Self-assembly of H2S-responsive nanoprodrugs based on natural rhein and geraniol for targeted therapy against Salmonella Typhimurium

Abstract

Salmonellosis is a globally extensive food-borne disease, which threatens public health and results in huge economic losses in the world annually. The rising prevalence of antibiotic resistance in Salmonella poses a significant global concern, emphasizing an imperative to identify novel therapeutic agents or methodologies to effectively combat this predicament. In this study, self-assembly hydrogen sulfide (H₂S)-responsive nanoprodrugs were fabricated with poly(α-lipoic acid)-polyethylene glycol grafted rhein and geraniol (PPRG), self-assembled into core-shell nanoparticles via electrostatic, hydrophilic and hydrophobic interactions, with hydrophilic exterior and hydrophobic interior. The rhein and geraniol are released from self-assembly nanoprodrugs PPRG in response to Salmonella infection, which is known to produce hydrogen sulfide (H₂S). PPRG demonstrated stronger antibacterial activity against Salmonella compared with rhein or geraniol alone in vitro and in vivo. Additionally, PPRG was also able to suppress the inflammation and modulate gut microbiota homeostasis. In conclusion, the as-prepared self-assembly nanoprodrug sheds new light on the design of natural product active ingredients and provides new ideas for exploring targeted therapies for specific Enteropathogens. Graphical illustration for construction of self-assembly nanoprodrugs PPRG and its antibacterial and anti-inflammatory activities on experimental Salmonella infection in mice.

Keywords: Geraniol; H2S-responsive nanoprodrugs; PPRG; Rhein; Salmonella infection; Self-assembly.

Graphical  illustration for construction of self-assembly nanoprodrugs PPRG and its antibacterial and anti-inflammatory activities on experimental Salmonella infection in mice

Fig. 1

Synthesis and characteristics of PPR, PEGylated PALA, and PPRG. A Structural formula of PPRG. B ¹H NMR. C FTIR. D GPC. E TEM. F particle size. G zeta potential. H nanoparticles deal with 1 mM·L⁻¹ Na₂S determined by Zetatrac particle size analyzer of PEGylated PALA, PPR, PPG, and PPRG. (I) Time-killing curve of S. Typhimurium ATCC 14028 under anaerobic condition for 24 h. (CON control, SAL Salmonella, RH rhein, PPR PEGylated PALA grafted rhein, GE geraniol, PPG PEGylated PALA grafted geraniol, and PPRG poly(α-lipoic acid)-polyethylene glycol grafted rhein and geraniol, respectively)

Fig. 2

The cell viability and LPS-stimulated RAW 264.7 cellls. Cell viability for A Caco-2. B 293 T.and C RAW 264.7. D Live/dead cell staining of Caco-2 cells using 40 μM·L⁻¹ test samples. LPS-stimulated RAW 264.7 cells incubated with test samples and measurements of E NO. F TNF-α. G IL-6. and H IL-1β

Fig. 3

Therapeutic efficacy in an acute Salmonella infection in vivo model. A The study protocol included streptomycin pretreatment followed by Salmonella infection, treatment with rhein (25 mg·kg⁻¹), PPR (250 mg·kg⁻¹), geraniol (12.5 mg·kg⁻¹), PPG (26 mg·kg⁻¹) and PPRG (21 mg·kg⁻¹), daily. B Variations of mouse body weights over time and normalized to the percentage of day zero body weight. C Cumulative food intake of per group from day 1 to 11. Quantification of bacterial burdens in D faeces. E heart. F spleen. and G lungs of infected mice with different treatments. Serum inflammatory factor levels of H TNF-α. I IL-6. J IL-10 in vivo for Salmonella-infected mice

Fig. 4

Intestinal morphology of mice in treatment. A The H&E staining of the jejunum and cecum in mice. B Histologic and immunohistochemical features of the cecum. C–E Villus length in the small intestine (duodenum, jejunum, and ileum)

Fig. 5

Gut microbiota analysis of infected mice after treatments. A Venn diagram indicating different OTU number for the four treatment groups. B Average bacterial taxonomic profiling of the gut microbiota in seven groups. C Nonmetric multidimensional scaling score plot based on Bray–Curtis distances. D UniFrac-based PCoA score plot based on weights. E Average bacterial taxonomic profiling of the gut microbiota in seven groups at phylum level. F–H Relative abundance of bacterial phylum obtained from LefSe results. I Average bacterial taxonomic profiling of the gut microbiota in seven groups at the genus level. J–L Relative abundance of the bacterial genera obtained from the LefSe results