Protective effects of silymarin in glioblastoma cancer cells through redox system regulation.

BACKGROUND: Glioblastoma multiforme, a deadly form of brain tumor, is characterized by aggressive growth and poor prognosis. Oxidative stress, a disruption in the balance between antioxidants and oxidants, is a crucial factor in its pathogenesis. Silymarin, a flavonoid extracted from milk thistle, has shown therapeutic potential in inhibiting cancer cell growth, promoting apoptosis, and reducing inflammation. It also regulates oxidative stress. This study aims to investigate the regulatory effects of silymarin on oxidative stress parameters, especially the transcription factor Nrf2 and its related enzymes in GBM cancer cells, to develop a new anti-cancer compound with low toxicity. METHODS AND RESULTS: First, the cytotoxicity of silymarin on U-87 MG cells was investigated by MTT and the results showed an IC50 of 264.6 µM. Then, some parameters of the redox system were measured with commercial kits, and the obtained results showed that silymarin increased the activity of catalase and superoxide dismutase enzymes, as well as the total antioxidant capacity levels; while the malondialdehyde level that is an indicator of lipid peroxidation was decreased by this compound. The expression level of Nrf2 and HO-1 and glutaredoxin and thioredoxin enzymes were checked by real-time PCR method, and the expression level increased significantly after treatment. CONCLUSIONS: Our findings suggest that silymarin may exert its cytotoxic and anticancer effects by enhancing the Nrf2/HO-1 pathway through antioxidant mechanisms in U-87 MG cells.

Therapeutic Potential of Silymarin in Mitigating Paclitaxel-Induced Hepatotoxicity and Nephrotoxicity: Insights into Oxidative Stress, Inflammation, and Apoptosis in Rats.

Background: Paclitaxel (PAX) is a widely used chemotherapy drug for various cancer types but often induces significant toxicity in multiple organ systems. Silymarin (SIL), a natural flavonoid, has shown therapeutic potential due to its multiple benefits. Aims: To evaluate the therapeutic efficacy of SIL in mitigating liver and kidney damage induced by PAX in rats, focusing on oxidative stress, inflammation, and apoptosis pathways. Study Design: Experimental animal model. Methods: The study included 28 male Wistar rats aged 12-14 weeks weighing 270-300 g. The rats were divided into four groups: control, SIL, PAX, and PAX + SIL, with seven in each group. The rats received intraperitoneal (i.p.) injections at a dose of 2 mg per kilogram of body weight of PAX for 5 successive days, followed by oral gavage with 200 mg/kg body mass of SIL for 10 uninterrupted days. We examined the effect of SIL on specific serum biochemical parameters using an autoanalyzer and rat-specific kits. The spectrophotometric methods was used to investigate oxidative stress indicators in kidney and liver tissues. Aquaporin-2 (AQP-2), B-cell lymphoma-2 (Bcl-2), cysteine aspartate-specific protease-3 (caspase-3), interleukin-6 (IL-6), nuclear factor kappa B (NF-κB), and streptavidin-biotin staining were used to assess immunoreactivity in PAX-induced liver and kidney injury models. Results: SIL treatment significantly reduced serum levels of alanine aminotransferase, aspartate aminotransferase, creatinine, urea, and C-reactive protein, indicating its effectiveness in treating PAX-induced liver and kidney injury. SIL treatment significantly reduced oxidative stress by increasing essential antioxidant parameters, such as superoxide dismutase, catalase, glutathione peroxidase, and glutathione. It also reduced malondialdehyde levels in liver and kidney tissues of SIL-PAX groups (p < 0.05). SIL administration reduced NF-κB, caspase-3, and IL-6 expression while increasing Bcl-2 and AQP2 levels in liver and kidney tissues of rats treated with SIL and PAX (p < 0.05). Conclusion: Our findings indicate the potential of SIL to alleviate PAX-induced liver and kidney damage in rats by reducing oxidative stress, inflammation, and apoptotic processes.

Formulation design and characterization of silymarin liposomes for enhanced antitumor activity.

Liposomes, a nanoscale carrier, plays an important role in the delivery of drug, affects the in vivo efficacy of drugs. In this paper, silymarin(SM)-loaded liposomes was optimized using the response surface method (RSM), with entrapment efficiency (EE%) as an index. The formulation was optimized as follow: lecithin (7.8mg/mL), SM/lecithin (1/26) and lecithin/cholesterol (10/1). The optimized SM liposomes had a high EE (96.58 ±3.06%), with a particle size of 290.3 ±10.5nm and a zeta potential of +22.98 ±1.73mV. In vitro release tests revealed that SM was released in a sustained-release manner, primarily via diffusion mechanism. In vitro cytotoxicity studies demonstrated that the prepared SM liposomes had stronger inhibitory effects than the model drug. Overall, these results indicate that this liposome system is suitable for intravenous delivery to enhance the antitumor effects of SM.

Nutraceutical blends predict enhanced health via microbiota reshaping improving cytokines and life quality: a Brazilian double-blind randomized trial.

Nutraceutical interventions supporting microbiota and eliciting clinical improvements in metabolic diseases have grown significantly. Chronic stress, gut dysbiosis, and metainflammation have emerged as key factors intertwined with sleep disorders, consequently exacerbating the decline in quality of life. This study aimed to assess the effects of two nutraceutical formulations containing prebiotics (fructooligosaccharides (FOS), galactooligosaccharides (GOS), yeast ß-glucans), minerals (Mg, Se, Zn), and the herbal medicine Silybum marianum L. Gaertn., Asteraceae (Milk thistle or Silymarin). These formulations, namely NSupple (without silymarin) and NSupple_Silybum (with silymarin) were tested over 180 days in overweight/obese volunteers from Brazil's southeastern region. We accessed fecal gut microbiota by partial 16S rRNA sequences; cytokines expression by CBA; anthropometrics, quality of life and sleep, as well as metabolic and hormonal parameters, at baseline (T0) and 180 days (T180) post-supplementation. Results demonstrated gut microbiota reshaping at phyla, genera, and species level post-supplementation. The Bacteroidetes phylum, Bacteroides, and Prevotella genera were positively modulated especially in the NSupple_Silybum group. Gut microbiota modulation was associated with improved sleep patterns, quality-of-life perception, cytokines expression, and anthropometric parameters post-supplementation. Our findings suggest that the nutraceutical blends positively enhance cardiometabolic and inflammatory markers. Particularly, NSupple_Silybum modulated microbiota composition, underscoring its potential significance in ameliorating metabolic dysregulation. Clinical trial registry number: NCT04810572. 23/03/2021.

The importance of integrated therapies on cancer: Silibinin, an old and new molecule.

In the landscape of cancer treatments, the efficacy of coadjuvant molecules remains a focus of attention for clinical research with the aim of reducing toxicity and achieving better outcomes. Most of the pathogenetic processes causing tumour development, neoplastic progression, ageing, and increased toxicity involve inflammation. Inflammatory mechanisms can progress through a variety of molecular patterns. As is well known, the ageing process is determined by pathological pathways very similar and often parallel to those that cause cancer development. Among these complex mechanisms, inflammation is currently much studied and is often referred to in the geriatric field as 'inflammaging'. In this context, treatments active in the management of inflammatory mechanisms could play a role as adjuvants to standard therapies. Among these emerging molecules, Silibinin has demonstrated its anti-inflammatory properties in different neoplastic types, also in combination with chemotherapeutic agents. Moreover, this molecule could represent a breakthrough in the management of age-related processes. Thus, Silibinin could be a valuable adjuvant to reduce drug-related toxicity and increase therapeutic potential. For this reason, the main aim of this review is to collect and analyse data presented in the literature on the use of Silibinin, to better understand the mechanisms of the functioning of this molecule and its possible therapeutic role.

Administration of silymarin in NAFLD/NASH: A systematic review and meta-analysis.

INTRODUCTION AND OBJECTIVES: Nonalcoholic fatty liver disease (NAFLD) is a chronic liver disease with a high prevalence worldwide and poses serious harm to human health. There is growing evidence suggesting that the administration of specific supplements or nutrients may slow NAFLD progression. Silymarin is a hepatoprotective extract of milk thistle, but its efficacy in NAFLD remains unclear. MATERIALS AND METHODS: Relevant studies were searched in PubMed, Embase, the Cochrane Library, Web of Science, clinicaltrails.gov, and China National Knowledge Infrastructure and were screened according to the eligibility criteria. Data were analyzed using Revman 5.3. Continuous values and dichotomous values were pooled using the standard mean difference (SMD) and odds ratio (OR). Heterogeneity was evaluated using the Cochran's Q test (I2 statistic). A P<0.05 was considered statistically significant. RESULTS: A total of 26 randomized controlled trials involving 2,375 patients were included in this study. Administration of silymarin significantly reduced the levels of TC (SMD[95%CI]=-0.85[-1.23, -0.47]), TG (SMD[95%CI]=-0.62[-1.14, -0.10]), LDL-C (SMD[95%CI]=-0.81[-1.31, -0.31]), FI (SMD[95%CI]=-0.59[-0.91, -0.28]) and HOMA-IR (SMD[95%CI]=-0.37[-0.77, 0.04]), and increased the level of HDL-C (SMD[95%CI]=0.46[0.03, 0.89]). In addition, silymarin attenuated liver injury as indicated by the decreased levels of ALT (SMD[95%CI]=-12.39[-19.69, -5.08]) and AST (SMD[95% CI]=-10.97[-15.51, -6.43]). The levels of fatty liver index (SMD[95%CI]=-6.64[-10.59, -2.69]) and fatty liver score (SMD[95%CI]=-0.51[-0.69, -0.33]) were also decreased. Liver histology of the intervention group revealed significantly improved hepatic steatosis (OR[95%CI]=3.25[1.80, 5.87]). CONCLUSIONS: Silymarin can regulate energy metabolism, attenuate liver damage, and improve liver histology in NAFLD patients. However, the effects of silymarin will need to be confirmed by further research.

Supplementation of Silymarin Alone or in Combination with Salvianolic Acids B and Puerarin Regulates Gut Microbiota and Its Metabolism to Improve High-Fat Diet-Induced NAFLD in Mice.

Silymarin, salvianolic acids B, and puerarin were considered healthy food agents with tremendous potential to ameliorate non-alcoholic fatty liver disease (NAFLD). However, the mechanisms by which they interact with gut microbiota to exert benefits are largely unknown. After 8 weeks of NAFLD modeling, C57BL/6J mice were randomly divided into five groups and fed a normal diet, high-fat diet (HFD), or HFD supplemented with a medium or high dose of Silybum marianum extract contained silymarin or polyherbal extract contained silymarin, salvianolic acids B, and puerarin for 16 weeks, respectively. The untargeted metabolomics and 16S rRNA sequencing were used for molecular mechanisms exploration. The intervention of silymarin and polyherbal extract significantly improved liver steatosis and recovered liver function in the mice, accompanied by an increase in probiotics like Akkermansia and Blautia, and suppressed Clostridium, which related to changes in the bile acids profile in feces and serum. Fecal microbiome transplantation confirmed that this alteration of microbiota and its metabolites were responsible for the improvement in NAFLD. The present study substantiated that alterations of the gut microbiota upon silymarin and polyherbal extract intervention have beneficial effects on HFD-induced hepatic steatosis and suggested the pivotal role of gut microbiota and its metabolites in the amelioration of NAFLD.

A Silymarin Antioxidant Serum Improves Facial Acne Alone and as Part of a Treatment Regimen.

BACKGROUND: Silymarin is an antioxidant that can protect against free radicals that cause premature signs of aging and oil oxidation that may contribute to breakouts. AIMS: The objective of these studies was to evaluate a silymarin antioxidant serum alone and in combination with a prescription acne treatment regimen in improving facial appearance in blemish-prone skin.  Methods: Two international studies were conducted. A 12-week study in Brazil enrolled 56 subjects to examine the effect of silymarin antioxidant serum on facial acne. Clinical grading on acne lesions, skin tone, clarity, and postinflammatory hyperpigmentation (PIH) were conducted. In addition, consumer self-assessment, analysis for markers of lipid peroxidation, and sebumeter analysis were completed. Another Unites States (US)/German study enrolled 40 subjects who were on topical prescription acne medications to which silymarin antioxidant serum was added. Acne lesion counts, tolerability, and facial appearance assessments were conducted in this study. RESULTS: The Brazilian study demonstrated a 45% reduction in inflammatory lesions and a 43% reduction in noninflammatory lesions after 12 weeks of silymarin antioxidant serum use. In addition, sebumeter testing showed a 16% reduction in oiliness at week 1. The US/German study showed the benefits of the serum in persons already on prescription acne therapy by reducing facial erythema by 60%, dryness by 49%, and scaling by 67%. CONCLUSION: Silymarin is shown in clinical testing to have significant benefits in reducing lipid peroxidation, oiliness, and PIH, and in improving key markers of skin aging. Additionally, the serum can be used alone or as an adjunctive treatment in acne therapy to further benefit aging, acne-prone skin. J Drugs Dermatol. 2024;23(4):     doi:10.36849/JDD.8120.

Inhibition of YAP/TAZ pathway contributes to the cytotoxicity of silibinin in MCF-7 and MDA-MB-231 human breast cancer cells.

Breast cancer is one of the most common cancers threatening women's health. Our previous study found that silibinin induced the death of MCF-7 and MDA-MB-231 human breast cancer cells. We noticed that silibinin-induced cell damage was accompanied by morphological changes, including the increased cell aspect ratio (cell length/width) and decreased cell area. Besides, the cytoskeleton is also destroyed in cells treated with silibinin. YAP/TAZ, a mechanical signal sensor interacted with extracellular pressure, cell adhesion area and cytoskeleton, is also closely associated with cell survival, proliferation and migration. Thus, the involvement of YAP/TAZ in the cytotoxicity of silibinin in breast cancer cells has attracted our interests. Excitingly, we find that silibinin inhibits the nuclear translocation of YAP/TAZ in MCF-7 and MDA-MB-231 cells, and reduces the mRNA expressions of YAP/TAZ target genes, ACVR1, MnSOD and ANKRD. More importantly, expression of YAP1 gene is negatively correlated with the survival of the patients with breast cancers. Molecular docking analysis reveals high probabilities for binding of silibinin to the proteins in the YAP pathways. DARTS and CETSA results confirm the binding abilities of silibinin to YAP and LATS. Inhibiting YAP pathway either by addition of verteporfin, an inhibitor of YAP/TAZ-TEAD, or by transfection of si-RNAs targeting YAP or TAZ further enhances silibinin-induced cell damage. While enhancing YAP activity by silencing LATS1/2 or overexpressing YAPS127/397A, an active form of YAP, attenuates silibinin-induced cell damage. These findings demonstrate that inhibition of the YAP/TAZ pathway contributes to cytotoxicity of silibinin in breast cancers, shedding lights on YAP/TAZ-targeted cancer therapies.

Silibinin solubilization: combined effect of co-solvency and inclusion complex formation.

OBJECTIVE: Belonging to the class II drugs according to the biopharmaceutics classification system, silibinin (SLB) benefits from high permeability but suffers poor solubility that negatively affects the development of any delivery system. This research aimed to improve SLB solubility by combined use of co-solvency and complexation phenomena. METHODS: Solubility studies were performed using the phase solubility analysis according to the shake-flask method in the presence of ethanol and 2-hydroxypropyl-ß-cyclodextrin (HP-ß-CD) as a co-solvent and inclusion complexing agent, respectively. SLB release studies from chitosan nanoparticles were carried out in double-wall, diffusion cells using the optimized drug release medium. RESULTS: SLB solubility was mathematically optimized constraining to using the lowest concentrations of ethanol and HP-ß-CD. SLB solubility increased linearly with the increase of HP-ß-CD concentration. The solubility in PBS-ethanol mixtures followed a log-linear model. SLB solubility in the presence of the ethanol co-solvent and HP-ß-CD complexing agent was optimized by adopting a genetic algorithm suggesting the phosphate buffer saline solution supplemented by 6%v/v ethanol and 8 mM HP-ß-CD as an optimized medium. The optimized solution was examined to study SLB release from chitosan nanoparticles (4.5 ± 0.2% drug loading) at 37 °C under static conditions. The sigmoidal release profile of SLB from the particles indicated a combination of erosion and diffusion mechanisms governing drug release from the nanoparticles. CONCLUSION: SLB solubility in a buffered solution supplemented by ethanol co-solvent and HP-ß-CD complexing agent is a function of free drug present in the semi-aqueous media, the drug-ligand binary complex, and the drug/ligand/co-solvent ternary complex.

Silymarin and MSC-exosomes ameliorate thioacetamide-evoked renal fibrosis by inhibiting TGF-ß/SMAD pathway in rats.

BACKGROUND: TGF-ß1 and SMAD3 are particularly pathogenic in the progression of renal fibrosis. AIM: This study aimed to evaluate the kidney protective potentials of silymarin (SM) and exosomes of mesenchymal stem cells against the nephrotoxin thioacetamide (TAA) in rats. METHODS: 32 female rats were randomly assigned into four groups: the control group, the TAA group, the TAA + SM group, and the TAA + Exosomes group. The kidney homogenates from all groups were examined for expression levels of TGF-ß receptors I and II using real-time PCR, expression levels of collagen type I and CTGF proteins using ELISA, and the expression levels of nuclear SMAD2/3/4, cytoplasmic SMAD2/3, and cytoplasmic SMAD4 proteins using the western blot technique. RESULTS: Compared to the control group, the injection of TAA resulted in a significant increase in serum levels of urea and creatinine, gene expression levels of TßRI and TßRII, protein expression levels of both collagen I and CTGF proteins, cytoplasmic SMAD2/3 complex, and nuclear SMAD2/3/4 (p-value < 0.0001), with significantly decreased levels of the co-SMAD partner, SMAD4 (p-value < 0.0001). Those effects were reversed considerably in both treatment groups, with the superiority of the exosomal treatment regarding the SMAD proteins and the expression levels of the TßRI gene, collagen I, and CTGF proteins returning to near-control values (p-value > 0.05). CONCLUSION: Using in vitro and in vivo experimental approaches, the research discovered a reno-protective role of silymarin and exosomes of BM-MSCs after thioacetamide-induced renal fibrosis in rats, with the advantage of exosomes.

Synthesis, physiochemical characterization, molecular docking study, and anti-breast cancer activity of silymarin loaded zein nanoparticles.

Breast cancer is a major cause of death in women worldwide leading to requirement of new therapeutic strategies. Silymarin demonstrated the anti-cancer activity however, due to low bioavailability its use is restricted. This study aimed to improve the solubility of silymarin by developing a silymarin loaded zein nanoparticles (SLNPs) which was stabilized by beta cyclodextrin. Comprehensive physiochemical characterization studies based on DLS, FTIR, UV-Vis Spectroscopy, FE-SEM, TEM, XRD, DSC, NMR and TGA confirmed the successful synthesis of SLNPs via an anti-solvent precipitation method. FE-SEM and TEM images demonstrated the uniform size and spherical shape of nanoparticles with encapsulation and loading efficiencies of 84.32 ± 1.9 % and 15.25 ± 2.4 % respectively. The zein protein interaction with silymarin, and ß-cyclodextrin was shown to be beneficial via the use of molecular simulations and binding energy calculations. Cellular studies demonstrated dose and time dependent cytotoxicity of SLNPs on MCF-7 breast cancer cell. FACS, qRT-PCR and Western blotting showed Bax (pro-apoptotic) upregulation while Bcl-2 (anti-apoptotic) downregulation. Our findings suggest that these loaded nanoparticles are more efficient than pure drug, enhancing its bioavailability and paving the path for developing it as a promising nutraceutical to treat breast cancer.

Camel milk or silymarin could improve the negative effects that experimentally produced by aflatoxin B1 on rat's male reproductive system.

BACKGROUND: Camel milk and silymarin have many different beneficial effects on several animal species. Meanwhile, Aflatoxins are mycotoxins with extraordinary potency that pose major health risks to several animal species. Additionally, it has been documented that aflatoxins harm the reproductive systems of a variety of domestic animals. The present design aimed to investigate the impact of aflatoxin B1 (AFB1) on rat body weight and reproductive organs and the ameliorative effects of camel milk and silymarin through measured serum testosterone, testes pathology, and gene expression of tumor necrosis factor (TNF-α), luteinizing hormone receptor (LHR), and steroidogenic acute regulatory protein (StAR) in the testes. A total of sixty mature male Wister white rats, each weighing an average of 83.67 ± 0.21 g, were used. There were six groups created from the rats. Each division had ten rats. The groups were the control (without any treatment), CM (1 ml of camel milk/kg body weight orally), S (20 mg silymarin/kg b. wt. suspension, orally), A (1.4 mg aflatoxin/kg diet), ACM (aflatoxin plus camel milk), and AS (aflatoxin plus silymarin). RESULTS: The results indicated the positive effects of camel milk and silymarin on growth, reproductive organs, and gene expression of TNF-α, LHR, and StAR with normal testicular architecture. Also, the negative effect of AFB1 on the rat's body weight and reproductive organs, as indicated by low body weight and testosterone concentration, was confirmed by the results of histopathology and gene expression. However, these negative effects were ameliorated by the ingestion of camel milk and silymarin. CONCLUSION: In conclusion, camel milk and silymarin could mitigate the negative effect of AFB1 on rat body weight and reproductive organs.

Synergistic effects of citicoline and silymarin nanomicelles in restraint stress-exposed mice.

This study evaluated the effects of citicoline and silymarin nanomicelles (SMnm) in repeated restraint stress (RRS). METHOD: Mice were exposed to RRS for four consecutive days, 2 hrs. daily. On day 5 of the study, SMnm (25 and 50 mg/kg, i.p.) and citicoline (25 and 75 mg/kg), and a combination of them (25 mg/kg, i.p.) were initiated. On day 18, anxiety-like behavior, behavioral despair, and exploratory behavior were evaluated. The prefrontal cortex (PFC) and the hippocampus were dissected measuring brain-derived neurotrophic factor (BDNF), cAMP response element-binding protein (CREB), and tumor necrosis factor-alpha (TNF-α) through Western Blot and ELISA, respectively. RESULTS: In RR-exposed mice, anxiety-like behavior in the elevated plus maze (EPM) was enhanced by reductions in open arm time (OAT%) P < 0.001, and open arm entry (OAE%) P < 0.001. In the forced swimming test (FST), the immobility increased P < 0.001 while the swimming and climbing reduced P < 0.001. In the open field test (OFT), general motor activity was raised P < 0.05. Further, body weights reduced P < 0.001, and tissue BDNF and pCREB expressions decreased P < 0.001 while TNF-α increased P < 0.001. Conversely, SMnm, citicoline and their combination could reduce anxiety-like behavior P < 0.001. The combination group reduced the depressive-like behaviors P < 0.001. Moreover, body weights were restored P < 0.001. Besides, BDNF and pCREB expressions increased while TNF-α reduced, P < 0.001. CONCLUSION: The combination synergistically improved emotion-like behaviors, alleviating the inflammation and upregulating the hippocampal BDNF-mediated CREB signaling pathway.

Effects of silymarin supplementation on liver and kidney functions: A systematic review and dose-response meta-analysis.

It is suggested that supplementation with silymarin (SIL) has beneficial impacts on kidney and liver functions. This systematic review and dose-response meta-analysis assessed the impact of SIL administration on certain hepatic, renal, and oxidative stress markers. A systematic search was conducted in various databases to identify relevant trials published until January 2023. Randomized controlled trials (RCTs) that evaluated the effects of SIL on kidney and liver markers were included. A random-effects model was used for the analysis and 41 RCTs were included. The pooled results indicated that SIL supplementation led to a significant reduction in serum levels of alkaline phosphatase, alanine transaminase, creatinine, and aspartate aminotransferase, along with a substantial elevation in serum glutathione in the SIL-treated group compared to their untreated counterparts. In addition, there was a nonsignificant decrease in serum levels of gamma-glutamyl transferase, malondialdehyde (MDA), total bilirubin, albumin (Alb), total antioxidant capacity, and blood urea nitrogen. Sub-group analyses revealed a considerable decline in MDA and Alb serum values among SIL-treated participants with liver disease in trials with a longer duration (≥12 weeks). These findings suggest that SIL may ameliorate certain liver markers with potential hepatoprotective effects, specifically with long-term and high-dose supplementation. However, its nephroprotective effects and impact on oxidative stress markers were not observed. Additional high-quality RCTs with longer durations are required to determine the clinical efficacy of SIL supplementation on renal and oxidative stress markers.

Silibinin is a suppressor of the metastasis-promoting transcription factor ID3.

BACKGROUND: ID3 (inhibitor of DNA binding/differentiation-3) is a transcription factor that enables metastasis by promoting stem cell-like properties in endothelial and tumor cells. The milk thistle flavonolignan silibinin is a phytochemical with anti-metastatic potential through largely unknown mechanisms. HYPOTHESIS/PURPOSE: We have mechanistically investigated the ability of silibinin to inhibit the aberrant activation of ID3 in brain endothelium and non-small cell lung cancer (NSCLC) models. METHODS: Bioinformatic analyses were performed to investigate the co-expression correlation between ID3 and bone morphogenic protein (BMP) ligands/BMP receptors (BMPRs) genes in NSCLC patient datasets. ID3 expression was assessed by immunoblotting and qRT-PCR. Luciferase reporter assays were used to evaluate the gene sequences targeted by silibinin to regulate ID3 transcription. In silico computational modeling and LanthaScreen TR-FRET kinase assays were used to characterize and validate the BMPR inhibitory activity of silibinin. Tumor tissues from NSCLC xenograft models treated with oral silibinin were used to evaluate the in vivo anti-ID3 effects of silibinin. RESULTS: Analysis of lung cancer patient datasets revealed a top-ranked positive association of ID3 with the BMP9 endothelial receptor ACVRL1/ALK1 and the BMP ligand BMP6. Silibinin treatment blocked the BMP9-induced activation of the ALK1-phospho-SMAD1/5-ID3 axis in brain endothelial cells. Constitutive, acquired, and adaptive expression of ID3 in NSCLC cells were all significantly downregulated in response to silibinin. Silibinin blocked ID3 transcription via BMP-responsive elements in ID3 gene enhancers. Silibinin inhibited the kinase activities of BMPRs in the micromolar range, with the lower IC50 values occurring against ACVRL1/ALK1 and BMPR2. In an in vivo NSCLC xenograft model, tumoral overexpression of ID3 was completely suppressed by systematically achievable oral doses of silibinin. CONCLUSIONS: ID3 is a largely undruggable metastasis-promoting transcription factor. Silibinin is a novel suppressor of ID3 that may be explored as a novel therapeutic approach to interfere with the metastatic dissemination capacity of NSCLC.

Pharmacokinetic Effects of Different Models of Nonalcoholic Fatty Liver Disease in Transgenic Humanized OATP1B Mice.

Organic anion transporting polypeptide (OATP) 1B1 and OATP1B3 (collectively, OATP1B) transporters encoded by the solute carrier organic anion transporter (SLCO) genes mediate uptake of multiple pharmaceutical compounds. Nonalcoholic steatohepatitis (NASH), a severe form of nonalcoholic fatty liver disease (NAFLD), decreases OATP1B abundance. This research characterized the pathologic and pharmacokinetics effects of three diet- and one chemical-induced NAFLD model in male and female humanized OATP1B mice, which comprises knock-out of rodent Oatp orthologs and insertion of human SLCO1B1 and SLCO1B3. Histopathology scoring demonstrated elevated steatosis and inflammation scores for all NAFLD-treatment groups. Female mice had minor changes in SLCO1B1 expression in two of the four NAFLD treatment groups, and pitavastatin (PIT) area under the concentration-time curve (AUC) increased in female mice in only one of the diet-induced models. OATP1B3 expression decreased in male and female mice in the chemical-induced NAFLD model, with a coinciding increase in PIT AUC, indicating the chemical-induced model may better replicate changes in OATP1B3 expression and OATP substrate disposition observed in NASH patients. This research also tested a reported multifactorial pharmacokinetic interaction between NAFLD and silymarin, an extract from milk thistle seeds with notable OATP-inhibitory effects. Males showed no change in PIT AUC, whereas female PIT AUC increased 1.55-fold from the diet alone and the 1.88-fold from the combination of diet with silymarin, suggesting that female mice are more sensitive to pharmacokinetic changes than male mice. Overall, the humanized OATP1B model should be used with caution for modeling NAFLD and multifactorial pharmacokinetic interactions. SIGNIFICANCE STATEMENT: Advanced stages of NAFLD cause decreased hepatic OATP1B abundance and increase systemic exposure to OATP substrates in human patients. The humanized OATP1B mouse strain may provide a clinically relevant model to recapitulate these observations and predict pharmacokinetic interactions in NAFLD. This research characterized three diet-induced and one drug-induced NAFLD model in a humanized OATP1B mouse model. Additionally, a multifactorial pharmacokinetic interaction was observed between silymarin and NAFLD.

Rifampin- and Silymarin-Mediated Pharmacokinetic Interactions of Exogenous and Endogenous Substrates in a Transgenic OATP1B Mouse Model.

Organic anion-transporting polypeptides (OATP) 1B1 and OATP1B3, encoded by the SLCO gene family of the solute carrier superfamily, are involved in the disposition of many exogenous and endogenous compounds. Preclinical rodent models help assess risks of pharmacokinetic interactions, but interspecies differences in transporter orthologs and expression limit direct clinical translation. An OATP1B transgenic mouse model comprising a rodent Slco1a/1b gene cluster knockout and human SLCO1B1 and SLCO1B3 gene insertions provides a potential physiologically relevant preclinical tool to predict pharmacokinetic interactions. Pharmacokinetics of exogenous probe substrates, pitavastatin and pravastatin, and endogenous OATP1B biomarkers, coproporphyrin-I and coproporphyrin-III, were determined in the presence and absence of known OATP/Oatp inhibitors, rifampin or silymarin (an extract of milk thistle [Silybum marianum]), in wild-type FVB mice and humanized OATP1B mice. Rifampin increased exposure of pitavastatin (4.6- and 2.8-fold), pravastatin (3.6- and 2.2-fold), and coproporphyrin-III (1.6- and 2.1-fold) in FVB and OATP1B mice, respectively, but increased coproporphyrin-I AUC0-24h only (1.8-fold) in the OATP1B mice. Silymarin did not significantly affect substrate AUC, likely because the silymarin flavonolignan concentrations were at or below their reported IC50 values for the relevant OATPs/Oatps. Silymarin increased the Cmax of pitavastatin 2.7-fold and pravastatin 1.9-fold in the OATP1B mice. The data of the OATP1B mice were similar to those of the pitavastatin and pravastatin clinical data; however, the FVB mice data more closely recapitulated pitavastatin clinical data than the data of the OATP1B mice, suggesting that the OATP1B mice are a reasonable, though costly, preclinical strain for predicting pharmacokinetic interactions when doses are optimized to achieve clinically relevant plasma concentrations.

Silymarin enriched gelatin methacrylamide bioink imparts hepatoprotectivity to 3D bioprinted liver construct against carbon tetrachloride induced toxicity.

Three-dimensional liver bioprinting is an emerging technology in the field of regenerative medicine that aids in the creation of functional tissue constructs that can be used as transplantable organ substitutes. During transplantation, the bioprinted donor liver must be protected from the oxidative stress environment created by various factors during the transplantation procedure, as well as from drug-induced damage from medications taken as part of the post-surgery medication regimen following the procedure. In this study, Silymarin, a flavonoid with the hepatoprotective properties were introduced into the GelMA bioink formulation to protect the bioprinted liver against hepatotoxicity. The concentration of silymarin to be added in GelMA was optimised, bioink properties were evaluated, and HepG2 cells were used to bioprint liver tissue. Carbon tetrachloride (CCl4) was used to induce hepatotoxicity in bioprinted liver, and the effect of this chemical on the metabolic activities of HepG2 cells was studied. The results showed that Silymarin helps with albumin synthesis and shields liver tissue from the damaging effects of CCl4. According to gene expression analysis, CCl4 treatment increased TNF-α and the antioxidant enzyme SOD expression in HepG2 cells while the presence of silymarin protected the bioprinted construct from CCl4-induced damage. Thus, the outcomes demonstrate that the addition of silymarin in GelMA formulation protects liver function in toxic environments.

A combination of silymarin and garlic extract enhances thyroid hormone activation and body metabolism in orally intoxicated male rats with atrazine: Impact on hepatic iodothyronine deiodinase type 1.

Atrazine is a widely applied herbicide to improve crop yield and maintain general health. It has been reported to impair thyroid function and architecture in experimental animals. Alterations in thyroid hormones disrupt normal body function and metabolism. Silymarin, a hepatoprotective flavonolignan, was found to improve thyroid function and body metabolism. Additionally, garlic displays several protective effects on body organs. Therefore, this study explored the prophylactic impact of natural compounds comprising silymarin and garlic extract on disrupted thyroid function, hepatic iodothyronine deiodinase type 1, and metabolic parameters in atrazine-intoxicated male rats. We found that daily pre- and co-treatment of atrazine-intoxicated male rats with silymarin (100 mg/kg, p.o) and/or garlic extract (10 mg/kg, p.o) significantly improved thyroid activation and hepatic functionality as evidenced by the re-establishment of T3, T3/T4, and TSH values as well as ALT and AST activities. Interestingly, individual or concurrent supplementation of the atrazine group with silymarin and garlic extract prevented the down-regulation in hepatic iodothyronine deiodinase type 1. These effects were coupled with the repletion of serum and hepatic antioxidants and the amelioration of lipid peroxidation. In addition, current natural products markedly alleviated weight gain, dyslipidemia, hyperglycemia, glucose intolerance, and insulin resistance. Notably, a cocktail of silymarin and garlic extract exerted superior protection against atrazine-triggered deterioration of thyroid, hepatic, and metabolic functioning to individual treatments. Present findings pinpoint the prophylactic and synergistic influence of silymarin and garlic extract combinatorial regimen on thyroid activation and body metabolism via enhancing antioxidant potential, maintaining hepatic function, and iodothyronine deiodinase type 1.