Hypericum perforatum L. Nanoemulsion Mitigates Cisplatin-Induced Chemobrain via Reducing Neurobehavioral Alterations, Oxidative Stress, Neuroinflammation, and Apoptosis in Adult Rats

Abstract

Cisplatin (Cis) is a potent chemotherapeutic agent; however, it is linked with oxidative stress, inflammation, and apoptosis, which may harmfully affect the brain. Hypericum perforatum L. (HP L.) is a strong medicinal plant, but its hydrophobic polyphenolic compounds limit its activity. Therefore, our study aimed to investigate the neuroprotective action of HP L. and its nanoemulsion (NE) against Cis-induced neurotoxicity. The prepared HP.NE was subjected to characterization. The droplet size distribution, surface charge, and morphology were evaluated. In addition, an in vitro dissolution study was conducted. Compared to Cis-intoxicated rats, HP L. and HP.NE-treated rats displayed improved motor activity and spatial working memory. They also showed an increase in their antioxidant defense system and a reduction in the levels of pro-inflammatory cytokines in the brain. Moreover, they showed an increase in the expression levels of the PON-3 and GPX genes, which are associated with a reduction in the brain levels of COX-2 and TP-53. These findings were confirmed by reducing the immunohistochemical expression of nuclear factor kappa (NF-ƘB) and enhanced Ki-67 levels. In conclusion, HP L. is a promising herb and could be used as an adjuvant candidate to ameliorate chemotherapeutic-induced neurotoxicity. Moreover, HP.NE has superior activity in lessening Cis-induced oxidative stress, inflammation, and apoptosis in brain tissue.

Keywords: Hypericum perforatum L.; apoptosis; cisplatin; inflammation; neurobehavior; neuroprotective; oxidative stress; toxicity.

Figure 1

Development of Hypericum perforatum L. nanoemulsion (HP.NE): (a) Ternary phase diagram represents the prepared 9 screening trials where the borderline separates the NE region. (b) Globule size, (c) polydispersity index, and (d) surface charge of unloaded (F1–3) and loaded (F4–6) selected NEs. (e) The in-vitro release profile of loaded NEs. (f) TEM image of F4 as the selected formulation for further investigations. * means significant at p < 0.05; **, significant at p < 0.01; ***, significant at p < 0.001).

Figure 2

Effect of HP L. and HP.NE administration on the motor activity and spatial working memory of Cis-intoxicated rats. (a) Open field test: number of crossing squares, (b) open field test: rearing activity, (c) Y-maze test: number of arm entries, and (d) Y-maze test: spontaneous alternation percentage (SAP). Data are expressed as mean ± standard error (SE)using one-way analysis of variance (one-way ANOVA) followed by Tukey’s post hoc test for seven rats in each group. * means significant from the Control group; @, significant from the Cis group; #, significant from the HP + Cis group.

Figure 3

Effect of HP L. and HP.NE administration on the brain levels of oxidative biomarkers of different groups. (a) SOD: super oxide dismutase, (b) CAT: catalase, (c) GST: glutathione -s- transferase, and (d) GSH: reduced glutathione. Data are expressed as mean ± SE (one-way ANOVA) followed by Tukey’s post hoc test for seven rats in each group. * means significant from the Control group; @, significant from the Cis group, #, significant from the HP + Cis group.

Figure 4

Effect of HP L. and HP.NE administration on the brain levels of malondialdehyde, 8-hydroxy-20-deoxyguanosine, and nitric oxide for different groups. (a) MDA: malondialdehyde, (b) 8-OHdG: 8-hydroxy-20-deoxyguanosine, and (c) NO: nitric oxide. Data are expressed as mean ± SE (one-way ANOVA) followed by Tukey’s post hoc test for seven rats in each group. * means significant from the Control group; @, significant from the Cis group; #, significant from the HP + Cis group.

Figure 5

Effect of HP L. and HP.NE administration on the brain levels of pro-inflammatory cytokines in different groups. (a) IL-6: interleukin 6, (b) IL1b: interleukin-1 beta, (c) TNFα: tumor necrosis factor α, and (d) Caspase-3. Data are expressed as mean ± SE (one-way ANOVA) followed by Tukey’s post hoc test for seven rats in each group. * means significant from the Control group; @, significant from the Cis group; #, significant from the HP + Cis group.

Figure 6

Effect of HP L. and HP.NE administration on the relative gene expression of PON-3, GPX COX-2, and TP-53 in brain tissues of different groups. (a) PON-3: paraoxonase 3, (b) GPX: glutathione peroxidase, (c) COX-2: cyclooxygenase-2, and (d) TP-53: tumor protein p53. Data are expressed as mean ± SE (one-way ANOVA) followed by Tukey’s post hoc test for seven rats in each group. * means significant from the Control group; @, significant from the Cis group; #, significant from the HP + Cis group.

Figure 7

Effect of HP L. and HP.NE administration on the histological structure of the brain. (a) Photomicrograph from control group presenting the normal structure of the hippocampal area, which is made up of Cornu Ammonis (CA1, CA2, CA3, and CA4) and Dentate gyrus (DG); scale bar denotes 200 m. (b) Photomicrograph of brain tissues stained with H&E; scale bars denote 10 and 20 m. Notable symbols on the figure point to neurophagia of degenerated neurons (curved arrows), dark stained neurons (arrows), loss of neurons (black arrowheads), areas void of neurons (*), and vacuolation (blue arrowheads). (c) Brain damage score. Results are expressed as median standard deviation (SD), determined using the Kruskal–Wallis test followed by Dunn’s multiple comparison test. * means significant from the control group; @, significant from the Cis group; #, significant from HP + Cis group.

Figure 8

Immunohistochemical staining of inflammatory Nuclear factor kappa B (NF-ƘB) and proliferative (Ki-67) markers in brain tissues of all groups. (a) Photomicrograph of brain tissues stained with NF-ƘB and Ki-67 (H&E); scale bars denote 10 µm. Remarkable arrows on the figure point to strong positive staining. (b) Statistical analysis of NF-ƘB expression (area%). (c) Statistical analysis of Ki-67 expression (area%). * means significant from the control group; @, significant from the Cis group; and #, significant from HP + Cis group.