Berberine-Loaded PVCL-PVA-PEG Self-Assembled Micelles for the Treatment of Liver Fibrosis

Abstract

Background: Liver fibrosis is a necessary pathological process in many chronic liver diseases. Studies have shown that the progression of chronic liver disease can be slowed by rational intervention in hepatic fibrosis. Berberine (BBR), a natural extract of Phellodendron amurense, inhibits the development of liver fibrosis through several mechanisms. However, the clinical application of BBR is limited due to its low solubility. Drug delivery systems have been developed to improve the solubility of hydrophobic drugs and increase their efficacy in treating the liver fibrosis.

Methods: In this study, a biocompatible nanomicelle was constructed by thin-film dispersion method using polyvinyl caprolactam-polyvinyl acetate-polyethylene glycol graft copolymer (PVCL-PVA-PEG) as a carrier to encapsulate BBR (PVCL-PVA-PEG/BBR-MCs) to improve the solubility of BBR and reduce the systemic side effects. The ability to inhibit HSC-T6 cell activation of PVCL-PVA-PEG/BBR-MCs was evaluated in vitro. The anti-hepatic fibrosis effects of PVCL-PVA-PEG/BBR-MCs were investigated in vivo.

Results: PVCL-PVA-PEG/BBR-MCs have a uniform spherical shape with a mean particle size of 60.04 ± 0.027 nm and a potential of 1.49 ± 0.32 mV. It had an encapsulation efficiency of 98.52% ± 0.70 and drug loading content of 6.16% ± 0.04. Compared to free BBR, PVCL-PVA-PEG/BBR-MCs significantly inhibited HSC-T6 cell activation and TGF-β 1-induced HSC-T6 cell migration in vitro. In vivo biodistribution experiments showed significantly improved hepatic distribution of PVCL-PVA-PEG/DiD-MCs compared to free DiD, suggesting that PVCL-PVA-PEG micelles enhance the ability of BBR to enter the liver and improve therapeutic efficacy. After treatment, PVCL-PVA-PEG/BBR-MCs significantly improved fibrotic liver structure and reduced collagen deposition in comparison to the CCl₄-treated group; the treatment outcome was more effective than that of the free BBR group.

Conclusion: Our results demonstrate the advantages of encapsulating BBR in PVCL-PVA-PEG micelles and highlight the potential of PVCL-PVA-PEG/BBR-MCs as a therapeutic strategy for the treatment of liver fibrosis.

Introduction

Liver fibrosis, a chronic liver disease characterized by hepatocyte morphological changes, extracellular matrix accumulation, intrahepatic obstruction and loss of functional hepatocyte clusters, is now a global public health problem.1–3 The etiology of liver fibrosis is complex, involving the interaction of multiple cells, mediators and signal pathways.4 Liver fibrosis is an inevitable pathological process in many chronic liver diseases.2 If not treated in time, it can lead to cirrhosis or even liver cancer. A number of studies have shown that interventions that target certain cells involved in liver fibrosis can slow down or even reverse the progression of liver fibrosis and can significantly reduce the development of severe chronic liver disease.5–7 Liver fibrosis is currently being treated by protecting normal liver cells,8 inhibiting hepatic stellate cells (HSCs) activation,9 decreasing extracellular matrix synthesis and secretion,10 and reducing liver inflammation. Interferons (INF; INF-γ),11 anti-HBV nucleotides and their analogues,12 endothelin receptor A antagonists,13,14 angiotensin receptor blockers, and extracts of natural products such as colchicine15 and silymarin16 are commonly used in the treatment of liver fibrosis, with some success. However, the current lack of clinical applications remains.

Berberine (BBR), an isoquinoline alkaloid found in the genus Phellodendron, has a wide range of pharmacological activities including antioxidant, antibacterial, anti-inflammatory, anticancer, anti-diabetic, anti-dyslipidaemic and anti-obesity properties.17–21 Recent studies have shown that it also plays an important role in the treatment of a wide range of fibrotic diseases such as liver fibrosis,22,23 lung fibrosis,24 kidney fibrosis25,26 and heart fibrosis.27,28 The metabolic process of BBR consists of two main steps, 1) demethylation, reduction and cleavage of the dioxymethylene in the molecular structure of BBR and 2) glucuronidation, sulfation and methylation.29 BBR is metabolized in the liver, where it provides the material and structural basis for treating hepatic fibrosis. However, the use of BBR in clinical practice is limited by its extremely low water solubility and bioavailability. To improve its solubility, many studies have attempted to make berberine hydrochloride, which is inevitably damaging to other organs.30

PVCL-PVA-PEG (soluplus®), a biocompatible amphiphilic block copolymer, can be used to improve the solubility of hydrophobic drugs.31,32 It has been reported that soluplus® has a very low critical micelle concentration (CMC) of about 7–8 mg/L, which contributes to the stability of the self-assembled micelles.31 Previously published studies have reported that soluplus® can be self-assembled into a drug delivery system for the encapsulation of water-insoluble drugs for the treatment of eye diseases and tumours.33,34

In this study, based on the perspective that nanodrug delivery systems can improve the solubility and biocompatibility of hydrophobic drugs,35,36 BBRs were prepared as homogeneously dispersed micelles that can be dissolved in aqueous solution. To improve berberine bioavailability, we used commercially available PVCL-PVA-PEG as a drug delivery vehicle to encapsulate berberine to prepare PVCL-PVA-PEG/BBR-MCs. The prepared PVCL-PVA-PEG/BBR-MCs improved the solubility of BBR APIs and further enhanced liver accumulation. The therapeutic efficacy of PVCL-PVA-PEG/BBR-MCs on fibrotic liver was evaluated by several physicochemical properties, providing a new potential approach for the treatment of liver fibrosis. The results showed that the prepared PVCL-PVA-PEG/BBR-MCs were efficiently taken up by HSC-T6 cell in vitro. Furthermore, they had a significant inhibitory effect on HSC-T6 cell activation in vitro. In vivo, the results showed that PVCL-PVA-PEG/BBR-MCs were able to reduce collagen deposition, improve the structure of the liver tissue as well as attenuate the progression of liver fibrosis.

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Materials

Berberine (BBR) and BCA protein assay kit were purchased from Beiing Solarbio Science &Technology Co.Ltd.. Polyvinyl caprolactam-polyvinyl acetate-polyethylene glycol graft copolymer (PVCL-PVA-PEG, Soluplus) was purchased from BASF (CABA, Argentina). HSC-T6 was purchased from Fenghui (Hunan). Dulbecco’s Modified Eagle’s Medium (DMEM) and fetal bovine serum (FBS) were purchased from Gibco. Alpha-Smooth Muscle antibody was purchased from SIGMA, Anti-Collagen I antibody was purchased from protein technology. DiD, HSP-90, and Tubulin were purchased from Beyotime Biotechnology. CCK-8 kit was purchased from Meilunbio.

Zha X, Hao Y, Ke Y, Wang Y, Zhang Y. Berberine-Loaded PVCL-PVA-PEG Self-Assembled Micelles for the Treatment of Liver Fibrosis. Int J Nanomedicine. 2024;19:10857-10872
https://doi.org/10.2147/IJN.S465214