Creation of fertility-restored materials for Ogura CMS in Brassica oleracea by introducing Rfo gene from Brassica napus via an allotriploid strategy.

KEY MESSAGE: Ogura CMS fertility-restored materials, with 18 chromosomes, normal seed setting, stable fertility and closer genetic background to the parent Chinese kale, were successfully developed in B. oleracea via a triploid strategy for the first time. Ogura cytoplasmic male sterility (CMS) is the most widely used sterile type in seed production for commercial hybrids of Brassica oleracea vegetables. However, the natural Ogura CMS restorer line has not been found in B. oleracea crops. In this study, the triploid strategy was used with the aim to create euploid B. oleracea progenies with the Rfo gene. The allotriploid AAC hybrid YL2 was used as a male parent to backcross with Ogura CMS Chinese kale. After successive backcrosses, the BC2 Rfo-positive individual 16CMSF2-11 and its BC3 progenies, with 18 chromosomes, were developed, which were morphologically identical to the parent Chinese kale. Compared with F1 and BC1 plants, it showed stable fertility performance, and regular meiosis behavior and could produce seeds normally under natural pollination. The genomic composition analysis of Rfo-positive progenies by using molecular markers showed that more than 87% of the C-genome components of BC3 Rfo-progenies recovered to the parent Chinese kale, while most or all of the An-genome segments were lost in 16CMSF2-11 and its progenies. The results suggested that the genetic background of Rfo-positive individuals was closer to that of the parent Chinese kale along with backcrossing. Hereof, the Ogura CMS fertility-restored materials of Chinese kale were successfully created via triploid strategy for the first time, providing a bridge for utilizing the Ogura CMS B. oleracea germplasm in the future. Moreover, our study indicates that the triploid strategy is effective for transferring genes from B. napus into B. oleracea.

Inheritance and expression of Bt cry1Ba3 gene in progeny from transformed cabbage plants.

Stable inheritance and expression of transgene are important parameters for successful use of a transgenic crop. We previously transformed a Bt cry1Ba3 gene into cabbage inbred line CA21-3. To evaluate the stability of our Bt cabbage lineages, transgene inheritance and expression were examined in four successive generations under greenhouse conditions. In our study, T1, T2 and T3 progenies of the three independent transgenic lineages (YA-1, YA-2 and YA-3) were generated and then the inheritance and expression of cry1Ba3 were analyzed in sexually derived progeny. Segregation ratio of 2.81:1, 3.27:1 and 3.07:1 was found in T1 progeny of lineages YA-1, YA-2 and YA-3, respectively. Chi-square analysis indicated that these segregation ratios of corresponding population fit the 3:1 ratio. Segregation ratios of the transgene in T2 progeny showed either 3:1 or all expression of cry1Ba3. These data suggest that cry1Ba3 in CA21-3 can be inherited in a Mendelian manner. ELISA analysis of transgenic plants from four generations demonstrated that cry1Ba3 had been stably transmitted to the T3 progeny. Additionally, under artificial infestation conditions, the homozygous T3-YA-1-2-1 line exhibited excellent resistance to Plutella xylostella as compared with un-transformed CA21-3. All these results imply that the three cabbage lineages are genetically stable and can be used to inhibit damage on cabbage caused by P. xylostella.

Antibacterial activity of 3-methylbenzo[d]thiazol-methylquinolinium derivatives and study of their action mechanism.

The increasing incidence of multidrug resistant bacterial infection renders an urgent need for the development of new antibiotics. To develop small molecules disturbing FtsZ activity has been recognized as promising approach to search for antibacterial of high potency systematically. Herein, a series of novel quinolinium derivatives were synthesized and their antibacterial activities were investigated. The compounds show strong antibacterial activities against different bacteria strains including MRSA, VRE and NDM-1 Escherichia coli. Among these derivatives, a compound bearing a 4-fluorophenyl group (A2) exhibited a superior antibacterial activity and its MICs to the drug-resistant strains are found lower than those of methicillin and vancomycin. The biological results suggest that these quinolinium derivatives can disrupt the GTPase activity and dynamic assembly of FtsZ, and thus inhibit bacterial cell division and then cause bacterial cell death. These compounds deserve further evaluation for the development of new antibacterial agents targeting FtsZ.

Berberine Protects against NEFA-Induced Impairment of Mitochondrial Respiratory Chain Function and Insulin Signaling in Bovine Hepatocytes.

Fatty liver is a major lipid metabolic disease in perinatal dairy cows and is characterized by high blood levels of non-esterified fatty acid (NEFA) and insulin resistance. Berberine (BBR) has been reported to improve insulin sensitivity in mice with hepatic steatosis. Mitochondrial dysfunction is considered a causal factor that induces insulin resistance. This study investigates the underlying mechanism and the beneficial effects of BBR on mitochondrial and insulin signaling in bovine hepatocytes. Revised quantitative insulin sensitivity check index (RQUICKI) of cows with fatty liver was significantly lower than that of healthy cows. Importantly, the Akt and GSK3ß phosphorylation levels, protein levels of PGC-1α and four of the five representative subunits of oxidative phosphorylation (OXPHOS) were significantly decreased in cows with fatty liver using Western Blot analysis. In bovine hepatocytes, 1.2 mmol/L NEFA reduced insulin signaling and mitochondrial respiratory chain function, and 10 and 20 umol/L BBR restored these changes. Furthermore, activation of PGC-1α played the same beneficial effects of BBR on hepatocytes treated with NEFA. BBR treatment improves NEFA-impaired mitochondrial respiratory chain function and insulin signaling by increasing PGC-1α expression in hepatocytes, which provides a potential new strategy for the prevention and treatment of fatty liver in dairy cows.

Genetics and fine mapping of a purple leaf gene, BoPr, in ornamental kale (Brassica oleracea L. var. acephala).

BACKGROUND: Due to its variegated and colorful leaves, ornamental kale (Brassica oleracea L. var. acephala) has become a popular ornamental plant. In this study, we report the fine mapping and analysis of a candidate purple leaf gene using a backcross population and an F2 population derived from two parental lines: W1827 (with white leaves) and P1835 (with purple leaves). RESULTS: Genetic analysis indicated that the purple leaf trait is controlled by a single dominant gene, which we named BoPr. Using markers developed based on the reference genome '02-12', the BoPr gene was preliminarily mapped to a 280-kb interval of chromosome C09, with flanking markers M17 and BoID4714 at genetic distances of 4.3 cM and 1.5 cM, respectively. The recombination rate within this interval is almost 12 times higher than the usual level, which could be caused by assembly error for reference genome '02-12' at this interval. Primers were designed based on 'TO1000', another B. oleracea reference genome. Among the newly designed InDel markers, BRID485 and BRID490 were found to be the closest to BoPr, flanking the gene at genetic distances of 0.1 cM and 0.2 cM, respectively; the interval between the two markers is 44.8 kb (reference genome 'TO1000'). Seven annotated genes are located within the 44.8 kb genomic region, of which only Bo9g058630 shows high homology to AT5G42800 (dihydroflavonol reductase), which was identified as a candidate gene for BoPr. Blast analysis revealed that this 44.8 kb interval is located on an unanchored scaffold (Scaffold000035_P2) of '02-12', confirming the existence of assembly error at the interval between M17 and BoID4714 for reference genome '02-12'. CONCLUSIONS: This study identified a candidate gene for BoPr and lays a foundation for the cloning and functional analysis of this gene.

Recessive male sterility in cabbage (Brassica oleracea var. capitata) caused by loss of function of BoCYP704B1 due to the insertion of a LTR-retrotransposon.

KEY MESSAGE: The LTR-retrotransposon insertion in BoCYP704B1 is proved to be the primary cause of the male sterility in cabbage. Effective allele-specific markers were developed for marker-assisted selection of male sterile gene. 83121A is a spontaneous male sterile mutant identified from cabbage. Genetic analysis indicated that male sterility is controlled by a single recessive gene. Pollen wall formation in the 83121A mutant was severely defective, with a lack of sporopollenin or exine. To understand the mechanisms of male sterility in 83121A, transcription analysis using RNA-Seq was carried out in the buds of the male sterile line 83121A and the male fertile line 83121B, which are near-isogenic lines differing only in the fertility trait. Via expression analysis of differentially expressed genes involved in pollen exine development before the bicellular pollen stage, BoCYP704B1 was identified as a candidate gene, which was approximately downregulated 30-fold in 83121A. BoCYP704B1 is a member of the evolutionarily conserved CYP704B family, which is essential for sporopollenin formation. The BoCYP704B1 transcript is specifically detected in the developing anthers of wild-type cabbage. Further sequence analysis revealed that a 5424-bp long terminal repeat-retrotransposon (LTR-RT) was inserted into the first exon of BoCYP704B1 in 83121A, which is not found in wild-type plants. The insertion of LTR-RT not only reduced the expression of BoCYP704B1 but also altered structure of protein encoded by BoCYP704B1. Moreover, linkage analysis showed that the homozygotic mutational BoCYP704B1 always cosegregated with male sterility. These data suggest that the LTR-RT insertion in BoCYP704B1 hinders sporopollenin formation in 83121A leading to male sterility. The allele-specific markers developed in this study were effective for marker-assisted selection of the male sterile gene.

Development of a novel allele-specific Rfo marker and creation of Ogura CMS fertility-restored interspecific hybrids in Brassica oleracea.

KEY MESSAGE: A novel allele-specific Rfo marker was developed and proved to be effective for MAS of Rfo gene in B. oleracea background and six Ogu-CMS fertility-restored interspecific hybrids were created for the first time. Ogura cytoplasmic male sterility (Ogu-CMS) has been extensively used for Brassica oleracea hybrid production. However, because of maternal inheritance, all the hybrids produced by CMS lines are male sterile and cannot be self-pollinated, which prohibits germplasm maintenance and innovation. This problem can be overcome by using the Ogu-CMS restorer line, but restorer material is absent in B. oleracea crops. Here, Rfo, a fertility-restored gene of Ogu-CMS, was transferred from rapeseed restorer lines into a Chinese kale Ogu-CMS line using interspecific hybridization combined with embryo rescue. Nine interspecific, triploid plant progenies were identified at morphological and ploidy level, with phenotypes intermediate between those of rapeseed and Chinese kale. Because the Rfo marker (Hu et al., Mol Breeding 22:663-674, 2008) cannot distinguish the Rfo and its homologies under a B. oleracea background, a novel allele-specific Rfo marker was developed based on the BLAST analysis of highly homologous Rfo sequences in B. oleracea. Screening using the novel Rfo marker found that six interspecific hybrids carrying Rfo were also fertile, although fertility varied during different flowering periods. Furthermore, BC1 offsprings with the Rfo gene were selected with the allele-specific Rfo marker and showed restored fertility. These results indicated that the novel allele-specific marker could be used for the MAS of Rfo gene in B. oleracea, and this study lays the foundation for the development of Ogu-CMS restorer material in cabbage and its related other subspecies.

Thermo-chemotherapy Induced miR-218 upregulation inhibits the invasion of gastric cancer via targeting Gli2 and E-cadherin.

Thermo-chemotherapy has been proven to reduce the invasion capability of cancer cells. However, the molecular mechanism underlying this anti-invasion effect is still unclear. In this study, the role of thermo-chemotherapy in the inhibition of tumor invasion was studied. The results demonstrated that expression of miR-218 was downregulated in gastric cancer tissues, which had a positive correlation with tumor invasion and metastasis. In vitro thermo-chemotherapy increased miR-218 expression in SGC7901 cells and inhibited both proliferation and invasion of cancer cells. Gli2 was identified as a downstream target of miR-218, and its expression was negatively regulated by miR-218. The thermo-chemotherapy induced miR-218 upregulation was also accompanied by increasing of E-cadherin expression. In conclusion, the present study indicates that thermo-chemotherapy can effectively decrease the invasion capability of cancer cells and increase cell-cell adhesion. miR-218 and its downstream target Gli2, as well as E-cadherin, participate in the anti-invasion process.

Personality Traits Affect the Cost-Effectiveness of Total Knee Arthroplasty.

OBJECTIVE: To assess the clinical benefit and compare the cost-effectiveness of total knee arthroplasty (TKA) in patients with different personality traits. METHODS: The present study was retrospectively conducted from January 2017 to May 2018. A total of 232 patients between 46 and 71 years old who underwent unilateral, primary TKA with the diagnosis of knee osteoarthritis were interviewed. Three types of data were required to compare the cost-effectiveness differences among groups: personality traits, postoperative clinical outcomes about health-related quality of life, and costs associated with TKA. Personality was assessed using the Eysenck Personality Questionnaire, functional outcome was assessed through the Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) questionnaire, and costs were evaluated. Besides, the marginal cost-effectiveness ratio (MCER) as the primary outcome, which relates the direct costs to the associated patient benefit as assessed by the clinical endpoint ($/quality-adjusted life years [QALY]), was compared among different personality traits. All information for this study was acquired by directly interviewing the patients and reviewing the medical computer records at our hospital. RESULTS: Two hundred and eleven patients completed the final analysis with an average of 24.6 months follow-up postoperatively. The choleric group, sanguine group, melancholic group, and phlegmatic group has 41, 70, 46, and 54 patients, respectively. A statistically significant difference in MECR, QALYs, and postoperative WOMAC existed among different personality traits (all P < 0.05). There was no significant difference in mean age (P = 0.588), body mass index (BMI) (P = 0.790), smoking (P = 0.934), heavy drinking (P = 0.994), chronic comorbidities (all P > 0.05), preoperative albumin <3.5 g/dL (P = 0.991), and American Society of Anaesthesiologists (ASA) score (P = 0.687) among personality traits. More women tend to be melancholic in comparison to other personality traits (P = 0.016). Melancholic patients attested inferiority of TKA compared with other personality traits, who would pay for the same QALYs at the highest costs (P < 0.05). By contrast, sanguine patients have a more cost-effective TKA than other personality traits, as they pay the least money for the same QALYs (P < 0.05). Although phlegmatic and choleric patients seemingly have moderate gains from TKA, in general, the extroversion (measured by the extroversion subscale) and stability (measured by the neuroticism subscale) displayed more pleasurable QALYs in comparison with introversion and instability (P < 0.05). Sensitivity analysis showed that the results mentioned above appeared not to be sensitive when varying key parameters (prosthesis survival and life expectancy) in a one-way sensitivity analysis. Sanguine and melancholic patients still have the lowest and highest MCER in comparison with choleric and phlegmatic traits (P < 0.05). The multivariate logistic regression showed that RA (adjusted OR = 1.3, 95% CI = 1.2-1.4, P < 0.01), ASA Class I-II (adjusted OR = 0.9, 95% CI = 0.8-1.0, P < 0.001), sanguine (adjusted OR = 0.8, 95% CI = 0.7-0.9, P < 0.001) and melancholic (adjusted OR = 1.2, 95% CI = 1.1-1.3, P < 0.001) were significantly associated with MCER. CONCLUSIONS: Before surgery, screening the melancholic patients would significantly reduce the economic burden, avoid unnecessary suffering, and shorten the recovery period.

Symmetry in Paraspinal Muscles as a Predictor of the Development of Pregnancy-Related Low Back and Pelvic Pain: A Prospective Study.

OBJECTIVE: To determine the asymmetry in the paraspinal muscle before pregnancy and evaluate its association with pregnancy-associated lumbopelvic pain (LPP). METHODS: This was a prospective case-control study conducted from January 2017 and December 2018. A total of 171 subjects (mean age ± SD, 27.4 ± 5.8 years) were finally divided into the LBP group, PGP group, and no LPP group. Each subject was asked to follow a standardized clinical imaging protocol before the pregnancy. The area of muscles (multifidus, erector spinae, and psoas muscles) on the axial slice at mid-disc of L4 -L5 and L5 -S1 were segmented and then the cross-sectional area (CSA) of a particular muscle was measured by outlining the innermost fascial border surrounding each muscle. The mean value of F-CSA's ratio to T-CSA (F/T CSA) was used to determine whether the bilateral paraspinal muscle was asymmetrical. Total muscle CSA (T-CSA) represents the sum of CSA of interested three muscles. The signal intensity can distinguish fat and muscle tissue in a different range. Based on this, functional CSA (F-CSA), represented by fat-free area, was evaluated quantitively by excluding the signal of the deposits of intramuscular fat. Total muscle CSA (T-CSA), functional CSA (F-CSA), and the ratio of F-CSA to T-CSA (F/T CSA) were measured unilaterally and compared between groups. Logistic regression was performed to determine the risk factors for pregnancy-associated LPP. The Pearson correlation coefficient was performed to test the relationship between asymmetry in F/T-CSA and pain rating. RESULTS: A total of 124 subjects (72.5%) (28.5 ± 5.2 years) had LPP during pregnancy. Forty-eight (38.7%) individuals had low back pain (LBP) and 76 (61.3%) had pelvic girdle pain (PGP). Seventy-six women (44.4%) were determined to have asymmetry in paraspinal muscle according to the definition in this methods section. The duration of follow-up was 24 months postpartum. A total of 39 (31.5%) women unrecovered from LPP. F/T-CSA was significantly decreased for LBP in the PGP group than in the and control group (0.03 ± 0.02 vs 0.05 ± 0.03 vs 0.12 ± 0.05, P < 0.001). Meanwhile, significant differences were detected in both groups (all P < 0.001). In patients with LBP, the level of paraspinal asymmetry, represented by the difference in F/T-CSA, was positively correlated with pain scores (r = 0.52, P < 0.01). However, no statistically significant correlation between pain scores and paraspinal asymmetry was found in PGP (r = 0.42, P > 0.05). Asymmetry in the paraspinal muscle (adjusted OR = 1.5), LBP (adjusted OR = 1.6), LPP in a previous pregnancy (adjusted OR = 1.4), sick leave ≥90 days (adjusted OR = 1.2), and heavy labor (adjusted OR = 1.2) were risk factors for the unrecovered LPP during pregnancy. CONCLUSIONS: Asymmetrical muscular compositions could lead to abnormal biomechanics for the segmental motions. Lateral-directed physical training and stretching may help decrease the occurrence and severity of this condition.

Nuciferine improves high-fat diet-induced obesity via reducing intestinal permeability by increasing autophagy and remodeling the gut microbiota.

Nuciferine (NF) has received extensive attention due to its medicinal value in the treatment of metabolic diseases, such as obesity; however, to date, the effects of NF on obesity-related intestinal permeability, autophagy and the gut microbiota have not been investigated. Herein, C57BL/6J mice were fed either a chow or a high-fat diet (HFD) with or without NF for 8 weeks. The results showed that NF supplement reduced weight gain, fat accumulation and intestinal permeability in the HFD mice accompanied by improved autophagy. Subsequently, an in vitro experiment was performed using Caco-2 and HT-29 cells, which showed that NF supplement not only promoted the formation of autophagosomes and autophagolysosomes, but also alleviated LPS-increased intestinal permeability. Importantly, NF supplement protected from LPS-induced paracellular permeability impairment after the administration of autophagy-related gene (Atg) 5 small-interfering RNA (siRNA). These results demonstrate that NF exerts beneficial effects on the intestinal permeability by improving autophagy. Furthermore, we also found that NF supplement lowered the abundance of Butyricimonas and increased the abundance of Akkermansia, an anti-obesity bacterium. Thus, overall, we demonstrated that NF supplement confers reduced intestinal permeability by improving autophagy and alters the composition of the gut microbiota in HFD-fed mice, thereby producing an anti-obesity effect.

Tanshinone IIA downregulates the CD40 expression and decreases MMP-2 activity on atherosclerosis induced by high fatty diet in rabbit.

Tanshinone IIA (Tan IIA) is a member of the major lipophilic components abstracted from the root of Salvia miltiorrhiza Bunge and has the capacity of anti-atherosclerosis. To investigate the potential mechanism, we established an animal model by giving high fatty diet to rabbits and Tan IIA was given in different dose. Then, superoxide dismutase (SOD) activity and the malondialdehyde (MDA) level in serum were detected using spectrophotometry; cluster of differentiation 40 (CD40) expression of cellular membrane fraction of aortas and matrix metalloproteinase-2 (MMP-2) activity of total protein extract of aortas were detected by Western Blotting and Zymography, respectively. Compared with the control group, the level of MDA, the expression of CD40 and the MMP-2 activity were increased while the SOD activity was decreased significantly in model group. After Tan IIA administration, the SOD activity was significantly increased while the level of MDA was decreased; both the expression of CD40 and the activity of MMP-2 were decreased. It is suggested that Tan IIA not only inhibits the oxidation but also suppresses the inflammation in atherosclerotic lesion. Our data suggest that not only anti-oxidation but also anti-inflammation by decrease the expression of CD40 and MMP-2 activity maybe the potential mechanisms by which Tan IIA anti-atherosclerosis.

Study of Benzofuroquinolinium Derivatives as a New Class of Potent Antibacterial Agent and the Mode of Inhibition Targeting FtsZ.

New generation of antibacterial agents are urgently needed in order to fight the emergence of multidrug-resistant bacteria. FtsZ is currently identified as a promising target for new types of antimicrobial compounds development because of its conservative characteristics and its essential role played in bacterial cell division. In the present study, the antibacterial activity of a series of benzofuroquinolinium derivatives was investigated. The results show that the compounds possess potent antibacterial activity against drug resistant pathogens including MRSA, VREF and NDM-1 Escherichia coli. Biological studies reveal that the compound is an effective inhibitor that is able to suppress FtsZ polymerization and GTPase activity and thus stopping the cell division and causing cell death. More importantly, this series of compounds shows low cytotoxicity on mammalian cells and therefore they could be new chemotypes for the development of new antibacterial agents targeting the cell-division protein FtsZ.

Tanshinone IIA inhibits atherosclerotic plaque formation by down-regulating MMP-2 and MMP-9 expression in rabbits fed a high-fat diet.

Tanshinone IIA (Tan IIA) is one of the major lipophilic components of Salvia miltiorrhiza Bunge (SM) and has an anti-atherosclerotic effect. To investigate the potential mechanism of this effect, we established an atherosclerotic animal model by feeding rabbits a high-fat diet, and Tan IIA was given at different doses. Intimal area of the aorta was measured by image analysis, serum levels of vascular adhesion molecule-1 (VCAM-1) and interleukin (IL-1beta) were measured by ELISA, while matrix metalloproteinase-2 and-9 (MMP-2, MMP-9) expression and their activities in atherosclerotic lesions were assessed by Western blotting and zymography respectively. Compared with the control group, the intimal area, serum levels of VCAM-1 and IL-1beta, the expression of MMP-2 and MMP-9 as well as their activities were increased significantly in the high-fat fed rabbit group. After Tan IIA administration, all of these parameters were decreased in a dose-dependent manner. Our results show that Tan IIA can inhibit atherosclerotic lesion formation in aorta and down-regulate protein expression and activities of MMP-2 and MMP-9 as well as serum VCAM-1 and IL-1beta in rabbits fed a high-fat diet.

The protective effect and mechanism of sevoflurane on LPS-induced acute lung injury in mice.

Acute lung injury (ALI) is a disturbance caused by infectious or non-infectious inflammation and lipopolysaccharide (LPS) could induce an artificial pathological ALI process. Sevoflurane has been demonstrated to be an inhaled anesthetic having anti-inflammatory and protective effects on inflammatory injury. To study the protective effects and mechanisms of sevoflurane on LPS-induced acute lung injury in mice. By assessing W/D ratio, sevofluranecan counteract the edema induced by LPS. The ELISA results showed that sevoflurane reduced IFN-γ production and increased IL-10 level. Elevation of PGE2 induced by sevofluraneand LPS in peritoneal macrophages was inhibited by NS-398, an inhibitor of the PGE2 regulator COX-2, indicating that NS-398 blocked COX-2 mediated PGE2 synthesis. NS-398 itself did not cause lung inflammation and mitigated the protective effect of sevoflurane on LPS-induced ALI in mice. LPS changes immune homeostasis, resulting in acute lung inflammatory injury. Inhaled sevoflurane regulates immune homeostasis, thereby playing a protective role in alleviating LPS-induced ALI.

MicroRNA-218 is upregulated in gastric cancer after cytoreductive surgery and hyperthermic intraperitoneal chemotherapy and increases chemosensitivity to cisplatin.

AIM: To investigate the molecular mechanisms of miRNA in advanced gastric cancers (AGCs) before and after cytoreductive surgery (CRS) + hyperthermic intraperitoneal chemotherapy (HIPEC). METHODS: A miRNA microarray containing human mature and precursor miRNA sequences was used to compare expression profiles in serum samples of 5 patients with AGC before and after CRS + HIPEC. The upregulation of miR-218 was confirmed by real-time reverse transcription polymerase chain reaction and its expression was analyzed in SGC7901 gastric cancer cells. RESULTS: miRNA microarray chip analysis found that the level of miR-218 expression was upregulated more than 8 fold after CRS + HIPEC. Furthermore, miR-218 increased gastric cancer cell chemosensitivity to cisplatin in vitro and inhibited gastric cell tumor growth in nude mice in vivo (0.5 vs 0.78, P < 0.05). CONCLUSION: Our results indicated that targeting miR-218 may provide a strategy for blocking the development of gastric cancer and reverse the multi-drug resistance of gastric cell lines.

[Inhibitor of tyrosine phosphorylation AG490 inhibits the proliferation of Jurkat T cells].

AIM: To study effect of AG490 on the activation, proliferation, cell cycle, apoptosis and the expression of protein ICBP90 of Jurkat T cells. To explore the possibility of inhibiting and the initial growth mechanism of Jurkat T cells by blocking JAK signaling pathway. METHODS: The expression level of CD69 and CD25 on the surface of Jurkat T cells was evaluated by fluorescin-conjugated monoclonal antibody double labeling technique. The proliferation of Jurkat T cells was analyzed by MTT method. The cycle of Jurkat T cell was analyzed by PI staining and Flow Cytometry. The apoptosis of Jurkat T cells was analyzed by Annexin V-FITC and PI staining. The expression of ICBP90 that correlated with the cell proliferation was checked by Western blot. RESULTS: The results came out that the proliferation capacity of Jurkat T cells was decreased with the increasing of AG490 ranging from 1 mmol/L to 30 mmol/L in a dose-dependent manner with the highest inhibitory rate of 27.37%. Cell-cycle analysis revealed that AG490 arrest it in G0/G1 phase and inhibited its entrance into S and G2/M phase, and reduced the expression of protein ICBP90, especially in the 24 h treatment. But the apoptosis of Jurkat T cells was not promoted by AG490. CONCLUSION: AG490 significantly inhibited the growth of Jurkat T cells, and its inhibition may reduce the expression of protein ICBP90, which was related to the cell cycle arrest rather than apoptosis.

[Tanshinone IIA down-regulates the expression of MMP-12 and TF in RAW 264.7 cells].

OBJECTIVE: To investigate the effect of tanshinone IIA (TanIIA) on the expression of tissue factor (TF) and matrix metalloproteinase-12 (MMP-12) in RAW264.7 cells and explore the possible mechanism. METHODS: RAW 264.7 cells were incubated with ox-LDL in the presence or absence of different concentrations of tanshinone IIA. At the end of the incubation, the cell proliferation was assessed by MTT assay, and superoxide dismutase (SOD) activity and malondialdehyde (MDA) and TF concentrations in the supernatant were detected by xanthine oxidase method, thiobarbituric acid method and ELISA, respectively. Western blotting was employed to determine MMP-12 expression in the cells. RESULTS: The cell proliferation was dose-dependently inhibited by TanIIA. SOD activity in the supernatant was increased significantly, while the MDA and TF concentration and MMP-12 expression in cells decreased after treatment of the cells with different concentrations of TanIIA. CONCLUSION: TanIIA inhibits the cell proliferation and TF and MMP-12 expressions in RAW264.7 cells stimulated by ox-LDL, and these effects may be related with the anti-oxidation property of TanIIA.

[Effect of Jagged-1 on the cytokine expression of murine myeloid-derived dendritic cells].

AIM: To investigate the in vitro effect of soluble Jagged-1/Fc on the cytokine expression of murine myeloid-derived dendritic cells induced by both rmGM-CSF and rmIL-4. METHODS: The DC culture model under rmGM-CSF plus rmIL-4 conditions was established. The influences of Jagged-1/Fc on the morphology of DC was checked, and the cytokine expression were analyzed by Luminex system and ELISA. The activation capacity of Jagged-1/Fc-induced DC for allogeneic lymphocytes was detected by MTT assay. RESULTS: Compared with LPS or Zymosan A, the amount of IL-4 expression of Jagged-1/Fc-induced DC increased significantly and the amount of TNF-alpha expression decreased markedly but gamma-secretase inhibitor DAPT reversed the inhibition of Jagged-1/Fc on TNF-alpha expression from DC and returned it to the control level. Jagged-1/Fc altered the amount of IL-10, IL-6, IL-2, IL-12 and IFN-gamma expression unconspicuously and had no effect on TGF-beta expression. The mixed lymphocyte reaction(MLR) revealed that Jagged-1/Fc-induced DC had the weakest ability to activate allogeneic lymphocytes and LPS-induced DC showed the strongest activation ability. CONCLUSION: The results indicate that Jagged-1/Fc-induced DC incline to induce immune tolerance and to prime naïve T cells to the Th2 cells.

Fenofibrate inhibits tumor necrosis factor-alpha-induced expression of CD40 and matrix metalloproteinase in human vascular endothelial cells.

OBJECTIVE: To investigate the regulatory effects of fenofibrate on TNF-alpha-induced CD40 expression and matrix metalloproteinase (MMP) activity in human vascular endothelial cells (HUVECs). METHODS: Quantitative RT-PCR and flow cytometry were employed to evaluate the effect of fenofibrate on TNF-alpha-induced CD40 mRNA and cell surface CD40 expression in HUVECs, and gelatin zymography was used to determine the effect of fenofibrate on the gelatinolytic activities of MMP-2 and MMP-9 in TNF-alpha-stimulated HUVECs. RESULTS: Fenofibrate at the concentrations of 5x10(-5), 1x10(-4) and 2x10(-4) mol/L significantly reduced TNF-alpha-induced increment of CD40 mRNA and cell surface CD40 expressions (P<0.01), with the maximal inhibition achieved at the concentration of 1x10(-4) mol/L. Fenofibrate at 2x10(-4) mol/L did not further decrease CD40 expression induced by TNF-alpha. Fenofibrate significantly inhibited the stimulatory effect of TNF-alpha on MMP-2 and MMP-9 activities in HUVECs. CONCLUSION: Fenofibrate reduces TNF-alpha-induced increment of CD40 expression and MMP-2 and MMP-9 activities in HUVECs.