Psoralen and Isopsoralen, Two Estrogen-Like Natural Products from Psoraleae Fructus, Induced Cholestasis via Activation of ERK1/2.

With the increasing use of oral contraceptives and estrogen replacement therapy, the incidence of estrogen-induced cholestasis (EC) has tended to rise. Psoralen (P) and isopsoralen (IP) are the major bioactive components in Psoraleae Fructus, and their estrogen-like activities have already been recognized. Recent studies have also reported that ERK1/2 plays a critical role in EC in mice. This study aimed to investigate whether P and IP induce EC and reveal specific mechanisms. It was found that P and IP increased the expression of esr1, cyp19a1b and the levels of E2 and VTG at 80 µM in zebrafish larvae. Exemestane (Exe), an aromatase antagonist, blocked estrogen-like activities of P and IP. At the same time, P and IP induced cholestatic hepatotoxicity in zebrafish larvae with increasing liver fluorescence areas and bile flow inhibition rates. Further mechanistic analysis revealed that P and IP significantly decreased the expression of bile acids (BAs) synthesis genes cyp7a1 and cyp8b1, BAs transport genes abcb11b and slc10a1, and BAs receptor genes nr1h4 and nr0b2a. In addition, P and IP caused EC by increasing the level of phosphorylation of ERK1/2. The ERK1/2 antagonists GDC0994 and Exe both showed significant rescue effects in terms of cholestatic liver injury. In conclusion, we comprehensively studied the specific mechanisms of P- and IP-induced EC and speculated that ERK1/2 may represent an important therapeutic target for EC induced by phytoestrogens.

Psoralen: a narrative review of current and future therapeutic uses.

Psoralen is a family of naturally occurring photoactive compounds found in plants that acquire potential cytotoxicity when activated by specific frequencies of electromagnetic waves. Psoralens penetrate the phospholipid cellular membranes and insert themselves between the pyrimidines of deoxyribonucleic acid (DNA). Psoralens are initially biologically inert and acquire photoreactivity when exposed to certain classes of electromagnetic radiation, such as ultraviolet light. Once activated, psoralens form mono- and di-adducts with DNA, leading to marked cell apoptosis. This apoptotic effect is more pronounced in tumor cells due to their high rate of cell division. Moreover, photoactivated psoralen can inhibit tyrosine kinase signaling and influence the immunogenic properties of cells. Thus, the cytotoxicity of photoactivated psoralen holds promising clinical applications from its immunogenic properties to potential anti-cancer treatments. This narrative review aims to provide an overview of the current understanding and research on psoralen and to explore its potential future pharmacotherapeutic benefits in specific diseases.

Spectroscopic view on the interaction between the psoralen derivative amotosalen and DNA.

Psoralens are eponymous for PUVA (psoralen plus UV-A radiation) therapy, which inter alia can be used to treat various skin diseases. Based on the same underlying mechanism of action, the synthetic psoralen amotosalen (AMO) is utilized in the pathogen reduction technology of the INTERCEPT® Blood System to inactivate pathogens in plasma and platelet components. The photophysical behavior of AMO in the absence of DNA is remarkably similar to that of the recently studied psoralen 4'-aminomethyl-4,5',8-trimethylpsoralen (AMT). By means of steady-state and time-resolved spectroscopy, intercalation and photochemistry of AMO and synthetic DNA were studied. AMO intercalates with a higher affinity into A,T-only DNA (KD = 8.9 × 10-5 M) than into G,C-only DNA (KD = 6.9 × 10-4 M). AMO covalently photobinds to A,T-only DNA with a reaction quantum yield of ΦR = 0.11. Like AMT, it does not photoreact following intercalation into G,C-only DNA. Femto- and nanosecond transient absorption spectroscopy reveals the characteristic pattern of photobinding to A,T-only DNA. For AMO and G,C-only DNA, signatures of a photoinduced electron transfer are recorded.

Psoralen synergizes with exosome-loaded SPC25 to alleviate senescence of nucleus pulposus cells in intervertebral disc degeneration.

OBJECTIVE: To explore the mechanism of psoralen synergized with exosomes (exos)-loaded SPC25 on nucleus pulposus (NP) cell senescence in intervertebral disc degeneration (IVDD). METHODS: IVDD cellular models were established on NP cells by tert-butyl hydroperoxide (TBHP) induction, followed by the treatment of psoralen or/and exos from adipose-derived stem cells (ADSCs) transfected with SPC25 overexpression vector (ADSCs-oe-SPC25-Exos). The viability, cell cycle, apoptosis, and senescence of NP cells were examined, accompanied by the expression measurement of aggrecan, COL2A1, Bcl-2, Bax, CDK2, p16, and p21. RESULTS: After TBHP-induced NP cells were treated with psoralen or ADSCs-oe-SPC25-Exos, cell proliferation and the expression of aggrecan, COL2A1, Bcl-2, and CDK2 were promoted; however, the expression of Bax, p16, p21, and inflammatory factors was decreased, and cell senescence, cycle arrest, and apoptosis were inhibited. Of note, psoralen combined with ADSCs-oe-SPC25-Exos further decelerated NP cell senescence and cycle arrest compared to psoralen or ADSCs-oe-SPC25-Exos alone. CONCLUSION: Combined treatment of psoralen and ADSCs-oe-SPC25-Exos exerted an alleviating effect on NP cell senescence, which may provide an insightful idea for IVDD treatment.

Phytochemical and Pharmacological Aspects of Psoralen - A Bioactive Furanocoumarin from Psoralea corylifolia Linn.

Since long ago, medicinal plants have played a vital role in drug discovery. Being blessed and rich in chemovars with diverse scaffolds, they have unique characteristics of evolving based on the need. The World Health Organization also mentions that medicinal plants remain at the center for meeting primary healthcare needs as the population relies on them. The plant-derived natural products have remained an attractive choice for drug development owing to their specific biological functions relevant to human health and also the high degree of potency and specificity they offer. In this context, one such esteemed phytoconstituent with inexplicable biological potential is psoralen, a furanocoumarin. Psoralen was the first constituent isolated from the plant Psoralea corylifolia, commonly known as Bauchi. Despite being a life-saver for psoriasis, vitiligo, and leukoderma, it also showed immense anticancer, anti-inflammatory, and anti-osteoporotic potential. This review brings attention to the possible application of psoralen as an attractive target for rational drug design and medicinal chemistry. It discusses the various methods for the total synthesis of psoralen, its extraction, the pharmacological spectrum of psoralen, and the derivatization done on psoralen.

Imaging the cellular response to an antigen tagged interstrand crosslinking agent.

Immunofluorescence imaging is a standard experimental tool for monitoring the response of cellular factors to DNA damage. Visualizing the recruitment of DNA Damage Response (DDR) components requires high affinity antibodies, which are generally available. In contrast, reagents for the display of the lesions that induce the response are far more limited. Consequently, DDR factor accumulation often serves as a surrogate for damage, without reporting the actual inducing structure. This limitation has practical implications given the importance of the response to DNA reactive drugs such as those used in cancer therapy. These include interstrand crosslink (ICL) forming compounds which are frequently employed clinically. Among them are the psoralens, natural products that form ICLs upon photoactivation and applied therapeutically since antiquity. However, despite multiple attempts, antibodies against psoralen ICLs have not been developed. To overcome this limitation, we developed a psoralen tagged with an antigen for which there are commercial antibodies. In this report we describe our application of the tagged psoralen in imaging experiments, and the unexpected discoveries they revealed.

Mutations in AcrR and RNA Polymerase Confer High-Level Resistance to Psoralen-UVA Irradiation.

DNA interstrand cross-links, such as those formed by psoralen-UVA irradiation, are highly toxic lesions in both humans and bacteria, with a single lesion being lethal in Escherichia coli. Despite the lack of effective repair, human cancers and bacteria can develop resistance to cross-linking treatments, although the mechanisms of resistance remain poorly defined. Here, we subjected E. coli to repeated psoralen-UVA exposure to isolate three independently derived strains that were >10,000-fold more resistant to this treatment than the parental strain. Analysis of these strains identified gain-of-function mutations in the transcriptional regulator AcrR and the alpha subunit of RNA polymerase that together could account for the resistance of these strains. Resistance conferred by the AcrR mutation is mediated at least in part through the regulation of the AcrAB-TolC efflux pump. Resistance via mutations in the alpha subunit of RNA polymerase occurs through a still-uncharacterized mechanism that has an additive effect with mutations in AcrR. Both acrR and rpoA mutations reduced cross-link formation in vivo. We discuss potential mechanisms in relation to the ability to repair and survive interstrand DNA cross-links. IMPORTANCE Psoralen DNA interstrand cross-links are highly toxic lesions with antimicrobial and anticancer properties. Despite the lack of effective mechanisms for repair, cells can become resistant to cross-linking agents through mechanisms that remain poorly defined. We derived resistant mutants and identified that two gain-of-function mutations in AcrR and the alpha subunit of RNA polymerase confer high levels of resistance to E. coli treated with psoralen-UVA. Resistance conferred by AcrR mutations occurs through regulation of the AcrAB-TolC efflux pump, has an additive effect with RNA polymerase mutations, acts by reducing the formation of cross-links in vivo, and reveals a novel mechanism by which these environmentally and clinically important agents are processed by the cell.

Psoralen prevents the inactivation of estradiol and treats osteoporosis via covalently targeting HSD17B2.

ETHNOPHARMACOLOGICAL RELEVANCE: Psoralea corylifolia L. seeds (P. corylifolia), popularly known as Buguzhi in traditional Chinese medicine, are often used to treat osteoporosis in China. Psoralen (Pso) is the key anti-osteoporosis constituent in P. corylifolia, however, its targets and mechanism of action are still unclear. AIM OF THE STUDY: The purpose of this study was to explore the interaction between Pso and 17-ß hydroxysteroid dehydrogenase type 2 (HSD17B2), an estrogen synthesis-related protein that inhibits the inactivation of estradiol (E2) to treat osteoporosis. MATERIALS AND METHODS: Tissue distribution of Pso was analyzed by in-gel imaging after oral administration of an alkynyl-modified Pso probe (aPso) in mice. The target of Pso in the liver was identified and analyzed using chemical proteomics. Co-localization and cellular thermal shift assays (CETSA) were used to verify the key action targets. To detect the key pharmacophore of Pso, the interaction of Pso and its structural analogs with HSD17B2 was investigated by CETSA, HSD17B2 activity assay, and in-gel imaging determination. Target competitive test, virtual docking, mutated HSD17B2 activity, and CETSA assay were used to identify the binding site of Pso with HSD17B2. A mouse model of osteoporosis was established by ovariectomies, and the efficacy of Pso in vivo was confirmed by micro-CT, H&E staining, HSD17B2 activity, and bone-related biochemical assays. RESULTS: Pso regulated estrogen metabolism by targeting HSD17B2 in the liver, with the α, ß-unsaturated ester in Pso being the key pharmacophore. Pso significantly suppressed HSD17B2 activity by irreversibly binding to Lys236 of HSD17B2 and preventing NAD+ from entering the binding pocket. In vivo studies in ovariectomized mice revealed that Pso could inhibit HSD17B2 activity, prevent the inactivation of E2, increase levels of endogenous estrogen, improve bone metabolism-related indices, and play a role in anti-osteoporosis. CONCLUSIONS: Pso covalently binds to Lys236 of HSD17B2 in hepatocytes to prevent the inactivation of E2, thereby aiding in the treatment of osteoporosis.

NEIL3-mediated proteasomal degradation facilitates the repair of cisplatin-induced DNA damage in human cells.

Anti-neoplastic effect of DNA cross-linking agents such as cisplatin, mitomycin C, and psoralen is attributed to their ability to induce DNA interstrand cross-links (ICLs), which block replication, transcription, and linear repair pathways by preventing DNA strand separation and trigger apoptosis. It is generally agreed that the Fanconi anemia (FA) pathway orchestrates the removal of ICLs by the combined actions of various DNA repair pathways. Recently, attention has been focused on the ability of the NEIL3-initiated base excision repair pathway to resolve psoralen- and abasic site-induced ICLs in an FA-independent manner. Intriguingly, overexpression of NEIL3 is associated with chemo-resistance and poor prognosis in many solid tumors. Here, using loss- and gain-of-function approaches, we demonstrate that NEIL3 confers resistance to cisplatin and participates in the removal of cisplatin-DNA adducts. Proteomic studies reveal that the NEIL3 protein interacts with the 26S proteasome in a cisplatin-dependent manner. NEIL3 mediates proteasomal degradation of WRNIP1, a protein involved in the early step of ICL repair. We propose that NEIL3 participates in the repair of ICL-stalled replication fork by recruitment of the proteasome to ensure a timely transition from lesion recognition to repair via the degradation of early-step vanguard proteins.

Individual and combined effects of chromium and ultraviolet-B radiation on defense system, ultrastructural changes, and production of secondary metabolite psoralen in a medicinal plant Psoralea corylifolia L.

The present study focuses on the effects of individual and combined stress of chromium (Cr) and ultraviolet-B (UV-B) radiation on Psoralea corylifolia L. The experiment comprised four sets: (i) control, (ii) eUV-B (elevated UV-B i.e., ambient + 7.2 kJ m-2 day-1 UV-B), (iii) Cr (chromium; 30 mg kg-1 soil), and (iv) Cr + eUV-B (chromium and elevated UV-B; Cr 30 mg kg-1 and ambient + 7.2 kJ m-2 day-1 UV-B). The eUV-B and Cr individually and in combination showed the variable responses on ultrastructure, physiology and biomass however, the impact was more prominent under individual Cr treatment followed by Cr + eUV-B and eUV-B. Higher bioconcentration factor and the lowered translocation factor consequently led to a higher reduction in the below ground biomass and the lesser reduction in above ground biomass under Cr + eUV-B treatment as compared to individual Cr treatment. In addition, higher induction in the enzymatic (glutathione reductase, ascorbate peroxidase, superoxide dismutase, and glutathione-S-transferase) and non-enzymatic antioxidants (glutathione reduced) were found to be responsible for efficient scavenging of hydrogen peroxide and superoxide radical leading to lowered MDA content under combined treatment as compared to Cr treatment. Deposition of Cr as electron dense granules in the cytoplasm, vacuoles, and cell wall under Cr and Cr + eUV-B is contemplated as one of the cellular mechanisms of P. corylifolia against the toxicity of Cr. Psoralen increased under all treatments with a maximum increase under Cr + eUV-B treatment. Taken together our results accentuated that P. corylifolia can be grown in an area contaminated with Cr and has a higher influx of UV-B for the attainment of psoralen considering its pharmaceutical perspectives.

Psoralen as a Photosensitizers for Photodynamic Therapy by Means of In Vitro Cherenkov Light.

Possible enhancements of DNA damage with light of different wavelengths and ionizing radiation (Rhenium-188-a high energy beta emitter (Re-188)) on plasmid DNA and FaDu cells via psoralen were investigated. The biophysical experimental setup could also be used to investigate additional DNA damage due to photodynamic effects, resulting from Cherenkov light. Conformational changes of plasmid DNA due to DNA damage were detected and quantified by gel electrophoresis and fluorescent staining. The clonogene survival of the FaDu cells was analyzed with colony formation assays. Dimethyl sulfoxide was chosen as a chemical modulator, and Re-188 was used to evaluate the radiotoxicity and light (UVC: λ = 254 nm and UVA: λ = 366 nm) to determine the phototoxicity. Psoralen did not show chemotoxic effects on the plasmid DNA or FaDu cells. After additional treatment with light (only 366 nm-not seen with 254 nm), a concentration-dependent increase in single strand breaks (SSBs) was visible, resulting in a decrease in the survival fraction due to the photochemical activation of psoralen. Whilst UVC light was phototoxic, UVA light did not conclude in DNA strand breaks. Re-188 showed typical radiotoxic effects with SSBs, double strand breaks, and an overall reduced cell survival for both the plasmid DNA and FaDu cells. While psoralen and UVA light showed an increased toxicity on plasmid DNA and human cancer cells, Re-188, in combination with psoralen, did not provoke additional DNA damage via Cherenkov light.

Psoralen suppresses the phosphorylation of amyloid precursor protein (APP) to inhibit myelosuppression.

This study aims to explore the effect of Psoralen on myelosuppression, and investigating the mechanism involved in. The mesenchymal stem cells (MSCs) were treated with CTX to construct cell model of myelosuppression, and then with APP knockdown or overexpression transfection. Cell proliferation, cell apoptosis, bone growth factors, and hematopoietic growth factors were identified. The animal model of myelosuppression syndrome was established by intraperitoneal injection of cyclophosphamide (CTX) into C57BL/6 mice, and then with APP knockdown transfection. The effect of Psoralen on myelosuppression mice with APP knockdown was explored, including observin the number of hematopoietic stem cells and bone marrow MSCs, detecting the degree of osteoporosis and the number of osteoclasts. The expression of phosphorylation-amyloid precursor protein (p-APP), bone growth factors, and hematopoietic growth factors were also examined. We found that CTX treatment inhibited cell proliferation, induced cell apoptosis, promoted p-APP/APP, and inhibited the expression of aph-1 homolog A (APH-1α), presenilin enhancer-2 (PEN-2), the receptor of advanced glycation endproducts (RAGE). Psoralen pretreatment effectively promoted cell proliferation, suppressed cell apoptosis, inhibited p-APP/APP and stimulated the expression of APH-1α, PEN-2, RAGE compared with CTX treatment. After APP knockdown, cell proliferation was inhibited, and cell apoptosis was increased. The release of bone growth factors and hematopoietic growth factors was decreased. Psoralen pretreatment could reverse the effect of APP knockdown on MSCs and myelosuppression mice. In conclusion, Psoralen treatment inhibited cell apoptosis and regulated bone growth factors and hematopoietic growth factors in myelosuppression syndrome by suppressing the phosphorylation of APP.

Psoralen downregulates osteoarthritis chondrocyte inflammation via an estrogen-like effect and attenuates osteoarthritis.

Estrogen and its receptor play a positive role in the development of osteoarthritis (OA). Psoralen is a plant-derived estrogen analog. This study aimed to verify whether psoralen inhibits OA through an estrogen-like effect. First, human primary chondrocytes in the late stage of OA were extracted to complete collagen type II immunofluorescence staining and cell proliferation experiments. Subsequently, estrogen, psoralen and estrogen receptor antagonists were co-cultured with OA chondrocytes, and RT-PCR was performed to detect the gene expression. A rabbit OA model was subsequently made by anterior cruciate ligament transection (ACLT). They were set as Sham group, OA group and Psoralen group, respectively. The articular cartilage samples were taken after 5 weeks of treatment, and the effect was observed by gross observation, histological staining, micro-CT scanning of subchondral bone. The results of cellular experiments displayed that the cultured cells were positive for collagen II fluorescence staining and 12 µg/mL psoralen was selected as the optimal concentration. In addition, psoralen had effects similar to estrogen, promoting the expression of estrogen tar-get genes CTSD, PGR and TFF1 and decreasing the expression of the inflammation-related gene TNF- α, IL-1ß and IL-6. The effect of psoralen was blocked after the use of an estrogen receptor antagonist. Further animal experiments indicated that the psoralen group showed less destruction of cartilage tissue and decreased OASRI scores compared with the OA group. A subchondral bone CT scan demonstrated that psoralen significantly increased subchondral bone mineral density (BMD), trabecular thickness and trabecular number and decreased trabecular separation. In summary, psoralen inhibits the inflammatory production of chondrocytes, which is related to estrogen-like effect, and can be used to attenuate the progression of OA.

Novel psoralen derivatives as anti-breast cancer agents and their light-activated cytotoxicity against HER2 positive breast cancer cells.

Psoralen derivatives are well known for their unique phototoxicity and also exhibits promising anti-breast cancer activity both in the presence and the absence of UVA irradiation. However, the structure-activity relationship on this scaffold remains lacking. Herein, a series of psoralen derivatives with various C-5 substituents were synthesized and evaluated for their in vitro dark and light-activated cytotoxicity against three breast cancer cell lines: MDA-MB-231, T47-D, and SK-BR-3. The type of substituents dramatically impacted the activity, with the 4-bromobenzyl amide derivative (3c) exhibiting the highest dark cytotoxicity against T47-D (IC50 = 10.14 µM), with the activity comparable to those of the reference drugs (doxorubicin, 1.46 µM; tamoxifen citrate, 20.86 µM; lapatinib 9.78 µM). On the other hand, the furanylamide 3g exhibits the highest phototoxicity against SK-BR-3 cells with the IC50 of 2.71 µM, which is almost tenfold increase compared to the parent compound, methoxsalen. Moreover, these derivatives showed exceptional selectivity towards HER2+ (SK-BR-3) over the HER2- (MDA-MB-231) breast cancer cell lines, which correlates well with the results from the molecular docking study, revealing that 3g formed favorable interactions within the active site of the HER2. Additionally, the cell morphology of SK-BR-3 cells suggested that the significant phototoxicity was related to induction of cell apoptosis. Most of the synthesized psoralen derivatives possess acceptable physicochemical properties and are suitable for being further developed as a novel anti-breast cancer agent in the future.

Psoralen induces hepatotoxicity by covalently binding to glutathione-S-transferases and the hepatic cytochrome P450.

BACKGROUND: Psoraleae Fructus has been widely used in China and its surroundings; however, Psoraleae Fructus and its compound preparation have been reported recently to cause liver injury in clinics. Thus, its safe use has attracted increasing attention. The possible mechanism is related to the metabolism of psoralen, but it still needs further clarification. PURPOSE: The present study was designed to evaluate the toxicity of psoralen and investigate the potentially related molecular mechanisms using chemical biology methods combined with animal experiments to provide evidence for the rational clinical use of psoralen. METHODS: An in vivo experiment was conducted with a time series of 20-80 mg/kg psoralen to verify its toxic performance. Target capture and click reactions were used to investigate direct targets of psoralen. Selectivity for different glutathione-S-transferase (GST) subtypes in the liver and inhibition of cytochrome P450 (CYP450) were also detected. RESULTS: Psoralen build-up in the liver is the primary cause of liver damage. Our study revealed the mechanism by which psoralen induces liver injury. Psoralen can bind directly to CYP2D6, CYP3A4, GST-α, and GST-µ and inhibit their activities, causing the depletion of glutathione (GSH) in vivo, which in turn induces hepatic damage. The special structure of α,ß-unsaturated lactones in psoralen facilitates its attachment to its target; therefore, complementing psoralen with GSH can efficiently protect the liver from damage. CONCLUSIONS: Psoralen causes a disorder in drug metabolism by inhibiting the activity of CYPs and GSTs, causing exhaustion of GSH, and subsequently leading to liver damage. The co-administration of GSH and psoralen is an effective way to avoid liver injury in clinical settings.

Psoralen alleviates radiation-induced bone injury by rescuing skeletal stem cell stemness through AKT-mediated upregulation of GSK-3ß and NRF2.

BACKGROUND: Repairing radiation-induced bone injuries remains a significant challenge in the clinic, and few effective medicines are currently available. Psoralen is a principal bioactive component of Cullen corylifolium (L.) Medik and has been reported to have antitumor, anti-inflammatory, and pro-osteogenesis activities. However, less information is available regarding the role of psoralen in the treatment of radiation-induced bone injury. In this study, we explored the modulatory effects of psoralen on skeletal stem cells and their protective effects on radiation-induced bone injuries. METHODS: The protective effects of psoralen on radiation-induced osteoporosis and irradiated bone defects were evaluated by microCT and pathological analysis. In addition, the cell proliferation, osteogenesis, and self-renewal of SSCs were explored. Further, the underlying mechanisms of the protective of psoralen were investigated by using RNA sequencing and functional gain and loss experiments in vitro and in vivo. Statistical significance was analyzed using Student's t test. The one-way ANOVA was used in multiple group data analysis. RESULTS: Here, we demonstrated that psoralen, a natural herbal extract, mitigated radiation-induced bone injury (irradiation-induced osteoporosis and irradiated bone defects) in mice partially by rescuing the stemness of irradiated skeletal stem cells. Mechanistically, psoralen restored the stemness of skeletal stem cells by alleviating the radiation-induced suppression of AKT/GSK-3ß and elevating NRF2 expression in skeletal stem cells. Furthermore, the expression of KEAP1 in skeletal stem cells did not significantly change in the presence of psoralen. Moreover, blockade of NRF2 in vivo partially abolished the promising effects of psoralen in a murine model of irradiation-induced osteoporosis and irradiated bone regeneration. CONCLUSIONS: In summary, our findings identified psoralen as a potential medicine to mitigate bone radiation injury. In addition, skeletal stem cells and AKT-GSK-3ß and NRF2 may thus represent therapeutic targets for treating radiation-induced bone injury.

Mechanistic Insights into the Ameliorating Effect of Melanogenesis of Psoralen Derivatives in B16F10 Melanoma Cells.

The objectives of this study were to investigate the melanogenetic potential of the psoralen derivatives 5-hydroxypsoralen, 5-methoxypsoralen, 8-hydroxypsoralen, 8-methoxypsoralen, and 5,8-dimethoxypsoralen in B16F10 melanoma cells. The results indicated that melanin production is significantly stimulated in B16F10 melanoma cells with 5-methoxypsoralen, 8-methoxypsoralen, and 5,8-dimethoxypsoralen, especially for 5-methoxypsoralen (bergapten), as reported previously. In addition, Western blot results showed that the protein levels of microphthalmia-associated transcription factor (MITF), tyrosinase, tyrosinase-related protein-1 (TRP-1), and tyrosinase-related protein-2 (TRP-2) increase after bergapten treatment for the first time. The results also showed that bergapten promotes the phosphorylation of Akt at Ser 473 and glycogen synthase kinase-3ß at Ser 9. Moreover, bergapten increased the content of ß-catenin in the cell cytoplasm and nucleus by reducing the phosphorylated ß-catenin (p-ß-catenin) content. The results also indicated that bergapten regulates melanogenesis by upregulating the phosphorylation of p38 and JNK-mitogen-activated protein kinase. Taken together, these findings suggest that the regulation of melanogenesis by bergapten may be mediated by the ß-catenin and MAPK signaling pathways and that bergapten might provide new insights into the pathogenesis of pigmented diseases.

Psoralen inhibits hepatitis B viral replication by down-regulating the host transcriptional machinery of viral promoters.

The hepatitis B virus (HBV) is a global public health challenge due to its highly contagious nature. It is estimated that almost 300 million people live with chronic HBV infection annually. Although nucleoside analogs markedly reduce the risk of liver disease progression, the analogs do not fully eradicate the virus. As such, new treatment options and drugs are urgently needed. Psoralen is a nourishing monomer of Chinese herb and is known to inhibit virus replication and inactivate viruses. In this study, we evaluated the potential of psoralen as an anti-HBV agent. Quantitative PCR and Southern blot analysis revealed that psoralen inhibited HBV replication in HepG2.2.15 â€‹cells in a concentration-dependent manner. Moreover, psoralen was also active against the 3TC/ETV-dual-resistant HBV mutant. Further investigations revealed that psoralen suppressed both HBV RNA transcription and core protein expression. The transcription factor FOXO1, a known target for PGC1α co-activation, binds to HBV pre-core/core promoter enhancer II region and activates HBV RNA transcription. Co-immunoprecipitation showed that psoralen suppressed the expression of FOXO1, thereby decreasing the binding of FOXO1 co-activator PGC1α to the HBV promoter. Overall, our results demonstrate that psoralen suppresses HBV RNA transcription by down-regulating the expression of FOXO1 resulting in a reduction of HBV replication.

Psoralen inhibits the proliferation and promotes apoptosis through endoplasmic reticulum stress in human osteosarcoma cells.

INTRODUCTION: Psoralen is a main active component of Psoralea corylifolia Linn. (Leguminosae). Psoralen has been reported to show antitumor effects and activity to accelerate osteoblastic proliferation. Nevertheless, the antitumor mechanism of psoralen in osteosarcoma has never been elucidated. The current study is aimed to investigate the therapeutic function of psoralen in human osteosarcoma cells and its potential regulatory mechanism. MATERIAL AND METHODS: Effects of psoralen (0-70 µg/mL) on the viability of two osteosarcoma cell lines cultured for 48 h was evaluated by MTT assays. The concentration of IC10 (8 µg/mL for MG-63 cells and 9 µg/mL for U2OS cells) was regarded to be a non-cytotoxic dose selected as the working concentration in the subsequent experiments. Effects of psoralen on cell proliferation for 48 h was assessed by colony formation assays. Flow cytometry analyses were performed to measure cell cycle and apoptosis. RT-qPCR and Western blotting were carried out to assess RNA expression and protein levels of endoplasmic reticulum (ER) stress associated factors. RESULTS: Psoralen inhibited osteosarcoma cell viability (IC50 25 µg/mL for MG-63 cells and IC50 40 µg/mL for U2OS cells) in a dose-dependent manner and growth inhibition rate reached the highest level when cells were treated with 70 µg/mL psoralen. Psoralen induced cell cycle arrest in the G0/G1 phase and promoted apoptosis of both MG-63 and U2OS cells. The treatment of psoralen resulted in an increase in ATF-6 and CHOP protein levels as well as a decrease in Bcl-2 protein level, indicating that cell apoptosis induced by psoralen was associated with ER stress. Treatment with 4-PBA, the ER stress inhibitor, attenuated the ability of psoralen to promote apoptosis of MG-63 and U2OS cells. CONCLUSIONS: Psoralen showed growth-inhibitory effects in osteosarcoma cells, and induced apoptosis via the ER stress pathway, which might be a potential drug to suppress the development of osteosarcoma.

RNF168 E3 ligase participates in ubiquitin signaling and recruitment of SLX4 during DNA crosslink repair.

SLX4/FANCP is a key Fanconi anemia (FA) protein and a DNA repair scaffold for incision around a DNA interstrand crosslink (ICL) by its partner XPF nuclease. The tandem UBZ4 ubiquitin-binding domains of SLX4 are critical for the recruitment of SLX4 to damage sites, likely by binding to K63-linked polyubiquitin chains. However, the identity of the ubiquitin E3 ligase that mediates SLX4 recruitment remains unknown. Using small interfering RNA (siRNA) screening with a GFP-tagged N-terminal half of SLX4 (termed SLX4-N), we identify the RNF168 E3 ligase as a critical factor for mitomycin C (MMC)-induced SLX4 foci formation. RNF168 and GFP-SLX4-N colocalize in MMC-induced ubiquitin foci. Accumulation of SLX4-N at psoralen-laser ICL tracks or of endogenous SLX4 at Digoxigenin-psoralen/UVA ICL is dependent on RNF168. Finally, we find that RNF168 is epistatic with SLX4 in promoting MMC tolerance. We conclude that RNF168 is a critical component of the signal transduction that recruits SLX4 to ICL damage.