Genome-Wide Characterization of B-Box Gene Family in Salvia miltiorrhiza.

B-box (BBX) is a type of zinc finger transcription factor that contains a B-box domain. BBX transcription factors play important roles in plant photomorphogenesis, signal transduction, as well as abiotic and biological stress responses. However, the BBX gene family of Salvia miltiorrhiza has not been systematically investigated to date. For this study, based on the genomic data of Salvia miltiorrhiza, 27 SmBBXs genes were identified and clustered into five evolutionary branches according to phylogenetic analysis. The promoter analysis suggested that SmBBXs may be involved in the regulation of the light responses, hormones, stress signals, and tissue-specific development. Based on the transcriptome data, the expression patterns of SmBBXs under different abiotic stresses and plant hormones were analyzed. The results revealed that the expressions of the SmBBXs genes varied under different conditions and may play essential roles in growth and development. The transient expression analysis implied that SmBBX1, SmBBX4, SmBBX9, SmBBX20, and SmBBX27 were in the nucleus. A transcriptional activation assay showed SmBBX1, SmBBX4, SmBBX20, and SmBBX24 had transactivation activities, while SmBBX27 had none. These results provided a basis for further research on the role of SmBBXs in the development of Salvia miltiorrhiza.

Novel histone deacetylase inhibitors derived from Magnolia officinalis significantly enhance TRAIL-induced apoptosis in non-small cell lung cancer.

Histone modifications play critical roles in the progression of non-small cell lung cancer (NSCLC), which accounts for almost 85% of all diagnosed lung cancers. Magnolol and polyphenol mixture (PM) derived from Magnolia officinalis exhibited remarkable antitumor activities in lung cancer. However, the epigenetic effects and molecular mechanisms of magnolol and PM in NSCLC have yet to be reported. In this study, the epigenetic effects of magnolol and PM in NSCLC were examined in vitro and in vivo. Results revealed that magnolol and PM significantly suppressed the expression levels and function of class I histone deacetylases (HDACs). In A549 and H1299 cells, magnolol and PM remarkably induced cell apoptosis by arresting the cell cycle in the G0/G1 phase while simultaneously activating various pro-apoptotic signals, including TRAIL-R2 (DR5), Bax, caspase 3, cleaved caspase 3, and cleaved PARP. However, these apoptosis-promoting effects could be attenuated by TSA, which is a specific class I HDACs inhibitor. ChIP assays also demonstrated that magnolol and PM significantly enriched the histone acetyl mark (H3K27ac) in the promoter region of DR5. In A549 xenograft model, magnolol and PM notably reduced tumor growth by 44.40% and 35.40%, respectively. Therefore, magnolol and PM, as potential inhibitors of class I HDACs, induced tumor cell apoptosis and suppressed tumor growth partially by epigenetically activating DR5, which is a key protein in death receptor signaling pathway.

Carnitine palmitoyltransferase 1A promotes mitochondrial fission by enhancing MFF succinylation in ovarian cancer.

Mitochondria are dynamic organelles that are important for cell growth and proliferation. Dysregulated mitochondrial dynamics are highly associated with the initiation and progression of various cancers, including ovarian cancer. However, the regulatory mechanism underlying mitochondrial dynamics is still not fully understood. Previously, our study showed that carnitine palmitoyltransferase 1A (CPT1A) is highly expressed in ovarian cancer cells and promotes the development of ovarian cancer. Here, we find that CPT1A regulates mitochondrial dynamics and promotes mitochondrial fission in ovarian cancer cells. Our study futher shows that CPT1A regulates mitochondrial fission and function through mitochondrial fission factor (MFF) to promote the growth and proliferation of ovarian cancer cells. Mechanistically, we show that CPT1A promotes succinylation of MFF at lysine 302 (K302), which protects against Parkin-mediated ubiquitin-proteasomal degradation of MFF. Finally, the study shows that MFF is highly expressed in ovarian cancer cells and that high MFF expression is associated with poor prognosis in patients with ovarian cancer. MFF inhibition significantly inhibits the progression of ovarian cancer in vivo. Overall, CPT1A regulates mitochondrial dynamics through MFF succinylation to promote the development of ovarian cancer. Moreover, our findings suggest that MFF is a potential therapeutic target for ovarian cancer.

Spica prunellae and its marker compound rosmarinic acid induced the expression of efflux transporters through activation of Nrf2-mediated signaling pathway in HepG2 cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Spica prunellae (SP) is a well-known traditional Chinese medicinal herb with properties of antihypertensive, antihyperglycemic, antiviral, anti-inflammatory, and antitumor activities. This herb is also popularly consumed as a food additive in some drinks or other food forms for treating pyreticosis. Rosmarinic acid (RA) is the marker compound from SP, which possesses anti-oxidative and anti-inflammatory functions. AIM OF THE STUDY: This study aims to investigate the regulatory effect of the water extract of SP (WESP) and RA on efflux transports (ETs), including P-glycoprotein (p-gp), multidrug resistance-associated protein 2 (MRP2), and breast cancer resistance protein (BCRP) in HepG2 cell line. Results would provide beneficial information for the proper application of SP in clinics. MATERIALS AND METHODS: HepG2 cells were treated with different doses of the tested drugs for 24 or 96h. MTT assay was used to examine cell viability. The protein and mRNA levels of the ETs were measured by using Western blot and real-time PCR, respectively. Reporter assay was used to study the antioxidant response element (ARE)-luciferin activity by using HepG2-C8 cells, which were generated by transfecting plasmid containing ARE-luciferin gene into HepG2 cells. The transport activities of ETs were tested by using substrate probes. RESULTS: WESP significantly (p<0.05) increased the expression of ETs in a dose-dependent manner. The increase caused by WESP was stronger than RA alone. Both WESP and RA promoted the translocation of nuclear factor E2-related factor-2 (Nrf2) from cytoplasm to the nucleus as well as significantly (p<0.05) enhanced the ARE-luciferin activity. WESP and RA also enhanced the efflux activity of P-gp and MRP2, accompanied by marked increase (p<0.05) in the intracellular ATP levels. CONCLUSIONS: WESP could significantly induce the expression of ETs through the activation of Nrf2-mediated signaling pathway in HepG2 cells. RA could be one of the active compounds responsible for the induction. WESP and RA also enhanced the efflux activity of P-gp and MRP2, and the increased intracellular ATP levels were likely involved in this induction. Results of this study provide a better understanding of the regulation of SP on ETs and the underlying molecular mechanism. Results indicated that potential drug-drug interactions may exist when SP is co-administered with other substrate drugs that are transported via the ETs, especially P-gp and MRP2, thereby providing beneficial information for appropriate use of SP for clinical therapy.

In Vivo Exposure of Kaempferol Is Driven by Phase II Metabolic Enzymes and Efflux Transporters.

Kaempferol is a well-known flavonoid; however, it lacks extensive pharmacokinetic studies. Phase II metabolic enzymes and efflux transporters play an important role in the disposition of flavonoids. This study aimed to investigate the mechanism by which phase II metabolic enzymes and efflux transporters determine the in vivo exposure of kaempferol. Pharmacokinetic analysis in Sprague-Dawley rats revealed that kaempferol was mostly biotransformed to conjugates, namely, kaempferol-3-glucuronide (K-3-G), kaempferol-7-glucuronide (K-7-G), and kaempferol-7-sulfate, in plasma. K-3-G represented the major metabolite. Compared with that in wild-type mice, pharmacokinetics in knockout FVB mice demonstrated that the absence of multidrug resistance protein 2 (MRP2) and breast cancer resistance protein (BCRP) significantly increased the area under the curve (AUC) of the conjugates. The lack of MRP1 resulted in a much lower AUC of the conjugates. Intestinal perfusion in rats revealed that the glucuronide conjugates were mainly excreted in the small intestine, but 7-sulfate was mainly excreted in the colon. In Caco-2 monolayers, K-7-G efflux toward the apical (AP) side was significantly higher than K-3-G efflux. In contrast, K-3-G efflux toward the basolateral (BL) side was significantly higher than K-7-G efflux. The BL-to-AP efflux was significantly reduced in the presence of the MRP2 inhibitor LTC4. The AP-to-BL efflux was significantly decreased in the presence of the BL-side MRPs inhibitor MK571. The BCRP inhibitor Ko143 decreased the glucuronide conjugate efflux. Therefore, kaempferol is mainly exposed as K-3-G in vivo, which is driven by phase II metabolic enzymes and efflux transporters (i.e., BCRP and MRPs).

Regulation of drug-metabolizing enzymes and efflux transporters by Astragali radix decoction and its main bioactive compounds: Implication for clinical drug-drug interactions.

ETHNOPHARMACOLOGICAL RELEVANCE: Astragali radix ("Huang Qi" in Chinese, HQ) is a well-known traditional Chinese herbal medicine that possesses various biological functions. Astragaloside IV (AS-IV), calycosin (CS), and formononetin (FMNT) are the three main bioactive compounds of HQ that are responsible for its pharmacological activities and therapeutic efficacy. AIM OF THE STUDY: This study aims to investigate the effects of HQ, AS-IV, CS, and FMNT on major human drug-metabolizing enzymes (DMEs), including CYP3A4, CYP2B6, CYP2E1, UGT1A, UGT1A6, SULT1A1, and SULT1A3, as well as efflux transporters (ETs), including P-gp, MRP2, BCRP, MRP1, and MRP3, by using HepG2 cell line. Results would provide beneficial information for the proper clinical application of HQ. MATERIALS AND METHODS: HepG2 cells were treated with HQ, AS-IV, CS, and FMNT for 96h. Cell viability was examined by MTT assay. The protein and mRNA levels of DMEs and ETs were measured using Western blot and real-time PCR, respectively. RESULTS: Compared with the control group, HQ considerably increased the expression levels of CYP3A4, CYP2B6, CYP2E1, UGT1A, P-gp, MRP2, BCRP, and MRP3 in a dose-dependent manner. Inversely, HQ significantly decreased the protein levels of UGT1A6, SULT1A1, and MRP1. Exposure to AS-IV induced the protein levels of UGT1A, P-gp, MRP1, and MRP3, but produced inhibitory effects on CYP3A4, CYP2B6, and BCRP. The expression levels of CYP3A4, UGT1A, SULT1A1, P-gp, MRP2, and MRP3 were remarkably increased in the CS-treated cells, whereas the protein levels of SULT1A3 and BCRP were decreased. FMNT treatment induced the protein levels towards CYP3A4, CYP2B6, UGT1A, P-gp, MRP1, MRP2, and MRP3, but inhibited the expression of CYP2E1, SULT1A1, and SULT1A3. CONCLUSIONS: HQ and its main bioactive compounds, including AS-IV, CS, and FMNT significantly regulated the expression of the major DMEs and ETs. HQ produced stronger regulations (induction or inhibition) on DMEs and ETs than AS-IV, CS, or FMNT alone. The results indicate that potential drug-drug interactions might exist when the tested drugs, specifically HQ, are co-administered with other substrate drugs that are metabolized or transported via the studied DMEs or ETs. This study provides beneficial information for appropriate use of HQ for clinical therapy.

Artemisinin and its derivatives can significantly inhibit lung tumorigenesis and tumor metastasis through Wnt/ß-catenin signaling.

Non-small-cell lung cancer (NSCLC) is the most prevalent malignancy worldwide given its high incidence, considerable mortality, and poor prognosis. The anti-malaria compounds artemisinin (ART), dihydroartemisinin (DHA), and artesunate (ARTS) reportedly have anti-cancer potential, although the underlying mechanisms remain unclear. In this work, we used flow cytometry to show that ART, DHA, and ARTS could inhibit the proliferation of A549 and H1299 cells by arresting cell cycle in G1 phase. Meanwhile, tumor malignancy including migration, invasion, cancer stem cells, and epithelial-mesenchymal transition were also significantly suppressed by these compounds. Furthermore, ART, DHA, and ARTS remarkably decreased tumor growth in vivo. By using IWP-2, the inhibitor of Wnt/ß-catenin pathway, and Wnt5a siRNA, we found that ART, DHA, and ARTS could render tumor inhibition partially dependent on Wnt/ß-catenin inactivation. These compounds could strikingly decrease the protein level of Wnt5-a/b and simultaneously increase those of NKD2 and Axin2, ultimately resulting in ß-catenin downregulation. In summary, our findings revealed that ART, DHA, and ARTS could suppress lung-tumor progression by inhibiting Wnt/ß-catenin pathway, thereby suggesting a novel target for ART, DHA, and ARTS in cancer treatment.