109-00483.
shu

Influence of reactive oxygen species responsive self-assembled nanomicelle loaded with pyroptosis inhibitor on full-thickness skin defects in diabetic rats

doi: 10.3760/cma.j.cn501225-20221109-00483
  • 1.

    Department of Nephrology, the First Affiliated Hospital of Wenzhou Medical University, Wenzhou 325000, China

  • 2.

    State Key Laboratory of Trauma, Burns and Combined Injury, Institute of Burn Research, the First Affiliated Hospital of Army Medical University (the Third Military Medical University), Chongqing Key Laboratory for Disease Proteomics, Chongqing 400038, China

Funds:

International (Regional) Cooperation and Exchange Program of National Natural Science Foundation of China 81920108022

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  • Corresponding author: Luo Gaoxing, Email: logxw@hotmail.com
  • Received Date: 2022-11-09
    Abstract
  •   Objective  To investigate the influence of reactive oxygen species (ROS) responsive self-assembled nanomicelle loaded with pyroptosis inhibitor on full-thickness skin defects in diabetic rats.  Methods  Experimental research methods were employed. A nucleotide-binding oligomerization domain (NOD) 1/2 inhibitor (NOD-IN-1) was encapsulated with nanomicelle polyethylene glycol-block-polypropylene sulfide (PEG-b-PPS), and the resulting product was called PEPS@NOD-IN-1. The morphology and hydration particle size of PEG-b-PPS and PEPS@NOD-IN-1 were observed by transmission electron microscope and particle size analyzer, respectively, and the encapsulation rate and drug loading rate of PEPS@NOD-IN-1 to NOD-IN-1 and the cumulative release rate of NOD-IN-1 by PEPS@NOD-IN-1 in phosphate buffer solution (PBS) alone and hydrogen peroxide-containing PBS within 40 h were measured and calculated by microplate reader, and the sample number was 3. Twenty-four male Sprague-Dawley rats aged 6-7 weeks were injected with streptozotocin to induce type 1 diabetes mellitus. Six full-thickness skin defect wounds were made on the back of each rat. The injured rats were divided into PBS group, NOD-IN-1 group, PEG-b-PPS group, and PEPS@NOD-IN-1 group with corresponding treatment according to the random number table, with 6 rats in each group. The wound healing was observed on post injury day (PID) 3, 7, and 12, and the wound healing rate was calculated. The ROS levels in wound tissue were detected by immunofluorescence method on PID 3. On PID 7, the granulation tissue thickness in wound was assessed by hematoxylin-eosin staining, the mRNA expressions of NOD1 and NOD2 were detected by real-time fluorescence quantitative reverse transcription polymerase chain reaction, and the protein expressions of NOD1, NOD2, and GSDMD-N terminals were detected by Western blotting. Six wounds from different rats in each group were taken for detection of the above indicators. Wound tissue (3 samples per group) was taken from rats in PBS group and PEPS@NOD-IN-1 group on PID 7, and transcriptome sequencing was performed using high-throughput sequencing technology platform. Differentially expressed genes (DEGs) significantly down-regulated in PEPS@NOD-IN-1 group as compared with PBS group were screened, and the enrichment analysis of Kyoto Encyclopedia of Genes and Genomes (KEGG) was performed. The DEG heatmap of the NOD-like receptor pathway, a pyroptosis-related pathway, was made. Protein-protein interaction (PPI) analysis of DEGs in heatmap was performed through the STRING database to screen key genes of PEPS@NOD-IN-1 regulating the NOD-like receptor pathway. Data were statistically analyzed with analysis of variance for repeated measurement, one-way analysis of variance, and Tukey test.  Results  PEG-b-PPS and PEPS@NOD-IN-1 were in spherical structures of uniform size, with hydration particle sizes of (134.2±3.3) and (143.1±2.3) nm, respectively. The encapsulation rate of PEPS@NOD-IN-1 to NOD-IN-1 was (60±5)%, and the drug loading rate was (15±3)%. The release of NOD-IN-1 from PEPS@NOD-IN-1 in PBS alone was slow, and the cumulative release rate at 40 h was only (12.4±2.3)%. The release of NOD-IN-1 from PEPS@NOD-IN-1 in hydrogen peroxide-containing PBS within 10 h was very rapid, and the cumulative release rate at 10 h reached (90.1±3.6)%. On PID 3 and 7, the wounds of rats in the four groups were gradually healed, and the healing in PEPS@NOD-IN-1 group was better than that in the other three groups. On PID 12, the wound scab area in PBS group was large, the wound epithelialization in NOD-IN-1 group and PEG-b-PPS group was obvious, and the wound in PEPS@NOD-IN-1 group was close to complete epithelialization. Compared with those in PBS group, NOD-IN-1 group, and PEG-b-PPS group, the wound healing rates on PID 7 and 12 in PEPS@NOD-IN-1 group were significantly increased (P<0.05), the level of ROS in wound tissue on PID 3 was significantly decreased (P<0.05), the thickness of granulation tissue in wound on PID 7 was significantly thickened (P<0.05), and the mRNA expressions of NOD1 and NOD2 and the protein expressions of NOD1, NOD2, and GSDMD-N terminals in wound tissue on PID 7 were significantly decreased (P<0.05). KEGG pathway analysis showed that DEGs significantly down-regulated in PEPS@NOD-IN-1 group as compared with PBS group were significantly enriched in NOD-like receptors, hypoxia-inducible factors, mitogen-activated protein kinases, and tumor necrosis factor (TNF) pathways. In the DEG heatmap of NOD-like receptor pathway, the genes regulating pyroptosis mainly involved NOD1, NOD2, NOD-like receptor thermoprotein domain-related protein 3, Jun, signal transduction and transcriptional activator 1 (STAT1), TNF-α-induced protein 3. The PPI results showed that NOD1, NOD2, and STAT1 were the key genes of PEPS@NOD-IN-1 regulating the NOD-like receptor pathway.  Conclusions  PEPS@NOD-IN-1 can down-regulate the level of local ROS in wounds and the expression of NOD1, NOD2, and GSDMD-N terminals, the key regulators of pyroptosis, thereby promoting the repair of full-thickness skin defect wounds in diabetic rats. PEPS@NOD-IN-1 can also significantly down-regulate the pyroptosis, inflammation, and hypoxia-related pathways of wounds, and regulate NOD-like receptor pathways by down-regulating key genes NOD1, NOD2, and STAT1.

     

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