Nine compounds were isolated and purified from 90% ethanol extract of Alstonia mairei Lévl by using various chromatographic methods, including silica gel, SephadexLH-20, MCI Gel and ODS column chromatography, combined with semi-preparative liquid phase separation methods. Modern spectroscopic methods (1D and 2D NMR, UV, IR, MS, etc.) were used to identify the structures of the isolated compounds. They were identified as mairoside A (1), 3′,6-di-O-sinaloylsucrose (2), myristic acid (3), methyl myristate (4), ethyl myristate (5), 3,4,5-trimethoxycinnamic acid (6), 3,4,5-trimethoxybenzoic acid (7), vinoline (8), kaempferol-3-O-rutinoside (9), among which compound 1 is a new glycoside, compounds 4 and 5 are new natural products, and the nuclear magnetic data of compound 4 were reported for the first time.

This study aims to investigate the potential effect of the water extract of fresh Rehmanniae Radix on hypercholesterolemia in mice that was induced by a high-fat and high-cholesterol diet and explore its possible mechanism from bile acid reabsorption. Male C57BL/6 mice were randomly assigned into the following groups: control, model, low-and high-dose(4 and 8 g·kg~(-1), respectively) fresh Rehmanniae Radix, and positive drug(simvastatin, 0.05 g·kg~(-1)). Other groups except the control group were fed with a high-fat and high-cholesterol diet for 6 consecutive weeks to induce hypercholesterolemia. From the 6th week, mice were administrated with corresponding drugs daily via gavage for additional 6 weeks, while continuing to be fed with a high-fat and high-cholesterol diet. Serum levels of total cholesterol(TC), triglycerides(TG), low density lipoprotein-cholesterol(LDL-c), high density lipoprotein-cholesterol(HDL-c), and total bile acid(TBA), as well as liver TC and TG levels and fecal TBA level, were determined by commercial assay kits. Hematoxylin-eosin(HE) staining, oil red O staining, and transmission electron microscopy were performed to observe the pathological changes in the liver. Three livers samples were randomly selected from each of the control, model, and high-dose fresh Rehmanniae Radix groups for high-throughput transcriptome sequencing. Differentially expressed genes were mined and KEGG pathway enrichment analysis was performed to predict the key pathways and target genes of the water extract of fresh Rehmanniae Radix in the treatment of hypercholesterolemia. RT-qPCR was employed to measure the mRNA levels of cholesterol 7α-hydroxylase(CYP7A1) and cholesterol 27α-hydroxylase(CYP27A1) in the liver. Western blot was employed to determine the protein levels of CYP7A1 and CYP27A1 in the liver as well as farnesoid X receptor(FXR), apical sodium-dependent bile acid transporter(ASBT), and ileum bile acid-binding protein(I-BABP) in the ileum. The results showed that the water extract of fresh Rehmanniae Radix significantly lowered the levels of TC and TG in the serum and liver, as well as the level of LDL-c in the serum. Conversely, it elevated the level of HDL-c in the serum and TBA in feces. No significant difference was observed in the level of TBA in the serum among groups. HE staining, oil red O staining, and transmission electron microscopy showed that the water extract reduced the accumulation of lipid droplets in the liver. Further mechanism studies revealed that the water extract of fresh Rehmanniae Radix significantly down-regulated the protein levels of FXR and bile acid reabsorption-related proteins ASBT and I-BABP. Additionally, it enhanced CYP7A1 and CYP27A1, the key enzymes involved in bile acid synthesis. Therefore, it is hypothesized that the water extract of fresh Rehmanniae Radix may exert an anti-hypercholesterolemic effect by regulating FXR/ASBT/I-BABP signaling, inhibiting bile acid reabsorption, and increasing bile acid excretion, thus facilitating the conversion of cholesterol to bile acids.
Animals ; Male ; Bile Acids and Salts/metabolism* ; Mice, Inbred C57BL ; Mice ; Diet, High-Fat/adverse effects* ; Cholesterol/metabolism* ; Drugs, Chinese Herbal/administration & dosage* ; Hypercholesterolemia/genetics* ; Receptors, Cytoplasmic and Nuclear/genetics* ; Rehmannia/chemistry* ; Liver/drug effects* ; Humans ; Cholesterol 7-alpha-Hydroxylase/genetics* ; Plant Extracts

This study aimed to explore the effects and mechanisms of total extracts from Anthriscus sylvestris on pulmonary hypertension in rats. Sixty male SD rats were divided into normal(NC) group, model(M) group, positive drug sildenafil(Y) group, low-dose A. sylvestris(ES-L) group, medium-dose A. sylvestris(ES-M) group, and high-dose A. sylvestris(ES-H) group. On day 1, rats were intraperitoneally injected with monocrotaline(60 mg·kg~(-1)) to induce pulmonary hypertension, and the rat model was established on day 28. From days 15 to 28, intragastric administration of the respective treatments was performed. After modeling and treatment, small animal echocardiography was used to detect the right heart function of the rats. Arterial blood gas was measured using a blood gas analyzer. Hematoxylin and eosin(HE) staining and Masson staining were performed to observe cardiopulmonary pathological damage. Flow cytometry was used to detect apoptosis in the lung and myocardial tissues and reactive oxygen species(ROS) levels. Western blot was applied to detect the expression levels of transforming growth factor-β1(TGF-β1), phosphorylated mothers against decapentaplegic homolog 3(p-Smad3), Smad3, tissue plasminogen activator(t-PA), and plasminogen activator inhibitor-1(PAI-1) in lung tissue. A blood routine analyzer was used to measure inflammatory immune cell levels in the blood. Enzyme-linked immunosorbent assay(ELISA) was used to detect the expression levels of P-selectin and thromboxane A2(TXA2) in plasma. The results showed that, compared with the NC group, right heart hypertrophy index, right ventricular free wall thickness, right heart internal diameter, partial carbon dioxide pressure(PaCO_2), apoptosis in cardiopulmonary tissue, and ROS levels were significantly increased in the M group. In contrast, the ratio of pulmonary blood flow acceleration time(PAT)/ejection time(PET), right cardiac output, change rate of right ventricular systolic area, systolic displacement of the tricuspid ring, oxygen partial pressure(PaO_2), and blood oxygen saturation(SaO_2) were significantly decreased in the M group. After administration of the total extract of A. sylvestris, right heart function and blood gas levels were significantly improved, while apoptosis in cardiopulmonary tissue and ROS levels significantly decreased. Further testing revealed that the total extract of A. sylvestris significantly decreased the levels of interleukin-1β(IL-1β), interleukin-6(IL-6), and PAI-1 proteins in lung tissue, while increasing the expression of t-PA. Additionally, the extract reduced the levels of inflammatory cells such as leukocytes, lymphocytes, granulocytes, and monocytes in the blood, as well as the levels of P-selectin and TXA2 in plasma. Metabolomics results showed that the total extract of A. sylvestris significantly affected metabolic pathways, including arginine biosynthesis, tyrosine metabolism, and taurine and hypotaurine metabolism. In conclusion, the total extract of A. sylvestris may exert an anti-pulmonary hypertension effect by inhibiting the TGF-β1/Smad3 signaling pathway, thereby alleviating immune-inflammatory responses and thrombosis.
Animals ; Male ; Smad3 Protein/metabolism* ; Transforming Growth Factor beta1/metabolism* ; Rats, Sprague-Dawley ; Rats ; Signal Transduction/drug effects* ; Hypertension, Pulmonary/genetics* ; Thrombosis/immunology* ; Drugs, Chinese Herbal/administration & dosage* ; Humans ; Apoptosis/drug effects*

OBJECTIVE: To investigate the effects of oridonin on platelet function and related mechanisms. METHODS: Washed platelets from healthy adults and mice were incubated with different concentrations of oridonin (2.5, 5 and 10 μmol/L) in vitro . The surface expression level of P-selectin and the activation of integrin αIIbβ3 in platelets were detected by flow cytometry, and the aggregation ability of platelets under the stimulation by various agonists was detected by light transmission aggregometry. The expression of P-AKT (Ser473) was detected by protein immunoblotting. Arterial thrombosis model was established in mice with mesenteric injury induced by ferric chloride, and tail hemorrhage model was established by cutting off the tail of mice. The effect of intraperitoneal injection of oridonin (10 mg/kg) on thrombosis and haemostasis was tested. RESULTS: Oridonin inhibited platelet P-selectin expression and integrin αIIbβ3 activation. In the presence of different stimulants, oridonin inhibited platelet aggregation in a concentration-dependent manner. The phosphorylation level of AKT Ser473 was reduced in the groups treated with different concentrations of oridonin. Oridonin significantly prolonged the time of mesenteric artery thrombosis in mice, but did not affect the tail bleeding time. CONCLUSION Oridonin inhibits platelet activation, aggregation, and thrombosis by inhibiting AKT phosphorylation, and may be used as a potential antiplatelet drug.
Diterpenes, Kaurane/pharmacology* ; Animals ; Mice ; Blood Platelets/drug effects* ; Platelet Aggregation/drug effects* ; P-Selectin/metabolism* ; Thrombosis ; Platelet Glycoprotein GPIIb-IIIa Complex/metabolism* ; Proto-Oncogene Proteins c-akt/metabolism* ; Humans ; Phosphorylation ; Platelet Activation/drug effects*

OBJECTIVE: To investigate the protective effect of achyranthes bidentata (AB) on sperm quality in mice with spermatogenic disorder through the glycolytic metabolic pathway and its action mechanism. METHODS: We equally randomized 40 Kunming mice into a normal control, a model control, a low-dose AB (3.5 g/kg) and a high-dose AB group (7.0 g/kg), and established the model of spermatogenic disorder in the latter three groups of mice by intraperitoneal injection of doxorubicin (30 mg/kg). Two days after modeling, we collected the testis and kidney tissues and blood samples from the mice for observation of the pathological changes in the testis tissue by HE staining, detection of perm motility with the sperm quality analyzer, examination of the apoptosis of testis cells by flow cytometry, measurement of the levels of testosterone (T), malondialdehyde (MDA), superoxide dismutase (SOD) and catalase (CAT) in the serum and testis tissue by ELISA, and determination of expressions of the key enzymes of glycolysis hexokinase Ⅱ (HK2), pyruvate kinase M2 (PKM2), platelet phosphofructokinase (PFKP), lactate dehydrogenase A (LDHA) and the meiosis proteins REC8 and SCP3 by Western blot, and the mRNA expressions of glycolytic phosphofructokinase 1 (PFK1), phosphoglycerate kinase 1 (PGK1), tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) by fluorescence quantitative PCR (FQ-PCR). RESULTS: Compared with the model controls, the mice in the AB groups showed significant increases in the testis coefficient, kidney index, sperm concentration, sperm motility, spermatogonia, primary spermatocytes, spermatids, sperm count and the serum T level (P<0.05 or P<0.01), but dramatic decreases in the apoptosis of testis cells and percentage of morphologically abnormal sperm (P<0.01). Achyranthes bidentata also significantly elevated the levels of SOD and CAT, and down-regulated the mRNA expressions of MDA, TNF-α and IL-1β (P<0.05 or P<0.01), and up-regulated the protein expressions of HK2, PKM2, PFKP, LDHA, REC8 and SCP3, and expressions of the glycolysis key genes Pfk1 and Pgk1 (P<0.05 or P<0.01). CONCLUSION Achyranthes bidentata ameliorates doxorubicin-induced spermatogenic disorder in mice by regulating the glycolytic pathway and reducing oxidative stress and the expressions of inflammatory factors.
Glycolysis/drug effects* ; Doxorubicin/toxicity* ; Spermatogenesis/drug effects* ; Random Allocation ; Male ; Animals ; Mice ; Disease Models, Animal ; Achyranthes/chemistry* ; Spermatozoa/pathology* ; Oxidative Stress/drug effects* ; Primary Cell Culture ; Apoptosis/drug effects* ; Sperm Motility/drug effects* ; Testis/pathology* ; Infertility, Male/prevention & control* ; Medicine, Chinese Traditional/methods* ; Animals, Outbred Strains

ObjectiveTo explore the mechanism of treadmill exercise against type 2 diabetes mellitus (T2DM) with non-alcoholic fatty liver disease (NAFLD) based on the regulator effects of exercise on ferroptosis. MethodsEight 8-week-old male m/m mice were used as control group (Con, n=8), and db/db mice of the matched age were randomly divided into T2DM model group (db/db, n=8), exercise group (db+Exe, n=8), p38 mitogen-activated protein kinase (MAPK) inhibitor group (db+SB203580, n=8) and exercise combined with p38 MAPK inhibitor group (db+Exe+SB203580, n=8). After one-week adaptive feeding, the mice in the db+Exe group and db+Exe+SB203580 group underwent moderate intensity treadmill exercise for 40 min/d, 5 d/week lasting 8 weeks. The db+SB203580 group and db+Exe+SB203580 group were treated with SB203580 (a specific inhibitor of p38 MAPK) with a dose of 5 mg/kg, 5 d/week for 8 weeks. And the exercise intervention was performed 2 h later after the intraperitoneal injection of SB203580. The body weight and fasting blood glucose of mice were measured regularly every week during the experiment. After 24 h of the last intervention, the mice were weighted, the liver tissues were taken, weighted and the liver index was calculated. The pathological changes of liver were determined by Oil Red O and hematoxylin-eosin (HE) staining. The levels of blood lipids, liver function, Fe²⁺ and oxidative stress markers of liver were measured by enzyme linked immunosorbent assay (ELISA). The related mRNA expression levels of lipogenesis and inflammation were evaluated by quantitative reverse transcriptase-mediated PCR (qRT-PCR). The related protein expression levels of lipogenesis and ferroptosis in liver were determined by immunohistochemical (IHC) staining and Western blot. ResultsThe body weight, fasting blood glucose, liver index, blood lipid and transaminase levels in the db/db group were significantly increased compared with the Con group. HE and Oil Red O staining showed severe lipid accumulation and ballooning change in the liver of db/db mice. Biochemical tests showed that Fe²⁺ and MDA level of liver constitution homogenate increased, while GSH level decreased significantly. The results of qRT-PCR showed that the mRNA levels of MCP-1, IL-6, SREBF1 and ACC1 in liver tissue of db/db mice were all significantly increased. Western blot results showed that the expression levels of SREBF1, ACC1 increased, ferroptosis relative proteins were significantly decreased. The 8 weeks of exercise significantly reduced the rise in body weight, blood glucose, liver index and blood lipid levels in db/db mice. Exercise intervention also alleviated hepatic steatosis and reduced the expression levels of Fe²⁺, MDA, MCP-1, IL-6, ACC1 and SREBF1, upregulated the expression levels of GSH, NRF2, HO-1, SLC7A11 and GPX4 in liver tissue of db/db mice. The intervention of exercise combined with SB203580 significantly down-regulated the mRNA expression levels of ACC1, MCP-1, IL-6, reduced the levels of Fe²⁺ and MDA, and up-regulated the level of GSH in db/db mice. Compared with the db+Exe group, the expression of Fe²⁺, MDA, MCP-1, and SREBF1 in the liver of the db+Exe+SB203580 group mice significantly increased, while the expression level of GSH and expression levels of ferroptosis relative proteins also significantly decreased. In addition, compared with db+SB203580 group, the iron accumulation and lipid peroxidation in the liver of db+Exe+SB203580 group were significantly improved. ConclusionThe8-week treadmill exercise can effectively alleviate liver injury and steatosis, and its mechanism may be related to the inhibition of hepatocyte ferroptosis through p38 MAPK signal.

Ten compounds were isolated and purified from the dichloromethane extract of stems of Ephedra intermedia by various chromatographic methods. Based on the analysis of physicochemical properties and spectral data, the structures of the ten compounds were identified as 1-allyl-3,4-dimethoxy-benzene-5-O-β-D-glucopyranoside (1), lyoniresinol (2), secoisolariciresinol (3), 4,3′,4′-trihydroxy-3-methoxylignan-9,9′-diyldiacetate (4), dehydroconiferyl alcohol (5), isocubebin (6), balanophonin B (7), sesquipinsapol B (8), crataegifin A (9), 1-(2-(4-hydroxy-3-methoxyphenyl)-3-(hydroxymethyl)-7-methoxy-2,3-dihydrobenzofuran-5-yl)ethan-1-one (10). Compound 1 is a new compound, named as 1-allyl-3,4-dimethhoxy-benzene-5-O-β-D-glucopyranosyl, compounds 2-10 are lignans and isolated from this plant for the first time. Compound 3, 4, 8 possess potentially anti-asthmatic activities.

Quercetin can mediate the activation of mitogen-activated protein kinase(MAPK)signaling pathways to prevent osteoporosis(OP).This paper comprehensively discusses the interrelationship between MAPK and osteoporosis-related cells based on the latest domestic and international research.Additionally,it elucidates the research progress of quercetin in mediating the MAPK signaling pathway for OP prevention.The aim is to provide an effective foundation for the clinical prevention and treatment of OP and the in-depth development of quercetin.

Platycodonis Radix is the dry root of Platycodon grandiflorum of Campanulaceae, which has a variety of pharmacological effects and is a commonly used bulk Chinese medicine. In this study, the chloroplast genome sequences of six P. grandiflorum from different producing areas has been sequenced with Illumina HiSeq X Ten platform. The specific DNA barcodes were screened, and the germplasm resources and genetic diversity were analyzed according to the specific barcodes. The total length of the chloroplast genome of 6 P. grandiflorum samples was 172 260-172 275 bp, and all chloroplast genomes showed a typical circular tetrad structure and encoded 141 genes. The comparative genomics analysis and results of amplification efficiency demonstrated that trnG-UCC and ndhG_ndhF were the potential specific DNA barcodes for identification the germplasm resources of P. grandiflorum. A total of 305 P. grandiflorum samples were collected from 15 production areas in 9 provinces, for which the fragments of trnG-UCC and ndhG_ndhF were amplificated and the sequences were analyzed. The results showed that trnG-UCC and ndhG_ndhF have 5 and 11 mutation sites, respectively, and 5 and 7 haplotypes were identified, respectively. The combined analysis of the two sequences formed 13 haplotypes (named Hap1-Hap13), and Hap4 is the main genotype, followed by Hap1. The unique haplotypes possessed by the three producing areas can be used as DNA molecular tags in this area to distinguish from the germplasm resources of P. grandiflorum from other areas. The haplotype diversity, nucleotide diversity and genetic distance were 0.94, 4.79×10^-3 and 0.000 0-0.020 3, respectively, suggesting that the genetic diversity was abundant and intraspecific kinship was relatively close. This study laid a foundation for the identification of P. grandiflorum, the protection and utilization of germplasm resources, and molecular breeding.

Objective To evaluate the patient-reported outcome(PRO)of patients with breast cancer who underwent autologous breast reconstruction and implant breast reconstruction.Methods Patients who underwent breast reconstruction in Shanghai Cancer Center,Fudan University from Jan 2020 to Jun 2021 were selected,including 111 patients who underwent autologous breast reconstruction and 108 patients who underwent implant breast reconstruction.Chinese version Breast-Q2.0 scale,breast cancer specificity scale QLQ-BR23 and EORTC quality of life scale QLQ-C30 were used to investigate the PRO of the two groups 18 months after operation.Results The rate of stage Ⅲ breast cancer in the self-weight construction group was higher than that in the implant reconstruction group(64.9%vs.44.4%,P<0.001).The preoperative neoadjuvant therapy and postoperative radiotherapy in the autologous reconstruction group were higher than those in the implant reconstruction group(P<0.001).Postoperative chemotherapy and endocrine therapy in the autologous reconstruction group were lower than those in the implant reconstruction group(P<0.001).The study based on Breast-Q scale showed that the breast satisfaction of autologous reconstruction group was higher than that of implant reconstruction(59.28±17.20 vs.54.94±14.48,P<0.05).The study based on QLQ-BR23 showed that the self-weight construction group was higher than the implant reconstruction group in the field of arm symptoms(20.02±20.80 vs.12.65±16.18,P<0.05).The study based on QLQ-C30 scale showed that there was no significant difference in all functional areas and symptom areas of patients.There was no significant difference in the number and time of social regression between the two groups.Conclusion Breast reconstruction can improve the PRO of breast cancer patients,and oncology factors will affect the choice of breast reconstruction.Patients with autologous breast reconstruction are more satisfied with breast appearance,but upper limb symptoms such as swelling and pain are more obvious than implant reconstruction,which is related to the higher proportion of axillary lymph node dissection in patients with autologous reconstruction.There is no significant difference in quality of life and social regression between the two groups.