The Cytoprotective Effect of C60 Derivatives in the Self-Microemulsifying Drug Delivery System against Triptolide-Induced Cytotoxicity In Vitro.

OBJECTIVE: The aim of this study was to optimize the formulation of a C60-modified self-microemulsifying drug delivery system loaded with triptolide (C60-SMEDDS/TP) and evaluate the cytoprotective effect of the C60-SMEDDS/TP on normal human cells. RESULTS: The C60-SMEDDS/TP exhibited rapid emulsification, an optimal particle size distribution of 50 ± 0.19 nm (PDI 0.211 ± 0.049), and a near-neutral zeta potential of -1.60 mV. The release kinetics of TP from the C60-SMEDDS/TP exhibited a sustained release profile and followed pseudo-first-order release kinetics. Cellular proliferation and apoptosis analysis indicated that the C60-SMEDDS/TP (with a mass ratio of TP: DSPE-PEG-C60 = 1:10) exhibited lower toxicity towards L02 and GES-1 cells. This was demonstrated by a higher IC50 (40.88 nM on L02 cells and 17.22 nM on GES-1 cells) compared to free TP (21.3 nM and 11.1 nM), and a lower apoptosis rate (20.8% on L02 cells and 26.3% on GES-1 cells, respectively) compared to free TP (50.5% and 47.0%) at a concentration of 50 nM. In comparison to the free TP group, L02 cells and GES-1 cells exposed to the C60-SMEDDS/TP exhibited a significant decrease in intracellular ROS and an increase in mitochondrial membrane potential (ΔψM). On the other hand, the C60-SMEDDS/TP demonstrated a similar inhibitory effect on BEL-7402 cells (IC50 = 28.9 nM) and HepG2 cells (IC50 = 107.6 nM), comparable to that of the free TP (27.2 nM and 90.4 nM). The C60-SMEDDS/TP group also exhibited a similar intracellular level of ROS and mitochondrial membrane potential compared to the SMEDDS/TP and free TP groups. METHOD: Fullerenol-Grafted Distearoyl Phosphatidylethanolamine-Polyethylene Glycol (DSPE-PEG-C60) was synthesized and applied in the self-microemulsifying drug delivery system. The C60-SMEDDS/TP was formulated using Cremophor EL, medium-chain triglycerides (MCT), PEG-400, and DSPE-PEG-C60, and loaded with triptolide (TP). The toxicity and bioactivity of the C60-SMEDDS/TP were assessed using normal human liver cell lines (L02 cells), normal human gastric mucosal epithelial cell lines (GES-1 cells), and liver cancer cell lines (BEL-7402 cells and HepG2 cells). The production of reactive oxygen species (ROS) after the C60-SMEDDS/TP treatment was assessed using 2',7'-dichlorofluorescein diacetate (DCFDA) staining. The alterations in mitochondrial membrane potential (ΔψM) were assessed by measuring JC-1 fluorescence. CONCLUSIONS: The cytoprotection provided by the C60-SMEDDS/TP favored normal cells (L02 and GES-1) over tumor cells (BEL-7402 and HepG2 cells) in vitro. This suggests a promising approach for the safe and effective treatment of TP.

Norcantharidin-Encapsulated C60-Modified Nanomicelles: A Potential Approach to Mitigate Cytotoxicity in Renal Cells and Simultaneously Enhance Anti-Tumor Activity in Hepatocellular Carcinoma Cells.

OBJECTIVE: The objective of this study was to examine the preparation process of DSPE-PEG-C60/NCTD micelles and assess the impact of fullerenol (C60)-modified micelles on the nephrotoxicity and antitumor activity of NCTD. METHOD: The micelles containing NCTD were prepared using the ultrasonic method and subsequently optimized and characterized. The cytotoxicity of micelles loaded with NCTD was assessed using the CCK-8 method on human hepatoma cell lines HepG2 and BEL-7402, as well as normal cell lines HK-2 and L02. Acridine orange/ethidium bromide (AO/EB) double staining and flow cytometry were employed to assess the impact of NCTD-loaded micelles on the apoptosis of the HK-2 cells and the HepG2 cells. Additionally, JC-1 fluorescence was utilized to quantify the alterations in mitochondrial membrane potential. The generation of reactive oxygen species (ROS) following micelle treatment was determined through 2',7'-dichlorofluorescein diacetate (DCFDA) staining. RESULTS: The particle size distribution of the DSPE-PEG-C60/NCTD micelles was determined to be 91.57 nm (PDI = 0.231). The zeta potential of the micelles was found to be -13.8 mV. The encapsulation efficiency was measured to be 91.9%. The in vitro release behavior of the micelles followed the Higuchi equation. Cellular experiments demonstrated a notable decrease in the toxicity of the C60-modified micelles against the HK-2 cells, accompanied by an augmented inhibitory effect on cancer cells. Compared to the free NCTD group, the DSPE-PEG-C60 micelles exhibited a decreased apoptosis rate (12%) for the HK-2 cell line, lower than the apoptosis rate observed in the NCTD group (36%) at an NCTD concentration of 75 µM. The rate of apoptosis in the HepG2 cells exhibited a significant increase (49%), surpassing the apoptosis rate observed in the NCTD group (24%) at a concentration of 150 µM NCTD. The HK-2 cells exhibited a reduction in intracellular ROS and an increase in mitochondrial membrane potential (ΔψM) upon exposure to C60-modified micelles compared to the NCTD group. CONCLUSIONS: The DSPE-PEG-C60/NCTD micelles, as prepared in this study, demonstrated the ability to decrease cytotoxicity and ROS levels in normal renal cells (HK-2) in vitro. Additionally, these micelles showed an enhanced antitumor activity against human hepatocellular carcinoma cells (HepG2, BEL-7402).

[Percutaneous permeability of rhein at shenque acupoint in vitro].

To study rhein's permeative properties of acupoint and non-acupoint and different species' transdermal administration in vitro. Cumulative permeation amount and steady-state infiltration rate were taken as evaluative indexes to assess the permeability difference. The Valia-Chien diffusion cell method was used to conduct the permeability test, with fresh acupoint and non-acupoint skin of rat, rabbit and swine in vitro as permeation barriers, and blank 20% EtOH saline as absorption liquid. HPLC was used to determine the rhein. The absorption difference was observed by confocal laser scanning microscopy. The 24-hour cumulative permeation amount through acupoint skin in rats was (102.63±9.60) µg•cm⁻², the steady-state infiltration rate was 4.307 µg•cm⁻²â€¢h⁻¹, both were higher than that through non-acupoint skin. The thickness of acupoint skin in rat was thinner than that in rabbit and swine. The cumulative permeation amount and steady-state infiltration rate of rhein in acupoint of rat were signally higher than those in rabbit and swine. The absorption difference can be clearly observed through an accumulation of fluorescence. In conclusion, species and acupoint all affect the permeability of rhein in vitro. The permeation amount and rate of rhein on Shenque acupoint were better than that on non-acupoint skin, which could verify that treatment through Shenque acupoint is superior to that through non-acupoint. The preliminary mechanism may be the drug delivery through Shenque acupoint as a channel and carrier, which is a visual verification the specificity and superiority of clinical application through Shenque acupoint in treating diseases.

Combined immunochemotherapy achieving targeted co-delivery of chlorogenic acid and doxorubicin by sialic acid-modified liposomes enhances anti-cancer efficacy.

Malignant melanoma is a high-grade aggressive skin tumor with an increasing incidence and mortality rates worldwide. Chemotherapeutic drugs such as doxorubicin have limited efficacy against melanoma due to their poor sensitivity, severe side effects, and drug resistance. Recent studies have shown that combinations of immunotherapy and chemotherapy have a synergistic effect in enhancing the anti-tumor effect. Here, we have developed liposomes co-loaded with chlorogenic acid (CA) and doxorubicin (DOX), modified with sialic acid-octadecylamine conjugate (SA-ODA), designated CA-DOX-SAL, that facilitate drug delivery by recognizing Siglec-1 receptor on TAMs. The physicochemical studies revealed the particle size and zeta potential of CA-DOX-SAL as 128.3 ± 0.8 nm and - 4.33 ± 0.50 mV, respectively. In vitro, CA-DOX-SAL demonstrated robust cellular uptake through SA receptor-mediated tumor-associated macrophages (TAM) targeting and exerted greater cytotoxicity on tumor cells. In vivo, targeted liposomes were found to accumulate in the tumor area, leading to an improvement in anti-tumor efficacy. In addition, CA-DOX-SAL effectively inhibited B16F10 melanoma tumor growth by stimulating the transition from tumor-promoting M2-type to anti-tumor M1-type and directly killing tumor cells. Overall, the co-delivery of immunomodulatory CA and chemotherapeutic DOX presents a promising therapeutic strategy to enhance clinical outcomes in the treatment of melanoma.

The effect of biofeedback training on patients with functional constipation.

The aim of this prospective quasi-experimental study was to explore the influence of biofeedback training on patients with functional constipation (FC). Changes in clinical symptoms, psychological status, quality of life, and autonomic nervous function in 21 FC patients before and after biofeedback training were investigated. The psychological status and quality of life were evaluated with the Zung Self-Rating Anxiety Scale (SAS), Zung Self-Rating Depression Scale (SDS), and a Chinese version of the MOS 36-Item Short-Form Health Survey. Autonomic nervous function was assessed on the basis of heart rate variability recorded with a HANS-1000 autonomic nervous biofeedback apparatus. After a complete course of training (10 sessions), clinical symptoms were greatly improved (p < .01), and the SAS and SDS scores were markedly decreased. There was a significant difference in the SAS and SDS scores before and after biofeedback (p < .01). The scores of general health perceptions, physical functioning, emotional role functioning, bodily pain, and vitality were increased significantly (p < .05), especially the scores of general health perceptions and emotional role functioning (p < .01), which indicated that quality of life in FC patients was significantly improved. No marked improvement of autonomic nervous function was found. Although a slight improvement in autonomic nervous activity was found, there was no significant statistical findings (p > .05). We conclude that biofeedback training can improve clinical symptoms, psychological status, and quality of life in FC patients, but further research is needed to determine whether biofeedback training can improve the autonomic nervous function in FC patients.

Preparation, Characterization, and Evaluation of Breviscapine Nanosuspension and Its Freeze-Dried Powder.

As a biopharmaceutics classification system (BCS) class IV drug, breviscapine (Bre) has low solubility in water, poor chemical stability, a short biological half-life and rapid removal from plasma. This paper prepared a Bre nanosuspension (Bre-NS) by an ultrasound-assisted anti-solvent precipitation method. Characterization of Bre-NS was studied using a Box-Behnken design concerning drug concentration in DMSO, an anti-solvent-to-solvent ratio, and sonication time. Under the optimized conditions of 170 mg/mL for the drug concentration, a 1:60 solvent-to-anti-solvent ratio, and a 9 min sonication time, the particle size of Bre-NS was 303.7 ± 7.3 nm, the polydispersity index was 0.178 ± 0.015, and the zeta potential was -31.10 ± 0.26 mV. Combined with the results from differential scanning calorimetry (DSC), powder X-ray diffraction (PXRD), and Fourier transform-infrared spectroscopy (FT-IR), the findings indicated that the crystal form and chemical structure of Bre-NS did not change during the entire process. The optimized formulation displayed good stability, increased solubility, and better in vitro release. Therefore, the results of this study can be a reference for the delivery system design of insoluble active components and effective parts in traditional Chinese medicine.

Recent Advances of D-α-tocopherol Polyethylene Glycol 1000 Succinate Based Stimuli-responsive Nanomedicine for Cancer Treatment.

D-α-tocopherol polyethylene glycol 1000 succinate (TPGS) is a pharmaceutical excipient approved by Chinese NMPA and FDA of USA. It's widely applied as a multifunctional drug carrier for nanomedicine. The advantages of TPGS include P-glycoprotein (P-gp) inhibition, penetration promotion, apoptosis induction via mitochondrial-associated apoptotic pathways, multidrug resistant (MDR) reversion, metastasis inhibition and so on. TPGS-based drug delivery systems which are responding to external stimulus can combine the inhibitory functions of TPGS towards P-gp with the environmentally responsive controlled release property and thus exerts a synergistic anti-cancer effect, through increased intracellular drug concentration in tumors cells and well-controlled drug release behavior. In this review, TPGS-based nano-sized delivery systems responsive to different stimuli were summarized and discussed, including pH-responsive, redoxresponsive and multi-responsive systems in various formulations. The achievements, mechanisms and different characteristics of TPGS-based stimuli-responsive drug-delivery systems in tumor therapy were also outlined.