Multi-Omics Analysis Reveals the Role of Changes in Platelet Activation Pathways in the Survival Outcome of Patients with Esophageal Squamous Cell Carcinoma.

Objective: The objective of this study was to integrate metabolomics and transcriptomics data to identify key diagnostic and prognostic markers for esophageal squamous cell carcinoma (ESCC). Plasma samples were collected from 85 ESCC patients at different stages and 50 healthy volunteers for non-targeted metabolomic analysis. Methods: Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was employed for non-targeted metabolomic analysis. Subsequently, we integrated the metabolomic data with transcriptomic data from the Gene Expression Omnibus (GEO) and prognosis data from The Cancer Genome Atlas Program (TCGA) to perform pathway analysis. Our focus was on pathways that involve both metabolites and upstream genes, as they often exhibit higher accuracy. Results: Through the integration of metabolomics and transcriptomics, we identified significant alterations in the platelet activation pathway in ESCC. This pathway involves the participation of both metabolites and genes, making it a more accurate reflection of pathological changes associated with the disease. Notably, metabolite arachidonic acid (AA) and chemokine receptor type 2(CXCR2) were significantly downregulated in ESCC, while genes collagen type I alpha 1(COL1A1), collagen type I alpha 2(COL1A2), collagen type III alpha 1(COL3A1), type 3 inositol 1,4,5-trisphosphate receptor (ITPR3), and insulin-like growth factor II mRNA binding protein 3(IGF2BP3) were significantly upregulated, indicating the presence of tumor-induced platelet activation in ESCC. Further analysis of prognosis data revealed that high expression of COL1A1, IGF2BP3, and ITPR3 was associated with a favorable prognosis for ESCC, while high CXCR2 expression was linked to an adverse prognosis. In addition, we combined COL1A1, ITPR3, IGF2BP3, CXCR2, and AA to form a diagnostic biomarker panel. The receiver operating characteristic curve (ROC) demonstrated excellent diagnostic capability (AUC=0.987). Conclusion: Our study underscores the significant role of platelet activation pathways and related genes in the diagnosis and prognosis of ESCC patients. These findings offer promising insights for improving the clinical management of ESCC.

Wnt10b regulates osteogenesis of adipose-derived stem cells through Wnt/ß-catenin signalling pathway in osteoporosis.

Our previous finding revealed that the Wnt10b RNA expression of osteoporotic adipose-derived stem cells (OP-ASCs) with impaired osteogenic capacity was significantly reduced than that of ASCs. There are no ideas that the relationship between the OP-ASCs' impaired osteogenic potential and Wnt10b expression. This study aimed to indicate the potential molecular mechanisms and functional role of Wnt10b in OP-ASCs, as well as to investigate a potential application to reverse the OP-ASCs' impaired osteogenic differentiation potential. The OP-ASCs and ASCs were harvested from the inguinal fat of osteoporosis (OP) mice with bilateral ovariectomy (OVX) and normal mice. qPCR and WB were used to detect the different levels of the expression of the Wnt10b RNA in both OP-ASCs and ASCs. Lentiviral-mediated regulation of Wnt10b expression was employed for OP-ASCs, and the detection of the expression levels of key molecules in the Wnt signalling pathway and key osteogenic factors was performed through qPCR and WB in vitro experiments. The capacity of OP-ASCs to osteogenesis was determined using alizarin red staining. Lastly, the repair effect of the BCP scaffolds incorporating modified OP-ASCs on the critical-sized calvarial defects (CSCDs) in OP mice was scanned and detected by micro-computed tomography, haematoxylin and eosin staining, Masson's trichrome staining and immunohistochemistry. First, we discovered that both the RNA and protein expression levels of Wnt10b were significantly lower in OP-ASCs than that in ASCs. In vitro experiments, upregulation of Wnt10b could activate the Wnt signalling pathway, and increase expression of ß-catenin, Lef1, Runx2 and osteopontin (Opn), thereby enhancing the osteogenic ability of OP-ASCs. In addition, the OP-ASCs with Wnt10b-overexpressing could promote the repair of CSCD in osteoporotic mice with increasing new bone volume, bone mineral density, and increased expression of Opn in new bone in vivo. Taken together, overexpression of Wnt10b could partially facilitate the differentiation of OP-ASCs towards osteogenesis and accelerated the healing of bone defects by activating the Wnt/ß-catenin signalling pathway in vitro and in vivo experiments. This study confirmed the important role of Wnt10b in regulating the osteogenic differentiation capability of OP-ASCs and indicated Wnt10b could be a potential therapeutic target for reversing the impaired osteogenic capabilities of OP-ASCs to therapy bone defects of OP patients.

A Biomimetic Se-nHA/PC Composite Microsphere with Synergistic Immunomodulatory and Osteogenic Ability to Activate Bone Regeneration in Periodontitis.

Accumulation of reactive oxygen species (ROS) in periodontitis exacerbates the destruction of alveolar bone. Therefore, scavenging ROS to reshape the periodontal microenvironment, alleviate the inflammatory response and promote endogenous stem cell osteogenic differentiation may be an effective strategy for treating bone resorption in periodontitis. In this study, sericin-hydroxyapatite nanoparticles (Se-nHA NPs) are synthesized using a biomimetic mineralization method. Se-nHA NPs and proanthocyanidins (PC) are then encapsulated in sericin/sodium alginate (Se/SA) using an electrostatic injection technique to prepare Se-nHA/PC microspheres. Microspheres are effective in scavenging ROS, inhibiting the polarization of macrophages toward the M1 type, and inducing the polarization of macrophages toward the M2 type. In normal or macrophage-conditioned media, the Se-nHA/PC microspheres effectively promoted the osteogenic differentiation of human periodontal ligament stem cells (hPDLSCs). Furthermore, the Se-nHA/PC microspheres demonstrated anti-inflammatory effects in a periodontitis rat model by scavenging ROS and suppressing pro-inflammatory cytokines. The Se-nHA/PC microspheres are also distinguished by their capacity to decrease alveolar bone loss, reduce osteoclast activity, and boost osteogenic factor expression. Therefore, the biomimetic Se-nHA/PC composite microspheres have efficient ROS-scavenging, anti-inflammatory, and osteogenic abilities and can be used as a multifunctional filling material for inflammatory periodontal tissue regeneration.

Hydrogel Drug Delivery Systems for Bone Regeneration.

With the in-depth understanding of bone regeneration mechanisms and the development of bone tissue engineering, a variety of scaffold carrier materials with desirable physicochemical properties and biological functions have recently emerged in the field of bone regeneration. Hydrogels are being increasingly used in the field of bone regeneration and tissue engineering because of their biocompatibility, unique swelling properties, and relative ease of fabrication. Hydrogel drug delivery systems comprise cells, cytokines, an extracellular matrix, and small molecule nucleotides, which have different properties depending on their chemical or physical cross-linking. Additionally, hydrogels can be designed for different types of drug delivery for specific applications. In this paper, we summarize recent research in the field of bone regeneration using hydrogels as delivery carriers, detail the application of hydrogels in bone defect diseases and their mechanisms, and discuss future research directions of hydrogel drug delivery systems in bone tissue engineering.

Advanced glycation end products inhibit the osteogenic differentiation potential of adipose-derived stem cells in mice through autophagy.

BACKGROUND: Diabetes mellitus (DM) microenvironment will accelerate the accumulation of Advanced glycation end products (AGEs), adipose-derived stem cells (ASCs) have poor osteogenesis in the DM microenvironment. Studies suggest autophagy plays a vital role in osteogenesis, but the mechanism of the altered osteogenic potential of ASCs has not been elucidated. Bone tissue engineering by ASCs is widely used in the treatment of bone defects with diabetic osteoporosis (DOP). Therefore, it is meaningful to explore the effect of AGEs on the osteogenic differentiation potential of ASCs and its potential mechanism for the repair of bone defects in DOP. MATERIALS AND METHODS: ASCs in C57BL/6 mice were isolated, cultured, then treated with AGEs, subsequently, cell viability and proliferation were detected through Cell Counting Kit 8 assay. 3-Methyladenine (3-MA), an autophagic inhibitor used to inhibit autophagic levels. Rapamycin (Rapa), an autophagy activator that further activated autophagy levels by inhibiting mTOR.The osteogenesis and autophagy changes of ASCs were analyzed by flow cytometry, qPCR, western blot, immunofluorescence, alkaline phosphatase (ALP) and alizarin red staining. RESULTS: AGEs reduced the autophagy level and osteogenic potential of ASCs. After 3-MA reduced autophagy, the osteogenic potential of ASCs also decreased. AGEs co-treatment with 3-MA, the levels of osteogenesis and autophagy reduced more significantly. When autophagy was activated by Rapa, it was found that it could rescue the reduced osteogenic potential of AGEs. CONCLUSIONS: AGEs reduce the osteogenic differentiation potential of ASCs through autophagy, and may provide a reference for the treatment of bone defects with diabetes osteoporosis.

The Protective Effect of Ursolic Acid on Unilateral Ureteral Obstruction in Rats by Activating the Nrf2/HO-1 Antioxidant Signaling Pathway.

Renal interstitial fibrosis is a common pathological feature of a variety of kidney diseases that progress to end-stage renal disease. The excessive deposition of extracellular matrix (ECM) is a typical pathological change of renal interstitial fibrosis. The production of reactive oxygen species in renal tubules is an important factor leading to the development of renal interstitial fibrosis. Ursolic acid (UA) is a natural pentacyclic triterpene carboxylic acid compound widely found in plants. It has anti-inflammatory, antioxidant, and antitumor cell proliferation effects. It can reduce the development of fibrosis by inhibiting the oxidative stress response of the liver; there is currently no relevant research on whether UA can protect the renal interstitial fibrosis by resisting oxidative stress in the kidneys. In this study, our purpose is to investigate the effect of ursolic acid on renal interstitial fibrosis after unilateral ureteral obstruction (UUO) in rats and its related mechanisms. We established a UUO model by surgically ligating the right ureter of the rat and instilling UA preparation (40 mg/kg/d) through the stomach after the operation, once a day for 7 days. We found that UUO caused impaired renal function, increased pathological damage, increased renal interstitial fibrosis, increased apoptosis, increased oxidative stress damage, and decreased antioxidants. However, after UA preparations were given, the abovementioned damage was significantly improved. At the same time, we also found that UA preparations can significantly increase the relative expression of Nrf2/HO-1 signaling pathway in kidney tissue after UUO. In order to further verify whether the Nrf2/HO-1 signaling pathway is involved in the development of renal interstitial fibrosis, we injected zinc protoporphyrin (ZnPP, 45 umol/kg), a specific blocker of the Nrf2/HO-1 signaling pathway, into the intraperitoneal cavity after UUO in rats and before the gastric perfusion of ursolic acid preparations. Subsequently, we observed that the protective effect of UA on renal interstitial fibrosis after UUO in rats was reversed. Combining all the research results, we proved that UA has a protective effect on renal interstitial fibrosis after UUO in rats, which may be achieved by activating the Nrf2/HO-1 signaling pathway.

Melatonin delays dark-induced leaf senescence by inducing miR171b expression in tomato.

Melatonin functions in multiple aspects of plant growth, development, and stress response. Nonetheless, the mechanism of melatonin in plant carbon metabolism remains largely unknown. In this study, we investigated the influence of melatonin on the degradation of starch in tomato leaves. Results showed that exogenous melatonin attenuated carbon starvation-induced chlorophyll degradation and leaf senescence. In addition, melatonin delayed leaf starch degradation and inhibited the transcription of starch-degrading enzymes after sunset. Interestingly, melatonin-alleviated symptoms of leaf senescence and starch degradation were compromised when the first key gene for starch degradation, α-glucan water dikinase (GWD), was overexpressed. Furthermore, exogenous melatonin significantly upregulated the transcript levels of several microRNAs, including miR171b. Crucially, the GWD gene was identified as a target of miR171b, and the overexpression of miR171b ameliorated the carbon starvation-induced degradation of chlorophyll and starch, and inhibited the expression of the GWD gene. Taken together, these results demonstrate that melatonin promotes plant tolerance against carbon starvation by upregulating the expression of miR171b, which can directly inhibit GWD expression in tomato leaves.

DA-6 improves sunflower seed vigor under Al3+ stress by regulating Al3+ balance and ethylene metabolic.

Aluminum (Al3+) stress restricts plant seed germination and seedling growth seriously. Here, the sunflower "S175″ variety was used to explore the technique of improving seed vigor under Al3+ stress and investigate the effect of diethyl aminoethyl hexanoate (DA-6) on physiological characteristics in sunflower seeds during germination under Al3+ stress. The results showed that 3.0 mmol·L-1 Al3+ treatment significantly suppressed the sunflower seed germination and seedling growth. Al3+ stress significantly increased Al3+ content and secretion rates of citric and malic acids in sunflower seeds during germination. Besides, endogenous ethylene content was increased in Al3+-treated seeds. DA-6 serves as a positive signal to regulate the sunflower seed germination under Al3+ stress. Moreover, DA-6 enhanced the activities of malic dehydrogenase, citrate synthase, and isocitrate dehydrogenase, up-regulated the expressions of organic acid transport-related genes (ALMT and MATE), resulting in reduced accumulation of Al3+. Furthermore, exogenous DA-6 mitigated excessive accumulation of ethylene by decreasing the 1-aminocyclopropane-1-dihydrodipicolinate synthase activity and related-gene expression. However, DA-6 treatment had no effect on abscisic acid or gibberellin metabolism in sunflower seeds under Al3+ stress. These results confirmed that DA-6 application enhanced the germination capacity through induction of the synthesis and transport of malic and citric acids, and suppression of the excessive accumulation of endogenous ethylene, thus contributing to alleviate Al3+ toxicity in sunflower seeds.

CSN improves seed vigor of aged sunflower seeds by regulating the fatty acid, glycometabolism, and abscisic acid metabolism.

Introduction: Sunflower seeds possess higher oil content than do cereal crop seeds. Storage of sunflower seeds is accompanied by loss of seed vigor and oxidation of storage and membrane lipids. Objectives: This study first reported that compound sodium nitrophenolate (CSN), a new plant growth modulator, improved the germination and seedling emergence of aged sunflower seeds. The present study provide a future reference as to the potential applications of CSN and the regulation mechanism of exogenous substances in increasing aged crop seed vigor. Methods: Phenotypic analysis was performed to investigate the effect of CSN on germination and seedling emergence from naturally- and artificially-aged sunflower seeds. The biochemical and enzyme activity analysis were conducted to test the CSN-induced effect on glycometabolism, fatty acid and abscisic acid metabolism. Meanwhile, gene expression analysis was carried out to detect the changes in the transcription level of sunflower seeds during early germination period after CSN treatment. Results: CSN application significantly increased the germination rate and seedling emergence rate of sunflower seeds under natural and artificial aging. Biochemical analysis indicated that, CSN treatment significantly enhanced the sucrose and fructose contents in aged sunflower seeds during early germination period. Moreover, the contents of several different fatty acids in CSN-treated sunflower seeds also significantly increased. Enzyme activity analysis revealed that CSN treatment remarkably up-regulated the activities of several critical enzymes related to triacylglycerol hydrolysis. Consequently, the transcription levels of the above key enzymes-related synthetic genes were also significantly up-regulated in CSN treatment. Furthermore, CSN treatment significantly decreased abscisic acid (ABA) content through the regulation of the gene expressions and activities of metabolism related-enzymes. Conclusion: Taken together, the contribution of CSN to the improvement of aged sunflower seed germination and seedling emergence might be closely related to the fatty acid, glycometabolism, and ABA metabolism.

LY3214996 relieves acquired resistance to sorafenib in hepatocellular carcinoma cells.

Background: Sorafenib, an oral multi-kinase inhibitor of rapidly accelerated fibrosarcoma; vascular endothelial growth factor receptor-2/3, platelet-derived growth factor receptor, c-Kit, and Flt-3 signaling, is approved for treatment of advanced hepatocellular carcinoma (HCC). However, the benefit of sorafenib is often diminished because of acquired resistance through the reactivation of ERK signaling in sorafenib-resistant HCC cells. In this work, we investigated whether adding LY3214996, a selective ERK1/2 inhibitor, to sorafenib would increase the anti-tumor effectiveness of sorafenib to HCC cells. Methods: The Huh7 cell line was used as a cell model for treatment with sorafenib, LY3214996, and their combination. Phosphorylation of the key kinases in the Ras/Raf/MAPK and PI3K/Akt pathways, protein expression of the cell cycle, and apoptosis migration were assessed with western blot. MTT and colony-formation assays were used to evaluate cell proliferation. Wound-healing assay was used to assess cell migration. Cell cycle and apoptosis analyses were conducted with flow cytometry. Results: LY3214996 decreased phosphorylation of the Ras/Raf/MAPK and PI3K/Akt pathways, including p-c-Raf, p-P90RSK, p-S6K and p-eIF4EBP1 activated by sorafenib, despite increased p-ERK1/2 levels. LY3214996 increased the anti-proliferation, anti-migration, cell-cycle progression, and pro-apoptotic effects of sorafenib on Huh7R cells. Conclusions: Reactivation of ERK1/2 appears to be a molecular mechanism of acquired resistance of HCC to sorafenib. LY3214996 combined with sorafenib enhanced the anti-tumor effects of sorafenib in HCC. These findings form a theoretical basis for trial of LY3214996 combined with sorafenib as second-line treatment of sorafenib-resistant in advanced HCC.

Normobaric hyperoxia plays a protective role against renal ischemia-reperfusion injury by activating the Nrf2/HO-1 signaling pathway.

Following renal ischemia-reperfusion injury (RIRI), because of the decrease in oxygen supply to the kidney, a large amount of oxygen-free radicals is generated, and in severe cases, tissue cells will undergo apoptosis or even die. Normobaric hyperoxia (NBHO) is a very common clinical adjuvant treatment. It restores the oxygen supply after renal ischemia and combats oxidative stress in tissues, thus playing a protective role. In this study, our aim is to elucidate the protective mechanism of NBHO inhalation in a rat RIRI model. We performed a surgical excision of the left kidney of the rat and established a right kidney solitary kidney model. Later, the right renal pedicle of the rat was clamped using a non-invasive vascular clamp for 45 min. After the vascular clamp was released and reperfused for 24 h, the rat was placed in a closed oxygen chamber. It was subjected to inhalation of high-concentration oxygen (50%-55%), 2 h daily, for 7 days.RIRI induces postoperative weight loss, impaired renal function, increased oxygen free radicals, reduced antioxidant substances, increased histopathological damage, and increased levels of apoptosis. These effects were significantly improved after treatment with NBHO. At the same time, NBHO significantly increased the expression levels of Nrf2 and HO-1 in the tissues after RIRI. To verify whether HO-1 induced by Nrf2 is involved in the resistance to oxidative stress, after the rat RIRI and before inhaling NBHO, we intraperitoneally injected HO-1 specific inhibitor zinc protoporphyrin (ZnPP) (45 µmol/Kg). However, we found that ZnPP reversed the protective effect of NBHO on RIRI in rats. Combining all the results, we have demonstrated the protective effect of NBHO on RIRI, which can be at least partially attributed to the activation of the Nrf2/HO-1 antioxidative stress pathway.

The PI3K/mTOR dual inhibitor BEZ235 nanoparticles improve radiosensitization of hepatoma cells through apoptosis and regulation DNA repair pathway.

Polymer materials encapsulating drugs have broad prospects for drug delivery. We evaluated the effectiveness of polyethylene glycol-poly (lactic-co-glycolic acid) (PLGA-PEG) encapsulation and release characteristics of PI3K/mTOR inhibitor NVP-BEZ235 (BEZ235). We proposed a strategy for targeting radiosensitization of liver cancer cells. The biocompatibility, cell interaction, and internalization of Glypican-3 (GPC3) antibody-modified, BEZ235-loaded PLGA-PEG nanoparticles (NP-BEZ235-Ab) in hepatoma cells in vitro were studied. Also, the cell killing effect of NP-BEZ235-Ab combined with γ-ray cell was evaluated. We used confocal microscopy to monitor nanoparticle-cell interactions and cellular uptake, conducted focus-formation experiments to analyze the synergistic biological effects of NP-BEZ235-Ab and priming, and studied synergy in liver cancer cells using molecular biological methods such as western blotting. We found that PLGA-PEG has good loading efficiency for BEZ235 and high selectivity to GPC3-positive HepG2 liver cancer cells, thus documenting that NP-BEZ235-Ab acts as a small-molecule drug delivery nanocarrier. At the nominal concentration, the NP-BEZ235-Ab nanoformulation synergistically kills liver cancer cells with significantly higher efficiency than does the free drug. Thus, NP-BEZ235-Ab is a potential radiosensitizer.

BEZ235 increases sorafenib inhibition of hepatocellular carcinoma cells by suppressing the PI3K/AKT/mTOR pathway.

BACKGROUND: Sorafenib is an oral multi-kinase inhibitor that inhibits hepatocellular carcinoma (HCC) via the Ras/Raf/MAPK pathway. However, sorafenib loses effectiveness because most tumors acquire drug resistance over time. As the PI3K/AKT/mTOR signaling pathway is also activated abnormally in HCC, we evaluated the effect of sorafenib, in combination with a dual PI3K/mTOR inhibitor, BEZ235, on HCC cell proliferation and survival in vitro. MATERIALS AND METHODS: Biological phenotypes were analysed in HCC cell lines, parental and sorafenib-resistant HepG2 cells (HepG2 and HepG2R), treated with sorafenib or BEZ235, alone or in combination. HCC cellular proliferation and apoptosis were investigated, and perturbations of the Ras/Raf/MAPK and PI3K/AKT/mTOR signaling/survival pathways were evaluated by western blot analysis. RESULTS: BEZ235 enhanced sorafenib inhibition of cellular proliferation, migration, and promotion of apoptosis in HepG2 and HepG2R cells. The combined effects were associated with inhibition of phosphorylation of AKT, mTOR and S6K in the PI3K/AKT/mTOR pathway, whereas the combination of sorafenib and BEZ235 did not significantly alter the Ras/Raf/MAPK pathway compared with the effect of sorafenib alone. CONCLUSION: Sorafenib/BEZ235 combination has potent anti-HCC cell activity. This anti-tumor activity is most likely multi-factorial, mainly involving PI3K down-regulation and AKT, mTOR and S6K dephosphorylation. Combined inhibition of PI3K/AKT/mTOR and Ras/Raf/MAPK pathways enhances sorafenib inhibition of HCC. The results of these in vitro studies suggest that trials of combined sorafenib and BEZ235 in the treatment of HCC should be considered.

BEZ235 enhances chemosensitivity of paclitaxel in hepatocellular carcinoma through inhibiting the PI3K/Akt/mTOR pathway.

Desensitization of hepatocellular carcinoma (HCC) to paclitaxel chemotherapy is a major deterrent to successful treatment of the cancer. Abnormal activation of the PI3K/Akt/mTOR, pathway is a common outcome of chemotherapy for HCC. Therefore, we investigated whether BEZ235, a dual PI3K and mTOR inhibitor, could increase the sensitivity of HCC to paclitaxel. In vitro results showed that paclitaxel, combined with BEZ235, inhibited HCC cell proliferation and migration, arrested the cell cycle in the G2/M phase, and promoted cell apoptosis by decreasing PI3K/Akt/mTOR activity. In vivo experiments confirmed that BEZ235 enhances the anti-tumor effect of paclitaxel by reducing PI3K/Akt/mTOR activity. Immunohistochemical staining showed that paclitaxel combined with BEZ235 reduced the numbers of Ki-67- and GPC3-positive HepG2 cells in tumor tissues. We conclude that BEZ235 enhanced the sensitivity of HCC to paclitaxel, and inhibition of PI3K/Akt/mTOR signaling might be a therapeutic strategy against paclitaxel-resistant HCC.

Effects of Porphyromonas gingivalis infection with different fimA genotypes on the secretion of IL-1β, IL-6, and TNF-α by human umbilical vein endothelial cells / 口腔疾病防治

Objective@#To study the effects of Porphyromonas gingivalis (P. g) infection with I, II and IV fimA genotypes on the expression of IL-1beta, IL-6 and TNF-alpha in human umbilical vein endothelial cells (HUVECs). @*Methods @#HUVECs infected with different fimA genotypes were divided into the fimA type I stimulation group, fimA type II stimulation group, fimA type IV stimulation group. In addition, a positive control group (E. coli LPS stimulation) and negative control group (cell culture medium only) were included. Cell proliferation was detected by MTT assay, and cell apoptosis was detected by flow cytometry. IL-1β, IL-6, TNF-α levels in the supernatant of HUVECs after P. g stimulation were assessed by ELISA at 2 h, 6 h and 24 h.@*Results @#HUVECs were infected by P. g with fimA type I,fimA type II and LPS for 24 h. Cell proliferation was inhibited compared with the negative control group (P < 0.05), but there was no significant difference in the apoptosis rate between P. g infection and the negative control group. IL-1β levels in cell culture supernatants were higher at 2th than 6 and 24 h after stimulation of HUVECs with different fimA genotypes, while the IL-6 levels were higher at 24 h than the other time, while the TNF-α levels were no significant difference at every time. After fimA type II and IV P. g infection, IL-1β, IL-6, and TNF-α levels were increased compared with fimA type I P. g (P < 0.05).@*Conclusion @# Different P. g fimA genotypes have different effects on stimulating HUVECs to induce dysfunction. Here, fimA type II and IV P. g exhibit a strong ability to upregulate the secretion of IL-1β, IL-6 and TNF-alpha.

Enhanced delivery of sorafenib with anti-GPC3 antibody-conjugated TPGS-b-PCL/Pluronic P123 polymeric nanoparticles for targeted therapy of hepatocellular carcinoma.

Although sorafenib (SFB) showed improved efficacy and much reduced the side effects in clinical liver cancer therapy, its therapeutic efficacy was still greatly limited due to short half-life in vivo as well as drug resistance. To solve these problems, we developed a novel SFB-loaded polymeric nanoparticle for targeted therapy of liver cancer. This polymeric nanoparticle, referred to NP-SFB-Ab, was fabricated from self-assembly of biodegradable block copolymers TPGS-b-poly(caprolactone) (TPGS-b-PCL) and Pluronic P123 and drug SFB, followed by conjugating the anti-GPC3 antibody. NP-SFB-Ab showed robust stability and achieve excellent SFB release in cell medium. The CLSM demonstrated that the Ab-conjugated NP exhibited much higher cellular uptake in HepG2 human liver cells than non-targeted NP. The MTT assay also confirmed that NP-SFB-Ab caused much greater cytotoxicity than non-targeted NP-SFB and free SFB. Finally, NP-SFB-Ab was proved to greatly inhibit the tumor growth of HepG2 xenograft-bearing nude mice without obvious side effects. Therefore, this NP-SFB-Ab provides a promising new approach for targeted therapy of hepatocellular carcinoma.

HsfA1a upregulates melatonin biosynthesis to confer cadmium tolerance in tomato plants.

Melatonin regulates broad aspects of plant responses to various biotic and abiotic stresses, but the upstream regulation of melatonin biosynthesis by these stresses remains largely unknown. Herein, we demonstrate that transcription factor heat-shock factor A1a (HsfA1a) conferred cadmium (Cd) tolerance to tomato plants, in part through its positive role in inducing melatonin biosynthesis under Cd stress. Analysis of leaf phenotype, chlorophyll content, and photosynthetic efficiency revealed that silencing of the HsfA1a gene decreased Cd tolerance, whereas its overexpression enhanced plant tolerance to Cd. HsfA1a-silenced plants exhibited reduced melatonin levels, and HsfA1a overexpression stimulated melatonin accumulation and the expression of the melatonin biosynthetic gene caffeic acid O-methyltransferase 1 (COMT1) under Cd stress. Both an in vitro electrophoretic mobility shift assay and in vivo chromatin immunoprecipitation coupled with qPCR analysis revealed that HsfA1a binds to the COMT1 gene promoter. Meanwhile, Cd stress induced the expression of heat-shock proteins (HSPs), which was compromised in HsfA1a-silenced plants and more robustly induced in HsfA1a-overexpressing plants under Cd stress. COMT1 silencing reduced HsfA1a-induced Cd tolerance and melatonin accumulation in HsfA1a-overexpressing plants. Additionally, the HsfA1a-induced expression of HSPs was partially compromised in COMT1-silenced wild-type or HsfA1a-overexpressing plants under Cd stress. These results demonstrate that HsfA1a confers Cd tolerance by activating transcription of the COMT1 gene and inducing accumulation of melatonin that partially upregulates expression of HSPs.

Melatonin enhances thermotolerance by promoting cellular protein protection in tomato plants.

Melatonin is a pleiotropic signaling molecule that provides physiological protection against diverse environmental stresses in plants. Nonetheless, the mechanisms for melatonin-mediated thermotolerance remain largely unknown. Here, we report that endogenous melatonin levels increased with a rise in ambient temperature and that peaked at 40°C. Foliar pretreatment with an optimal dose of melatonin (10 µmol/L) or the overexpression of N-acetylserotonin methyltransferase (ASMT) gene effectively ameliorated heat-induced photoinhibition and electrolyte leakage in tomato plants. Both exogenous melatonin treatment and endogenous melatonin manipulation by overexpression of ASMT decreased the levels of insoluble and ubiquitinated proteins, but enhanced the expression of heat-shock proteins (HSPs) to refold denatured and unfolded proteins under heat stress. Meanwhile, melatonin also induced expression of several ATG genes and formation of autophagosomes to degrade aggregated proteins under the same stress. Proteomic profile analyses revealed that protein aggregates for a large number of biological processes accumulated in wild-type plants. However, exogenous melatonin treatment or overexpression of ASMT reduced the accumulation of aggregated proteins. Aggregation responsive proteins such as HSP70 and Rubisco activase were preferentially accumulated and ubiquitinated in wild-type plants under heat stress, while melatonin mitigated heat stress-induced accumulation and ubiquitination of aggregated proteins. These results suggest that melatonin promotes cellular protein protection through induction of HSPs and autophagy to refold or degrade denatured proteins under heat stress in tomato plants.

Genistein-loaded nanoparticles of star-shaped diblock copolymer mannitol-core PLGA-TPGS for the treatment of liver cancer.

The purpose of this research is to develop nanoparticles (NPs) of star-shaped copolymer mannitol-functionalized PLGA-TPGS for Genistein delivery for liver cancer treatment, and evaluate their therapeutic effects in liver cancer cell line and hepatoma-tumor-bearing nude mice in comparison with the linear PLGA nanoparticles and PLGA-TPGS nanoparticles. The Genistein-loaded M-PLGA-TPGS nanoparticles (MPTN), prepared by a modified nanoprecipitation method, were observed by FESEM and TEM to be near-spherical shape with narrow size distribution. The nanoparticles were further characterized in terms of their size, size distribution, surface charge, drug-loading content, encapsulation efficiency and in vitro drug release profiles. The data showed that the M-PLGA-TPGS nanoparticles were found to be stable, showing almost no change in particle size and surface charge during 3-month storage of their aqueous solution. In vitro Genistein release from the nanoparticles exhibited biphasic pattern with burst release at the initial 4days and sustained release afterwards. The cellular uptake efficiency of fluorescent M-PLGA-TPGS nanoparticles was 1.25-, 1.22-, and 1.29-fold higher than that of the PLGA-TPGS nanoparticles at the nanoparticle concentrations of 100, 250, and 500µg/mL, respectively. In the MPTN group, the ratio of apoptotic cells increased with the drug dose increased, which exhibited dose-dependent effect and a significant difference compared with Genistein solution group (p<0.05). The data also showed that the Genistein-loaded M-PLGA-TPGS nanoparticles have higher antitumor efficacy than that of linear PLGA-TPGS nanoparticles and PLGA nanoparticles in vitro and in vivo. In conclusion, the star-shaped copolymer M-PLGA-TPGS could be used as a potential and promising bioactive material for nanomedicine development for liver cancer treatment.