Organoid forming potential as complementary parameter for accurate evaluation of breast cancer neoadjuvant therapeutic efficacy.

BACKGROUND: 13-15% of breast cancer/BC patients diagnosed as pathological complete response/pCR after neoadjuvant systemic therapy/NST suffer from recurrence. This study aims to estimate the rationality of organoid forming potential/OFP for more accurate evaluation of NST efficacy. METHODS: OFPs of post-NST residual disease/RD were checked and compared with clinical approaches to estimate the recurrence risk. The phenotypes of organoids were classified via HE staining and ER, PR, HER2, Ki67 and CD133 immuno-labeling. The active growing organoids were subjected to drug sensitivity tests. RESULTS: Of 62 post-NST BC specimens, 24 were classified as OFP-I with long-term active organoid growth, 19 as OFP-II with stable organoid growth within 3 weeks, and 19 as OFP-III without organoid formation. Residual tumors were overall correlated with OFP grades (P < 0.001), while 3 of the 18 patients (16.67%) pathologically diagnosed as tumor-free (ypT0N0M0) showed tumor derived-organoid formation. The disease-free survival/DFS of OFP-I cases was worse than other two groups (Log-rank P < 0.05). Organoids of OFP-I/-II groups well maintained the biological features of their parental tumors and were resistant to the drugs used in NST. CONCLUSIONS: The OFP would be a complementary parameter to improve the evaluation accuracy of NST efficacy of breast cancers.

[Effect and mechanism of Danggui Buxue Decoction-containing serum in mitigating H9c2 cell injury caused by exposure to intermittent low oxygen].

This study aims to explore the effect and mechanism of Danggui Buxue Decoction(DBD)-containing serum in alleviating the H9c2 cell injury caused by the exposure to intermittent low oxygen. H9c2 cells were assigned into five groups: control(CON) group, intermittent low oxygen(IH) group, intermittent low oxygen plus DBD-containing serum(IH+DBD) group, intermittent low oxygen plus the autophagy enhancer rapamycin(IH+RAPA) group, and intermittent low oxygen plus DBD-containing serum and the autophagy inhibitor 3-methyladenine(IH+DBD+3-MA) group. Monodansylcadaverine(MDC) staining was employed to detect the changes of autophagosomes. Cell counting kit-8(CCK-8) assay was employed to determine the activity of myocardial cells, and lactate dehydrogenase(LDH) and creatine kinase(CK) kits were used to measure the LDH and CK levels in the cell culture, which would reflect the degree of cell damage. TdT-mediated dUTP nick-end labeling(TUNEL) staining was used to detect the apoptosis of myocardial cells, and JC-1 fluorescence probe to detect the changes in mitochondrial membrane potential. Western blot was employed to determine the expression levels of the autophagy-related proteins microtubule-associated proteins light chain 3Ⅱ(LC3Ⅱ), microtubule-associated proteins light chain 3Ⅰ(LC3Ⅰ), P62, Parkin and apoptosis related proteins pro caspase-3, caspase-3, B-cell lymphoma-2(Bcl-2), Bcl-2-associated X(Bax). The results showed that compared with the CON group, the IH group showed decreased fluorescence intensity of MDC staining, decreased LC3Ⅱ/LC3Ⅰ ratio, down-regulated Parkin expression, and up-regulated expression of P62. In addition, the IH group showed decreased cell survival rate, increased content of LDH and CK in the culture medium, increased number of TUNEL positive cells, and decreased pro caspase-3/caspase-3 and Bcl-2/Bax ratios and mitochondrial membrane potential. Compared with the IH group, the IH+DBD and IH+RAPA groups showed increased fluorescence intensity of MDC staining, increased LC3Ⅱ/LC3Ⅰ ratio, up-regulated Parkin expression, and down-regulated P62 expression. In addition, the two groups showed increased cell survival rate, reduced content of LDH and CK in the culture medium, decreased number of TUNEL positive cells, and increased pro caspase-3/caspase-3 and Bcl-2/Bax ratios and mitochondrial membrane potential. The IH+DBD+3-MA and IH groups showed no significant differences in the above indicators. Compared with the IH+DBD group, the IH+DBD+3-MA group showed decreased fluorescence intensity of MDC staining, decreased LC3Ⅱ/LC3Ⅰ ratio, down-regulated Parkin expression, and up-regulated P62 expression. In addition, the group had decreased cell survival rate, increased content of LDH and CK in the culture medium, increased number of TUNEL positive cells, decreased pro caspase-3/caspase-3 and Bcl-2/Bax ratios, and declined mitochon-drial membrane potential. To sum up, DBD could promote the mitophagy, inhibit the apoptosis, and alleviated the injury of H9c2 cells exposed to low oxygen.

Low-temperature plasma modification, structural characterization and anti-diabetic activity of an apricot pectic polysaccharide.

To fully research the anti-diabetic activity of apricot polysaccharide, low temperature plasma (LTP) was used to modify apricot polysaccharide. The modified polysaccharide was isolated and purified using column chromatography. It was found that LTP modification can significantly improve the α-glucosidase glucosidase inhibition rate of apricot polysaccharides. The isolated fraction FAPP-2D with HG domain showed excellent anti-diabetic activity in insulin resistance model in L6 cell. We found that FAPP-2D increased the ADP/ATP ratio and inhibited PKA phosphorylation, activating the LKB1-AMPK pathway. Moreover, FAPP-2D activated AMPK-PGC1α pathway, which could stimulated mitochondrial production and regulate energy metabolism, promoting GLUT4 protein transport to achieve an anti-diabetic effect. The Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy data showed that the LTP modification could increase the CH bond content while decreasing the C-O-C/C-O bond content, indicating that LTP destroyed the C-O-C/C-O bond, which enhanced the anti-diabetes activity of the modified apricot pectin polysaccharide. Our findings could pave the way for the molecular exploitation of apricot polysaccharides and the application of low-temperature plasma.

Sappanone A: A natural PDE4 inhibitor with dual anti-inflammatory and antioxidant activities from the heartwood of Caesalpinia sappan L.

ETHNOPHARMACOLOGICAL RELEVANCE: Sumu (Lignum sappan), the dry heartwood of Caesalpinia sappan L., is a traditional Chinese medicine used as an analgesic and anti-inflammatory agent. AIM OF THE STUDY: The study aspired to discover natural phosphodiesterase 4 (PDE4) inhibitors with dual anti-inflammatory and antioxidant activities from Sumu for the treatment of chronic obstructive pulmonary disease (COPD). MATERIALS AND METHODS: To accurately and efficiently identify natural PDE4 inhibitors from Sumu, molecular docking and molecular dynamics (MD) analysis methods were used for structure-based virtual screening of a self-built database of primary polyphenols in Sumu. According to the previous studies of Sumu and the free radical scavenging mechanism of polyphenols, the reported antioxidant components from Sumu and the potential antioxidants with the antioxidant pharmacophore of catechol and π-conjugated moieties were selected from the potential PDE4 inhibitors predicted by docking. Sappanone A, a potential PDE4 inhibitor with antioxidant activity from Sumu, was selected, calculated and synthesized to evaluate its dual anti-inflammatory and antioxidant functions in vitro and in vivo studies. Herein sappanone A was assayed for its inhibitory effects against PDE4 enzyme activity, tumor necrosis factor-alpha (TNF-α) production induced by lipopolysaccharide (LPS) in RAW264.7 macrophages and malondialdehyde (MDA) production induced by Fe2+ in mouse lung homogenate; sappanone A was also assayed for its abilities of radical (DPPH) scavenging, reducing Fe3+ and complexing Fe2+ in vitro. Additionally, LPS-induced acute lung injury (ALI) in mice was used to evaluate its anti-inflammatory activity as a PDE4 inhibitor in vivo, and the levels of TNF-α and total protein in bronchoalveolar lavage fluid (BALF) and myeloperoxidase (MPO) activity in the lung were assayed. RESULTS: The present study predicted and validated that sappanone A was a promising PDE4 inhibitor from Sumu with dual anti-inflammation and antioxidant activities from Sumu. In vitro, sappanone A remarkably inhibited PDE4 enzyme activity and reduced TNF-α production induced by LPS in RAW264.7 macrophages and MDA production induced by Fe2+ in mouse lung homogenate. Meanwhile, it showed outstanding abilities of scavenging DPPH radicals, reducing Fe3+ and complexing Fe2+. In vivo, sappanone A (25 mg/kg and 50 mg/kg, i.p., twice daily for 7 days) distinctly prevented LPS-induced ALI in mice by reducing the levels of TNF-α and total protein in BALF and MPO activity in the lung. CONCLUSION: Sappanone A is a natural PDE4 inhibitor with dual anti-inflammatory and antioxidant activities from the traditional Chinese medicine Sumu, which may be a promising therapeutic agent to prevent the vicious cycle of COPD inflammation and oxidative stress.

[Preparation of microemulsion gel loading enriched ingredients of Epimedii Folium and its pharmacodynamics evaluation].

On the basis of previous studies, this study prepared and evaluated microemulsion gel loading enriched ingredients of Epimedii Folium and investigated its protective effect against peripheral nervous system damage caused by chemotherapeutics. The preparation method and the type and dosage of the matrix were investigated from rheology, preparation difficulty, and drug loading. Then the optimal prescription was determined and the microemulsion gel loading enriched ingredients of Epimedii Folium was prepared. The in vitro release and transdermal behaviors of the gel were investigated in the Franz diffusion cell with epimedin A1,A,B,C, and icariin as evaluation indicators. The oxaliplatin-induced peripheral neuropathy(OIPN) model was established in Wistar rats. The protective effect of the microemulsion gel loading enriched ingredients of Epimedii Folium against peripheral nervous system damage caused by chemotherapeutics was evaluated by behavioral measurement after drug administration and histopathological examination of dorsal root ganglia and sciatic nerve. The preparation process of the microemulsion gel loading enriched ingredients of Epimedii Folium was stable, and the release of the five components was consistent with the Hixson-Crowell cube root law. Behavioral indicators intuitively showed that the drug could effectively relieve mechanical allodynia caused by oxaliplatin. The histopathological examination showed that the drug can improve neuron damage in the dorsal root ganglia, axon degeneration, and demyelination caused by oxaliplatin. Therefore, the preparation process of the microemulsion gel loading enriched ingredients of Epimedii Folium is feasible, which can achieve stable drug release. It has a certain therapeutic effect on chemotherapy-induced peripheral neuropathy(CIPN).

[Danggui Buxue Decoction improves heart function of chronic intermittent hypoxia mice by promoting mitochondrial autophagy and inhibiting cardiomyocyte apoptosis].

The study established a chronic intermittent hypoxia(CIH) model in mice to investigate the effects of Danggui Buxue Decoction(DBD) on mitochondrial autophagy and cardiomyocyte apoptosis and explore its protective effect and mechanism on cardiac function of CIH mice. Forty C57 BL/6 N male mice were randomly divided into the control(CON) group, CIH group, CIH+DBD group, and DBD group, with 10 mice in each group. CIH was induced by filling the hypoxic chamber with N_2(90 s) to reduce the O_2 concentration to 5% and then filling the hypoxic chamber with O_2(90 s) to restore O_2 concentration to 21%, 3 min per cycle, and the CIH treatment continued for 35 d, 8 h per day. Mice in the CIH+DBD and DBD groups were treated with intragastric administration of DBD every day, while those in the CON and CIH groups with the same volume of normal saline. The cardiac function of mice was measured by echocardiography. The pathological changes in myocardium were observed after HE staining, followed by the observation of cardiomyocyte apoptosis by Tunel staining. The expression of apoptosis-related proteins pro-caspase-3, caspase-3, Bcl-2, and Bax and autophagy-related proteins LC3Ⅱ, LC3Ⅰ, P62, parkin, and cytochrome C(Cytc) was detected by Western blot. The mitochondrial membrane potential was observed using JC-1 fluorescent probe. Compared with the CON group, the CIH group exhibited remar-kably lowered left ventricular ejection fraction(LVEF) and left ventricular fractional shortening(LVFS), elevated left ventricular end-systolic volume(LVESV) and end-diastolic volume(LVEDV), disordered myocardial fiber arrangement, increased number of TUNEL-positive cells, decreased pro-caspase-3/caspase-3, Bcl-2/Bax, and LC3Ⅱ/LC3Ⅰ ratios, parkin, mitochondrial Cytc expression, and mitochondrial membrane potential, and up-regulated P62 and Cytc expression. Compared with the CIH group, DBD increased LVEF, LVFS, pro-caspase-3/caspase-3, Bcl-2/Bax, and LC3Ⅱ/LC3Ⅰ ratios, and parkin expression, as well as mitochond-rial Cytc expression, and mitochondrial membrane potential, decreased LVESV, LVEDV, and the number of Tunel-positive cells, and improved the myocardial fiber arrangement. DBD has a protective effect on the heart function of CIH mice. It improves the heart function possibly by promoting mitochondrial autophagy to ameliorate mitochondrial function and inhibiting the cardiomyocyte apoptosis.

Higher efficacy of resveratrol against advanced breast cancer organoids: A comparison with that of clinically relevant drugs.

The lack of reliable drugs is a therapeutic challenge of advanced breast cancers (ABCs). Resveratrol (Res) exerts inhibitory effects on breast cancer cell lines and animal models, while its efficacy against individual breast cancer cases remains unknown. This study aims to use ABC-derived organoids (ABCOs) as the ex vivo therapeutic platform to clarify the effectiveness of resveratrol against different ABC subtypes. Immunohistochemical staining confirmed that the ABCOs maintained their original tumors' ER, PR, HER2, and Ki67 expression patterns. ABCO proliferation and viability tests showed >50% cell death rates in 79.2% (19/24) of Res-treated, 28.6% (2/7) fulvestrant-treated, 66.7% (4/6) paclitaxel-treated, and 66.7% (6/9) gemcitabine-treated ABCOs. pSTAT3 nuclear translocation was more frequent in Res-sensitive (17/19; 89.47%) than that (1/5; 20%) of Res-insensitive ABCOs, which were suppressed upon Res treatment. Statistical analysis revealed a close correlation of STAT3 activation with the efficacy of Res, but not related to tumor receptor expression patterns (ER, PR, HER2) and pathological classification. We demonstrate for the first time the higher efficacy and broader spectrum of Res against different subtypes of ABCOs in comparison with that of conventional antibreast cancer drugs, providing an alternative approach for better management of ABCs.

[Analysis of CiteSpace knowledge map for traditional Chinese medicine prevention and treatment of cerebral small vessel diseases].

This study aims to analyze the research on the prevention and treatment of cerebral small vessel diseases(CSVDs) with traditional Chinese medicine(TCM) based on knowledge map, and to preliminarily explore the research hotspots and trends. To be specific, articles on TCM treatment of CSVDs in CNKI, Wanfang, and VIP(from establishment to November 2021) were retrieved, followed by bibliometric analysis. Then CiteSpace 5.7 R4 and Gephi were employed for generation of maps on annual number of articles, author cooperation, institution cooperation, keyword co-occurrence, keyword clustering, and keyword emergence. A total of 106 eligible articles were screened out, and the annual number of articles presented a steady upward trend. A total of 277 authors were included in the author cooperation network, among whom CHEN Zhigang published the most articles. A total of 87 institutions were included in the institution cooperation network, among which Dongfang Hospital of Beijing University of Chinese Medicine showed the most frequent cooperation with other institutions. Keyword clustering showed that research on the TCM treatment of CSVDs mainly focused on five aspects: related disease research, neurological function deficits, disease nature and location in TCM, TCM treatment methods, and formulas. The prevention and treatment of CSVDs with TCM in China has been developing steadily in the past ten years, and TCM has unique advantages in the prevention and treatment of this disease. The knowledge maps vividly demonstrated the development and research hotspots and trends in this field. The result is expected to provide a reference for further research in this field.

Capsaicin ameliorates intermittent high glucose-mediated endothelial senescence via the TRPV1/SIRT1 pathway.

BACKGROUND: Patients with diabetes have accelerated vascular aging when compared with healthy individuals. Hyperglycemia, especially intermittent high glucose (IHG), is the main cause of vascular endothelial senescence. Capsaicin, a major component of chili pepper is thought to contribute to cardiovascular protection by spicy food. OBJECTIVE: To investigate the pathway related with the effects of capsaicin on endothelial cell senescence induced by IHG. METHODS: HUVECs were exposed to IHG (5 mM or 33 mM glucose, alternating every 12 hours for 3 days) and treated with capsaicin at 0.3, 1 and 3 µM. To determine endothelial cell senescence, we examined the senescence-related ß-galactosidase staining, cell cycle arrest, cell viability, as well as production of reactive oxygen species (ROS). To evaluate the involvement of TRPV1/[Ca2+]i/CaMKII/AMPK/SIRT1 pathway in anti- senescence effects of capsaicin, HUVECs were treated with CAPZ (a TRPV1 antagonist), BAPTA-AM (an intracellular calcium chelator), KN62 (a CaMKII antagonist), compound C (an AMPK inhibitor), or EX527 (a SIRT1 inhibitor). To knockdown TRPV1, HUVECs were transfected with shRNA lentivirus targeting TRPV1. The levels of SIRT1, p21, TRPV1, AMPK and phospho-AMPK were evaluated by western blotting. RESULTS: IHG suppressed the levels of SIRT1 and enhanced endothelial senescence. Capsaicin upregulated SIRT1 expression and downregulated the senescence marker, p21, thereby protecting endothelial cells from IHG-induced senescence as indicated by relieved G0/G1 phase arrest, improved cell viabilities, and reduced counts of senescent cells and ROS production. Pre-treatment with CAPZ, BAPTA-AM, KN62 or compound C abrogated the anti-senescence effects of capsaicin. Capsaicin restored AMPK phosphorylation and IHG-inhibited TRPV1 expression. Moreover, TRPV1 silencing suppressed SIRT1 expression and abolished the anti-senescence effects of capsaicin. CONCLUSION: Capsaicin elevates SIRT1 levels through TRPV1/[Ca2+]i/CaMKII/AMPK pathway and suppresses IHG-mediated endothelial cell senescence. This study provides initial evidence that capsaicin is a potential candidate for the prevention of vascular aging in diabetes.

Lactobacillus huangpiensis sp. nov. and Lactobacillus laiwuensis sp. nov., isolated from the gut of honeybee (Apis mellifera).

Four Gram-stain-positive bacterial strains were isolated from the gut of honeybee (Apis mellifera) in China. These strains were characterized using a polyphasic taxonomic approach. The data demonstrated that three of the four strains represented two novel species of the genus Lactobacillus, strains F306-1T and F551-2T were designated as the type strains. Results of 16S rRNA gene sequence analysis indicated that strains F306-1T, F447 and F551-2T were phylogenetically related to the type strains of Lactobacillus kimbladii and Lactobacillus kullabergensis, having 99.1-99.7 % 16S rRNA gene sequence (about 1400 bp) similarities. The phylogenetic tree based on concatenated pheS, rpoA, gyrB, hsp60, recA, rpoB and tuf sequences (4114 bp) and the phylogenomic tree based on whole genome sequences indicated that strains F306-1T and F447 were most closely related to L. kullabergensis Biut2NT, and strain F551-2T was most closely related to L. kimbladii Hma2NT. Strains F306-1T and F447 shared 99.9 % average nucleotide identity (ANI), 99.7 % digital DNA-DNA hybridization (dDDH) and 99.9 % average amino acid identity (AAI) values, indicating that they belong to the same species. Strain F306-1T exhibited the highest ANI (94.4 %), dDDH (56.7 %) and AAI (94.7 %) values to L. kullabergensis Biut2NT. Strain F551-2T had the highest ANI (94.0 %), dDDH (54.3 %) and AAI (95.8 %) values with L. kimbladii Hma2NT. Acid production from amygdalin, maltose, starch, gentiobiose and turanose, activity of esterase (C4) and α-glucosidase, growth with 3 % NaCl at 37 °C under strict anaerobic condition (on mMRS agar plates), and growth with 1-6% NaCl at 37 °C under aerobic condition (on mMRS agar plates supplemented with 0.05 % cysteine or with 1 % cysteine and 2 % fructose) could differentiate strains F306-1T and F447 from L. kullabergensis DSM 26262T. Acid production from d-glucose, arbutin and gentiobiose, growth with 3 % NaCl at 37 °C under strict anaerobic condition (on mMRS agar plates), and growth at 45 °C under strict anaerobic condition (on mMRS agar plates) could differentiate strain F551-2T from L. kimbladii DSM 26263T. Based upon the data obtained in the present study, two novel species, Lactobacillus huangpiensis sp. nov. and Lactobacillus laiwuensis sp. nov., are proposed and the type strains are F306-1T (=LMG 32144T=JCM 34361T=CCTCC AB 2020300T) and F551-2T (=JCM 34502T=CCTCC AB 2021027T), respectively.

Age-dependent alteration in metabolism of vitamin B6 , neurotransmitters, and amino acids after 4'-O-methylpyridoxine administration in rats.

4'-O-methylpyridoxine (MPN), a recognized antivitamin B6 compound, is a potentially poisonous substance found in Ginkgo biloba L. In this work, the effects of MPN on the metabolism of vitamin B6 , neurotransmitters, and amino acids were compared in the plasma and brain of young and adult rats under various administration times. Results showed that the contents of MPN residues in the plasma and brain of young rats were 12.72 and 14.76 µM higher than adult rats, respectively. Moreover, the levels of 5-hydroxytryptamine and dopamine in the brain of young rats have decreased by 13.78% and 7.19%, respectively, compared with the control group, at 2 h after MPN administration. Furthermore, the principal component analysis revealed that MPN was an important contributor to the amino acid composition in the brain of young rats. These results suggest that age may lead to different toxic effects of MPN. PRACTICAL APPLICATION: 4'-O-methylpyridoxine is primarily responsible for poisoning due to overconsumption of Ginkgo biloba seeds. This study will provide an exploratory understanding of the age-dependent toxicity of 4'-O-methylpyridoxine.

[Effects of Bu Zhong Yi Qi decoction on CIH-induced interstitial lung fibrosis in mice].

OBJECTIVE: To investigate the effects of chronic intermittent hypoxia (CIH) on the expression of transforming growth factor-ß (TGF-ß), P-samd3, serum laminin (LN) and hyaluronidase (HA) in mouse lung tissues and the protective effects of Bu Zhong Yi Qi decoction on lung interstitial deposition damage in CIH mice. METHODS: Fifty SPF-grade C57BL mice were randomly divided into five groups (n=10): blank control group, CIH model group, and CIH+ low, medium and high doses of Bu Zhong Yi Qi decoction group. Mice were placed under normoxia or CIH conditions, respectively. The Chinese medicine group was given the corresponding doses of drugs. HE staining was performed to assess pathological changes and Masson staining was performed to assess collagen deposition. Western blot was performed to detect the expressions of channel proteins such as TGF-ß1, P-smad3 and down stream α-SMA and Collagen I. ELISA was performed to detect the serum levels of TGF-ß1, LN and HA. RESULTS: HE staining showed alveolar collapse, septal thickening and epithelial cell necrosis in CIH mice, Masson showed massive collagen fiber proliferation and deposition in lung interstitium, while the above changes in lung tissues were significantly improved in the CIH + Bu Zhong Yi Qi decoction groups compared with the CIH group. TGF-ß1, P-smad3 and Collagen I, Collagen Ⅲ, and α-SMA expression levels were increased compared with the blank control group (P＜0.05), and the expressions of TGF-ß1 and LN in serum were upregulated (P＜0.05). The expressions of TGF-ß1, P-smad3, Collagen I protein and SMA-α in the lung tissues of the CIH+ Bu Zhong Yi Qi decoction groups were downregulated significantly compared with those of the CIH group (P＜0.05), and the improvement of multiple indexes in the CIH+high-dose CIH intervention group was better than those of the low-dose group (P＜0.05). CONCLUSION: Bu Zhong Yi Qi decoction can inhibit alveolar structural changes and excessive collagen deposition in the interstitium of CIH mice, and then improve lung function in CIH mice. The mechanism may be related to the down-regulation of protein expression related to TGF-ß/smads signaling pathway by Bu Zhong Yi Qi decoction.

In vivo toxicity assessment of 4'-O-methylpyridoxine from Ginkgo biloba seeds: Growth, hematology, metabolism, and oxidative parameters.

4'-O-methylpyridoxine (MPN), a recognized antivitamin B6 compound, is a potentially poisonous substance found in Ginkgo biloba seeds and leaves. In this work, the body weights, histopathological changes, plasma vitamin B6 (VB6), biochemical parameters, oxidative stress responses, and amino acids of rats were investigated after intragastric administration of MPN for 15 days. Results showed that intragastric administration of 50 mg/kg BW MPN caused pathological changes in the brain and heart tissues of rats. Administration of 10 mg/kg and 30 mg/kg BW MPN can significantly increase VB6 analogs in the plasma of rats, such as pyridoxal-5'-phosphate, pyridoxal. Results of biochemical parameters indicated that MPN can damage brains and hearts by changing the enzyme activity of these organs. These results suggest that consumption of Ginkgo biloba seeds for the long term, even in a small quantity, may lead to poisoning.

[Systematic review and Meta-analysis of clinical efficacy and safety of Ginkgo Leaf Tablets in treatment of acute cerebral infarction].

To systematically evaluate the clinical efficacy and safety of Ginkgo Leaf Tablets(GLT) in the treatment of acute cerebral infarction(ACI). Seven databases both at home and abroad were systematically retrieved from their establishment to March 2020. The data of the included studies were extracted after review and screening. The quality of the included studies was assessed with the Cochrane risk bias assessment tool, and then the included studies were put into Meta-analysis by RevMan 5.3 to evaluate the total cli-nical efficiency, neurological function score, blood lipids and incidence of adverse reactions in treatment of ACI by GLT. Finally, the GRADE system was adopted to evaluate the evidence quality of each outcome indicator and form recommendations. Ten studies involving 886 participants were included, all of which were of low quality. Meta-analysis results showed that,(1)in terms of the total clinical efficiency, GLT+Western medicine was superior to Western medicine alone(RR_(NDS)=1.20, 95%CI[1.06, 1.36], P=0.005; RR_(NIHSS)=1.35, 95%CI[1.09, 1.69], P=0.007), and there was no statistical difference between GLT+Xuesaitong Injection+Wes-tern medicine and Xuesaitong Injection+Western medicine(RR=1.16, 95%CI[1.00, 1.35], P=0.05).(2)In terms of improving neurological function score, GLT+Western medicine was superior to Western medicine alone(MD_(NIHSS[moderate(severe)])=-1.55, 95%CI[-2.22,-0.88], P<0.000 01; MD_(NIHSS(severe))=-7.51, 95%CI[-8.00,-7.02], P<0.000 01; MD_(NDS)=-1.36, 95%CI[-2.39,-0.33], P=0.01), and GLT+Danshen Injection+Western medicine was superior to Danshen Injection+Western medicine(MD_(NDS)=-3.09, 95%CI[-3.84,-2.34], P<0.000 01).(3)In terms of regulating blood lipids, GLT+Western medicine was superior to Wes-tern medicine alone(MD_(TC)=-1.40, 95%CI[-2.13,-0.66], P=0.000 2; MD_(TG)=-1.29, 95%CI[-1.86,-0.73], P<0.000 01; MD_(LDL-C)=-1.48, 95%CI[-2.91,-0.04], P=0.04; MD_(HDL-C)=0.07, 95%CI[0.02, 0.12], P=0.009).(4)In terms of incidence of adverse reactions, there was no statistical difference between GLT+Western medicine and Western medicine alone(RR=0.63, 95%CI[0.30, 1.32], P=0.22). The results of the evaluation showed that the evidence level of each outcome indicator was low, and the recommendation was at weak level. In conclusion, GLT+Western medicine could improve the total clinical efficiency, neurological function score, and blood lipid status, with a low incidence of adverse reactions. However, due to the small amount of included stu-dies, low study quality and low level of evidence, it is expected to carry out clinical studies with standardized design and large sample size in the future to further investigate the clinical efficacy and safety of GLT in the treatment of ACI.

4'-O-methylpyridoxine: Preparation from Ginkgo biloba Seeds and Cytotoxicity in GES-1 Cells.

Ginkgo biloba seeds are wildly used in the food and medicine industry. It has been found that 4'-O-methylpyridoxine (MPN) is responsible for the poisoning caused by G. biloba seeds. The objective of this study was to explore and optimize the extraction method of MPN from G. biloba seeds, and investigate its toxic effect on human gastric epithelial cells (GES-1) and the potential related mechanisms. The results showed that the extraction amount of MPN was 1.933 µg/mg, when extracted at 40 °C for 100 min, with the solid-liquid ratio at 1:10. MPN inhibited the proliferation of GES-1 cells, for which the inhibition rate was 38.27% when the concentration of MPN was 100 µM, and the IC50 value was 127.80 µM; meanwhile, the cell cycle was arrested in G2 phase. High concentration of MPN (100 µM) had significant effects on the nucleus of GES-1 cells, and the proportion of apoptotic cells reached 43.80%. Furthermore, the Western blotting analysis showed that MPN could reduce mitochondrial membrane potential by increasing the expression levels of apoptotic proteins Caspase 8 and Bax in GES-1 cells. In conclusion, MPN may induce apoptosis in GES-1 cells, which leads to toxicity in the human body.

Analysis of the potential and mechanism of Ginkgo biloba in the treatment of Alzheimer's disease based on network pharmacology.

Objective: Study the principle and possible mechanism of Ginkgo biloba in the treatment of Alzheimer's disease (AD) which is based on network pharmacology. Methods: The potential targets of active ingredients of Ginkgo biloba were collected by Traditional Chinese Medicine Integrated Database platform (TCMSP). TCMSP is a pharmacological system for drug discovery from Chinese herbal medicine. The disease targets of AD were searched and collected by the database of gene-disease associations (DisGeNET) and literature. The obtained targets were standardized by the UniProt database. STING network platform and Cytoscape were used to construct protein-protein interaction network (PPI) of the key targets. According to Materscape, we clarify the possible mechanism of action including Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway and Gene Ontology (GO) enrichment analysis. Results: The compound-target network contains 27 active ingredients, 191 related targets, 18 key targets, including PLAU, HMOX1, TNF, INSR, MPO, MAOB, IGF2, IL1B, ESR1, BCL2, ACHE, BAX, GSK3B, PPARG, SLC2A4, NOS3, CASP3, VEGFA. GO enrichment analysis has got a total of 640 GO items, including 609 biological process (BP) items (95.1%), 16 molecular function (MF) items (2.5%) and 15 cellular component (CC) items (2.4%). After KEGG enrichment, 44 pathways were obtained. Conclusion: Through the construction of "component-target-pathway", GO biological function and KEGG pathway enrichment analysis were performed on core targets, and the possibility of Ginkgo biloba for the treatment of AD was explored from multiple targets and pathways, which provided a new approach for multi-target treatment.

Improvement of the stabilities and antioxidant activities of polyphenols from the leaves of Chinese star anise (Illicium verum Hook. f.) using ß-cyclodextrin-based metal-organic frameworks.

BACKGROUND: Polyphenols extracted from plants are usually highly unstable and rapidly transformed into various reaction products during food and drug processing, thus limiting their applications. To improve the stability and solubility of polyphenols from the leaves of Chinese star anise (Illicium verum Hook. f.), and hence to expand their application to food and medicine, the extracted anise leaf polyphenols (ALPs) were microencapsulated using ß-cyclodextrin (ß-CD) and cyclodextrin-based metal-organic frameworks (ß-CD-MOFs). RESULTS: The optimum inclusion rate of ALP/ß-CD-MOFs was 97.80% at a core-wall ratio of 1:10. Meanwhile, the stabilities, solubilities and antioxidant activities of the polyphenols before and after inclusion were compared. The results showed both the stabilities and solubilities of ALP/ß-CD-MOFs were significantly improved compared with those of ALPs and ALP/ß-CD, suggesting the potential of ß-CD-MOFs as newer and better carriers than ß-CD for polyphenols in food industry applications. The free radical (including superoxide, hydroxyl and DPPH radicals) scavenging activities were also improved by microencapsulation. Superoxide radical scavenging reaction also showed slow-release property of ALP/ß-CD-MOFs. The formation of the inclusion complex was further confirmed using Fourier transform infrared spectral characterization. CONCLUSIONS: Microencapsulation with ß-CD-MOFs could expand the application scope of ALPs, and it is more effective than encapsulation with ß-CD. This is important for a better understanding and application of this useful traditional Chinese plant. As a new material with high efficiency and edibility, ß-CD-MOFs are not limited to the chemical field, but also have potential in new areas of food, medicine and healthcare products. © 2020 Society of Chemical Industry.

Amelioration by Idesia polycarpa Maxim. var. vestita Diels. of Oleic Acid-Induced Nonalcoholic Fatty Liver in HepG2 Cells through Antioxidant and Modulation of Lipid Metabolism.

Idesia polycarpa Maxim. var. vestita Diels (I. polycarpa) is well known as an edible oil plant which contains abundant linoleic acid and polyphenols. The objective of this study was to maximize the by-product of defatted fruit of I. polycarpa. We found that the fraction D of ethyl acetate extract (EF-D) contained more polyphenols, which contribute to its strong antioxidant activity by antioxidant assays (DPPH, ABTS, and FRAP). Meanwhile, EF-D showed a significant lipid-lowering effect on oleic acid- (OA-) induced hepatic steatosis in HepG2 cells through enhancing antioxidant activity, reducing liver damage, and regulating lipid metabolism, antioxidant, and inflammation-related gene expression. The SOD and T-AOC levels significantly increased, but the levels of MDA, AST, and ALT decreased obviously when treated with EF-D. In general, EF-D improved the antioxidant enzyme activities and decreased the hepatic injury activities. Besides, treatment with EF-D for NAFLD influenced lipid metabolism and inflammation by activating PPARα which was associated with the increased expression of CPT1 and decreased expression of SCD, NF-κB, and IL-1. Moreover, EF-D improved the oxidative stress system through activation of the Nrf2 antioxidant signal pathways and upregulated its target genes of HO-1, NQO1, and GSTA2. The results highlighted the EF-D from the defatted fruit of I. polycarpa regarding lipid-lowering, proving it to be a potential drug resource of natural products for treating the nonalcoholic fatty liver disease (NAFLD).

Mu-Xiang-You-Fang protects PC12 cells against OGD/R-induced autophagy via the AMPK/mTOR signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Mu-Xiang-You-Fang (MXYF) is a classic prescription of Hui medicine. It is composed of five herbs and has been used to treat ischemic stroke for many years. However, the potential pharmacological mechanisms of MXYF remain unclear. The present research is aimed to investigate the protective effect and possible mechanisms of MXYF treatment in an in vitro model of cerebral ischemia-reperfusion injury. MATERIALS AND METHODS: An oxygen-glucose deprivation and reperfusion (OGD/R) model of PC12 cells was established. The effect of MXYF on the cell viability after OGD/R injury was determined using a cell counting kit (CCK-8) assay. The colorimetric method was used to determine the lactate dehydrogenase (LDH) leakage rate. The calcium concentration was determined by the chemical fluorescence method, and mitochondrial membrane potential was determined using flow cytometry. Monodansylcadaverine (MDC) staining and electron microscopic analysis were then conducted to detect autophagy after oxygen-glucose deprivation and reperfusion in PC12 cells. Immunofluorescence and western blot analyses were used to detect the expression of proteins associated with autophagy. RESULTS: It was found that MXYF (1, 2, 4 µg/mL) could significantly increase cell viability and mitochondrial membrane potential and decrease the calcium concentration and LDH release rate in PC12 cells. After OGD/R injury in PC12 cells, the number of autophagosomes and autophagolysosome significantly increased. MXYF (4 µg/mL) inhibited the autophagy induced by OGD/R and inhibited the expression of LC3, beclin1, p-AMPK, and ULK1. In contrast, the expression of p-mTOR, p-p70s6k, and p62 was significantly enhanced. CONCLUSIONS: These findings suggest that MXYF inhibits autophagy after OGD/R-induced PC12 cell injury through the AMPK-mTOR pathway. Thus, MXYF might have therapeutic potential in treating ischemic stroke.