[Research on mechanism of hypolipidemic effect of Massa Medicata Fermentata based on metabolomics].

This paper aims to study the therapeutic effect of Massa Medicata Fermentata on hyperlipidemia model rats and investigate its mechanism of hypolipidemic effect with the help of non-targeted metabolomics. The mixed hyperlipidemia model rats were constructed by giving high-fat chow. After successful modeling, the rats were divided into the model group, pravastatin sodium group(4.4 mg·kg~(-1)), lipotropic group(0.1 g·kg~(-1)), high-dose group(2.4 g·kg~(-1)), medium-dose group(1.2 g·kg~(-1)), and low-dose group(0.6 g·kg~(-1)) of Massa Medicata Fermentata, and they were administered for four weeks once daily. An equal volume of ultrapure water was given to the blank group and model group. Serum lipid level and liver hematoxylin-eosin(HE) staining were used as indicators to estimate the intervention effect of Massa Medicata Fermentata on mixed hyperlipidemia, and the changes in metabolites in plasma of mixed hyperlipidemia model rats were analyzed by non-targeted metabolomics. The mechanism of the hypolipidemic effect of Massa Medicata Fermentata was analyzed through metabolite pathway enrichment. The results showed that compared with the model group, the Massa Medicata Fermentata administration group, especially the high-dose group, could significantly reduce the content of total cholesterol(TC), triglyceride(TG), and low-density lipoprotein cholesterol(LDL-c)(P<0.05 or P<0.01), and liver HE staining revealed that the number of adipocytes in the high-dose group was reduced to some extent. The potential biomarkers obtained by non-targeted metabolomics screening included glycerol 3-phosphate, sphingomyelin, sphingosine 1-phosphate, and deoxyuridine, which were mainly involved in the sphingolipid metabolism process, glycerophospholipid metabolism process, glycerol ester metabolism pathway, and pyrimidine metabolism pathway, totaling four possible metabolic pathways related to lipid metabolism. This study provides a reference for an in-depth investigation of the hypolipidemic mechanism of Massa Medicata Fermentata, which is of great significance for further promoting the clinical application of Massa Medicata Fermentata and increasing the indications.

[Inhibitory effect and molecular mechanism of sinomenine on human hepatocellular carcinoma HepG2 and SK-HEP-1 cells].

This study aimed to investigate the effect and molecular mechanism of sinomenine on proliferation, apoptosis, metastasis, and combination with inhibitors in human hepatocellular carcinoma HepG2 cells and SK-HEP-1 cells. The effect of sinomenine on the growth ability of HepG2 and SK-HEP-1 cells were investigated by CCK-8 assay, colony formation assay, and BeyoClick~(TM) EdU-488 staining. The effect of sinomenine on DNA damage was detected by immunofluorescence assay, and the effect of sinomenine on apoptosis of human hepatocellular carcinoma cells was clarified by Hoechst 33258 staining and CellEvent~(TM) Cystein-3/7Green ReadyProbes~(TM) reagent assay. Cell invasion assay and 3D tumor cell spheroid invasion assay were performed to investigate the effect of sinomenine on the invasion ability of human hepatocellular carcinoma cells in vitro. The effect of sinomenine on the regulation of protein expression related to the protein kinase B(Akt)/mammalian target of rapamycin(mTOR)/signal transducer and activator of transcription 3(STAT3) signaling pathway in HepG2 and SK-HEP-1 cells was examined by Western blot. Molecular docking was used to evaluate the strength of affinity of sinomenine to the target cysteinyl aspartate specific proteinase-3(caspase-3) and STAT3, and combined with CCK-8 assay to detect the changes in cell viability after combination with STAT3 inhibitor JSI-124 in combination with CCK-8 assay. The results showed that sinomenine could significantly reduce the cell viability of human hepatocellular carcinoma cells in a concentration-and time-dependent manner, significantly inhibit the clonogenic ability of human hepatocellular carcinoma cells, and weaken the invasive ability of human hepatocellular carcinoma cells in vitro. In addition, sinomenine could up-regulate the cleaved level of poly ADP-ribose polymerase(PARP), a marker of apoptosis, and down-regulate the protein levels of p-Akt, p-mTOR, and p-STAT3 in human hepatocellular carcinoma cells. Molecular docking results showed that sinomenine had good affinity with the targets caspase-3 and STAT3, and the sensitivity of sinomenine to hepatocellular carcinoma cells was diminished after STAT3 was inhibited. Therefore, sinomenine can inhibit the proliferation and invasion of human hepatocellular carcinoma cells and induce apoptosis, and the mechanism may be attributed to the activation of caspase-3 signaling and inhibition of the Akt/mTOR/STAT3 pathway. This study can provide a new reference for the in-depth research and clinical application of sinomenine and is of great significance to further promote the scientific development and utilization of sinomenine.

[Morin induces autophagy and apoptosis in hepatocellular carcinoma cells through Akt/mTOR/STAT3 pathway].

This study investigated the effect and mechanism of morin in inducing autophagy and apoptosis in hepatocellular carcinoma cells through the protein kinase B(Akt)/mammalian target of rapamycin(mTOR)/signal transducer and activator of transcription protein 3(STAT3) pathway. Human hepatocellular carcinoma SK-HEP-1 cells were stimulated with different concentrations of morin(0, 50, 100, 125, 200, and 250 µmol·L~(-1)). The effect of morin on the viability of SK-HEP-1 cells was detected by Cell Counting Kit-8(CCK-8). The effect of morin on the proliferation and apoptosis of SK-HEP-1 cells was investigated using colony formation assay, flow cytometry, and BeyoClick~(TM) EdU-488 with different concentrations of morin(0, 125, and 250 µmol·L~(-1)). The changes in the autophagy level of cells treated with morin were examined by transmission electron microscopy and autophagy inhibitors. The impact of morin on the expression levels of proteins related to the Akt/mTOR/STAT3 pathway was verified by Western blot. Compared with the control group, the morin groups showed decreased viability of SK-HEP-1 cells in a time-and concentration-dependent manner, increased number of apoptotic cells, up-regulated expression level of apoptosis marker PARP, up-regulated phosphorylation level of apoptosis-regulating protein H2AX, decreased number of positive cells and the colony formation rate, an upward trend of expression levels of autophagy-related proteins LC3-Ⅱ, Atg5, and Atg7, and decreased phosphorylation levels of Akt, mTOR, and STAT3. These results suggest that morin can promote apoptosis, inhibit proliferation, and induce autophagy in hepatocellular carcinoma cells, and its mechanism of action may be related to the Akt/mTOR/STAT3 pathway.

Effect of Shengmai Yin on Epithelial-Mesenchymal Transition of Nasopharyngeal Carcinoma Radioresistant Cells.

OBJECTIVE: To investigate the mechanism by which Chinese medicine Shengmai Yin (SMY) reverses epithelial-mesenchymal transition (EMT) through lipocalin-2 (LCN2) in nasopharyngeal carcinoma (NPC) cells CNE-2R. METHODS: Morphological changes in EMT in CNE-2R cells were observed under a microscope, and the expressions of EMT markers were detected using quantitative real-time PCR (RT-qPCR) and Western blot assays. Through the Gene Expression Omnibus dataset and text mining, LCN2 was found to be highly related to radiation resistance and EMT in NPC. The expressions of LCN2 and EMT markers following SMY treatment (50 and 100 µ g/mL) were detected by RT-qPCR and Western blot assays in vitro. Cell proliferation, migration, and invasion abilities were measured using colony formation, wound healing, and transwell invasion assays, respectively. The inhibitory effect of SMY in vivo was determined by observing a zebrafish xenograft model with a fluorescent label. RESULTS: The CNE-2R cells showed EMT transition and high expression of LCN2, and the use of SMY (5, 10 and 20 µ g/mL) reduced the expression of LCN2 and reversed the EMT in the CNE-2R cells. Compared to that of the CNE-2R group, the proliferation, migration, and invasion abilities of SMY high-concentration group were weakened (P<0.05). Moreover, SMY mediated tumor growth and metastasis in a dose-dependent manner in a zebrafish xenograft model, which was consistent with the in vitro results. CONCLUSIONS: SMY can reverse the EMT process of CNE-2R cells, which may be related to its inhibition of LCN2 expression. Therefore, LCN2 may be a potential diagnostic marker and therapeutic target in patients with NPC.

Bioaccessibility of carotenoids and antioxidant capacity of seed-used pumpkin byproducts powders as affected by particle size and corn oil during in vitro digestion process.

Powders made from seed-used pumpkin flesh (SUPF) are potential sources of carotenoids. In this study, unexplored effects of particle size and corn oil on bioaccessible amounts of carotenoids and antioxidant capacity of SUPF powders during in vitro digestion process were investigated. Overall, total carotenoid relative bioaccessibility (TCRB) of 100 mesh-sized powder (100 MP, 15.46%) was higher than that of 18 mesh-sized powder (18 MP, 12.94%). With the addition of 2% corn oil, TCRB increased 108.35% (18 MP) and 88.55% (100 MP), respectively. Lutein (≥27160 µg/100 g) and ß-carotene (≥5192 µg/100 g) were main carotenoid monomers in SUPF and significantly correlated with DPPH radical scavenging activity of digestive supernatant (p < 0.05). Notably, DPPH radical scavenging activity of 18 MP increased 96.54% with corn oil. These results implied that smaller particle size and oil addition could improve bioaccessible amounts of carotenoids and antioxidant capacity of SUPF powders.

[Inheritance and development of traditional Mongolian medicine processing].

The processing of Mongolian medicine,which is called " mort harl" in Mongolian language,refers to a traditional processing technology to " tame" some toxic,aggressive,ineffective or inconvenient Mongolian medicines,so as to make it " compliant" to clinical needs. It is the summary of long-term experience in drug preparation by Mongolian medicine experts,one of the bridges for the dialectical unity of Mongolian medicine,the essential content in evaluation of the clinical efficacy of Mongolian medicine and the study of Mongolian medicine modernization,and also the important soft power carrier of " intangible cultural heritage" and " grassland culture" in Inner Mongolia autonomous region. In this study,the processing history,purpose,crafts,mechanism,processing standards and quality standards of Mongolian medicine were explained,and some suggestions were proposed for the problems of the Mongolian medicine processing and development: focus on the basic theory of Mongolian medicine and the clinical experience of Mongolian medicine in the development of traditional Mongolian medicine processing; strengthen the literature research on the processing method of Mongolian medicine; establish comprehensive and systematic Mongolian medicine concocts standards and quality standards; enhance the research and development of special processing equipment and process quality control instruments for Mongolian medicine; and strengthen the training of professional technicians,the protection of copyright in Mongolian medicine processing,and scientific research on Mongolian medicine processing. In the inheritance of the tradition,the latest achievements of modern scientific development can be also absorbed to provide reference for the further development of traditional Mongolian medicine processing technology.

[Effects of Shengmai Jianghuang San on intestinal flora in nude mice with radio resistant cells of nasopharyngeal carcinoma].

Modern pharmacological studies have shown that Shengmai San has the effects of enhancing immunity and improving blood circulation, and Curcumae Longae Rhizoma(Jianghuang) has anti-inflammatory, anti-cancer, anti-oxidation and other functions. Shengmai San combined with Jianghuang is a new research direction in the study of anti-tumor of traditional Chinese medicines. The main treatment for nasopharyngeal carcinoma is radiation therapy, but radiation therapy can cause a variety of side effects, and it also changes the composition of the intestinal flora. In this study, the 16 s rDNA sequencing platform was used to perform macro-sequence sequencing of the intestinal flora samples of nude mice bearing the veins of Shengmai Jianghuang San, and then the results of intestinal flora data were analyzed to investigate the effect of Shengmai Jianghuang San on tumors. The results showed that Shengmai Jianghuang San combined with irradiation could enhance the therapeutic effect of tumor treatment. Radiation therapy would reduce the total number and diversity of intestinal flora in nude mice, and also change the structure of the flora. Shengmai Jianghuang San could protect the diversity of colonies, and also partially restore the colony imbalance caused by irradiation. This study provides a research idea for Shengmai Jianghuang San as a sensitizing adjuvant for radiotherapy of nasopharyngeal carcinoma.

Fructus Ligustri Lucidi ethanol extract inhibits osteoclastogenesis in RAW264.7 cells via the RANKL signaling pathway.

Fructus ligustri Lucidi (FLL) is the fruit of Ligustrum lucidum Ait and a traditional Chinese medicine, primarily known for its role in osteoporosis prevention and treatment. The present study aimed to elucidate the effect and underlying mechanism of action of ethanol extract of FLL on osteoclast differentiation and bone resorption, and to identify the active compounds within it. RAW264.7 murine monocyte/macrophage cells were stimulated with the receptor activator of nuclear factor κB ligand (RANKL) to induce osteoclast differentiation in vitro. The present study demosntrated that FLL extract and its two primary components, oleanolic acid (OA) and ursolic acid (UA), significantly suppressed RANKL­induced tartrate resistant acid phosphatase (TRAP) activity and multinucleate osteoclast formation without inducing cytotoxicity; however, no effect was observed on the apoptosis of mature osteoclasts. Additionally, RANKL­induced mRNA expression levels of the key transcription factors, tumor necrosis factor receptor associated factor­6, nuclear factor of activated T cell­c1 and c­Fos, and the osteoclast markers, TRAP, cathepsin K and matrix metalloproteinase­9 were suppressed by FLL, OA and UA. However, no effect was observed on RANKL­induced mRNA expression levels of Src. These results demonstrated that FLL may inhibit osteoclastogenesis in RAW264.7 cells via RANKL signaling pathways. OA and UA are active compounds in inducing this effect; however, their specific roles remain to be elucidated.

Fructus ligustri lucidi ethanol extract improves bone mineral density and properties through modulating calcium absorption-related gene expression in kidney and duodenum of growing rats.

Optimizing peak bone mass in early life is one of key preventive strategies against osteoporosis. Fructus ligustri lucidi (FLL), the fruit of Ligustrum lucidum Ait., is a commonly prescribed herb in many kidney-tonifying traditional Chinese medicinal formulas to alleviate osteoporosis. Previously, FLL extracts have been shown to have osteoprotective effect in aged or ovariectomized rats. In the present study, we investigated the effects of FLL ethanol extract on bone mineral density (BMD) and mechanical properties in growing male rats and explored the underlying mechanisms. Male weaning Sprague-Dawley rats were randomized into four groups and orally administrated for 4 months an AIN-93G formula-based diet supplementing with different doses of FLL ethanol extract (0.40, 0.65, and 0.90 %) or vehicle control, respectively. Then calcium balance, serum level of Ca, P, 25(OH)2D3, 1,25(OH)2D3, osteocalcin (OCN), C-terminal telopeptide of type I collagen (CTX-I), and parathyroid hormone, bone microarchitecture, and calcium absorption-related genes expression in duodenum and kidney were analyzed. The results demonstrated that FLL ethanol extract increased BMD of growing rats and improved their bone microarchitecture and mechanical properties. FLL ethanol extract altered bone turnover, as evidenced by increasing a bone formation maker, OCN, and decreasing a bone resorption maker, CTX-I. Intriguingly, both Ca absorption and Ca retention rate were elevated by FLL ethanol extract treatment, possibly through the mechanisms of up-regulating the transcriptions of calcitropic genes in kidney (1α-hydroxylase) and duodenum (vitamin D receptor, calcium transporter calbindin-D9k, and transient receptor potential vanilloid 6). In conclusion, FLL ethanol extract increased bone mass gain and improved bone properties via modulating bone turnover and up-regulating calcium absorption-related gene expression in kidney and duodenum, which could then activate 1,25(OH)2D3-dependent calcium transport in male growing rats.

Fructus Ligustri Lucidi (FLL) ethanol extract increases bone mineral density and improves bone properties in growing female rats.

Osteoporosis is a chronic disease affecting millions of people worldwide. It is generally accepted that acquisition of a high peak bone mass (PBM) early in life can reduce the risk of osteoporosis later in life. The aims of this study were to investigate the effects of Fructus Ligustri Lucidi (FLL) ethanol extract on bone mineral density and its mechanical properties in growing female rats and to explore the underlying mechanisms. The rats were given different doses of FLL extract mixed with AIN-93G formula (0.40, 0.65 and 0.90 %), and a group given AIN-93G diet treatment only was used as control. The intervention lasted for 16 weeks until the animals were about 5 months old, the time when the animals almost reach their PBM. Our results showed that FLL treatment increased bone mineral density and improved bone mechanical properties in the growing female rats in a dose-dependent manner. In addition, FLL treatment significantly decreased the serum bone-resorbing marker, CTX-I, while significantly increasing serum 25(OH)D3 and thereby increasing Ca absorption and Ca retention. Intriguingly, both in vivo and in vitro results demonstrated that FLL treatment could reduce the RANKL/OPG ratio. In conclusion, FLL ethanol extract exerted beneficial effects on peak bone mass acquisition and the improvement of bone mechanical properties by favoring Ca metabolism and decreasing the RANKL/OPG ratio.