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Anti-Infectivity against Herpes Simplex Virus and Selected Microbes and Anti-Inflammatory Activities of Compounds Isolated from Eucalyptus globulus Labill - PubMed

  • ️Mon Jan 01 2018

Anti-Infectivity against Herpes Simplex Virus and Selected Microbes and Anti-Inflammatory Activities of Compounds Isolated from Eucalyptus globulus Labill

Viliam Brezáni et al. Viruses. 2018.

Abstract

Herpes simplex virus (HSV) causes numerous mild-to-serious human diseases, including mucocutaneous herpes infections and life-threatening herpes encephalitis. Moreover, herpes viral lesions can be complicated by inflammation and secondary bacterial infections. The development of resistance to antiviral drugs along with the undesirable side effects of these drugs are relevant argue for the development of new anti-HSV drugs with diverse mechanisms of action. Eucalyptus extracts have been used for decades to combat various infectious diseases. We isolated and studied 12 pure compounds and one mixture of two constitutional isomers from the leaves and twigs of E. globulus. The structures were identified by spectroscopic methods (NMR, HR-MS, IR) and all of them were tested for antiherpetic activity against the replication of antigen types HSV-1 and HSV-2. Tereticornate A (12) (IC50: 0.96 μg/mL; selectivity index CC50/IC50: 218.8) showed the strongest activity in the anti-HSV-1 assay, even greater than acyclovir (IC50: 1.92 μg/mL; selectivity index CC50/IC50: 109.4), a standard antiviral drug. Cypellocarpin C (5) (EC50: 0.73 μg/mL; selectivity index CC50/EC50: 287.7) showed the most potent anti-HSV-2 activity, also more intensive than acyclovir (EC50: 1.75 μg/mL; selectivity index CC50/EC50: 120.0). The antimicrobial activity of the isolated compounds was also evaluated against the bacteria Staphylococcus aureus, Bacillus cereus, Escherichia coli, and Pseudomonas aeruginosa and the yeast Candida albicans. The anti-inflammatory potential was examined using LPS-stimulated THP-1-XBlue™-MD2-CD14 and THP-1 macrophages and focusing on the influences of the NF-κB/AP-1 activity and the secretion of pro-inflammatory cytokines IL-1β and TNF-α.

Keywords: Eucalyptus; HSV-1; HSV-2; IL-1β; NF-κB/AP-1; ROS; TNF-α; anti-inflammatory; antibacterial; antiherpetic.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1

Structures of isolated compounds.

Figure 1
Figure 1

Structures of isolated compounds.

Figure 2
Figure 2

Effect of compound 4 on the viability of human leukemic monocytes THP-1. The cells were treated with compound 4 dissolved in DMSO at increasing concentrations 0.25–0.5–1–1.25–2.5–5–10–20 μM or with only the vehicle and their viability was measured after 24 h. Untreated control cells were used to verify the non-toxic effect of the vehicle. The viability of the cells treated with compound 4 was compared to that of the vehicle-treated cells. The results are expressed as the mean ± SE for three independent assays. The concentration of DMSO never exceeded 0.1%.

Figure 3
Figure 3

Effect of compounds 1–3, 5–12, and 13 (a + b) on NF-κB/AP-1 activity. THP-1-XBlue™-MD2-CD14 cells were pre-treated for 1 h with the test compounds (5 μM dissolved in DMSO), prednisone (P; 5 μM dissolved in DMSO), with only the vehicle (V). Subsequently, LPS (1 μg/mL) was added (to all but C, control cells) to trigger the activation of NF-κB/AP-1. After 24 h, the activity of NF-κB/AP-1 was evaluated based on the amount of secreted alkaline phosphatase measured spectrophotometrically. The results are expressed as the mean ± SE for six independent experiments measured in triplicate. ** indicates a significant difference (p < 0.01) in comparison with the vehicle-treated cells and **** indicates a significant difference (p < 0.0001) in comparison with the vehicle-treated cells.

Figure 4
Figure 4

The effects of compounds 8 and 12 on the secretion of pro-inflammatory cytokines IL-1β (a) and TNF-α (b). PMA-differentiated macrophages were pre-treated with test compounds 8 and 12 at a concentration of 5 μM, with prednisone (P) 1 μM dissolved in DMSO, or with only the vehicle (V). LPS 1 μg/mL was added (to all but C, control cells) after 1 h of incubation. After 24 h, the supernatants were collected and the secretions of IL-1β and TNF-α in each culture medium were evaluated using ELISA. Results are expressed as the mean ± SE for three independent experiments. * indicates a significant difference (p < 0.05) in comparison with the vehicle-treated cells, ** indicates a significant difference (p < 0.01), and **** p < 0.0001.

Figure 5
Figure 5

Effect of selected E. globulus compounds on the production of ROS. THP-1 monocytes were pre-treated with compounds 1–3, 5–12, and 13 (a + b) at concentrations of 5 μM, with 1 μM quercetin (Q) or with only the vehicle (V) for 30 min. Pyocyanin at a final concentration of 100 μM was then added (to all but C, control cells) to trigger the generation of ROS. After 30 min, DCFH-DA (5 μg/mL) dissolved in DMF was introduced into the cell medium. The intracellular fluorescence of dichlorofluorescein produced in this oxidative reaction was measured spectrophotometrically. The results in Figure 5 show the mean ± SE for three independent experiments. *** indicates a significant difference (p < 0.001) in comparison with the vehicle-treated cells.

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