This study was designed to investigate the effect of 5,2',4'-trihydroxy-6,7,5'-trimethoxy flavone nanoparticle (TTF1-NP) on inducing apoptosis of implanted tumour cells in nude mice and the mechanism of endoplasmic reticulum stress pathway. The implanted hepatoma model was established in nude mice, and used to test the drug TTF1-NP in five groups (vehicle, 5 μmol·kg^-1 TTF1-NP, 10 μmol·kg^-1 TTF1-NP, 20 μmol·kg^-1 TTF1-NP and adriamycin). The nude mice were killed after the treatment to determine the tumor growth inhibition rate (IR). Morphological changes of implanted tumor cells were observed by HE staining; apoptosis of tumor cells was detected by TUNEL; the protein expression of GRP78, p-JNK and caspase 12 were analyzed using immunocytochemistry staining and Western blotting. We tested the effects of TTF1-NP on implanted HepG-2 cell tumor growth in nude mice. TTF1-NP-treated mice showed volume of tumor smaller than that of the vehicle-treated mice. The tumor mass of the TTF1-NP-treated mice were significantly reduced than those of the vehicle-treated mice. In addition, the tumor growth rate of the TTF1-NP-treated mice was significantly lower than that of the vehicle-treated mice, and the tumor growth inhibition ratio of the TTF1-NP-treated mice was significantly higher than that of the vehicle-treated mice. TTF1-NP exhibited an inhibitory effect on implanted tumor cells in the model. The IR was 51.2%, 54.2%, 61.8% and 65.9%, respectively. In comparison with the vehicle group, the treated groups exhibited alteration in cell morphology and apoptosis of tumor cells, and expression of GRP78, p-JNK and caspase 12, which were observed by immunocytochemistry staining and Western blotting. Taken together, our results suggest that TTF1-NP induces apoptosis of implanted tumor cells in nude mice and the main mechanism is related to activation of endoplasmic reticulum stress.

This study was designed to investigate the mechanism of 5,2',4'-trihydroxy-6,7,5'-trimethoxy flavone nanoparticle (TTF1-NP) in the induction of apoptosis of human hepatocellular carcinoma HepG-2 cells. MTT assay, immunocytochemical staining and flow cytometry with Annexin V-FITC/PI were used to demonstrate inhibition of proliferation of HepG-2 cells and cell apoptosis. The inhibition was studied in a dose- and time-dependent manner. Western blot results showed that TTF1-NP down-regulated the signals of survivin, p-STAT3 and STAT3, but up-regulated the expression level of cleaved caspase-3. Taken together, our results showed that TTF1-NP induced HepG-2 cell apoptosis through inhibition of the STAT3 expression.

The study was designed to test the role of 5,2',4'-trihydroxy-6,7,5'-trimethoxy flavone nanoparticle (TTF1-NP) on lipopoiysaccharide (LPS)-induced inflammatory response, and to explore the anti-inflammatory mechanism in human hepatocellular carcinoma cells. Inflammatory responses were induced in human hepato-cellular carcinoma HepG2 cells with LPS; Proliferation effect of TTF1-NP in LPS-stimulated HepG2 cells were detected by MTT assay; The expression of TLR4, AKT/mTOR signaling related proteins and IL-6 were detected by Western blot assay. The results showed that TTF1-NP inhibited the proliferation of HepG2 cells induced by LPS in a dose-dependent manner; TTF1-NP inhibited the expression of TLR4, the activation of AKT and mTOR, and expression of IL-6 in a dose-dependent manner; TTF1-NP inhibited the activation of AKT/mTOR signaling pathway and TLR4 proteins leading to suppression of IL-6 expression in HepG2 cells stimulated by insulin. These results suggest that TTF1-NP inhibited inflammatory responses from LPS treatment with a potential mechanisms in the inhibition of AKT/mTOR pathway.

This study aimed to investigate apoptosis induction of ginsenoside compound K (ginsenoside CK) in human liver cancer SMMC-7721 cells and the involvement of TGF-β1/Smads signaling pathway. MTT assay was used to detect cell viability following ginsenoside CK treatment in SMMC-7721 cells. Annexin V-FITC/PI assay was used to detect apoptosis. After ginsenoside CK, or TGF-β1/Smads pathway activator TGFβ1 and inhibitor LY2109761 treatment, the TGF-β1/Smads pathway proteins and apoptosis proteins were detected by Western blot. The results showed that ginsenoside CK inhibited the proliferation of SMMC-7721 cells in a dose- and time-dependent manner. Annexin V-FITC/PI showed that ginsenoside CK induced apoptosis in SMMC-7721 cells. Meanwhile, ginsenoside CK inhibited the expression of Smad2/3, p-Smad2/3, Smad4, but promoted Smad7 expression, cleavage of caspase-3 and down-regulated Bcl-2/Bax. Compared with TGFβ1 treatment alone, levels of Smad2/3, p-Smad2/3, Smad4 and the ratio of Bcl-2/Bax were down-regulated, whereas Smad7 or cleaved caspase-3 was up-regulated in the ginsenoside CK+TGF-β1 group. In addition, Smad2/3, p-Smad2/3 and Smad4 expression were decreased in LY2109761 group. Compared with LY2109761 group, cleaved caspase-3 expression and Bcl-2/Bax have no significant change in ginsenoside CK+LY2109761 group. Taken together, our results showed that ginsenoside CK induced apoptosis in SMMC-7721 cells, and such induction is related to inhibiting TGF-β1/Smads signaling pathway.

Objective:To explore the environmental risk factors of different categories of congenital heart defects ( CHD) and provide evidence for further risk factors and prevention research of CHD pheno-types. Methods:Data of Guangdong CHD Register Study from 2004 to 2012 were used. In the study, 3 038 CHD cases and 3 038 paired controls from 34 hospitals distributed in 17 cities were registered and related information were collected using uniform, and structured questionnaires. All the CHD phenotypes were coded according to the International Classification of Diseases 10th Revision (ICD-10) and classified into 6 categories according to their pathological features. Univariate analyses were adopted to filter poten-tial risk factors for each category of CHD. Then multivariate conditional Logistic regression was used to calculate the odds ratios of the risk factors for each category of CHD. Results:The risk factors for left-to-right shunt CHD included low ( OR=2 . 63 , 95%CI:2 . 04 -3 . 39 ) or over birth weight ( OR =2 . 21 , 95%CI:1 . 47-3 . 32 ) , premature delivery ( OR=1 . 95 , 95%CI:1 . 53-2 . 49 ) , polyembryony ( OR=1. 99, 95%CI: 1. 22 -3. 26), maternal low education, mother as factory worker (OR =1. 62, 95%CI:1 . 32-1 . 98 ) , parity≥2 ( OR =1 . 38 , 95%CI: 1 . 13 -1 . 69 ) , maternal abnormal reproduction history ( OR=2 . 29 , 95%CI:1 . 75-3 . 01 ) , fever ( OR=2 . 38 , 95%CI:1 . 26-4 . 48 ) , virus infection ( OR=1 . 80 , 95%CI:1 . 29 -2 . 51 ) , medicine usage ( OR=1 . 73 , 95%CI:1 . 11 -2 . 69 ) , passive smoking ( OR=1 . 69 , 95%CI:1 . 26-2 . 29 ) , chemical agent contact ( OR=8 . 71 , 95%CI:2 . 33 -32 . 58 ) , living in newly decorated houses ( OR=2 . 56 , 95%CI:1 . 60-4 . 09 ) or room close to the main road ( OR=1 . 40 , 95%CI:1 . 14-1 . 72 ) in the first 3 months of pregnancy and father as factory worker ( OR=1 . 46 , 95%CI:1 . 23-1 . 73 ) . The risk factors for pulmonary outflow tract obstruction CHD in-cluded low ( OR =5 . 98 , 95% CI: 2 . 88 -12 . 44 ) or over birth weight ( OR = 6 . 56 , 95% CI:1. 19-36. 26), maternal low education, parity≥2 (OR=2. 08, 95%CI:1. 03-4. 22), virus infection in the first 3 months of pregnancy ( OR =4 . 30 , 95%CI: 1 . 27 -13 . 45 ) . The risk factors for left ventricular outflow tract obstruction CHD included father as factory worker ( OR=6 . 01 , 95%CI:1 . 05-34. 59). The risk factors for transposition of the great arteries included low birth weight (OR=12. 93, 95%CI:1. 14-146. 26), maternal low education, mother as factory worker (OR=3. 69, 95%CI:1. 53-8. 91). The risk factors for conditions with intra cardiac mixing of oxygenated and deoxygenated blood in-cluded parity=2 ( OR=3 . 45 , 95%CI:1 . 42-8 . 38 ) . The risk factors for other CHD included over birth weight (OR=4. 87, 95%CI:1. 19-19. 94), maternal abnormal reproduction history (OR=2. 96, 95%CI:1. 14 - 7. 68 ), virus infection ( OR = 4. 92, 95% CI: 1. 56 - 15. 47 ), medicine usage (OR=4. 90, 95%CI:1. 22-19. 77) or passive smoking (OR=10. 31, 95%CI:1. 25-85. 05) in the first 3 months of pregnancy. Conclusion:The environmental risk factors were discrepant among different categories of CHD. Further risk factors study of CHD phenotypes should be performed specially. To prevent CHD, attention should be paid to the risk factors which are related to multi or complex categories of CHD.

Objective To study the technology for purification of total alkaloids from Rhizoma Coptidis by maeroreticular adsorbent resin.Methods LD605,D101,DA201,NKA-9,and AB-8 types of macroreticular adsorbent resins werre used to separate and purify the total alkaloid from Rhizoma Coptidis.The yields and purities of the products were compared as indexes.Results AB-8 Type macroreticular adsorbent resin had optimum adsorption and elution parameters with its dynamic saturated adsorption ratio up to 1.23 mg/g.After eluted with 2 BV of distilled water and 2 BV 40% ethanol,the yields of total alkaloid was 85%,and content was 80%.Conclusion The AB-8 type macroreticular resin showsa better comprehensive adsorption property and can be used to separate and purify the total alkaloids from Rhizoma Coptidis.

ObjectiveTo investigate the value of intestinal fatty acid binding protein （I-FABP） in predicting the development and progression of acute-on-chronic liver failure （ACLF）. MethodsA retrospective analysis was performed for the clinical data of 168 patients with decompensated liver cirrhosis who were admitted to The Affiliated Hospital of Yanbian University from September 2020 to March 2023. The conditions of the patients with ACLF on admission were observed， and the patients were followed up for 6 months to identify new-onset ACLF cases. ELISA was used to measure the serum level of I-FABP on admission. The Mann-Whitney U test was used for comparison of non-normally distributed continuous data between two groups， and the Kruskal-Wallis H rank sum test was used for comparison between multiple groups； the chi-square test was used for comparison of categorical data between groups； the Jonckheere-Terpstra test was used for trend analysis. The Spearman correlation analysis was used to investigate the correlation between two variables， and the multivariate Cox regression analysis was used to investigate the influencing factors for new-onset ACLF during follow-up. The Kaplan-Meier curve was used to analyze the onset of ACLF in different groups， and the log-rank test was used for the analysis of such differences. The receiver operating characteristic （ROC） curve and the area under the ROC curve （AUC） were used to investigate the performance of I-FABP in predicting the development and progression of ACLF. ResultsAmong the 168 patients enrolled in this study， there were 43 patients with ACLF and 125 patients without ACLF， among whom 19 developed ACLF during follow-up. The patients with ACLF on admission had a significantly higher level of I-FABP than those without ACLF （Z=4.359， P<0.001）. The patients with new-onset ACLF had a significantly higher level of I-FABP than those without new-onset ACLF （Z=3.414， P<0.001）. The level of I-FABP increased with the increase in ACLF severity grade （H=17.385， P<0.001，P_trend<0.001）. The multivariate Cox regression analysis showed that I-FABP was independently associated with new-onset ACLF during follow-up （hazard ratio=2.138， 95% confidence interval ［CI］： 1.297 — 3.525， P=0.003）， and the tertile of I-FABP showed a good discriminatory ability （χ²=12.16， P<0.001）. The ROC curve showed that I-FABP had a good performance in predicting the development and progression of ACLF， with an area under the ROC curve of 0.854 （95%CI： 0.791 — 0.903） and 0.747 （95%CI： 0.661 — 0.820）， respectively， and an optimal cut-off value of 2.07 μg/L and 1.86 μg/L， respectively. ConclusionI-FABP can be used as a biomarker to predict the development and progression of ACLF， and it may help to identify high-risk patients and improve clinical management.

Diabetic kidney disease (DKD) is the most common complication of diabetes mellitus (DM). Qianjin Wenwu decoction (QWD), a well-known traditional Korean medicine, has been used for the treatment of DKD, with satisfactory therapeutic effects. This study was designed to investigate the active components and mechanisms of action of QWD in the treatment of DKD. The results demonstrated that a total of 13 active components in five types were found in QWD, including flavonoids, flavonoid glycosides, phenylpropionic acids, saponins, coumarins, and lignins. Two key proteins, TGF-β1 and TIMP-1, were identified as the target proteins through molecular docking. Furthermore, QWD significantly suppressed Scr and BUN levels which increased after unilateral ureteral obstruction (UUO). Hematoxylin & eosin (H&E) and Masson staining results demonstrated that QWD significantly alleviated renal interstitial fibrosis in UUO mice. We also found that QWD promoted ECM degradation by regulating MMP-9/TIMP-1 homeostasis to improve renal tubulointerstitial fibrosis and interfere with the expression and activity of TGF- β1 in DKD treatment. These findings explain the underlying mechanism of QWD for the treatment of DKD, and also provide methodological reference for investigating the mechanism of traditional medicine in the treatment of DKD.
Rats ; Mice ; Animals ; Ureteral Obstruction/metabolism* ; Kidney/metabolism* ; Tissue Inhibitor of Metalloproteinase-1/metabolism* ; Molecular Docking Simulation ; Rats, Sprague-Dawley ; Kidney Diseases/drug therapy* ; Extracellular Matrix/metabolism* ; Flavonoids/metabolism* ; Fibrosis

OBJECTIVE: To investigate the biological function of Cysteine rich (CysR) domain of a disintegrin and metalloprotease with thrombospondin type 1 repeats-13 (ADAMTS13) on cleavage of von Willebrand factor (vWF) and provide experimental evidence for exploring the pathogenesis of thrombotic thrombocytopenic purpura (TTP). METHODS: The six amino acids (EDGTLS) in ADAMTS13 CysR domain were point mutated one by one, and the mutant ADAMTS13 proteins were expressed and purified. The cleavage products of vWF polymer by wild-type or mutant ADAMTS13 under denaturing condition or shear stress were separated by 1% SeaKem HGT agarose gel and detected by Western blot. RESULTS: The mutant ADAMTS13 plasmids (M1: Glu515Ala; M2: Asp516Ala; M3: Gly517Ala; M4: Thr518Ala; M5: Leu519Ala; M6: Ser520Ala) were successfully constructed and the proteins of wild-type and mutant ADAMTS13 were purified. Wild-type ADAMTS13 almost completely cleaved the vWF polymer under denaturing condition, while the cleavage activity of M1 mutant was significantly reduced in the same condition (P<0.01). The cleavage activity of M1 mutant of ADAMTS13 was also significantly reduced compared with that of the wild-type under shear stress (P<0.01). The activity of M1 mutant to cleave the FRETS-vWF73 was dramatically reduced compared with that of wild-type ADAMTS13. However, the binding ability of M1 mutant to vWF was similar with that of wild-type ADAMTS13. CONCLUSION The CysR domain of ADAMTS13 plays an important role in the digestion of vWF under denaturing condition and shear stress. The Glu515 amino acid residue might be an important site for substrate recognition.
ADAM Proteins ; ADAMTS13 Protein/genetics* ; Humans ; Purpura, Thrombotic Thrombocytopenic/genetics* ; von Willebrand Factor/genetics*

The raphe of the human penis and scrotum is considered to develop secondarily after disappearance of the initial midline seam by fusion of the bilateral genital folds. However, the fetal development was still obscure. We examined histological sections of 30 fetuses (17 males and 13 females) at 10-15 weeks. In male fetuses, the scrotum was not yet clearly identified because of no descent of testis. The perineal raphe was thin and wavy at 10 weeks, and it was continuous with and took a direction same as the inferior wall of the closed penile urethra after physiological hypospadias. Depending on growth of the bulbospongiosus muscle and corpus spongiosus penis, the midline intermuscular septum obtained a connection to the subcutaneous wavy raphe and made the latter thick and straight at 12-15 weeks. Notably, the perineal raphe extended posteriorly to attach to the external anal sphincter. In female fetuses, an epithelial fusion occurred along a short distance at the posterior end of the vestibule. However, in front of the external anal sphincter, a large midline mesenchymal tissue from the urorectal septum did not contain a raphe-like structure. Moreover, since the bilateral bulbospongiosus muscles were separated widely by the vestibule, they did not provide a midline septum. Fetal development of the perineal raphe was accelerated by reinforcement from the muscular septum. In contrast, without such a muscular support, the female raphe could not maintain its growth even if the seed appeared at the posterior end of the vestibule.
Anal Canal* ; Female ; Fetal Development ; Fetus* ; Humans* ; Hypospadias ; Male ; Muscles ; Penis ; Scrotum ; Testis ; Urethra