Egyptian mandarin peel oil's anti-scabies potential via downregulation-of-inflammatory/immune-cross-talk: GC-MS and PPI network studies

Abstract

The current study investigated the scabicidal potential of Egyptian mandarin peel oil (Citrus reticulata Blanco, F. Rutaceae) against sarcoptic mange-in-rabbits. Analysis of the oil's GC-MS identified a total of 20 compounds, accounting for 98.91% of all compounds found. Mandarin peel oil topical application improved all signs of infection, causing a scabicidal effect three days later, whereas in vitro application caused complete mite mortality one day later. In comparison to ivermectin, histopathological analysis showed that the epidermis' inflammatory-infiltration/hyperkeratosis-had disappeared. In addition to TIMP-1, the results of the mRNA gene expression analysis showed upregulation of I-CAM-1-and-KGF and downregulation of ILs-1, 6, 10, VEGF, MMP-9, and MCP-1. The scabies network was constructed and subjected to a comprehensive bioinformatic evaluation. TNF-, IL-1B, and IL-6, the top three hub protein-coding genes, have been identified as key therapeutic targets for scabies. From molecular docking data, compounds 15 and 16 acquired sufficient affinity towards the three screened proteins, particularly both possessing higher affinity towards the IL-6 receptor. Interestingly, it achieved a higher binding energy score than the ligand of the docked protein rather than displaying proper binding interactions like those of the ligand. Meanwhile, geraniol (15) showed the highest affinity towards the GST protein, suggesting its contribution to the acaricidal effect of the extract. The subsequent, MD simulations revealed that geraniol can achieve stable binding inside the binding site of both GST and IL-6. Our findings collectively revealed the scabicidal ability of mandarin peel extract for the first time, paving the way for an efficient, economical, and environmentally friendly herbal alternative for treating rabbits with Sarcoptes mange.

Figure 1

General outflow of the study.

Figure 2

GC/MS spectrum for Citrus reticulata peels oil.

Figure 3

Structures of identified compounds, using GC/MS analysis, from Citrus reticulata oil isolated from peels.

Figure 4

The H₂O₂ scavenging activity of both the mandarin peel oil and the standard increased in a concentration-dependent manner (Fig. 4A). Interestingly, at a concentration of 1000 μg/mL, mandarin peel oil exhibited the highest superoxide removal action, with an IC50 value of 176.2 μg/mL (Fig. 4B). This indicates that the oil was more effective at scavenging the superoxide radicals than the standard, ascorbic acid.

Figure 5

Inspection of mange-infected rabbits under a microscope, (A) control group (paraffin oil), (B) mandarin peels oil group (20% peels oil in paraffin oil), (C) ivermectin group (5% ivermectin).

Figure 6

Microscopical-examination-of-skin from-different-groups of-animals, (A) normal-architecture-of the-skin: e; epidermis, d; dermis, h.f.; hair-follicles, (B) control-group-showing-skin-damage-with-hyperkeratosis (red arrows), mites-remnants-embedded-in the-skin (blue arrows), hypergranulation-of-dermis (green-arrows), severe-akanthosis-with-cellular-infiltration (black-arrows), (C) mandarin-peels-oil group showing-restoration of-normal-architecture, with-mild-infiltration (red-arrow), healthy-sebaceous-glands (yellow-arrow) and hair-follicles (black-arrows), (D) ivermectin-group showing-moderate damage-with-hyperkeratosis (red-arrow), mature-mites with-eggs-remnants-embedded in-the-dermis (black-arrow) surrounded-by-cellular-infiltration (green-arrow), and some-sebaceous-adenitis (yellow-arrows).

Figure 7

Relative gene expression in skin tissue of different animal groups using qRT-PCR After normalisation to GAPDH, (A) I-CAM, (B) IL-1, (C) IL-10, (D) MCP-1, (E) TIMP-1, (F) MMP-9, (G) KGF, (H) IL-6, and (I) VEGF. In comparison to the healthy control group, the data show an increase in expression by a factor of two. The mean ± SD are shown as bars. A one-way ANOVA test is used to determine whether there's a significant difference between categories, via (a) p < 0.05 in contrast to the normal control grouping and (b) p < 0.05 in contrast to the marketplace drug-induced category.

Figure 8

(A) Through a circular network design, the margins represent interactions between proteins, and the nodes serve as the hub protein criteria. Each protein's connectivity is represented by the dimension of the nodes; the larger the node, the greater its connection with other nodes in the network, (B) Functional enrichment analysis of filtered 16 protein coding genes by ShinyGO (https://www.genome.jp/kegg/, accessed on 12 September 2022); (http://bioinformatics.sdstate.edu/go/, accessed on 13 September 2022, a graphical gene set enrichment tool).

Figure 9

(A) 2D actions and 3D docking represent compound 15 (geraniol) in the successful pocket location of IL-1 (PDB: 6Y8M), (B) 2D actions and 3D docking represent compound 16 (1-decanol) in the successful pocket location of IL-1 (PDB: 6Y8M), (C) 2D actions and 3D docking present in compound 15 in the successful pocket location of IL-6 (PDB: 1ALU), (D) 2D relationships and 3D docking present in compound 16 in the successful pocket location of IL-6 (PDB: 1ALU), (E) 2D relationships and 3D docking present in compound 17 in the successful pocket location of IL-6 (PDB: 1ALU), (E) 2D relationships and 3D docking represent compound 16 in TNF- successful pocket location (PDB: 2AZ5), and (F) 2D relationships and 3D docking present compound 15 in GST successful pocket location (PDB: 2AZ5) (PDB: 3EIN).

Figure 9

(A) 2D actions and 3D docking represent compound 15 (geraniol) in the successful pocket location of IL-1 (PDB: 6Y8M), (B) 2D actions and 3D docking represent compound 16 (1-decanol) in the successful pocket location of IL-1 (PDB: 6Y8M), (C) 2D actions and 3D docking present in compound 15 in the successful pocket location of IL-6 (PDB: 1ALU), (D) 2D relationships and 3D docking present in compound 16 in the successful pocket location of IL-6 (PDB: 1ALU), (E) 2D relationships and 3D docking present in compound 17 in the successful pocket location of IL-6 (PDB: 1ALU), (E) 2D relationships and 3D docking represent compound 16 in TNF- successful pocket location (PDB: 2AZ5), and (F) 2D relationships and 3D docking present compound 15 in GST successful pocket location (PDB: 2AZ5) (PDB: 3EIN).

Figure 10

RMSDs of geraniol inside both GST and IL-6 in comparison with the co-crystalized ligand of each protein [(A) and (B) respectively] over the course of 50 ns-long MD simulation.

Figure 11

In silico druglikeness (Lipinski and Veber rules) of compounds 15 and 16.

Figure 12

The suggested mechanism for the effect of mandarin peel oil on scabies-infected rabbits.