Betulinic acid induces apoptosis of HeLa cells via ROS-dependent ER stress and autophagy in vitro and in vivo.

Betulinic acid (BA), a naturally occurring lupane-type triterpenoid, possesses a wide range of potential activities against different types of cancer. However, the molecular mechanisms involved in anti-cervical cancer about BA were rarely investigated. Herein, the role of BA in cervical cancer suppression by ROS-mediated endoplasmic reticulum stress (ERS) and autophagy was deeply discussed. The findings revealed that BA activated Keap1/Nrf2 pathway and triggered mitochondria-dependent apoptosis due to ROS production. Furthermore, BA increased the intracellular Ca2+ levels, inhibited the expression of Beclin1 and promoted the expression of GRP78, LC3-II, and p62 associated with ERS and autophagy. Besides, BA initiated the formation of autophagosomes and inhibited autophagic flux by the co-administration of BA with 3-methyladenine (3-MA) and chloroquine (CQ), respectively. The in vivo experiment manifested that hydroxychloroquine (HCQ) enhanced the apoptosis induced by BA. For the first time, we demonstrated that BA could initiate early autophagy, inhibit autophagy flux, and induce protective autophagy in HeLa cells. Thus, BA could be a potential chemotherapy drug for cervical cancer, and inhibition of autophagy could enhance the anti-tumor effect of BA. However, the interactions of signaling factors between ERS-mediated and autophagy-mediated apoptosis deserve further attention.

Tanshinone IIA reverses gefitinib resistance in EGFR-mutant lung cancer via inhibition of SREBP1-mediated lipogenesis.

BACKGROUND AND AIM: Gefitinib resistance is an urgent problem to be solved in the treatment of non-small cell lung cancer (NSCLC). Tanshinone IIA (Tan IIA) is one of the main active components of Salvia miltiorrhiza, which exhibits significant antitumor effects. The aim of this study is to explore the reversal effect of Tan IIA on gefitinib resistance in the epidermal growth factor receptor (EGFR)-mutant NSCLC and the underlying mechanism. EXPERIMENTAL PROCEDURE: CCK-8, colony formation assay, and flow cytometry were applied to detect the cytotoxicity, proliferation, and apoptosis, respectively. The changes in lipid profiles were measured by electrospray ionization-mass spectrometry (MS)/MS. Western blot, real-time q-PCR, and immunohistochemical were used to detect the protein and the corresponding mRNA levels. The in vivo antitumor effect was validated by the xenograft mouse model. KEY RESULTS: Co-treatment of Tan IIA enhanced the sensitivity of resistant NSCLC cells to gefitinib. Mechanistically, Tan IIA could downregulate the expression of sterol regulatory element binding protein 1 (SREBP1) and its downstream target genes, causing changes in lipid profiles, thereby reversing the gefitinib-resistance in EGFR-mutant NSCLC cells in vitro and in vivo. CONCLUSIONS AND IMPLICATIONS: Tan IIA improved gefitinib sensitivity via SREBP1-mediated lipogenesis. Tan IIA could be a potential candidate to enhance sensitivity for gefitinib-resistant NSCLC patients.

Hypersampsonone H attenuates ulcerative colitis via inhibition of PDE4 and regulation of cAMP/PKA/CREB signaling pathway.

BACKGROUND AND OBJECTIVES: Ulcerative colitis (UC) is a recurrent intestinal inflammatory disease which poses a serious threat to the life of patients. However, there are no specific drugs for UC yet. Hypericum sampsonii Hance (HS) is a Chinese herbal medicine traditionally used to treat enteritis and dysentery. Our previous studies have demonstrated that HS holds potential anti-UC effects, and a novel compound named Hypersampsonone H (HS-1) isolated from HS possesses significant anti-inflammatory activity. However, the beneficial effects of HS-1 on UC remain unclear. This study aimed to investigate the therapeutic effects of HS-1 on UC and its potential mechanisms, both in vitro and in vivo. METHODS: The in vitro model was employed using LPS-induced RAW264.7 cells to investigate the anti-inflammatory effects of HS-1 and its possible mechanisms. Furthermore, the therapeutic efficacy and potential mechanisms of HS-1 against dextran sulfate sodium (DSS)-induced acute colitis were assessed through histopathological examination, biochemical analysis, and molecular docking. RESULTS: In vitro, HS-1 significantly reduced LPS-induced inflammatory responses, as indicated by inhibiting NO production, down-regulating the overexpression of COX-2 and iNOS, as well as regulating the imbalanced levels of IL-6, TNF-α, and IL-10. Moreover, HS-1 also inhibited the expression of PDE4, elevated the intracellular cAMP level, and promoted the phosphorylation of CREB, thereby activating the PKA/CREB pathway in RAW264.7 cells. In vivo, HS-1 demonstrated therapeutic capacity against DSS-induced colitis by alleviating the symptoms of colitis mice, regulating the abnormal expression of inflammatory mediators, protecting the integrity of intestinal epithelial barrier, and reducing tissue fibrosis. Consistently, HS-1 was found to decrease the expression of PDE4 isoforms, subsequently activating the cAMP/PKA/CREB signaling pathway. Furthermore, the molecular docking results indicated that HS-1 exhibited a high affinity for PDE4, particularly PDE4D. Further mechanistic validation in vitro demonstrated that HS-1 possessed a synergistic effect on forskolin and an antagonistic effect on H-89 dihydrochloride, thereby exerting anti-inflammatory effects through the cAMP/PKA/CREB signaling pathway. CONCLUSION: We disclose that HS-1 serves as a promising candidate drug for the treatment of UC by virtue of its ability to reduce DSS-induced colitis via the inhibition of PDE4 and the activation of cAMP/PKA/CREB signaling pathway.

Novel 6/7/6 ring system diterpenoids and cytochalasins from the fungus Eutypella scoparia GZU-4-19Y and their anti-inflammatory activity.

Two new compounds eutyditerpenoid A (1) and seco-phenochalasin B (5), together with seven known compounds diaporthein A (2), aspergillon A (3), phenochalasin B (4), cytochalasins Z24 and Z25 (6 and 7), scoparasins A and B (8 and 9) were isolated from marine-derived Eutypella scoparia GZU-4-19Y. Among them, eutyditerpenoid A (1) with a rare 6/7/6 ring system possesing an anhydride moiety was the first example in the pimarane-type diterpenoids. Their structures were determined based on spectroscopic methods and the electronic circular dichroism (ECD) calculations. In the bioassays, all of the isolates were evaluated for their inhibitory activity against NO production induced by lipopolysaccharide in RAW 264.7 cells. Compounds 3 and 7 showed potent NO inhibition activity with IC50 values of 2.1 and 17.1 µM respectively, and the former also significantly suppressed the protein expression of iNOS and COX-2 at the concentration of 2.5 µM.

Two new 2-(2-phenylethyl)chromone derivatives and two sesquiterpenes from agarwood of Aquilaria sinensis with anti-inflammatory activity.

Two new 2-(2-phenylethyl)chromones (1-2), two new sesquiterpenes (12-13), and twelve known compounds (3-11, 14-16) were isolated from agarwood of Aquilaria sinensis. These structures were confirmed by HRESIMS, 1D and 2D NMR spectra. The absolute configurations of two new sesquiterpenes were determined by comparing the experimental and calculated ECD spectra. Among them, 7,8-dihydroxy-2-[2-(4'-methoxyphenyl)ethyl]chromone (2) was the first time found that the hydroxyl groups at both C-7/C-8 in agarwood. And Aseudesm B (13), the aldehyded methyl group at C-5 of eucalyptane sesquiterpenes was first discovered in natural products. In the bioassays, all compounds were evaluated for their inhibitory activity against lipopolysaccharide-activated nitric oxide (NO) production in RAW264.7 cells. Compounds 2-5, 7, 9-10, and 13-14 revealed notable inhibitory effects against NO production with IC50 values ranging from 4.0 to 13.0 µM.

Rotundic acid alleviates hyperlipidemia in rats by regulating lipid metabolism and gut microbiota.

Disturbances in lipid metabolism and dysbiosis of the gut microbiota play an important role in the progression of hyperlipidemia. Previous study indicated that Ilicis Rotundae Cortex possesses anti-hyperlipidemic activity, and rotundic acid (RA) identified as a key active compound to be incorporated into the body. The study aimed to evaluate the anti-hyperlipidemia effects of RA and explored its impact on gut microbiota and lipid metabolism, as well as its possible mechanisms for improving hyperlipidemia. The study methodology included a comprehensive evaluation of the effects of RA on steatosis markers of hyperlipidemia, lipid metabolism, and gut microbiota by assessing biochemical parameters and histopathology, lipidomics, 16S rRNA gene sequencing, and short-chain fatty acid (SCFA) assays. The results showed that RA effectively reduced body weight and the steatosis markers in serum and liver. Moreover, the lipidomic analysis revealed significant changes in plasmatic and hepatic lipid levels, and these were restored by RA. According to the results of 16S rRNA gene sequencing, RA supplementation raised the relative abundance of Bacteroidetes and Proteobacteria while decreasing the relative abundance of Firmicutes. RA significantly boosted the relative abundance of SCFAs by increasing SCFAs-producing bacteria such as Bacteroides, Alloprevotella, Desulfovibrio, etc. In summary, RA could regulate triglyceride metabolism and glycerophospholipid metabolism, restore gut microbiota structure, and increase the relative abundance of SCFAs-producing bacteria to exert its hypolipidemic effects. These findings suggest RA to be a promising therapeutic agent for hyperlipidemia.

Polycyclic polyprenylated acylphloroglucinols from Hypericum sampsonii Hance and their anti-inflammatory activity.

Phytochemical investigation of Hypericum sampsonii Hance resulted in the isolation of thirty-five polycyclic polyprenylated acylphloroglucinols including six new ones (1, 3, 5, and 15-17). Their structures were elucidated by UV, IR, NMR, HRESIMS, and calculated ECD analysis. Some compounds were evaluated for their anti-inflammatory effects in LPS-induced RAW264.7 cells. Compounds 1 and 26 showed significant inhibitory effects on LPS-induced NO production, and markedly suppressed the protein expression of iNOS and COX-2 in LPS-activated RAW264.7 cells.

Effect of Salvia miltiorrhiza Bunge extracts on improving the efficacy and reducing the toxicity of Tripterygium wilfordii polyglycosides in the treatment of rheumatoid arthritis.

ETHNOPHARMACOLOGICAL RELEVANCE: Tripterygium wilfordii polyglycosides (TWP), extracted from the traditional Chinese herb Tripterygium wilfordii, has been widely used in the treatment of rheumatoid arthritis (RA). However, the toxicity of TWP to a variety of organs such as liver, kidney and testis greatly limits its clinical application. Salvia miltiorrhiza Bunge is often used in the treatment of RA due to its blood circulation promoting, stasis resolving, and anti-inflammatory effects. Salvia miltiorrhiza Bunge has also been reported to possess multiple organ protective effects. AIM OF THE STUDY: To investigate the influences of two main components of Salviorrhiza miltiorrhiza Bunge, hydrophilic salvianolic acids (SA) and lipophilic tanshinones (Tan), on the efficacy and toxicity of TWP in treating RA and to explore the underlying mechanisms. MATERIALS AND METHODS: SA and Tan were extracted from Salvia miltiorrhiza Bunge and the extracts were quantitated by HPLC and identified by UPLC-Q/TOF-MS. Then, a collagen-induced arthritis (CIA) rat model was established using bovine type II collagen (CII) and incomplete Freund's adjuvant (IFA). CIA rats were treated with TWP and/or SA/Tan. After 21 days of continuous treatment, arthritis symptoms and organs toxicity were evaluated. Meanwhile, serum metabolomics were investigated by the UPLC-Q/TOF-MS to understand the underlying mechanism. RESULTS: SA and Tan extracts could significantly alleviate arthritis symptoms in CIA rats and decrease the serum levels of inflammatory factors TNF-α, IL-1ß and IL-6 when combined with TWP. Meanwhile, both extracts alleviated injury of liver, kidney and testis caused by TWP, and the hydrophilic extract SA was superior. Moreover, a total of 38 endogenous differential metabolites were identified between the CIA model group and the TWP group, among which 33 metabolites were significantly recovered after the combination of SA or Tan. Metabolic pathway analysis showed that SA and Tan can affect metabolic pathways including linoleic acid metabolism, glycerophospholipid metabolism, sphingolipid metabolism and steroid biosynthesis metabolism pathway. CONCLUSIONS: Our findings indicated for the first time that two Salviorrhiza miltiorrhiza Bunge extracts could improve the efficacy and reduce the toxicity of TWP in the treatment of RA by adjusting metabolic pathways, and the hydrophilic extract SA was superior.

Seven new meroterpenoids from the fungus Penicillium sclerotiorum GZU-XW03-2.

Seven previously undescribed meroterpenoids, peniscmeroterpenoids H - N (1-7), were isolated from the marine-derived fungus Penicillium sclerotiorum GZU-XW03-2. Their structures were established by the spectroscopic methods and the electronic circular dichroism (ECD) calculations. Peniscmeroterpenoid H was a 6/6/6/5/6 rearranged pentacyclic meroterpenoid, featuring a unique 2-oxaspiro[5.5] undeca-4,7-dien-3-one motif. Peniscmeroterpenoids I and J (2 and 3) owned rare 6(D)/5(E) fused rings were not common in natural products, and compound 2 was the second example of a berkeleyone analogue stripped of the methyl ester fragment. Peniscmeroterpenoid K (4) was the first case where the C-24 was oxidized. In bioassay, compound 5 showed moderate anti-inflammatory activity.

New chemical structures and liver-protective activity of the diterpenoids from Callicarpa rubella.

Callicarpa rubella is a characteristic folk herb in the genus Callicarpa, and has abundant ethnobotanical usage as indigenous medicine in Lingnan area of P. R. China. However, the phytochemical and pharmacological properties of C. rubella was rarely investigated. Now, three new diterpenoids, named rubellapene A-C (1-3), along with five known analogues (4-8), were isolated from C. rubella. Their structures were determined by spectroscopic methods, quantum chemical electronic circular dichroism calculations and single-crystal X-ray diffraction analysis. Notably, the norditerpenoids C18 of clerodane type (rubellapene B) was rarely found in the genus Callicarpa. The liver protective effects of all of the isolates (1-8) were evaluated by the changes of cell viability and transaminase content of AST and ALT in H2O2-induced BRL cells. Compound 1, 3-8 exhibited that potent liver protective effects at different levels.

Integration of metabolomics and transcriptomics to reveal ferroptosis is involved in Tripterygium wilfordii polyglycoside tablet-induced testicular injury.

ETHNOPHARMACOLOGICAL RELEVANCE: Tripterygium wilfordii polyglycoside tablet (TWP), a traditional Chinese medicine preparation, has multiple pharmacological properties, including anti-inflammatory, immune-modulatory and anti-proliferative activities. However, the reproductive toxicity of TWP greatly limits its clinical application and the mechanism of TWP-induced reproductive toxicity is not fully understood yet. AIM OF THE STUDY: This study was designed to explore the mechanism of TWP-induced testis injury in male rats. MATERIALS AND METHODS: The mechanism underlying TWP-induced rat testicular injury was firstly investigated by integration of metabolomics and transcriptomics. Meanwhile, histopathological analysis, Western blot and RT-qPCR were performed to confirm the damaging effects and mechanisms of TWP on rat testis. RESULTS: Histopathological analysis revealed that TWP had significant testicular damage, which severely reduced the testis's tubular diameter and epithelium height. Further, TWP caused the protein level of ZO-1, CLDN11, PLZF, and OCT4 significantly downregulate, suggesting the blood-testis barrier function and spermatogenesis were damaged. Differentially expressed genes (DEGs), including 4952 upregulated and 2626 downregulated, were found in TWP-exposed testis compared to the normal group. Moreover, 77 changed metabolites were identified from testis samples. With integrated analysis of DEGs and changed metabolites, we found that glutathione metabolism and ferroptosis played an essential role in testicular injury. Additionally, the levels of ferroptosis-related protein GPX4, SLC7A11, and NRF2 were significantly downregulated, and the protein level of 4-HNE, a leading product of lipid peroxidation and oxidative stress, was upregulated. The changes in ferroptosis-related genes indicated that TWP might promote ferroptosis in rat testis. CONCLUSION: These results suggested that ferroptosis was involved in the testicular damage caused by TWP, which might provide a new strategy to alleviate TWP- induced testicular injury.

Protective effects of 4-geranyloxy-2,6-dihydroxybenzophenonel on DSS-induced ulcerative colitis in mice via regulation of cAMP/PKA/CREB and NF-κB signaling pathways.

Hypericum sampsonii Hance has traditionally been used to treat enteritis and diarrhea. As one of the main benzophenones isolated from H. sampsonii, 4-geranyloxy-2,6-dihydroxybenzophenonel (4-GDB) has been shown to possess anti-inflammatory effects. However, the therapeutic effect and potential mechanisms of 4-GDB in ulcerative colitis (UC) remain unclear. This study aimed to evaluate the role of 4-GDB in UC using a dextran sulfate sodium-induced colitis mouse model. Intragastric administration of 4-GDB (20 mg/kg/day) for 8 days significantly attenuated colonic injury, reduced the expression of inflammatory mediators, and improved colonic barrier function in mice with colitis. Furthermore, in vivo and in vitro experiments indicated that 4-GDB could activate cAMP/PKA/CREB and inhibit the NF-κB pathway. Collectively, 4-GDB may be a potential agent for treating UC by regulating the cAMP/PKA/CREB and NF-κB pathways.

Neoline, fuziline, songorine and 10-OH mesaconitine are potential quality markers of Fuzi: In vitro and in vivo explorations as well as pharmacokinetics, efficacy and toxicity evaluations.

ETHNOPHARMACOLOGICAL RELEVANCE: Fuzi, the lateral roots of Aconitum carmichaelii Debx, plays an irreplaceable role in treating Yang deficiency and cold coagulation syndromes. However, Fuzi has a narrow margin of safety since its pharmacological constituents, Aconitum alkaloids, have potential cardiotoxicity and neurotoxicity. The current quality markers (Q-markers) for the control of Fuzi's efficacy and toxicity are 3 monoester-diterpenoid alkaloids, namely, benzoylaconine (BAC), benzoylhypaconine and benzoylmesaconine (BMA) and 3 diester-diterpenoid alkaloids, namely, aconitine (AC), hypaconitine and mesaconitine (MA). However, mounting evidence indicates that the current 6 Q-markers may not be efficacy- or toxicity-specific enough for Fuzi. AIM OF THE STUDY: The aim of this study was to explore and evaluate efficacy- or toxicity-specific potential quality markers (PQ-markers) of Fuzi. MATERIALS AND METHODS: PQ-markers were explored by analyzing 30 medicinal samples and alkaloids exposed in mouse. Pharmacokinetics of PQ-markers on C57BL/6J mice were determined. Anti-inflammatory effects of PQ-markers were evaluated by λ-carrageenan-induced paw edema model and lipopolysaccharide-induced RAW264.7 cell inflammatory model, while analgesic effects were assessed by acetic acid-induced pain model and Hargreaves test. Cardiotoxicity and neurotoxicity of PQ-markers were assessed by histological and biochemical analyses, while acute toxicity was evaluated by modified Kirschner method. RESULTS: After in vitro and in vivo explorations, 7 PQ-markers, namely, neoline (NE), fuziline (FE), songorine (SE), 10-OH mesaconitine (10-OH MA), talatizamine, isotalatizidine and 16ß-OH cardiopetalline, were found. In the herbal medicines, NE, FE, SE and 10-OH MA were found in greater abundance than many other alkaloids. Specifically, the amounts of NE, FE and SE in the Fuzi samples were all far higher than that of BAC, and the contents of 10-OH MA in 56.67% of the samples were higher than that of AC. In mouse plasma and tissues, NE, FE, SE, talatizamine, isotalatizidine and 16ß-OH cardiopetalline had higher contents than the other alkaloids, including the 6 current Q-markers. The pharmacokinetics, efficacy and toxicity of NE, FE, SE and 10-OH MA were further evaluated. The average oral bioavailabilities of NE (63.82%), FE (18.14%) and SE (49.51%) were higher than that of BMA (3.05%). Additionally, NE, FE and SE produced dose-dependent anti-inflammatory and analgesic effects, and their actions were greater than those of BMA. Concurrently, the toxicities of NE, FE and SE were lower than those of BMA, since no cardiotoxicity or neurotoxicity was found in mice after NE, FE and SE treatment, while BMA treatment notably increased the creatine kinase activity and matrix metalloproteinase 9 level in mice. The average oral bioavailability of 10-OH MA (7.02%) was higher than that of MA (1.88%). The median lethal dose (LD50) of 10-OH MA in mice (0.11 mg/kg) after intravenous injection was close to that of MA (0.13 mg/kg). Moreover, 10-OH MA produced significant cardiotoxicity and neurotoxicity, and notable anti-inflammatory and analgesic effects that were comparable to those of MA. CONCLUSIONS: Seven PQ-markers of Fuzi were found after in vitro and in vivo explorations. Among them, NE, FE and SE were found to be more efficacy-specific than BMA, and 10-OH MA was as toxicity-specific as MA.

Triterpenoids from the genus Ilex attenuate free fatty acid-induced lipid accumulation in HepG2 cells by regulating lipid metabolism disorder and the AMPK signalling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Various traditional Chinese medicines from the genus Ilex (Aquifoliaceae) have been reported to have excellent hypolipidaemic effects. Although triterpenoids have been found to be the main active components, the underlying mechanisms have not been clarified. AIM OF THE STUDY: This study aimed to investigate the lipid-lowering effect, structure-activity relationship and action mechanism of triterpenoids from the genus Ilex. MATERIALS AND METHODS: FFA was used to induce HepG2 cells to establish a classical lipid-lowering activity screening model for the activities of 31 triterpenoids, and the contents of intracellular lipids, TC, and TG were measured. Furthermore, the structure-activity relationship was discussed. Mechanistically, UPLC-Q/TOF-MS-based metabolomics and lipidomics studies were performed, and metabolic pathways were analysed to investigate the lipid-lowering mechanism. Moreover, western blotting was performed to analyse the expression of key proteins of lipid metabolism and predict the targets of action. RESULTS: Thirteen triterpenoids significantly reduced intracellular lipid accumulation and decreased the levels of TG and TC. Among them, rotundic acid (RA) showed stronger lipid-lowering activity than the simvastatin-positive group, and structure-activity relationship analysis indicated that the hydroxyl groups at C-3 and C-19, hydroxymethyl groups at C-23, and carboxyl groups at C-28 may be the key groups for biological activity. Twenty-two metabolites in the metabolomics study and 19 metabolites in the lipidomics study were identified. The identified biomarkers were primarily glycerophosphocholine, LysoPCs, PCs, TAGs, LysoPEs, LysoPIs and sphingolipids, which are involved in glycerophospholipid and sphingolipid metabolism. Moreover, western blotting analysis showed that the expression of SREBP-1 and HMGCR decreased, while AMPK and ACC phosphorylation and the expression of CPT1A and CYP7A1 increased in the RA-treated group. CONCLUSION: The results suggested that triterpenoids from the genus Ilex showed significant lipid-lowering effects and that RA may be a novel hypolipidaemic drug candidate. Moreover, the underlying mechanism indicated that RA showed a lipid-lowering effect by regulating glycerophospholipid and sphingolipid metabolism and activating the AMPK pathway.

New chromone compounds from the marine derived fungus Diaporthe sp. XW12-1.

Four new chromone compounds diaporspchromanones A-C (1-3) and diaporspchromanone H (4), together with three known compounds (5-7) were separated from the marine derived fungus Diaporthe sp. XW12-1. The structures of the new compounds, including their absolute configurations, were elucidated by extensive spectroscopic analysis and the Mosher's ester method. Among them, diaporspchromanones A-C (1-3) possess a 3-substituted-chroman-4-one skeleton, which are rarely found in natural sources. In the bioassays, all compounds were evaluated for their inhibitory activity against lipopolysaccharide-activated nitric oxide (NO) production in RAW264.7 cells. Compounds 2 and 3 showed potent anti-inflammatory effects than the positive control (indomethacin, IC50, 70.33 ± 0.95 µM) (p < 0.05) with IC50 values of 19.06 ± 3.60 and 9.56 ± 0.18 µM, respectively.

A systematic review: Botany, phytochemistry, traditional uses, pharmacology, toxicology, quality control and pharmacokinetics of Ilex rotunda Thunb.

ETHNOPHARMACOLOGICAL RELEVANCE: Ilex rotunda Thunb. (I. rotunda) is an Ilex species of Aquifoliaceae, widely distributed in East Asia. Its dried bark is commonly used as a medicinal part in the field of traditional Chinese medicine (TCM), named Ilicis Rotundae Cortex. This medicinal plant is commonly used for clearing heat and removing toxin, draining dampness and relieving pain in TCM to treat tonsillitis, acute gastroenteritis, gastric and duodenal ulcer, rheumatism, traumatic injury, and so on. It also has significant development value on lipid-lowering, hepatoprotection and anti-inflammation, but the potential mechanism needs to be further explored. AIM OF THE REVIEW: More and more medicinal substances are being discovered in I. rotunda with multiple biological activities, which help to advance the ethno-pharmacological research in I. rotunda. However, to date there is a lack of a systematic summary of research progress on I. rotunda. This review aims to provide a critical summary of the current studies on I. rotunda. The progress in research on botany, phytochemistry, traditional uses, pharmacology, toxicology, quality control and pharmacokinetics of the plant is discussed. It hopes to provide useful references and guidance for the future directions of research on I. rotunda. MATERIALS AND METHODS: Studies of I. rotunda were collected via Google Scholar and Baidu Scholar, PubMed, ScienceDirect, SciFinder, Web of Science, China National Knowledge Infrastructure (CNKI), WANFANG DATA and libraries. Some local books, official websites, PhD or MS's dissertations were also included. The literature cited in this review covered the period from 1956 to January 2022. RESULTS: Analysis of the literature indicates that I. rotunda is a potentially valuable herbal medicine for the therapeutic of various diseases. To date, 120 compounds were found and identified in I. rotunda, mainly including triterpenoids, phenylpropanoids, etc. Modern pharmacological studies also found that the plant has the activities of protecting the cardiovascular system, lowering lipids and protecting the liver, as well as being an anti-inflammatory, anti-tumor and antibacterial. CONCLUSIONS: This review summarizes the results from current studies of I. rotunda. However, the current explanation seems insufficient and unsatisfactory, in terms of the relationships between the traditional uses and the modern pharmacological activities, the mechanisms and the material basis. Thus, a critical and comprehensive evaluation is necessary to explore its future research prospects and development direction.

Absorption characteristics of ilexgenin A and ilexsaponin B1 in human umbilical vein endothelial cells after administration of the total triterpenoid saponins from Ilex pubescens.

Ilex pubescens is a famous Chinese herbal medicine, frequently used to treat cardiovascular disease in South China. In this study, we aim to explore the absorption properties of ilexgenin A (C1) and ilexsaponin B1 (C3) in vascular endothelial cells after administration of the total triterpenoid saponins from Ilex pubescens (IPTS) and clarify the possible transport mechanisms. A UPLC-qTOF-MS/MS system was used to identify the components in IPTS that could be intracellularly transported by human umbilical vein endothelial cells (HUVECs). Afterwards, a rapid, highly selective and sensitive method was established to simultaneously quantify the concentration of C1 and C3 in HUVECs after administration of IPTS. The results demonstrate that pretreatment with IPTS could promote the survival of HUVECs and reduce the damage caused by TNF-α to HUVECs. Among the main 11 components in IPTS, eight components could be absorbed by HUVECs, including seven triterpenoids and one phenolic acid. The uptake of C1 and C3 by HUVECs occurred in a time-, temperature- and concentration-dependent manner, indicating the participation of passive diffusion and active transportation mechanisms. The two triterpenoid saponins all exhibited rapid absorption and a bimodal phenomenon in their concentration-time profiles, and equilibrium could be achieved after 6 h. Furthermore, C1 and C3 intracellular transportation was regulated by serum proteins, sodium-dependent glucose transporter 1 and P-glycoprotein. The current research for the first time demonstrates the in vitro pharmacokinetics characteristics of C1 and C3 in HUVECs lines, which could supply a new way of understanding the treatment of cardiovascular diseases.

Hypericumsampsonii attenuates inflammation in mice with ulcerative colitis via regulation of PDE4/PKA/CREB signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Hypericum sampsonii Hance (Yuanbaocao), a traditional herbal medicine with various pharmacological properties, is traditionally used to treat diarrhea and enteritis in China for hundreds of years. Investigations have uncovered its anti-inflammatory effects and corresponding bioactive constituents in H. sampsonii, however, the mechanisms of action for the treatment of enteritis are still unclear. AIMS OF THE STUDY: This study aims to investigate the therapeutic effects and molecular mechanisms of H. sampsonii in a dextran sulfate sodium (DSS)-induced ulcerative colitis (UC) mice model. MATERIALS AND METHODS: The major ingredients of the ethyl acetate extract (HS) in H. sampsonii were analyzed by UPLC-QTOF-MS. The inflammatory state of UC mice was caused by 3% DSS once daily for seven days. During DSS treatment, the mice in the positive drug group and the other three groups were orally administered 5-ASA (positive control) or HS daily. After treatment with HS or 5-ASA for a week, colonic pathological observation and the molecular biological index were performed for therapeutic evaluation, including visual inspection in the length and weight of colons and spleens, pathological morphology by hematoxylin and eosin (HE) staining, determination of oxidative markers, inflammatory cytokines and tumor necrosis factor-alpha (TNF-α) levels in colonic tissues as well as spleen index. Gene expression levels of inflammatory cytokines, antioxidant enzymes and PDE4 were detected using kits and PCR, while the expression of colonic tight junction proteins and relative signals of PKA/CREB signaling pathway were analyzed by Western blot. RESULTS: The main components in HS were found to be polycyclic polyprenylated acylphloroglucinols (PPAPs). HS distinctly alleviated DSS-stimulated UC-like lesions symptoms as evidenced by a significant recovery from body weight, colon lengths, and histological injuries of colons. HS reduced the accumulation of pro-inflammatory cytokines and improved the mRNA level of IL-10. Simultaneously, the colonic mRNA expression levels of IL-1ß, IL-17, iNOS and COX-2 were all significantly suppressed by HS in a dose-dependent manner. Furthermore, HS restored the protein expression of tight junction-associated protein (ZO-1 and occluding). Besides, HS significantly inhibited the protein level of PDE4 and decreased the expressions of PKA and phosphorylated CREB. CONCLUSION: This is the first work about main composition and anti-UC effect of Hypericum sampsonii Hance. For the first time, this study reveals HS is not toxic in a single dose and exert significantly protective effect in DSS-colitis mice. The underlying mechanisms may involve the improvement to inflammatory status, the protection for intestinal barrier function, the inhibition of PDE4, and the activation of PKA/CREB signaling pathway. This study provided an experimental basis for the traditional application of H. sampsonii Hance in the treatment of diarrhea and dysentery.

Triterpenoid saponins from Ilex pubescens promote blood circulation in blood stasis syndrome by regulating sphingolipid metabolism and the PI3K/AKT/eNOS signaling pathway.

BACKGROUND: Blood stasis syndrome (BSS) is a severe disorder involving disturbances in glycerophosphocholine metabolism. Ilex pubescens (IP) can regulate the levels of lipids, such as lysophosphatidylcholine (LPC) and lysophosphatidylethanolamine (LPE); however, the main active constituent of IP and its corresponding mechanism in BSS treatment are still unclear. PURPOSE: To explore the mechanisms by which triterpenoid saponins of IP (IPTS) promote blood circulation using system pharmacology-based approaches. METHODS: Sprague-Dawley (SD) rat BSS model was prepared by oral administration of IPTS for 7 days followed by adrenaline hydrochloride injection before immersion in ice water. Coagulation parameters in plasma and thromboxane B2 (TXB2), endothelin (ET) and 6-keto-PGF1α in serum were measured. The possible influence on abdominal aortas was evaluated by histopathology assessment. Human vein endothelial cells (HUVECs) were incubated with ox-LDL, and the effects of IPTS on cell viability and LDH release were investigated. UPLC-QTOF-MS/MS was used for metabolic profile analysis of lipid-soluble components in rat plasma and intracellular metabolites in HUVECs. Network pharmacology was used to predict the relevant targets and model pathways of BSS and the main components of IPTS. Molecular docking, molecular dynamics (MD) simulation and biochemical assays were used to predict molecular interactions between the active components of IPTS and target proteins. RT-PCR was used to detect the mRNA level of target proteins. Western blotting and immunohistochemistry (IHC) were used to verify the mechanisms by which IPTS promotes blood circulation in BSS. RESULTS: IPTS improved blood biochemical function in the process of BSS and played a role in vascular protection and maintenance of the normal morphology of blood vessels. Furthermore, metabolite pathways involved in steroid biosynthesis and sphingolipid metabolism were significantly perturbed. Both metabolomics analysis and network pharmacology results showed that IPTS ameliorates vascular injury and that lipid accumulation may be mediated by PI3K/AKT signaling pathway activation. MD simulation and enzyme inhibitory activity results suggested that the main components of IPTS can form stable complexes with PI3K, AKT and eNOS and that the complexes have significant binding affinity. PI3K, AKT, p-AKT, and eNOS mRNA and protein levels were considerably elevated in the IPTS-treated group. Thus, IPTS protects the vasculature by regulating the PI3K/AKT signaling pathway, activating eNOS and increasing the release of NO. CONCLUSION: A possible mechanism by which IPTS prevents BSS is proposed: IPTS can promote blood circulation by modulating sphingolipid metabolism and activating the PI3K/AKT/eNOS signaling pathways.

Bioactive pterocarpans from the root of Astragalus membranaceus var. mongholicus.

Eleven undescribed and three known pterocarpans were isolated and identified from the traditional Chinese medicine "Huang-qi", Astragali Radix (the root of Astragalus membranaceus var. mongholicus (Bunge) P.K.Hsiao). The structures of these pterocarpans were determined using spectroscopic, X-ray crystallographic, quantum chemical calculation, and chemical methods. Pterocarpans, almost exclusively distributed in the family of Leguminosae, are the second largest subgroup of isoflavanoids. However, pterocarpan glycoside number is limited, most of which are glucosides, and only one pterocarpan apioside was isolated from nature. Notably, nine rare apiosyl-containing pterocarpan glycosides were isolated and identified. The hypoglycemic activities of all these compounds were evaluated using α-glucosidase and DPP-IV inhibitory assays respectively, and some isolates displayed the α-glucosidase inhibitory function. The antioxidant activities of all compounds were evaluated using the ORAC and DPPH radical scavenging assays, respectively. All compounds exhibited varying degrees of oxygen radical absorbance capacity, and some compounds displayed DPPH radical scavenging ability.