In Vitro Anti-Rotaviral Activity of Bavachin Isolated from Psoralea corylifolia L. (Fabaceae).

Rotavirus is the main causative agent of viral gastroenteritis among young animals worldwide. Currently, no clinically approved or effective antiviral drugs are available to combat rotavirus infections. Herein, we evaluated the anti-rotaviral activities of extracts and bavachin isolated from Psoralea corylifolia L. (Fabaceae) (P. corylifolia) against the bovine rotavirus G8P[7] and porcine rotavirus G5P[7] in vitro. Two assay strategies were performed: (1) a virucidal assay to reduce viral infectivity by virus neutralization and (2) a post-treatment assay to assess viral replication suppression. The results from the virucidal assay showed that the extracts and bavachin did not exert anti-rotaviral activities. In the follow-up analysis after treatment, bavachin exhibited robust antiviral efficacy, with 50% effective concentration (EC50) values of 10.6 µM (selectivity index [SI] = 2.38) against bovine rotavirus G8P[7] and 13.0 µM (SI = 1.94) against porcine rotavirus G5P[7]. Bavachin strongly suppressed viral RNA synthesis in the early (6 h) and late stages (18 h) after rotaviral infection. These findings strongly suggest that bavachin may have hindered the virions by effectively inhibiting the early stages of the virus replication cycle after rotaviral infection. Furthermore, confocal imaging showed that bavachin suppressed viral protein synthesis, notably that of the rotaviral protein (VP6). These results suggest that bavachin has strong antiviral activity against rotaviruses, inhibits viral replication, and is a candidate natural therapeutic drug targeting rotaviral infection. The utilization of bavachin isolated from P. corylifolia may contribute to decreased mortality rates, lower medication expenses, and enhanced economic viability in domestic farms.

[Pseudo-targeted metabolomics study of immune stress-mediated idiosyncratic liver injury induced by synergistic effects of bavachin and epimedin B].

Chinese patent medicine preparations containing Epimedii Folium and Psoraleae Fructus have been associated with the occurrence of idiosyncratic drug-induced liver injury(IDILI). However, the specific toxic biomarkers and mechanisms underlying these effects remain unclear. This study aimed to comprehensively assess the impact of bavachin and epimedin B, two principal consti-tuents found in Psoraleae Fructus and Epimedii Folium, on an IDILI model induced by tumor necrosis factor-α(TNF-α) treatment, both in vitro and in vivo. To evaluate the extent of liver injury, various parameters were assessed. Lactate dehydrogenase(LDH) release in the cell culture supernatant, as well as the levels of alanine aminotransferase(ALT) and aspartate transaminase(AST) in mouse plasma were measured. Additionally, histological analysis employing hematoxylin-eosin staining was performed to observe liver tissue changes indicative of the severity of liver injury. Furthermore, a pseudo-targeted metabolomics approach was employed, followed by multivariate analysis, to identify differential metabolites. These identified metabolites were subsequently subjected to Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment analysis. The results showed that at the cellular level, after 2 hours of TNF-α stimulation, bavachin significantly increased the release of LDH in HepG2 cells compared to the normal group and the group treated alone; after the combination of bavachin and epimedin B, the release of LDH further significantly increased on the original basis. Similarly, although the individual or combination treatments of bavachin and epimedin B did not induce liver injury in normal mice, the combination of both drugs induced marked liver injury in TNF-α treated mice, leading to a significant elevation in plasma AST and ALT levels and substantial infiltration of inflammatory immune cells in the liver tissue. Pseudo-targeted metabolomics analysis identified seven common differential metabolites. Among these, D-glucosamine-6-phosphate, N1-methyl-2-pyridone-5-carboxamide, 17beta-nitro-5a-androstane, irisolidone-7-O-glucuronide, and N-(1-deoxy-1-fructosyl) valine emerged as potential biomarkers, with an area under the curve(AUC) exceeding 0.9. Furthermore, our results suggest that the metabolism of nicotinic acid and nicotinamide, as well as the linoleic acid metabolic pathway, may play pivotal roles in bavachin and epimedin B-induced IDILI. In conclusion, within an immune-stressed environment mediated by TNF-α, bavachin and epimedin B appear to induce IDILI through disruptions in metabolic processes.

Bavachin Rejuvenates Sensitivity of Colistin against Colistin-Resistant Gram-Negative Bacteria.

The emergence of plasmid-mediated colistin resistance threatens the efficacy of colistin as a last-resort antibiotic used to treat infection caused by Gram-negative bacteria (GNB). Given the shortage of new antibiotics, the discovery of adjuvants to existing antibiotics is a promising strategy to combat infections caused by multidrug-resistant (MDR) GNB. This study was designed to investigate the potential synergistic antibacterial activity of bavachin, a bioactive compound extracted from the Psoralea Fructus, combined with colistin against MDR GNB. Herein, the synergistic efficacy in vitro and the therapeutic efficacy of colistin combined with bavachin in vivo were evaluated. The synergistic mechanism was detected by fluorescent probe and the transcript levels of mcr-1. Bavachin combined with colistin showed an excellent synergistic activity against GNB, as the FICI ≤ 0.5. In contrast to colistin alone, combination therapy dramatically increased the survival rate of Galleria mellonella and mice in vivo. Moreover, the combination of bavachin and colistin significantly reduced the amount of bacterial biofilm formation, improved the membrane disruption of colistin and inhibited mcr-1 transcription. These findings show that bavachin is a potential adjuvant of colistin, which may provide a new strategy to combat colistin-resistant bacteria infection with lower doses of colistin.

Bavachin ameliorates neuroinflammation and depressive-like behaviors in streptozotocin-induced diabetic mice through the inhibition of PKCδ.

Depression and diabetes are closely linked; however, the pathogenesis of depression associated with diabetes is unclear, and there are no clinically effective antidepressant drugs for diabetic patients with depression. Bavachin is an important active ingredient in Fructus Psoraleae. In this study, we evaluated the anti-neuroinflammatory and antidepressant effects associated with diabetes and the molecular mechanisms of bavachin in a streptozotocin-induced diabetes mouse model. We found that bavachin clearly decreased streptozotocin (STZ)-induced depressive-like behaviors in mice. It was further found that bavachin significantly inhibited microglia activation and the phosphorylation level of PKCδ and inhibited the activation of the NF-κB pathway in vivo and in vitro. Knockdown of PKCδ with siRNA-PKCδ partially reversed the inhibitory effect of bavachin on the NF-κB pathway and the level of pro-inflammatory factors. We further found that PKCδ directly bound to bavachin based on molecular docking and pull-down assays. We also found that bavachin improved neuroinflammation-induced neuronal survival and functional impairment and that this effect may be related to activation of the ERK and Akt pathways mediated by the BDNF pathway. Taken together, these data suggested that bavachin, by targeting inhibition PKCδ to inhibit the NF-κB pathway, further reduced the inflammatory response and oxidative stress and subsequently improved diabetic neuronal survival and function and finally ameliorated diabetes-induced depressive-like behaviors in mice. For the first time, we found that bavachin is a potential agent for the treatment of diabetes-associated neuroinflammation and depression and that PKCδ is a potential target for the treatment of diabetes-associated neuroinflammation, including depression.

Bavachin combined with epimedin B induce idiosyncratic liver injury under immunological stress conditions.

Reports on Chinese patent medicines preparations containing Epimedii Folium (EF) and Psoraleae Fructus (PF) resulting in idiosyncratic drug-induced liver injury (IDILI) have received widespread attention. Previous studies have shown that bavachin and epimedin B-two active ingredients derived from both EF and PF-are potential components associated with IDILI, but the underlying mechanism remains unclear. We evaluated bavachin and epimedin B-induced IDILI under TNF-α-mediated immunological stress conditions and generated liver lipid metabolism profiles using lipidomics and multivariate statistical analysis. We next applied transcriptomics to identify the differential gene expression on the transcription level. Our results showed that co-exposure to bavachin, epimedin B under immunological stress conditions resulted in obvious liver injury. The differential metabolites screened in our study were closely related to the immune homeostasis of the liver. Sixteen differentially expressed genes were found, Zc3h6 and R3hdml were upregulated, while Sumo2, Cd74, Banp, Oas3, Oas2, Gbp8, Slfn8, Gbp2b, Serpina3g, Zbtb40, H2-Ab1, Osgin1, Tgtp1 and Hspa1b were all downregulated. These differentially expressed genes were associated with biological processes concerning metabolic process and immune system process. Further integrative analysis indicated that bavachin combined with epimedin B affected genes that were not only related to immune system processes, but also to lipid metabolism. Ultimately, this led to an imbalance in the immune microenvironment in the liver and may have contributed to the observed liver injury.

Bavachin induces liver injury and cell apoptosis by targeting Wnt/ß-catenin/DRP1 signaling pathway mediated mitochondrial dysfunction.

Psraleae Fructus (PF) is a well-known traditional Chinese medicine in China. While numerous liver injury reports caused by PF limits its clinical application. Bavachin, a flavonoid compound isolated from the fruits of Psoralea corylifolia L., has been validated to induce direct apoptosis in hepatocytes and liver tissues in our previous studies. However, the subcellular mechanisms of bavachin induced liver injury is still elusive. Here, utilizing 6-week-old C57BL/6 J mice and human embryonic hepatocytes (L02 cells), we report that bavachin activates dynamic-related protein 1 (DRP1) mediated excess mitochondrial fission and endoplasmic reticulum (ER) stress related apoptosis via Wnt/ß-catenin signaling pathway. Notably, DRP1 knockdown or XAV-939 induced Wnt/ß-catenin inhibition decreased bavachin-induced ER stress and cell apoptosis in L02 cells. In addition, bavachin impaired mitochondrial structural and function in the mice liver tissues. Mdivi-1, a mitochondrial fission inhibitor targeting DRP1, prevented bavachin-induced mitochondrial and ER structural damage, ER stress, and liver injury. Our results demonstrated that bavachin induced mitochondrial fission plays a crucial role in bavachin induced ER stress related liver injury, via the mechanism that involved activation of Wnt/ß-catenin signaling pathway.

Undesirable ER stress induced by bavachin contributed to follicular atresia in zebrafish ovary.

Fructus psoraleae (FP) is a commonly used herb with potential reproductive toxicity. Bavachin (BV), one of essential active ingredients of FP, was found to exhibit estrogenic activity, but its effect on female reproductive system remains unknown. In this study, the impact of BV on the female zebrafish reproductive system and underlying molecular mechanism were determined in vivo and ex vivo. The results showed that BV could accumulate in zebrafish ovary, leading to obvious follicular atresia and increase in gonadal index and vitellogenin content. Endoplasmic reticulum (ER) swelling and hypertrophy were observed in the BV-treated zebrafish ovary, accompanied by an increase in the expressions of ER stress and unfolded protein response (UPR) related genes, namely atf6, ire-1α and xbp1s. In the ex vivo study, BV was found to decrease the survival rate and maturation rate of oocytes, while increasing the expression of Ca2+. Additionally, BV led to an elevation in the level of estrogen receptor ESR1 and the expressions of genes involved in ER stress and UPR, including atf6, ire-1α, xbp1s, chop and perk. Moreover, molecular docking revealed that BV could directly bind to immunoglobulin heavy chain binding protein (BiP) and estrogen receptor 1 (ESR1). Besides, the alterations induced by BV could be partially reversed by fulvestrant (FULV) and 4-phenylbutyric acid (4-PBA), respectively. Thus, long-termed BV-containing medicine treatment could generate reproductive toxicity in female zebrafish by causing follicular atresia through BiP- and ESR-mediated ER stress and UPR, providing a potential target for the prevention of reproductive toxicity caused by BV.

A Scd1-mediated metabolic alteration participates in liver responses to low-dose bavachin.

Hepatotoxicity induced by bioactive constituents in traditional Chinese medicines or herbs, such as bavachin (BV) in Fructus Psoraleae, has a prolonged latency to overt drug-induced liver injury in the clinic. Several studies have described BV-induced liver damage and underlying toxicity mechanisms, but little attention has been paid to the deciphering of organisms or cellular responses to BV at no-observed-adverse-effect level, and the underlying molecular mechanisms and specific indicators are also lacking during the asymptomatic phase, making it much harder for early recognition of hepatotoxicity. Here, we treated mice with BV for 7 days and did not detect any abnormalities in biochemical tests, but found subtle steatosis in BV-treated hepatocytes. We then profiled the gene expression of hepatocytes and non-parenchymal cells at single-cell resolution and discovered three types of hepatocyte subsets in the BV-treated liver. Among these, the hepa3 subtype suffered from a vast alteration in lipid metabolism, which was characterized by enhanced expression of apolipoproteins, carboxylesterases, and stearoyl-CoA desaturase 1 (Scd1). In particular, increased Scd1 promoted monounsaturated fatty acids (MUFAs) synthesis and was considered to be related to BV-induced steatosis and polyunsaturated fatty acids (PUFAs) generation, which participates in the initiation of ferroptosis. Additionally, we demonstrated that multiple intrinsic transcription factors, including Srebf1 and Hnf4a, and extrinsic signals from niche cells may regulate the above-mentioned molecular events in BV-treated hepatocytes. Collectively, our study deciphered the features of hepatocytes in response to BV insult, decoded the underlying molecular mechanisms, and suggested that Scd1 could be a hub molecule for the prediction of hepatotoxicity at an early stage.

Bavachin alleviates diabetic nephropathy in db/db mice by inhibition of oxidative stress and improvement of mitochondria function.

Diabetic nephropathy (DN) is a major complication of diabetes mellitus. Psoralea corylifolia L. seed (PCS) is a traditional medicine effective against various diseases. In this study, we aimed to investigate the effect of bavachin, the major active component of PCS, on DN in db/db mice. Bavachin (10 mg/kg/day) was administered orally to 12-week-old male db/db mice for 6 wk. For in vitro experiments, SV40 MES13 cells were treated with bavachin in the presence of 25 mM glucose. Food and water intake and urine mass were significantly increased in db/db mice compared to wild-type CON mice, but bavachin administration significantly reduced these increases. Urinary microalbumin, blood urea nitrogen, and creatinine clearance which were significantly increased in db/db mice, were also decreased by bavachin administration. Glomerular area and collagen deposition in the kidney were significantly decreased in db/db mice following bavachin administration. Increased renal levels of fibrotic factors, fibronectin, COL1A1, and α-SMA, were reduced following bavachin administration. Protein expressions of antioxidant enzymes, namely SOD2, catalase, and HO-1, and mitochondrial function-related factors, namely SIRT1, PGC1α, Nrf1, and mtTFA, were reduced in the kidney tissues of db/db mice compared to wild-type CON mice, and bavachin administration upregulated these protein expressions. In vitro studies also showed that bavachin decreases mitochondria ROS production, increases the expression of PGC-1α and SIRT1, and decreases the expression of α-SMA in high glucose-treated SV40 MES13 cells. Based on these results, bavachin improved DN by inhibiting oxidative stress and enhancing mitochondrial function.

Bavachin induces apoptosis in colorectal cancer cells through Gadd45a via the MAPK signaling pathway.

Bavachin is a dihydroflavonoid compound isolated from Psoralea corylifolia, and exhibits anti-bacterial, anti-inflammatory, anti-tumor and lipid-lowering activities. Recent attention has gradually drawn on bavachin-induced apoptosis in many human cancer cell lines. However, the anti-cancer effects and related mechanisms in colorectal cancer remain unknown. Here, we investigated the effects of bavachin on colorectal cancer in vivo and in vitro. The results showed that bavachin inhibited the proliferation of human colorectal cancer cells and induce apoptosis. These changes were mediated by activating the MAPK signaling pathway, which significantly up-regulated the expression of Gadd45a. Furthermore, Gadd45a silencing obviously attenuated bavachin-mediated cell apoptosis. Inhibition of the MAPK signaling pathway by JNK/ERK/p38 inhibitors also weakened the up-regulation of Gadd45a by bavachin. The anticancer effect of bavachin was also validated using a mouse xenograft model of human colorectal cancer. In conclusion, these findings suggest that bavachin induces the apoptosis of colorectal cancer cells through activating the MAPK signaling pathway.

Bavachin protects against diet-induced hepatic steatosis and obesity in mice.

Non-alcoholic fatty liver disease (NAFLD) is a major health concern worldwide, and the incidence of metabolic disorders associated with NAFLD is rapidly increasing because of the obesity epidemic. There are currently no approved drugs that prevent or treat NAFLD. Recent evidence shows that bavachin, a flavonoid isolated from the seeds and fruits of Psoralea corylifolia L., increases the transcriptional activity of PPARγ and insulin sensitivity during preadipocyte differentiation, but the effect of bavachin on glucose and lipid metabolism remains unclear. In the current study we investigated the effects of bavachin on obesity-associated NAFLD in vivo and in vitro. In mouse primary hepatocytes and Huh7 cells, treatment with bavachin (20 µM) significantly suppressed PA/OA or high glucose/high insulin-induced increases in the expression of fatty acid synthesis-related genes and the number and size of lipid droplets. Furthermore, bavachin treatment markedly elevated the phosphorylation levels of AKT and GSK-3ß, improving the insulin signaling activity in the cells. In HFD-induced obese mice, administration of bavachin (30 mg/kg, i.p. every other day for 8 weeks) efficiently attenuated the increases in body weight, liver weight, blood glucose, and liver and serum triglyceride contents. Moreover, bavachin administration significantly alleviated hepatic inflammation and ameliorated HFD-induced glucose intolerance and insulin resistance. We demonstrated that bavachin protected against HFD-induced obesity by inducing fat thermogenesis and browning subcutaneous white adipose tissue (subWAT). We revealed that bavachin repressed the expression of lipid synthesis genes in the liver of obese mice, while promoting the expression of thermogenesis, browning, and mitochondrial respiration-related genes in subWAT and brown adipose tissue (BAT) in the mice. In conclusion, bavachin attenuates hepatic steatosis and obesity by repressing de novo lipogenesis, inducing fat thermogenesis and browning subWAT, suggesting that bavachin is a potential drug for NAFLD therapy.

Bavachin induces apoptosis in colorectal cancer cells through Gadd45a via the MAPK signaling pathway / 中国天然药物

Bavachin is a dihydroflavonoid compound isolated from Psoralea corylifolia, and exhibits anti-bacterial, anti-inflammatory, anti-tumor and lipid-lowering activities. Recent attention has gradually drawn on bavachin-induced apoptosis in many human cancer cell lines. However, the anti-cancer effects and related mechanisms in colorectal cancer remain unknown. Here, we investigated the effects of bavachin on colorectal cancer in vivo and in vitro. The results showed that bavachin inhibited the proliferation of human colorectal cancer cells and induce apoptosis. These changes were mediated by activating the MAPK signaling pathway, which significantly up-regulated the expression of Gadd45a. Furthermore, Gadd45a silencing obviously attenuated bavachin-mediated cell apoptosis. Inhibition of the MAPK signaling pathway by JNK/ERK/p38 inhibitors also weakened the up-regulation of Gadd45a by bavachin. The anticancer effect of bavachin was also validated using a mouse xenograft model of human colorectal cancer. In conclusion, these findings suggest that bavachin induces the apoptosis of colorectal cancer cells through activating the MAPK signaling pathway.

Prevention of LPS-Induced Acute Kidney Injury in Mice by Bavachin and Its Potential Mechanisms.

Acute kidney injury (AKI) is a serious complication of sepsis with a rapid onset and high mortality rate. Bavachin, an active component of Psoralea corylifolia L., reportedly has antioxidant, anti-apoptotic, and anti-inflammatory effects; however, its beneficial effects on AKI remain undetermined. We investigated the protective effect of bavachin on lipopolysaccharide (LPS)-induced AKI in mice and elucidated the underlying mechanism in human renal tubular epithelial HK-2 cells. Increased serum creatinine and blood urea nitrogen levels were observed in LPS-injected mice; however, bavachin pretreatment significantly inhibited this increase. Bavachin improved the kidney injury score and decreased the expression level of tubular injury markers, such as neutrophil gelatinase-associated lipocalin (NGAL) and kidney injury molecule-1 (KIM-1), in both LPS-injected mice and LPS-treated HK-2 cells. LPS-induced oxidative stress via phosphorylated protein kinase C (PKC) ß and upregulation of the NADPH oxidase (NOX) 4 pathway was also significantly decreased by treatment with bavachin. Moreover, bavachin treatment inhibited the phosphorylation of MAPKs (P38, ERK, and JNK) and nuclear factor (NF)-κB, as well as the increase in inflammatory cytokine levels in LPS-injected mice. Krüppel-like factor 5 (KLF5) expression was upregulated in the LPS-treated HK-2 cells and kidneys of LPS-injected mice. However, RNAi-mediated silencing of KLF5 inhibited the phosphorylation of NF-kB, consequently reversing LPS-induced KIM-1 and NGAL expression in HK-2 cells. Therefore, bavachin may ameliorate LPS-induced AKI by inhibiting oxidative stress and inflammation via the downregulation of the PKCß/MAPK/KLF5 axis.

Bavachin Suppresses Alpha-Hemolysin Expression and Protects Mice from Pneumonia Infection by Staphylococcus aureus.

Staphylococcus aureus (S. aureus) infection causes dramatic harm to human health as well as to livestock development. As an important virulence factor, alpha-hemolysin (hla) is critical in the process of S. aureus infection. In this report, we found that bavachin, a natural flavonoid, not only efficiently inhibited the hemolytic activity of hla, but was also capable of inhibiting it on transcriptional and translational levels. Moreover, further data revealed that bavachin had no neutralizing activity on hla, which did not affect the formation of hla heptamers and exhibited no effects on the hla thermal stability. In vitro assays showed that bavachin was able to reduce the S. aureus-induced damage of A549 cells. Thus, bavachin repressed the lethality of pneumonia infection, lung bacterial load and lung tissue inflammation in mice, providing potent protection to mice models in vivo. Our results indicated that bavachin has the potential for development as a candidate hla inhibitor against S. aureus.

Ginsenoside Rb1 Ameliorated Bavachin-Induced Renal Fibrosis via Suppressing Bip/eIF2α/CHOP Signaling-Mediated EMT.

The nephrotoxicity of Fructus Psoraleae, an effective traditional Chinese medicine for vitiligo treatment, has been reported. As one of the main toxic components in Fructus Psoraleae, bavachin (BV) was considered to be related to Fructus Psoraleae-caused adverse outcomes, but the direct evidence and molecular mechanism underlying BV-induced nephrotoxicity are not well elucidated. Therefore, this study was designed to confirm whether BV would cause toxic effects on the kidney and explore the possible mode of action. Our results demonstrated that days' treatment with 0.5 µM BV indeed caused obvious renal fibrosis in the zebrafish kidney. The obvious E- to N-cadherin switch and the expressions of proteins promoting epithelial-mesenchymal transition (EMT) were observed in BV-treated human renal tubular epithelial and zebrafish kidneys. In addition, elevated reactive oxygen species (ROS) levels and Bip/eIF2α/CHOP-mediated endoplasmic reticulum (ER) stress and the unfolded protein response (UPR) were caused by BV, both of which could be reversed by ROS scavenger N-acetyl-L-cysteine (NAC). Also, blocking ER stress-caused cytoplasmic Ca2+ overload with 4-PBA notably alleviated BV-induced alterations in key molecular events related to EMT and renal fibrosis. Furthermore, of the natural compounds subjected to screening, ginsenoside Rb1 significantly downregulated BV-induced ER stress by inhibiting ROS generation and following the activation of Bip/eIF2α/CHOP signaling in HK2 cells. Subsequently, BV-triggered EMT and renal fibrosis were both ameliorated by ginsenoside Rb1. In summary, our findings suggested that BV-induced ROS promoted the appearance of EMT and renal fibrosis mainly via Bip/eIF2α/CHOP-mediated ER stress. This ER stress-related toxic pathway might be a potential intervention target for BV-caused renal fibrosis, and ginsenoside Rb1 would be a promising drug against BV- or Fructus Psoraleae-induced nephrotoxicity.

Bavachin inhibits IL-4 expression by downregulating STAT6 phosphorylation and GATA-3 expression and ameliorates asthma inflammation in an animal model.

IL-4 plays a key role in many human diseases, including allergic asthma, autoimmunity, allergies, and cancer. Therefore, the investigation of small compounds that regulate IL-4 secretion is of great interest for fighting these diseases. Natural flavonoids are useful compounds reported to have therapeutic effects in diseases involving IL-4. This study aimed to investigate small compounds that inhibit the effect of IL-4 with the lowest cell toxicity. We found that Bavachin had the greatest ability to downregulate IL-4 in the spleen of T cells from 4get IL-4-GFP mice. To identify the underlying molecular mechanisms, we found that Bavachin could decrease the IL-4 levels by downregulating the level of Gata-3 expression and STAT6 phosphorylation. These findings reveal the potential use of Bavachin as a tool for selectively controlling diseases involving IL-4 and, more generally, STAT6-dependent responses.

Bavachin produces immunoadjuvant activity by targeting the NFAT signaling pathway.

BACKGROUND: Bavachin, a flavonoid compound isolated from the seeds and fruits of Psoralea corylifolia l. (family Fabaceae), is used as a traditional medicine in Asia. Indeed, it is reported to have various medicinal functions such as estrogenic and antiinflammatory activities among others. However, to date, the effects of bavachin on T cell activation have yet to be reported. PURPOSE AND STUDY DESIGN: We aimed to determine the effects of bavachin on the activation of a human T cell line in vitro and on antigen-specific immune responses in mice in vivo. METHODS: In a nuclear factor of activated T cells (NFAT) activity assay, the Jurkat T cell line expressing a luciferase reporter driven by an NFAT-response element was stimulated with antihuman CD3/CD28 antibody and bavachin. Furthermore, the level of cytokine production was measured in the Jurkat T cell line stimulated with phorbol 12-myristate 13-acetate/ionomycin and bavachin using an IL-2 ELISA and a cytometric bead array assay. For in vivo analyses, mice were subcutaneously immunized with an antigen (ovalbumin protein) and bavachin, and the immune responses of mice were analyzed by FACS analysis, a T cell proliferation assay, a cytokine ELISA, and an antiovalbumin-specific antibody ELISA. RESULTS: We found that bavachin activated NFAT-mediated transcription in the human T cell line in vitro. In mice, when bavachin was administered with the antigen, an increase in T cell responses and antibody production specific to the antigen was observed. CONCLUSION: Our results suggest that bavachin has immunoadjuvant and immunomodulation effects, which arise through activation of the NFAT signaling pathway.

Bavachin exerted anti-neuroinflammatory effects by regulation of A20 ubiquitin-editing complex.

Neuroinflammation is a major pathophysiological contributor to the progression of the central nervous system disorders. Bavachin is a natural product belonging to the flavonoid class. The anti-neuroinflammatory effect and the molecular mechanisms are not well understood. In this study, we found bavachin can exert anti-neuroinflammatory effect via inhibition of nuclear factor-kappa B (NF-κB) signaling. We found that bavachin can obviously upregulate the expression of A20 (TNFAIP3) in microglial cells. Further studies suggested siRNA-A20 knockdown treatment can attenuate the inhibitory effects of bavachin on neuroinflammation. We further found bavachin can increase the interaction of ubiquitin-editing enzyme A20 complex including A20, Tax1-binding protein 1 (TAX1BP1) and Itch, the subsequently downregulated the K63-ubiquitination of TNF receptor associated factor 6 (TRAF6) and NF-κB signaling pathway. Altogether, our results indicated that bavachin exerted anti-neuroinflammatory effects through inhibition of NF-κB signaling mediated by regulation of ubiquitin-editing enzyme A20 complex. Our finding has important clinical significance for the potential application of bavachin in the treatment of neurological disorders.

Bavachin enhances NLRP3 inflammasome activation induced by ATP or nigericin and causes idiosyncratic hepatotoxicity.

Psoraleae Fructus (PF) is a well-known traditional herbal medicine in China, and it is widely used for osteoporosis, vitiligo, and other diseases in clinical settings. However, liver injury caused by PF and its preparations has been frequently reported in recent years. Our previous studies have demonstrated that PF could cause idiosyncratic drug-induced liver injury (IDILI), but the mechanism underlying its hepatotoxicity remains unclear. This paper reports that bavachin isolated from PF enhances the specific stimuli-induced activation of the NLRP3 inflammasome and leads to hepatotoxicity. Bavachin boosts the secretion of IL-1ß and caspase-1 caused by ATP or nigericin but not those induced by poly(I:C), monosodium urate crystal, or intracellular lipopolysaccharide. Bavachin does not affect AIM2 or NLRC4 inflammasome activation. Mechanistically, bavachin specifically increases the production of nigericin-induced mitochondrial reactive oxygen species among the most important upstream events in the activation of the NLRP3 inflammasome. Bavachin increases the levels of aspartate transaminase and alanine aminotransferase in serum and hepatocyte injury accompanied by the secretion of IL-1ß via a mouse model of lipopolysaccharide-mediated susceptibility to IDILI. These results suggest that bavachin specifically enhances the ATP- or nigericin-induced activation of the NLRP3 inflammasome. Bavachin also potentially contributes to PF-induced idiosyncratic hepatotoxicity. Moreover, bavachin and PF should be evaded among patients with diseases linked to the ATP- or nigericin-mediated activation of the NLRP3 inflammasome, which may be a dangerous factor for liver injury.

Sestrin2 protects against bavachin induced ER stress through AMPK/mTORC1 signaling pathway in HepG2 cells.

Bavachin (BV), a natural flavonoid compound derived from Psoralea corylifolia L, has been reported to be a potential hepatotoxin. Our previous studies have found that BV can induce endoplasmic reticulum (ER) stress-related cell apoptosis, but the molecular mechanism underlying BV-induced ER stress remains obscure. Sestrin2, a highly conserved stress-inducible protein, is involved in the cellular responses of various stress conditions and homeostatic regulation. However, whether Sestrin2 participated in the ER stress related hepatotoxicity against BV is still elusive. In the present study, we aim to investigate the role of BV on liver injury of mice and the impact of Sestrin2 on BV-induced ER stress in HepG2 cells. The results in mice showed that BV induced ER stress related liver injury with increased Sestrin2 expression involvement. Knockdown of Sestrin2 with siRNA aggravated BV-induced ER stress significantly in HepG2 cells. Further mechanistic study uncovered that inhibition of mTORC1 with rapamycin blocked BV-induced ER stress, and treatment with Sestrin2 siRNA blocked the inhibition effect of AMPK to mTORC1. Therefore, constant mTORC1 would lead to accumulation of misfolded or unfolded proteins and aggravated ER stress. Collectively, our study indicates that Sestrin2 confers protection against BV-induced ER stress via activating of the AMPK/mTORC1 pathway.