Anti-depressant effect of Naringenin-loaded hybridized nanoparticles in diabetic rats via PPARγ/NLRP3 pathway

Abstract

Naringenin (NAR) has various biological activities but low bioavailability. The current study examines the effect of Naringenin-loaded hybridized nanoparticles (NAR-HNPs) and NAR on depression induced by streptozotocin (STZ) in rats. NAR-HNPs formula with the highest in vitro NAR released profile, lowest polydispersity index value (0.21 ± 0.02), highest entrapment efficiency (98.7 ± 2.01%), as well as an acceptable particle size and zeta potential of 415.2 ± 9.54 nm and 52.8 ± 1.04 mV, respectively, was considered the optimum formulation. It was characterized by differential scanning calorimetry, examined using a transmission electron microscope, and a stability study was conducted at different temperatures to monitor its stability efficiency showing that NAR-HNP formulation maintains stability at 4 °C. The selected formulation was subjected to an acute toxicological test, a pharmacokinetic analysis, and a Diabetes mellitus (DM) experimental model. STZ (50 mg/kg) given as a single i.p. rendered rats diabetic. Diabetic rat groups were allocated into 4 groups: one group received no treatment, while the remaining three received oral doses of unloaded HNPs, NAR (50 mg/kg), NAR-HNPs (50 mg/kg) and NAR (50 mg/kg) + peroxisome proliferator-activated receptor-γ (PPAR-γ) antagonist, GW9662 (1mg/kg, i.p.) for three weeks. Additional four non-diabetic rat groups received: distilled water (normal), free NAR, and NAR-HNPs, respectively for three weeks. NAR and NAR-HNPs reduced immobility time in forced swimming test and serum blood glucose while increasing serum insulin level. They also reduced cortical and hippocampal 5-hydroxyindoeacetic acid, 3,4-Dihydroxy-phenylacetic acid, malondialdehyde, NLR family pyrin domain containing-3 (NLRP3) and interleukin-1beta content while raised serotonin, nor-epinephrine, dopamine and glutathione level. PPAR-γ gene expression was elevated too. So, NAR and NAR-HNPs reduced DM-induced depression by influencing brain neurotransmitters and exhibiting anti-oxidant and anti-inflammatory effects through the activation PPAR-γ/ NLRP3 pathway. NAR-HNPs showed the best pharmacokinetic and therapeutic results.

Figure 1

Schematic summary of the experimental study.

Figure 2

In vitro NAR release profiles of different hybridized nanoparticle formulations in comparison with the free drug in phosphate buffer saline pH 6.8 at 37 °C.

Figure 3

DSC thermograms of (A) NAR, (B) lipoid S45, (C) P407, (D) CS and (E) NAR-HNP6.

Figure 4

Transmission electron micrographs of NAR-HPN6.

Figure 5

Storage effect on the particle size (a), polydispersity index (b), entrapment efficiency (c), and the zeta potential (d) of NAR-HNP6 at 4 °C, 25 °C and 40 °C. Results are stated as mean ± SD (n = 3). * vs correspondent o day.

Figure 6

Effect of 2 weeks treatment of NAR and NAR-HNPs on (a) ALT, (b) AST, (c) Creatinine and (d) Urea in male and female rats. Results are expressed as mean ± SEM. HNPs, hybridized nanoparticles; NAR-HNPs, naringenin- loaded hybridized nanoparticles.

Figure 7

Effect of 2 weeks treatment of unloaded HNPs and the selected NAR-HNPs on (a) RBC, (b) HGB, (c) HCT and (d) PLT and (e) WBC in male rats. Results are stated as mean ± SEM. HNPs, hybridized nanoparticles; NAR-HNPs, naringenin- loaded hybridized nanoparticles.

Figure 8

Effect of 2 weeks treatment of unloaded HNPs and the selected NAR-HNPs on (a) RBC, (b) HGB, (c) HCT and (d) PLT and (e) WBC in female rats. Results are stated as mean ± SEM. HNPs, hybridized nanoparticles; NAR-HNPs, naringenin-loaded hybridized nanoparticles.

Figure 9

Linearity of NAR by HPLC verification method.

Figure 10

Linearity of Brain monoamines namely (a) NE, (b) 5-HT, and (c) DA by HPLC verification method.

Figure 11

Plasma concentration–time curve of NAR after oral injection of NAR and NAR-HNPs.

Figure 12

Effect of NAR and NAR-HNPs on body weight changes in diabetic rats. Results are stated as mean ± SEM. ^avs normal group, ^bvs control STZ group, ^cvs STZ + NAR (50 mg/kg), ^dvs STZ + NAR-HNPs (50 mg/kg) at p < 0.05. NAR, naringenin; STZ, streptozotocin; NAR-HNPs, naringenin- loaded hybridized nanoparticles.

Figure 13

Effect of NAR and NAR-HNPs on spontaneous locomotor activity in diabetic rats subjected to activity cage test. Results are stated as mean ± SEM. NAR, naringenin; STZ, streptozotocin; NAR-HNPs, naringenin- loaded hybridized nanoparticles.

Figure 14

Effect of NAR and NAR-HNPs on (a) immobility time, (b) latency to immobility time on diabetic rats subjected to FST. Results are stated as mean ± SEM. ^avs normal group, ^bvs control STZ group, ^cvs STZ + NAR (50 mg/kg), ^dvs STZ + NAR-HNPs (50 mg/kg) at p < 0.05. NAR, naringenin; STZ, streptozotocin; NAR-HNPs, naringenin- loaded hybridized nanoparticles.

Figure 15

Effect of NAR and NAR-HNPs on (a) GSH, (b) MDA, (c) NLRP3, (d) IL-1β cortical contents in diabetic rats. Results are stated as mean ± SEM. ^avs normal group, ^bvs control STZ group, ^cvs STZ + NAR (50 mg/kg), ^dvs STZ + NAR-HNPs (50 mg/kg) at p < 0.05. NAR, naringenin; STZ, streptozotocin; NAR-HNPs, naringenin- loaded hybridized nanoparticles.

Figure 16

Effect of NAR and NAR-HNPs on (a) GSH, (b) MDA, (c) NLRP3, (d) IL-1β hippocampal contents in diabetic rats. Results are stated as mean ± SEM. ^avs normal group, ^bvs control STZ group, ^cvs STZ + NAR (50 mg/kg), ^dvs STZ + NAR-HNPs (50 mg/kg) at p < 0.05. NAR, naringenin; STZ, streptozotocin; NAR-HNPs, naringenin- loaded hybridized nanoparticles.