Dual roles of the MPK3 and MPK6 mitogen-activated protein kinases in regulating Arabidopsis stomatal development.

An Arabidopsis (Arabidopsis thaliana) mitogen-activated protein kinase (MAPK) cascade composed of YODA (YDA)-MKK4/MKK5-MPK3/MPK6 plays an essential role downstream of the ERECTA (ER)/ER-LIKE (ERL) receptor complex in regulating stomatal development in the leaf epidermis. STOMAGEN (STO), a peptide ligand produced in mesophyll cells, competes with EPIDERMAL PATTERNING FACTOR2 (EPF2) for binding ER/ERL receptors to promote stomatal formation. In this study, we found that activation of MPK3/MPK6 suppresses STO expression. Using MUTE and STO promoters that confer epidermis- and mesophyll-specific expression, respectively, we generated lines with cell-specific activation and suppression of MPK3/MPK6. The activation or suppression of MPK3/MPK6 in either epidermis or mesophyll cells is sufficient to alter stomatal differentiation. Epistatic analyses demonstrated that STO overexpression can rescue the suppression of stomatal formation conferred by the mesophyll-specific expression of the constitutively active MKK4DD or MKK5DD, but not by the epidermis-specific expression of these constitutively active MKKs. These data suggest that STO is downstream of MPK3/MPK6 in mesophyll cells, but upstream of MPK3/MPK6 in epidermal cells in stomatal development signaling. This function of the MPK3/MPK6 cascade allows it to coordinate plant epidermis development based on its activity in mesophyll cells during leaf development.

Fermentation of Polygonati Rhizoma aqueous extract using Lactiplantibacillus plantarum under the condition of eutrophication.

In this experiment, the eutrophication system was established by adding sucrose and yeast powder, and the pH and dissolved oxygen were measured in a bioreactor in real time to study the effect of aerobic environment on the fermentation process of Polygonati Rhizoma extract by Lactiplantibacillus plantarum. To further analyze metabolic changes, UPLC-Q-Exactive MS was used for metabolomic analysis and metabolic profiling. Multivariate analysis was performed using principal component analysis and Orthogonal projections to latent structures discriminant analysis. Finally, 313 differential metabolites were selected, 196 of which were annotated through database matching. After fermentation, the content of short-chain fatty acids, lactic acid, and their derivatives increased significantly, and there were 13 kinds and 4 kinds, respectively. Both compounds and their derivatives are beneficial to the intestinal flora. Consequently, incorporating L. plantarum into the aerobic fermentation process of Polygonati Rhizoma extract within the eutrophic system is potentially advantageous in enhancing the impact of its fermentation solution on the gut microbiota and its effects on human health. Our findings for this kind of edible and medicinal material research and development offer useful insights.

Ginsenoside Rg1 attenuates D-galactose-induced neural stem cell senescence via the Sirt1-Nrf2-BDNF pathway.

With the ageing of society's population, neurodegenerative diseases have become an important factor affecting the quality of life and mortality in the elderly. Since its physiopathological processes are complex and the authorized medications have recently been shown to have several adverse effects, the development of safe and efficient medications is urgently needed. In this study, we looked at how ginsenoside Rg1 works to postpone neural stem cell ageing and brain ageing, giving it a solid scientific foundation for use as a therapeutic therapy for neurodegenerative diseases.

Complexation of Apigenin and Oxymatrine Leading to Enhanced Anti-inflammatory Activity.

Apigenin (APG) is a well-known dietary flavonoid with multiple bioactivities, but its poor aqueous solubility may result in low oral bioavailability and thus compromised therapeutic effects. In the present study, APG was complexed with oxymatrine (OMT), a natural quinolizidine alkaloid, for enhanced anti-inflammatory activity, and the related mechanisms in the interaction of APG with OMT were investigated. Fourier transform-infrared spectroscopy, fluorescence spectroscopy, Raman spectroscopy, and proton nuclear magnetic resonance spectroscopy characterizations demonstrated the occurrence of an APG-OMT complex formed at a molar ratio of 1:2. Then, molecular dynamics simulations and quantum chemical calculations were utilized to elucidate that hydrogen bonding, van der Waals forces, and hydrophobic effects were the main forces acting in the formation of the APG-OMT complex. Pharmacokinetic studies in rats demonstrated that the oral bioavailability of APG in the APG-OMT complex was significantly higher than that of APG alone. Finally, bioactivity evaluation in the lipopolysaccharide-induced acute inflammatory injury mouse models showed that the APG-OMT complex exhibited more potent anti-inflammatory effects than APG alone. This study confirmed that APG and OMT exerted enhanced anti-inflammatory effects through self-complexation, which may provide a novel strategy for improving the bioavailability and bioactivity of natural product mixtures.

The reliability of analytical reference lines for determining esthetically pleasing lip position: An assessment of consistency, sensitivity, and specificity.

INTRODUCTION: This study aimed to identify a simple yet reliable soft-tissue parameter for the clinical determination of esthetic lip position by investigating the most consistent reference lines and assessing their sensitivity and specificity. METHODS: A total of 5745 records from Chinese patients aged >18 years were screened. In part I of the study, lateral view photographs of 96 subjects (33 males, 63 females) with esthetic facial profiles were selected. The profile esthetics of each photograph was first scored by 52 dental students, followed by 97 laypeople on a 5-point attractiveness scale. For the top 25% of photographs with the highest score for each sex (8 males, 16 females), the consistency of 6 commonly used reference lines were assessed to determine the esthetic lip position. In part II of the study, lip positions relative to Steiner's (S) and Ricketts' (E) lines in the profile photographs of 86 patients (43 males, 43 females) deemed to have an esthetically unpleasing profile were compared with those in 86 Chinese movie star idols (43 males, 43 females). RESULTS: In part I of the study, the S, E, and Burstone's (B) lines exhibited the lowest standard deviations for the upper and lower lips. B line was excluded from further analysis because of its higher mean absolute values, and S and E lines were used for the subjective assessment in part II of the study. In part II, the S line showed a sensitivity of 86.0% and 86.0% and a specificity of 81.4% and 83.7% for males and females, respectively. In contrast, the E line presented a sensitivity of 88.4% and 93.0% and a specificity of 79.1% and 74.4% for males and females, respectively. CONCLUSIONS: S, E, and B lines were the most consistent soft-tissue parameters among both sexes; however, because of the smaller absolute values, the S line would be more convenient among the 3 for a quick clinical assessment of lip position. Moreover, the performance of both S and E lines was similar among both sexes, which supports using these lines in assessing the esthetic lip position.

Angelica sinensis polysaccharides alleviate the oxidative burden on hematopoietic cells by restoring 5-fluorouracil-induced oxidative damage in perivascular mesenchymal progenitor cells.

CONTEXT: 5-Fluorouracil (5-FU)-injured stromal cells may cause chronic bone marrow suppression; however, the underlying mechanism remains unclear. Angelica sinensis polysaccharide (ASP), the main biologically active ingredient of the Chinese herb, Angelica sinensis (Oliv.) Diels (Apiaceae), may enrich the blood and promote antioxidation. OBJECTIVE: This study investigated the protective antioxidative effects of ASP on perivascular mesenchymal progenitors (PMPs) and their interactions with hematopoietic cells. MATERIALS AND METHODS: PMPs were dissociated from C57BL/6 mouse femur and tibia and were subsequently divided into the control, ASP (0.1 g/L), 5-FU (0.025 g/L), and 5-FU + ASP (pre-treatment with 0.1 g/L ASP for 6 h, together with 0.025 g/L 5-FU) then cultured for 48 h. Hematopoietic cells were co-cultured on these feeder layers for 24 h. Cell proliferation, senescence, apoptosis, and oxidative indices were detected, along with stromal osteogenic and adipogenic differentiation potentials. Intercellular and intracellular signaling was analyzed by real-time quantitative reverse transcription polymerase chain reaction and Western blotting. RESULTS: ASP ameliorated the reactive oxygen species production/scavenge balance in PMPs; improved osteogenic differentiation; increased SCF, CXCL12, VLA-4/VCAM-1, ICAM-1/LFA1, and TPO/MPL, Ang-1/Tie-2 gene expression. Further, the ASP-treated feeder layer alleviated hematopoietic cells senescence (from 21.9 ± 1.47 to 12.1 ± 1.13); decreased P53, P21, p-GSK-3ß, ß-catenin and cyclin-D1 protein expression, and increased glycogen synthase kinase (GSK)-3ß protein expression in co-cultured hematopoietic cells. DISCUSSION AND CONCLUSIONS: ASP delayed oxidative stress-induced premature senescence of 5-FU-treated feeder co-cultured hematopoietic cells via down-regulation of overactivated Wnt/ß-catenin signaling. These findings provide a new strategy for alleviating myelosuppressive stress.

[Effect of apigenin in combination with oxymatrine on non-small cell lung cancer and mechanism].

This study explores the effect of apigenin(APG), oxymatrine(OMT), and APG+OMT on the proliferation of non-small cell lung cancer cell lines and the underlying mechanisms. Cell counting kit-8(CCK-8) assay was used to detect the vitality of A549 and NCI-H1975 cells, and colony formation assay to evaluate the colony formation ability of the cells. EdU assay was employed to examine the proliferation of NCI-H1975 cells. RT-qPCR and Western blot were performed to detect the mRNA and protein expression of PLOD2. Molecular docking was carried out to explore the direct action ability and action sites between APG/OMT and PLOD2/EGFR. Western blot was used to study the expression of related proteins in EGFR pathway. The viability of A549 and NCI-H1975 cells was inhibited by APG and APG+OMT at 20, 40, and 80 µmol·L~(-1) in a dose-dependent manner. The colony formation ability of NCI-H1975 cells was significantly suppressed by APG and APG+OMT. The mRNA and protein expression of PLOD2 was significantly inhibited by APG and APG+OMT. In addition, APG and OMT had strong binding activity with PLOD2 and EGFR. In APG and APG+OMT groups, the expression of EGFR and proteins in its downstream signaling pathways was significantly down-regulated. It is concluded that APG in combination with OMT could inhibit non-small lung cancer, and the mechanism may be related to EGFR and its downstream signaling pathways. This study lays a new theoretical basis for the clinical treatment of non-small cell lung cancer with APG in combination with OMT and provides a reference for further research on the anti-tumor mechanism of APG in combination with OMT.

Nasal asymmetry changes during growth and development in 6- to 12-year-old children with repaired unilateral cleft lip and palate: A 3D computed tomography analysis.

Repaired unilateral cleft lip and palate (UCLP) is often accompanied by the deformity and asymmetry of the nasal region. Three-dimensional analysis was performed to investigate the relationship between nasal soft- and hard-tissue asymmetries, as well as the changes in nasal asymmetry with age, among children with repaired UCLP (age: 6-12 years). Forty-seven patients were included in this study. Their computed tomography records were retrieved for analysis of the 3D asymmetry of 10 landmarks of the nasal soft and hard tissues. We observed that asymmetry was more severe in nasal hard tissues than in soft tissues, particularly in the sagittal dimension. Compared with patients aged 6-9 years old, patients aged 10 to 12 years old had significantly increased vertical asymmetry at the base of the alar groove (Gbase, p = 0.027) and the lateral point of the piriform aperture (LPA), (p < 0.001). The correlation between the LPA and the alar region was weak to moderate (r = 0.290 to 0.488). In conclusion, we found no evidence of growth and development in nasal hard-tissue asymmetry among 6- to 12-year-old children with repaired UCLP, except for the vertical dimension. Nasal soft tissue exhibited a more preferable symmetry than hard tissue, and this could be attributed to the compensatory growth of nasal soft tissue, particularly in the vertical and sagittal dimensions. The weak to moderate correlations between nasal soft-tissue asymmetry and hard-tissue asymmetry were observed in the three dimensions. Surgeons should consider these factors when repositioning the nasal alar and controlling the size of the nostrils.

[Active fractions of Camellia nitidissima inhibit non-small cell lung cancer via suppressing epidermal growth factor receptor].

The present study explored the effects and its underlying mechanisms of four active fractions of Camellia nitidissima(leaf polyphenols, leaf saponins, flower polyphenols, and flower saponins in C. nitidissima) in inhibiting the proliferation and migration of non-small cell lung cancer(NSCLC) by suppressing the epidermal growth factor receptor(EGFR). MTT assay was used to detect the effect of four active fractions on the proliferation of NCI-H1975 and HCC827 cells. Wound healing assay and Transwell assay were adopted to evaluate the effect of four active fractions on the migration of NSCLC. The effect of four active fractions on the enzyme activity of EGFR was detected. Molecular docking was carried out to explore the direct action capacity and action sites between representative components of the four active fractions and EGPR. Western blot assay was employed to investigate the effect of four active fractions on the protein expression in EGFR downstream signaling pathways. The results of the MTT assay indicated that the cell viability of NCI-H1975 and HCC827 cells was significantly inhibited by four active fractions at 50, 100, 150, and 200 µg·mL~(-1) in a dose-dependent manner. Wound healing assay and Transwell assay revealed that the migration of NCI-H1975 and HCC827 cells was significantly suppressed by four active fractions. In addition, the results of the protein activity assay showed that the enzyme activity of EGFR was significantly inhibited by four active fractions. The molecular docking results confirmed that various components in four active fractions possessed strong binding activity to EGFR enzymes. Western blot assay revealed that four active fractions down-regulated the protein expression of EGFR and its downstream signaling pathways. It is concluded that the four active fractions of C. nitidissima can inhibit NSCLC. The mechanism may be related to EGFR and its downstream signaling pathways. This study provides a new scientific basis for the clinical treatment of NSCLC with active fractions of C. nitidissima, which is of reference significance for further research on the anti-tumor mechanism of C. nitidissima.

Survival and immune response of white shrimp Litopenaeus vannamei following single and concurrent infections with WSSV and Vibrio parahaemolyticus.

The survival and immune responses of Litopenaeus vannamei were evaluated during white spot syndrome virus (WSSV) or Vibrio parahaemolyticus single and concurrent infections. The mortality, WSSV load, activities of 4 immune enzymes: acid phosphatase (ACP), alkaline phosphatase (AKP), peroxidase (POD) and superoxide dismutase (SOD), and the transcription of Evolutionarily Conserved Signaling Intermediate in Toll pathways of L.vannamei (LvECSIT) were quantified at 0, 3, 6, 12, 24, 48, 72 and 96 h post-infection (pi). The results showed: (i) the cumulative mortality of the co-infection group (WSSV and V. Parahaemolyticus 83%) was significantly lower than the WSSV infection group (97%) (P < 0.05) at 96 hpi; (ii) copies of WSSV in the co-infection group were significantly lower than that of the single infection group from 24 to 96 hpi (P < 0.05); (iii) ACP, AKP,POD and SOD activity in the gills of the co-infection group was higher than that of the WSSV group at12, 48 and 96 hpi (P < 0.05).The expression of LvECSIT mRNA in the co-infection group was significantly higher than in the WSSV infection group from 12 to 72 hpi (P < 0.05).The results indicate that proliferation of WSSV is inhibited by V.parahaemolyticus infection. In addition, infection with WSSV alone causes a significant reduction in some immune responses of shrimp than co-infection with WSSV and V.parahaemolyticus occurs at 26 °C. Third, LvECSIT, an essential member of TLR signaling pathway might play a crucial role in shrimp defense against WSSV - Vibrio co-infection.

Orthodontic Treatment Need and the Psychosocial Impact of Malocclusion in 12-Year-Old Hong Kong Children.

OBJECTIVES: To determine the prevalence of orthodontic treatment need in 12-year-old children in Hong Kong and its relationship with the psychosocial impact of malocclusion and to assess their associations with sociodemographic factors. MATERIALS AND METHODS: A random sample of 687 12-year-old children was recruited from 45 secondary schools in Hong Kong. Orthodontic treatment need was assessed on study models by five indices: the Dental Health Component of the Index of Orthodontic Treatment Need (IOTN-DHC), the Aesthetic Component of the IOTN (IOTN-AC), the Dental Aesthetic Index (DAI), the Index of Complexity Outcome and Need (ICON), and the Peer Assessment Rating (PAR). The psychosocial impact of malocclusion on participants and sociodemographic factors were obtained from a questionnaire. Logistic regression was used to examine the correlations between treatment need and the psychosocial impact of malocclusion as well as their associations with sociodemographic factors. RESULTS: The final number of participants was 667 (339 boys and 328 girls, participation rate 667/687 = 97.1%). The prevalence of orthodontic treatment need varied depending on the indices used (10.9-47.8%), but significant correlations were found among the five indices (p < 0.01). The uptake of treatment among the cohort was 2.3%. Boys had higher IOTN-DHC (p < 0.05), DAI (p < 0.05), and PAR (p = 0.05) scores than girls. IOTN-AC was significantly associated with the psychosocial impact of malocclusion (p < 0.05). Parents' level of education and household income were not significantly associated with either treatment need or the psychosocial impact of malocclusion (p > 0.05). CONCLUSION: The need for orthodontic treatment in 12-year-old children in Hong Kong remained high, and the uptake of treatment was low. Boys had a higher normative treatment need than girls. Among the five indices, IOTN-AC appears to be the best indicator of the psychosocial impact of malocclusion.

Ginsenoside Rg1 Decreases Oxidative Stress and Down-Regulates Akt/mTOR Signalling to Attenuate Cognitive Impairment in Mice and Senescence of Neural Stem Cells Induced by D-Galactose.

Adult hippocampal neurogenesis plays a pivotal role in learning and memory. The suppression of hippocampal neurogenesis induced by an increase of oxidative stress is closely related to cognitive impairment. Neural stem cells which persist in the adult vertebrate brain keep up the production of neurons over the lifespan. The balance between pro-oxidants and anti-oxidants is important for function and surviving of neural stem cells. Ginsenoside Rg1 is one of the most active components of Panax ginseng, and many studies suggest that ginsenosides have antioxidant properties. This research explored the effects and underlying mechanisms of ginsenoside Rg1 on protecting neural stem cells (NSCs) from oxidative stress. The sub-acute ageing of C57BL/6 mice was induced by subcutaneous injection of D-gal (120 mg kg-1 day-1) for 42 day. On the 14th day of D-gal injection, the mice were treated with ginsenoside Rg1 (20 mg kg-1 day-1, intraperitoneally) or normal saline for 28 days. The study monitored the effects of Rg1 on proliferation, senescence-associated and oxidative stress biomarkers, and Akt/mTOR signalling pathway in NSCs. Compared with the D-gal group, Rg1 improved cognitive impairment induced by D-galactose in mice by attenuating senescence of neural stem cells. Rg1 also decreased the level of oxidative stress, with increased the activity of superoxide dismutase and glutathione peroxidase in vivo and in vitro. Rg1 furthermore reduced the phosphorylation levels of protein kinase B (Akt) and the mechanistic target of rapamycin (mTOR) and down-regulated the levels of downstream p53, p16, p21 and Rb in D-gal treated NSCs. The results suggested that the protective effect of ginsenoside Rg1 on attenuating cognitive impairment in mice and senescence of NSCs induced by D-gal might be related to the reduction of oxidative stress and the down-regulation of Akt/mTOR signaling pathway.

Inhibitory effect and molecular mechanism of mesenchymal stem cells on NSCLC cells.

Non-small-cell lung cancer (NSCLC) is still the main threat of cancer-associated death. Current treatment of NSCLC has limited effectiveness, and unfortunately, the prognosis of NSCLC remains poor. Therefore, a novel strategy for cancer therapy is urgently needed. Stem cell therapy has significant potential for cancer treatment. Mesenchymal stem cells (MSCs) with capacity for self-renewal and differentiation into various cells types exhibit the feature of homing to tumor site and immunosuppression, have been explored as a new treatment for various cancers. Studies revealed that the broad repertoire of trophic factors secreted by MSCs extensively involved in the interplay between MSCs and tumor cells. In this study, we confirmed that MSCs do have the paracrine effect on proliferation and migration of NSCLC cells (A549, NCI-H460, and SK-MES-1). Co-culture system and conditioned medium experiments results showed that soluble factors secreted by MSCs inhibited the proliferation of NSCLC cells in vitro. The scratch assay showed that conditioned medium of MSCs could suppress the migration of NSCLC cells in vitro. Western blot results showed that the expression of proteins relevant to cell proliferation, anti-apoptosis, and migration was remarkably decreased via MAPK/eIF4E signaling pathway. We speculated that soluble factors secreted by MSCs might be responsible for inhibitory mechanism of NSCLC cells. By Human Gene Expression Microarray Assay and recombinant Vascular Endothelial Growth Factor 165 (VEGF165) neutralizing experiment, we verified that VEGF might be responsible for the down-regulation of proteins related to cell proliferation, anti-apoptosis, and migration by suppressing translation initiation factor eIF4E via MAPK signaling pathway. Taken together, our study demonstrated that a possible trophic factor secreted by MSCs could manipulate translation initiation of NSCLC cells via MAPK signaling pathway, and significantly affect the fate of tumor cells, which will be a new strategy for cancer therapy.

Single-chain antibody-delivered Livin siRNA inhibits human malignant melanoma growth in vitro and in vivo.

Although gene therapy has brought new insights into the treatment of malignant melanoma, targeting delivery of nucleic acid which targets critical oncogene/anti-oncogene in vivo is still a bottleneck in the therapeutic application. Our previous in vitro studies have found that the oncogene Livin could serve as a potential molecular target by small interfering RNA for gene therapy of malignant melanoma. However, how to transport Livin small interfering RNA into malignant melanoma cells specifically and efficiently in vivo needs further investigation. Cumulative evidence has suggested that single-chain antibody-mediated small interfering RNA targeted delivery is an effective way to silence specific genes in human cancer cells. Indeed, this study designed a protamine-single-chain antibody fusion protein, anti-MM scFv-tP, to deliver Livin small interfering RNA into LiBr cells. Further experiments confirmed the induction of cell apoptosis and suppression of cell proliferation by anti-MM scFv-tP in LiBr cells, along with efficient silence of Livin gene both in vitro and in vivo. Altogether, our findings provide a feasible approach to transport Livin small interfering RNA to malignant melanoma cells which would be a new therapeutic strategy for combating malignant melanoma.

Perovskite-type La0.8Sr0.2Co0.8Fe0.2O3 with uniform dispersion on N-doped reduced graphene oxide as an efficient bi-functional Li-O2 battery cathode.

A composite cathode including N-rGO with homogeneously dispersed perovskite La0.8Sr0.2Co0.8Fe0.2O3 on the surface is studied. Li-O2 batteries with LSCF@N-rGO cathode show better performance than those with LSCF-SP or N-rGO cathode. EIS and morphology analysis indicate that LSCF is beneficial to remold the shape of Li2O2 and catalyze the decomposition of Li2O2.

Interleukin-24 Concentrations Not Related with Age, but Affect Pro- and Anti-Inflammatory Cytokines Differently in Healthy Donors.

BACKGROUND: Interleukin 24 (IL-24) is expressed at different levels in a variety of tumor tissues and matched normal tissues and is regarded as a potential tumor biomarker as its expression levels in tumor tissues are associated with tumor patient prognosis. At present, the expression level of IL-24 in healthy human peripheral blood is unknown. METHODS: In this study, 1940 blood samples were collected using different processing methods from healthy donors. ELISA was used to detect IL-24 concentrations. RESULTS: The results showed that processing methods had the greatest influence on test results, with the highest IL24 concentration in EDTA plasma and the lowest in sodium citrate plasma. Lengths of storage time at 4°C had no obvious effect on IL-24 test results, and IL-24 in peripheral blood was stable for 15 days. IL-24 concentration in the sera of healthy donors showed no associations with age, blood glucose, hemoglobin, total cholesterol, carcinoembryonic antigen, absolute lymphocyte counts, alpha fetoprotein, white blood cells, thyroid stimulating hormone, or cereal third transaminase. We also confirmed that IL-24 expression level in the blood of healthy subjects was positively correlated with pro-inflammatory cytokines, tumor necrosis factor-alpha (TNF-α), and interleukin 6 (IL-6), but negatively correlated with anti-inflammatory cytokine, IL-10. CONCLUSIONS: We observed that sample processing methods influence the detection of IL-24 levels as EDTA plasma had the highest IL-24 concentration, and citric acid sodium, the lowest. Age, gender, and physical and chemical indicators were not related to IL-24 concentrations. We confirmed the IL-24 concentration was positively related to IL-6 and TNF-α and negatively to IL-10.

Inhibitory effect of genetically engineered mesenchymal stem cells with Apoptin on hepatoma cells in vitro and in vivo.

Hepatocellular carcinoma (HCC) is an aggressive tumor and has become one of the most frequent causes of cancer death in the world. The rate of post-operative recurrence and metastasis are still high even though after surgical resection. It is a difficult problem with extraordinary importance for the clinical treatment. So stem cell therapy becomes one of the anti-tumor biotherapy methods which is exploring. Due to the feature of homing to tumor site and immunosuppressive, mesenchymal stem cells (MSCs) have the capacity of gene treatment to tumor as a vehicle. Apoptin derived from chicken anemia virus is one kind of protein with an inherent ability to lyse cancer cells while leaving normal cells unharmed. Adenovirus (Ad) vectors can be modified to deliver therapeutic genes with the advantages of low toxicity and high transfer capacity. Now it has not been reported that combining MSCs and Adenovirus with Apoptin are used in HCC treatment. This study intends to construct recombinant adenovirus which expresses Apoptin and then infects human bone marrow MSCs, and explore the migration of MSCs to the hepatoma cells and inhibitory effect of genetically engineered mesenchymal stem cells with Apoptin on hepatoma cells in vitro and in vivo. Our research successfully established the recombinant Ad which was constructed by Ad system, and obtained MSCs which could secrete Apoptin. We found that both the modified MSCs with Apoptin and their conditional medium significantly inhibited the proliferation of liver cancer cells HepG2, which provided a novel means and experimental basis for stem cell treatment for HCC. This study tries to search for a stem cell therapy for cancers, which will provide a new approach and experimental basis for the clinical treatment of cancer. At the same time, this research will also provide experimental basis for a novel in vivo drug delivery system through stem cells as vehicle, which will resolve immune rejection induced by repeated applications of drug directly delivered by Ad vectors and reduce the high cost of a large-scale production and purification of exogenous drugs.

Inhibitory effect and mechanism of mesenchymal stem cells on liver cancer cells.

Mesenchymal stem cells (MSCs), with their capacity for self-renewal and differentiation into various cell types, are important seed cells for stem cell therapy. MSCs exhibit potent pathotropic migratory properties that make them attractive for use in tumor prevention and therapy. However, little is known about the underlying molecular mechanisms that link MSCs to the targeted tumor cells. This study investigated the inhibitory effect and mechanism of MSCs on human hepatoma HepG2 cells using co-culture and conditioned medium system and animal transplantation model. The HepG2 cells were co-cultured with MSCs or treated with conditional media derived from MSCs cultures in vitro. Results of methylthiazolyldiphenyl tetrazolium assay and flow cytometric assay showed that the proliferation and apoptosis of HepG2 cells decreased and increased, respectively. Reverse transcription polymerase chain reaction analysis showed that the expression levels of bcl-2, c-Myc, ß-catenin, and survivin were downregulated. The results of enzyme-linked immunosorbent assay and Western blot proved that MSCs secreted Dkk-1 to inhibit the expression of Wnt signaling pathway-related factors (bcl-2, c-Myc, ß-catenin, and survivin) in tumor cells, consequently inhibiting the proliferation and promoting the apoptosis of HepG2 cells. Animal transplantation experiment showed that tumor growth was significantly inhibited when HepG2 cells were co-injected with MSCs into nude mice. These results suggested that MSCs inhibited the growth and promoted the apoptosis of HepG2 cells in a dose-dependent manner. This study provided a new approach and experimental basis for cancer therapy. This study also proved that the Wnt signaling pathway may have a function in MSC-mediated tumor cell inhibition.

The Effect of Low HBV-DNA Viral Load on Recurrence in Hepatocellular Carcinoma Patients Who Underwent Primary Locoregional Treatment and the Development of a Nomogram Prediction Model.

(1) Background: HBV-DNA is an essential clinical indicator of primary hepatocellular carcinoma (HCC) prognosis. Our study aimed to investigate the prognostic implication of a low load of HBV-DNA in HCC patients who underwent local treatment. Additionally, we developed and validated a nomogram to predict the recurrence of patients with low (20-100 IU/mL) viral loads (L-VL). (2) Methods: A total of 475 HBV-HCC patients were enrolled, including 403 L-VL patients and 72 patients with very low (<20 IU/mL) viral loads (VL-VL). L-VL HCC patients were randomly divided into a training set (N = 282) and a validation set (N = 121) at a ratio of 7:3. Utilizing the Lasso-Cox regression analysis, we identified independent risk factors for constructing a nomogram. (3) Results: L-VL patients had significantly shorter RFS than VL-VL patients (38.2 m vs. 23.4 m, p = 0.024). The content of the nomogram included gender, BCLC stage, Glob, and MLR. The C-index (0.682 vs. 0.609); 1-, 3-, and 5-year AUCs (0.729, 0.784, and 0.783, vs. 0.631, 0.634, the 0.665); calibration curves; and decision curve analysis (DCA) curves of the training and validation cohorts proved the excellent predictive performance of the nomogram. There was a statistically significant difference in RFS between the low-, immediate-, and high-risk groups both in the training and validation cohorts (p < 0.001); (4) Conclusions: Patients with L-VL had a worse prognosis. The nomogram developed and validated in this study has the advantage of predicting patients with L-VL.

The water-soluble subfraction from Artemisia argyi alleviates LPS-induced inflammatory responses via multiple pathways and targets in vitro and in vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: As a traditional Chinese medicine, Artemisia argyi has been used medicinally and eaten for more than 2000 years in China. It is widely reported in treating inflammatory diseases such as eczema, dermatitis, arthritis, allergic asthma and colitis. Although several studies claim that its volatile oil and organic reagent extracts have certain anti-inflammatory effects, the water-soluble fractions and molecular mechanisms have not been studied. AIM OF THE STUDY: To evaluate the therapeutic effect of A. argyi water extract (AAWE) on lipopolysaccharide (LPS)-induced inflammatory responses and to identify the most effective water-soluble subfractions. Moreover, the relevant pharmacological and molecular mechanisms by which the active subfraction mitigates inflammation were further investigated. MATERIALS AND METHODS: Firstly, RAW 264.7 cells stimulated with LPS were treated with AAWE (50, 100, and 200 µg/mL) or the water-soluble subfractions separated by D101 macroporous resin (AAWE1-AAWE4, 100 µg/mL), and NO production and mRNA levels of inflammatory genes were evaluated to determine the most effective water-soluble subfractions. Secondly, the chemical components of the active subfraction (AAWE4) were analyzed by UPLC-QTOF-MS. Thirdly, transcriptome and network pharmacology analysis, RT-qPCR and Western blotting assays were conducted to explore the underlying anti-inflammatory mechanism and active compounds of AAWE4. Subsequently, the binding ability of the potential active components in AAWE4 to the core targets was further determined by molecular docking. Eventually, the in vivo anti-inflammatory activity of AAWE4 (1.17, 2.34 and 4.68 g/kg, administered per day for 7 d) was evaluated in mice with LPS-induced systemic inflammation. RESULTS: In this study, AAWE showed excellent anti-inflammatory effects, and its water-soluble subfraction AAWE4 exhibited the strongest inhibitory effect on NO concentration and inflammatory gene mRNA expression after LPS stimulation, indicating that it was the most effective subfraction. Thereafter, four main compounds in AAWE4 were confirmed or tentatively identified by UPLC-QTOF-MS, including three flavonoid glycosides and one phenolic acid. Furthermore, the transcriptome and network pharmacology analysis showed that AAWE4 inhibited inflammation via multiple pathways and multiple targets. Based on the RT-qPCR and Western blotting results, AAWE4 downregulated not only the p38, PI3K, CCL5, MMP9, AP-1, and BCL3 mRNA expression levels activated by LPS but also their upstream and downstream protein expression levels and protein phosphorylation (p-AKT/AKT, p-p38/p38, p-ERK/ERK, p-JNK/JNK). Moreover, four identified compounds (isochlorogenic acid A, vicenin-2, schaftoside and isoschaftoside) could significantly inhibit NO content and the overexpression of inflammatory factors TNF-α, IL-1ß, iNOS and COX-2 mRNA induced by LPS, and the molecular docking confirmed the high binding activity of four active compounds with selected core targets (p38, AKT1, MMP9, and CCL5). In addition, the mRNA expression and immunohistochemical analysis showed that AAWE44 could inhibit lung inflammation via multiple pathways and multiple targets in vivo. CONCLUSIONS: The findings of this study suggest that the water-soluble subfraction AAWE4 from A. argyi ameliorated the inflammation caused by LPS through multiple pathways and multiple targets in vitro and in vivo, providing scientific support for the medicinal use of A. argyi. Importantly, it shows that the A. argyi subfraction AAWE4 can be developed as an anti-inflammatory drug.