Low-Temperature Signaling Pathways and Their Signaling Factors in Plant

Low temperature as abiotic stress adversely impacts plant growth and development, and limits the ecological distribution of plants as well. Throughout their long evolutionary history, plants have developed a range of complicated and precise molecular regulatory mechanisms to deal with low-temperature stress, involving the activation of signal transduction pathways and the regulation of related genes. In this review, we provide a systematic summary of the most recent research findings regarding three hypotheses of cellular perception of low-temperature signals and two major intracellular low-temperature signaling pathways, including CBF-dependent signaling pathways and CBF-independent signaling pathways. Focus is placed on the functions of each component of the ICE-CBF-COR signaling cascade as well as their interrelationships. This review concludes that although some progress has been made in the identification, function, and mechanism of low-temperature response genes, their roles in the low-temperature regulatory network and molecular mechanisms still need to be studied in detail, which will be of great significance for improving the low-temperature tolerance of plants and adapting to climate change.

Application of Transgenic Technology in Identification for Gene Function on Grasses

Perennial grasses have developed intricate mechanisms to adapt to diverse environments,enabling their resistance to various biotic and abiotic stressors.These mechanisms arise from strong natural selection that contributes to enhancing the adaptation of forage plants to various stress conditions.Methods such as antisense RNA technology,CRISPR/Cas9 screening,virus-induced gene silencing,and transgenic technology,are commonly utilized for investigating the stress response functionalities of grass genes in both warm-season and cool-season varieties.This review focuses on the functional identification of stress-resistance genes and regulatory elements in grasses.It synthesizes recent studies on mining functional genes,regulatory genes,and protein kinase-like signaling factors involved in stress responses in grasses.Additionally,the review outlines future research directions,providing theoretical support and references for further exploration of(i)molecular mechanisms underlying grass stress responses,(ii)cultivation and domestication of herbage,(iii)development of high-yield varieties resistant to stress,and(iv)mechanisms and breeding strategies for stress resistance in grasses.

Blocking VEGF signaling augments interleukin-8 secretion via MEK/ERK/1/2 axis in human retinal pigment epithelial cells

AIM:To identify proangiogenic factors engaged in neovascular age-related macular degeneration(AMD)except vascular endothelial growth factor(VEGF)from human retinal pigment epithelial(h RPE)cells and investigate the underlying mechanisms.METHODS:VEGF receptor 2(VEGFR2)in ARPE-19 cells was depleted by si RNA transfection or overexpressed through adenovirus infection.The m RNA and the protein levels of interleukin-8(IL-8)in ARPE-19 cells were measured by quantitative real-time polymerase chain reaction and enzyme-linked immunosorbent assay respectively.The protein levels of AKT,p-AKT,MEK,p-MEK,ERK1/2,p-ERK1/2,JNK,p-JNK,p38 and p-p38 were detected by Western blotting.A selective chemical inhibitor,LY3214996,was employed to inhibit phosphorylation of ERK1/2.Cell viability was determined by MTT assay.RESULTS:Knockdown of VEGFR2 in ARPE-19 cells robustly augmented IL-8 production at both the m RNA and the protein levels.Silencing VEGFR2 substantially enhanced phosphorylation of MEK and ERK1/2 while exerted no effects on phosphorylation of AKT,JNK and p38.Inhibiting ERK1/2 phosphorylation by LY3214996 reversed changes in VEGFR2 knockdown-induced IL-8 upregulation at the m RNA and the protein levels with no effects on cell viability.VEGFR2 overexpression significantly reduced IL-8 generation at the m RNA and the protein levels.CONCLUSION:Blockade of VEGF signaling augments IL-8 secretion via MEK/ERK1/2 axis and overactivation of VEGF pathway decreases IL-8 production in h RPE cells.Upregulated IL-8 expression after VEGF signaling inhibition in h RPE cells may be responsible for being incompletely responsive to anti-VEGF remedy in neovascular AMD,and IL-8 may serve as an alternative therapeutic target for neovascular AMD.

Inhibition of epidermal growth factor receptor signaling protects human malignant glioma cells from hypoxia - induced cell death

Epidermal growth factor receptor(EGFR)signaling has become an importanttarget for drug development becauseEGFR signaling enhances tumor cell proliferation,migration,and invasion and inhibits apoptosis.However,theresults of clinical trials using EGFR inhibitors in patients with solid tumors have been disappointing.Here,wereport a protective effect of the EGFR inhibitors AG1478 and PD153035 against cell death induced by acute hy-poxia,which contrasts with their proapoptotic effects under normoxia.Under hypoxic conditions,both agents re-

The therapeutic mechanism of Polygonatum sibiricum polysaccharide on T2DM rats based on the Nrf2 signaling pathway

Background:This study will be aimed at investigating the effects of Polygonatum sibiricum polysaccharide(PSP)in a rat model of type 2 diabetes mellitus(T2DM).Methods:Thirty Sprague–Dawley male rats were evenly distributed into three groups:normal,T2DM,and PSP groups,by applying a random sample table method.The typical cluster was served balanced food every day,whereas the others were supplied with a high-fat diet and streptozotocin injections to make T2DM rat models.The changes in humor organic chemistry indicators and liver histopathology were determined following the model institution for every cluster.After PSP intervention,western blotting was applied to research the expression levels of crucial transcription factors concerned within the nuclear factor erythroid 2-related factor 2(Nrf2)signal pathway,as well as Nrf2,glutamate-cysteine ligase chemical process fractional monetary unit,NQO1 and HO-1 within the liver tissues of the rat models,and to seem into the therapeutic edges of PSP and the way it happens in T2DM rats.Results:PSP intervention considerably reduced the concentration levels of aldohexose,lipids,liver-operated indicators,and alternative organic chemistry indicators within the humor of T2DM rat models and improved the histopathological changes within the liver.In addition,the activity of SOD and GSH-Px were increased,and the levels of MDA were decreased in the liver tissues of T2DM rat models following PSP intervention.Western blotting revealed that the expression levels of the Nrf2,HO-1,glutamate-cysteine ligase chemical,and NQO1 proteins were increased in rat liver tissues following PSP intervention.Conclusion:PSP has therapeutic effects in T2DM rat models,and its mechanism of motion might also be related to regulating the Nrf2 signaling pathway in liver tissues and alleviating oxidative stress.

Regenerating gene 4 promotes chemoresistance of colorectal cancer by affecting lipid droplet synthesis and assembly

BACKGROUND Regenerating gene 4(REG4)has been proved to be carcinogenic in some cancers,but its manifestation and possible carcinogenic mechanisms in colorectal cancer(CRC)have not yet been elucidated.Our previous study found that the drug resistance of CRC cells may be closely linked to their fat metabolism.AIM To explore the role of REG4 in CRC and its association with lipid droplet formation and chemoresistance.METHODS We conducted a meta-analysis and bioinformatics and pathological analyses of REG4 expression in CRC.The effects of REG4 on the phenotypes and related protein expression were also investigated in CRC cells.We detected the impacts of REG4 on the chemoresistance and lipid droplet formation in CRC cells.Finally,we analyzed how REG4 regulated the transcription and proteasomal degradation of lipogenic enzymes in CRC cells.RESULTS Compared to normal mucosa,REG4 mRNA expression was high in CRC(P<0.05)but protein expression was low.An inverse correlation existed between lymph node and distant metastases,tumor-node-metastasis staging or short overall survival and REG4 mRNA overexpression(P<0.05),but vice versa for REG4 protein expression.REG4-related genes included:Chemokine activity;taste receptors;protein-DNA and DNA packing complexes;nucleosomes and chromatin;generation of second messenger molecules;programmed cell death signals;epigenetic regulation and DNA methylation;transcription repression and activation by DNA binding;insulin signaling pathway;sugar metabolism and transfer;and neurotransmitter receptors(P<0.05).REG4 exposure or overexpression promoted proliferation,antiapoptosis,migration,and invasion of DLD-1 cells in an autocrine or paracrine manner by activating the epidermal growth factor receptor-phosphoinositide 3-kinase-Akt-nuclear factor-κB pathway.REG4 was involved in chemoresistance not through de novo lipogenesis,but lipid droplet assembly.REG4 inhibited the transcription of acetyl-CoA carboxylase 1(ACC1)and ATP-citrate lyase(ACLY)by disassociating the complex formation of anti-acetyl(AC)-acetyl-histone 3-AC-histone 4-inhibitor of growth protein-5-si histone deacetylase;-sterol-regulatory element binding protein 1 in their promoters and induced proteasomal degradation of ACC1 or ACLY.CONCLUSION REG4 may be involved in chemoresistance through lipid droplet assembly.REG4 reduces expression of de novo lipid synthesis key enzymes by inhibiting transcription and promoting ubiquitination-mediated proteasomal degradation.

Dihydroergotamine ameliorates liver fibrosis by targeting transforming growth factor β type Ⅱ receptor

BACKGROUND The transforming growth factor β(TGFβ) signaling pathway plays a crucial role in the development of liver fibrosis by activating TGFβ type Ⅱ receptor(TGFβR2), followed by the recruitment of TGFβR1 finally triggering downstream signaling pathway.AIM To find drugs targeting TGFβR2 that inhibit TGFβR1/TGFβR2 complex formation, theoretically inhibit TGFβ signaling pathway, and thereby ameliorate liver fibrosis.METHODS Food and Drug Administration-approved drugs were screened for binding affinity with TGFβR2 by virtual molecular docking. We identified 6 candidates and further explored their potential by Cell Counting Kit-8(CCK-8) cell cytotoxic experiment to validate toxicity and titrated the best cellular working concentrations. Next, we further demonstrated the detailed molecular working mechanisms using mutagenesis analysis. Finally, we used a mouse model to investigate its potential anti-liver fibrosis effect.RESULTS We identified 6 drug candidates. Among these 6 drugs, dihydroergotamine(DHE) shows great ability in reducing fibrotic gene expressions such as collagen, p-SMAD3, and α-SMA in TGFβ induced cellular model of liver fibrosis in LX-2 cells. Furthermore, we demonstrated that DHE binds to TGFβR2. Moreover, mutation of Leu27, Phe30, Thr51, Ser52, Ile53, and Glu55 of TGFβR2 disrupted the binding of TGFβR2 with DHE. In addition, DHE significantly improved liver fibrosis, as evidenced by Masson’s trichrome staining of liver sections. This is further supported by the width and the velocity of the portal vein, and serum markers of liver function. In line with those observations, DHE also decreased macrophages infiltration and extracellular matrix deposition in the liver.CONCLUSION DHE alleviates liver fibrosis by binding to TGFβR2 thereby suppressing TGFβ signaling pathway. We show here that as far as drug repurposing, DHE has great potential to treat liver fibrosis.

Multiomics reveal human umbilical cord mesenchymal stem cells improving acute lung injury via the lung-gut axis

BACKGROUND Acute lung injury(ALI)and its final severe stage,acute respiratory distress syndrome,are associated with high morbidity and mortality rates in patients due to the lack of effective specific treatments.Gut microbiota homeostasis,including that in ALI,is important for human health.Evidence suggests that the gut microbiota improves lung injury through the lung-gut axis.Human umbilical cord mesenchymal cells(HUC-MSCs)have attractive prospects for ALI treatment.This study hypothesized that HUC-MSCs improve ALI via the lung-gut microflora.AIM To explore the effects of HUC-MSCs on lipopolysaccharide(LPS)-induced ALI in mice and the involvement of the lung-gut axis in this process.METHODS C57BL/6 mice were randomly divided into four groups(18 rats per group):Sham,sham+HUC-MSCs,LPS,and LPS+HUC-MSCs.ALI was induced in mice by intraperitoneal injections of LPS(10 mg/kg).After 6 h,mice were intervened with 0.5 mL phosphate buffered saline(PBS)containing 1×10^(6) HUC-MSCs by intraperitoneal injections.For the negative control,100 mL 0.9%NaCl and 0.5 mL PBS were used.Bronchoalveolar lavage fluid(BALF)was obtained from anesthetized mice,and their blood,lungs,ileum,and feces were obtained by an aseptic technique following CO_(2) euthanasia.Wright’s staining,enzyme-linked immunosorbent assay,hematoxylin-eosin staining,Evans blue dye leakage assay,immunohistochemistry,fluorescence in situ hybridization,western blot,16S rDNA sequencing,and non-targeted metabolomics were used to observe the effect of HUC-MSCs on ALI mice,and the involvement of the lung-gut axis in this process was explored.One-way analysis of variance with post-hoc Tukey’s test,independent-sample Student’s t-test,Wilcoxon rank-sum test,and Pearson correlation analysis were used for statistical analyses.RESULTS HUC-MSCs were observed to improve pulmonary edema and lung and ileal injury,and decrease mononuclear cell and neutrophil counts,protein concentrations in BALF and inflammatory cytokine levels in the serum,lung,and ileum of ALI mice.Especially,HUC-MSCs decreased Evans blue concentration and Toll-like receptor 4,myeloid differentiation factor 88,p-nuclear factor kappa-B(NF-κB)/NF-κB,and p-inhibitorαof NF-κB(p-IκBα)/IκBαexpression levels in the lung,and raised the pulmonary vascular endothelial-cadherin,zonula occludens-1(ZO-1),and occludin levels and ileal ZO-1,claudin-1,and occludin expression levels.HUC-MSCs improved gut and BALF microbial homeostases.The number of pathogenic bacteria decreased in the BALF of ALI mice treated with HUCMSCs.Concurrently,the abundances of Oscillospira and Coprococcus in the feces of HUS-MSC-treated ALI mice were significantly increased.In addition,Lactobacillus,Bacteroides,and unidentified_Rikenellaceae genera appeared in both feces and BALF.Moreover,this study performed metabolomic analysis on the lung tissue and identified five upregulated metabolites and 11 downregulated metabolites in the LPS+MSC group compared to the LPS group,which were related to the purine metabolism and the taste transduction signaling pathways.Therefore,an intrinsic link between lung metabolite levels and BALF flora homeostasis was established.CONCLUSION This study suggests that HUM-MSCs attenuate ALI by redefining the gut and lung microbiota.

Insights into the regulation of C-repeat binding factors in plant cold signaling

Cold temperatures, a major abiotic stress, threaten the growth and development of plants, worldwide. To cope with this adverse environmental cue, plants from temperate climates have evolved an array of sophisticated mechanisms to acclimate to cold periods, increasing their ability to tolerate freezing stress. Over the last decade, significant progress has been made in determining the molecular mechanisms underpinning cold acclimation, including following the identification of several pivotal components, including candidates for cold sensors, protein kinases, and transcription factors. With these developments, we have a better understanding of the CBF-dependent cold-signaling pathway. In this review, we summarize recent progress made in elucidating the cold-signaling pathways, especially the C-repeat binding factor-dependent pathway, and describe the regulatory function of the crucial components of plant cold signaling. We also discuss the unsolved questions that should be the focus of future work.

Effects of Haobie Yangyin Ruanjian Decoction on hepatic fibrosis induced by carbon tetrachloride in rats

AIM:To explore the anti-fibrotic effect of Haobie Yangyin Ruanjian Decoction(HYRD)on CCl4-induced hepatic fibrosis in rats and its modulation on the transforming growth factor(TGF)β-Smad signaling pathway.METHODS:Fifty-six healthy Wistar rats were randomly divided into five groups:normal control group(n=6),CCl4-induced hepatic fibrosis group(n=14) and three treatment groups(the treated rats received HYRD via oral administration at daily dosages of 8.2,2.5 and 0.82 g/kg,respectively)of HYRD(n=12,respectively).Experimental hepatic fibrosis was induced by subcutaneous injection of carbon tetrachloride solution(CCl4 dissolved in peanut oil,4:6,V/V)with 0.5 mL/100 g body weight for the first time,and then 0.3 mL/100 g body weight twice a week for 8 wk.In the former 2 wk,rats were raised by feedstuffⅠ(80% corn meal,20%lard,0.5%cholesterol).After 2 wk,they were raised by feedstuffⅡ(corn meal and 0.5% cholesterol).Except for the control group,30%alcohol solution was given orally to each rat every other day from the beginning,1 mL for each rat.Liver function parameters and hepatic hydroxyproline content were detected by chromatometry.Serum levels of hyaluronic acid(HA),typeⅣcollagen(CIV),typeⅢprecollagen(PCⅢ)and laminin(LN)were assayed with radioimmunoassay.Deposition of collagen was observed with hematoxylin-eosin staining and collagen staining.Gene expression of TGFβ1 and Smad3 were detected with real-time reverse transcriptase-polymerase chain reaction and Western blotting,respectively.RESULTS:The serum levels of alanine transaminase and aspartate transaminase were increased in the model group compared with the control group(P<0.01),and they were decreased in the three treatment groups compared with the model group.The serum levels of total protein and albumin were decreased in the model group and increased in the three treatment groups.The hepatic hydroxyproline content and serum levels of PCⅢ,HA,LN and CIV were markedly increased in the model group compared with the control group,and decreased in the treatment groups.The gene expression of TGFβ1 and Smad3 was enhanced in the model group compared with the control group,and HYRD could down regulate their expression.CONCLUSION:HYRD can inhibit hepatic fibrosis induced by CCl4 in rats,which is probably associated with its down-regulation on fibrogenic signal transduction of TGFβ-Smad pathway.

Silencing of signal transducer and activator of transcription 3 expression by RNA interference suppresses growth of human hepatocellular carcinoma in tumor-bearing nude mice

AIM： To explore the effect of silencing of signal transducer and activator of transcription 3 （STAT3） expression by RNA interference （RNAi） on growth of human hepatocellular carcinoma （HCC） in tumorbearing nude mice in vivo.METHODS： To construct the recombinant plasmid of pSilencer 3.0-H1-STAT3-siRNA-GFP （pSHI-siRNA- STAT3） and establish the tumor-bearing nude mouse model of the HCC cell line SMMC7721, we used intratumoral injection together with electroblotting to transfect the recombinant plasmid pSHI-siRNA- STAT3 into the transplanted tumor. The weight of the nude mice and tumor volumes were recorded. STAT3 gene transcription was detected by semi-quantitative reverse transcription polymerase chain reaction （RT- PCR）. Level of protein expression and location of STAT3 were determined by Western blotting and immunohistochemical staining. STAT3-related genes such as survivin, c-myc, VEGF, p53 and caspase3 mRNA and protein expression were detected in tumor tissues at the same time. The terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling （TUNEL） assay was used to detect apoptosis of tumor cells.RESULTS： The weight of the treated nude mice increased, and the tumor volume decreased markedly compared with those of the mock-treated and negative control groups （P 〈 0.01）. The results of RT-PCR and Western blotting showed that mRNA and protein levels of STAT3 declined markedly in the treated group. The change in STAT3-related gene expression in tumor tissues at the mRNA and protein level also varied, the expression of survivin, VEGF and c-myc were obviously reduced, and expression of p53 and caspase3 increased （P 〈 0.01）. Most of the tumor tissue ceils in the treated group developed apoptosis that was detected by TUNEL assay.CONCLUSION： Silencing of STAT3 expression by RNAi significantly inhibits expression of STAT3 mRNA and protein, and suppresses growth of human HCC in tumor-bearing nude mice. The mechanism may be related to down-regulation of survivin, VEGF and c-myc and up-regulation of p53 and caspase3 expression. Accordingly, the STAT3 gene may act as an important and effective target in gene therapy of HCC.

Association of Down-regulation of CD109 Expression with Up-expression of Smad7 in Pathogenesis of Psoriasis

Transforming growth factor（TGF）-β signaling plays an important role in the pathogenesis of psoriasis. CD109, a novel TGF-β co-receptor, which inhibits TGF-β signaling by enhancing Smad7-dependent degradation of TGF-β type Ⅰ receptor（TGF-β RⅠ）, is abnormally expressed in psoriasis. To date, the expression of Smad7 and the correlation between CD109 and Smad7 expression in psoriasis have not been fully elucidated. This study was designed to investigate the expression and the correlation of CD109 and TGF-β signaling associated proteins in psoriasis and their roles in the pathogenesis of psoriasis. Thirty-two psoriasis specimens were subjected to immunohistochemical staining for CD109, Smad7, TGF-β RⅠ and Ki67. Ten normal skin（NS） specimens served as controls. The positive expression rate（% positive cells） of Smad7 and Ki67 in psoriasis was significantly higher than in NS（62.6%±19.9% vs. 17.2%±4.4%, and 50.7%±14.3% vs. 19.5%±3.2%, respectively, P〈0.001）, and the expression levels of CD109 and TGF-β RⅠ were reduced significantly in psoriasis as compared with NS（8.1%±6.7% vs. 35.8%±6.7% and 27.3%±3.4% vs. 3.0%±3.4%, respectively, P〈0.001）. There were significantly negative correlations between CD109 and Smad7（r=-0.831, P〈0.01）. These findings indicated that CD109 might play a certain role in the pathogenesis of psoriasis. Lower expression of CD109 and TGF-β RⅠ was highly correlated with higher expression of Smad7 and Ki67, suggesting that CD109 may induce the pathogenesis of psoriasis through Smad7-mediated degradation of TGF-β RⅠ, and lead to the termination of TGF-β signaling.

Axonal remodeling in the corticospinal tract after stroke： how does rehabilitative training modulate it？

Stroke causes long-term disability, and rehabilitative training is commonly used to improve the consecutive functional recovery. Following brain damage, surviving neurons undergo morphological alterations to reconstruct the remaining neural network. In the motor system, such neural network remodeling is observed as a motor map reorganization. Because of its significant correlation with functional recovery, motor map reorganization has been regarded as a key phenomenon for functional recovery after stroke. Although the mechanism underlying motor map reorganization remains unclear, increasing evidence has shown a critical role for axonal remodeling in the corticospinal tract. In this study, we review previous studies investigating axonal remodeling in the corticospinal tract after stroke and discuss which mechanisms may underlie the stimulatory effect of rehabilitative training. Axonal remodeling in the corticospinal tract can be classified into three types based on the location and the original targets of corticospinal neurons, and it seems that all the surviving corticospinal neurons in both ipsilesional and contralesional hemisphere can participate in axonal remodeling and motor map reorganization. Through axonal remodeling, corticospinal neurons alter their output selectivity from a single to multiple areas to compensate for the lost function. The remodeling of the corticospinal axon is influenced by the extent of tissue destruction and promoted by various therapeutic interventions, including rehabilitative training. Although the precise molecular mechanism underlying rehabilitation-promoted axonal remodeling remains elusive, previous data suggest that rehabilitative training promotes axonal remodeling by upregulating growth-promoting and downregulating growth-inhibiting signals.

Biology of tooth replacement in amniotes

Tooth replacement is a common trait to most vertebrates, including mammals. Mammals, however, have lost the capacity for continuous tooth renewal seen in most other vertebrates, and typically have only 1-2 generations of teeth. Here, we review the mechanisms of tooth replacement in reptiles and mammals, and discuss in detail the current and historical theories on control of timing and pattern of tooth replacement and development.

Baicalin Antagonizes Prostate Cancer Stemness via Inhibiting Notch1/NF-κB Signaling Pathway

Objective To investigate the molecular mechanisms underlying the effect of baicalin on prostate cancer(PCa)progression both in vivo and in vitro.Methods The in situ PCa stem cells(PCSCs)-injected xenograft tumor models were established in BALB/c nude mice.Tumor volume and weight were respectively checked after baicalin(100 mg/kg)treatment.Hematoxylin-eosin(HE)staining was used to observe the growth arrest and cell necrosis.mRNA expression levels of acetaldehyde dehydrogenase 1(ALDH1),CD44,CD133 and Notch1 were determined by reverse transcription-polymerase chain reaction.Protein expression levels of ALDH1,CD44,CD133,Notch1,nuclear factorκB(NF-κB)P65 and NF-κB p-P65 were detected by Western blot.Expression and subcellular location of ALDH1,CD44,CD133,Notch1 and NF-κB p65 were detected by immunofluorescence analysis.In vitro,cell cycle distribution and cell apoptosis of PC3 PCSCs was assessed by flow cytometry after baicalin(125µmol/L)treatment.The migration and invasion abilities of PCSCs were assessed using Transwell assays.Transmission electron microscopy scanning was utilized to observe the structure and autophagosome formation of baicalin-treated PCSCs.In addition,PCSCs were infected with lentiviruses expressing human Notch1.Results Compared with the control group,the tumor volume and weight were notably reduced in mice treated with 100 mg/kg baicalin(P<0.05 or P<0.01).Histopathological analysis showed that baicalin treatment significantly inhibited cell proliferation and promoted cell apoptosis.Furthermore,baicalin treatment reduced mRNA and protein expression levels of CD44,CD133,ALDH1,and Notch1 as well as the protein expression of NF-κB p-P65 in the xenograft tumor(P<0.01).In vitro,the cell proliferation of PCSCs was significantly attenuated after treatment with 125µmol/L baicalin for 72 h(P<0.01).The cell migration and invasion rates were decreased following treatment with baicalin for 48 and 72 h(P<0.01).Baicalin notably induced cell apoptosis and seriously damaged the structure of PCSCs.The mRNA and protein expressions of CD133,CD44,ALDH1 and Notch1 in PCSCs were significantly downregulated following baicalin treatment(P<0.01).Importantly,the inhibitory effects of baicalin on PCa progression and stemness were reversed by Notch1 overexpression(P<0.05 or P<0.01).Conclusion Mechanistically,baicalin exhibited a potential therapeutic effect on PCa via inhibiting the Notch1/NF-κB signaling pathway and its mediated cancer stemness.

Functional evaluation of the insulin/insulin-like growth factor signaling pathway in determination of wing polyphenism in pea aphid

Wing polyphenism is a common phenomenon that plays key roles in environmental adaptation of insects.Insulin/insulin-like growth factor signaling(IIS)pathway is a highly conserved pathway in regulation of metabolism,development,and growth in metazoans.It has been reported that IS is required for switching of wing morph in brown planthopper via regulating the development of the wing pad.However,it remains elusive whether and how IIS pathway regulates transgenerational wing dimorphism in aphid.In this study,we found that pairing and solitary treatments can induce pea aphids to produce high and low percentage winged offspring,respectively.The expression level of ILP5(insulin-like peptide 5)in maternal head was significantly higher upon solitary treatment in comparison with pairing,while silencing of ILP5 caused no obvious change in the winged offspring ratio.RNA interference-mediated knockdown of FoxO(Forkhead transcription factor subgroup O)in stage 20 embryos significantly increased the winged offspring ratio.The results of pharmacological and quantitative polymerase chain reaction experiments showed that the embryonic insulin receptors may not be involved in wing polyphenism.Additionally,ILP4 and ILP11 exhibited higher expression levels in 1st wingless offspring than in winged offspring.We demonstrate that FoxO negatively regulates the wing morph development in embryos.ILPs may regulate aphid wing polyphenism in a developmental stage-specific manner.However,the regulation may be not mediated by the canonical IIS pathway.The findings advance our understanding of IIS pathway in insect transgenerational wing polyphenism.

Huangqin Decoction Delays Progress of Colitis-Associated Carcinogenesis by Regulating Nrf2/HO-1 Antioxidant Signal Pathwayin Mice

Objective:To investigate the effect of Huangqin Decoction(HQD)on nuclear factor erythroid 2 related-factor 2(Nrf2)/heme oxygenase(HO-1)signaling pathway by inducing the colitis-associated carcinogenesis(CAC)model mice with azoxymethane(AOM)/dextran sodium sulfate(DSS).Methods:The chemical components of HQD were analyzed by liquid chromatography-quadrupole-time-of-flight mass spectrometry(LC-Q-TOF-MS/MS)to determine the molecular constituents of HQD.Totally 48 C57BL/6J mice were randomly divided into 6 groups by a random number table,including control,model(AOM/DSS),mesalazine(MS),low-,medium-,and high-dose HQD(HQD-L,HQD-M,and HQD-H)groups,8 mice in each group.Except for the control group,the mice in the other groups were intraperitoneally injected with AOM(10 mg/kg)and administrated with 2.5%DSS orally for 1 week every two weeks(totally 3 rounds of DSS)to construct a colitis-associated carcinogenesis mouse model.The mice in the HQD-L,HQD-M and HQD-H groups were given HQD by gavage at doses of 2.925,5.85,and 11.7 g/kg,respectively;the mice in the MS group was given a suspension of MS at a dose of 0.043 g/kg(totally 11 weeks).The serum levels of malondialdehyde(MDA)and superoxide dismutase(SOD)were measured by enzyme-linked immunosorbent assay.The mRNA and protein expression levels of Nrf2,HO-1,and inhibitory KELCH like ECH-related protein 1(Keap1)in colon tissue were detected by quantitative real-time PCR,immunohistochemistry,and Westem blot,respectively.Results:LC-Q-TOF-MS/MS analysis revealed that the chemical constituents of HQD include baicalin,paeoniflorin,and glycyrrhizic acid.Compared to the control group,significantly higher MDA levels and lower SOD levels were observed in the model group(P<0.05),whereas the expressions of Nrf2 and HO-1 were significantly decreased,and the expression of Keap1 increased(P<0.01).Compared with the model group,serum MDA level was decreased and SOD level was increased in the HQD-M,HQD-H and MS groups(P<0.05).Higher expressions of Nrf2 and HO-1 were observed in the HQD groups.Conclusion:HQD may regulate the expression of Nrf2 and HO-1 in colon tissue,reduce the expression of MDA and increase the expression of SOD in serum,thus delaying the progress of CAC in AOM/DSS mice.

Electroacupuncture Attenuates Immune-Inflammatory Response in Hippocampus of Rats with Vascular Dementia by Inhibiting TLR4/MyD88 Signaling Pathway

Objective:To investigate whether electroacupuncture(EA)alleviates cognitive impairment by suppressing the toll-like receptor 4(TLR4)/myeloid differentiation factor 88(MyD88)signaling pathway,which triggers immune-inflammatory responses in the hippocampus of rats with vascular dementia(VaD).Methods:The experiments were conducted in 3 parts and in total the Sprague-Dawley rats were randomly divided into 8 groups by a random number table,including sham,four-vessel occlusion(4-VO),4-VO+EA,4-VO+non-EA,sham+EA,4-VO+lipopolysaccharide(LPS),4-VO+LPS+EA,and 4-VO+TAK-242 groups.The VaD model was established by the 4-VO method.Seven days later,rats were treated with EA at 5 acupoints of Baihui(DV 20),Danzhong(RN 17),Geshu(BL 17),Qihai(RN 6)and Sanyinjiao(SP 6),once per day for 3 consecutive weeks.Lymphocyte subsets,lymphocyte transformation rates,and inflammatory cytokines interleukin-6(IL-6)and tumor necrosis factorα(TNF-α)were measured to assess immune function and inflammation in VaD rats.Transmission electron microscopy was used to observe the ultrastructure of nerve cells in the hippocampus.The levels of TLR4,MyD88,IL-6,and TNF-αwere detected after EA treatment.TLR4/MyD88 signaling and cognitive function were also assessed after intracerebroventricular injection of TLR4 antagonist TAK-242 or TLR4 agonist LPS with or without EA.Results:Compared with the 4-VO group,EA notably improved immune function of rats in the 4-VO+EA group,inhibited the protein and mRNA expressions of TLR4 and MyD88 in the hippocampus of rats,reduced the expressions of serum IL-6 and TNF-α(all P<0.05 or P<0.01),and led to neuronal repair in the hippocampus.There were no significant differences between the 4-VO+LPS+EA and 4-VO+EA groups,nor between the 4-VO+TAK-242 and 4-VO+EA groups(P>0.05).Conclusions:EA attenuated cognitive impairment associated with immune inflammation by inhibition of the TLR4/MyD88 signaling pathway.Thus,EA may be a promising alternative therapy for the treatment of VaD.

Analysis of epidermal growth factor signaling in nasal mucosa epithelial cell proliferation involved in chronic rhinosinusitis

Background Aberrant epithelial repair has been observed in chronic rhinosinusitis （CRS） patients; however, the mechanism of epithelial cell repair regulation is unclear. Epidermal growth factor （EGF） plays an important role in regulating epithelial cell repair in lower airway and may be a critical factor in the remodeling processes of CRS. The objective of our research is to evaluate the differences between CRS and normal subjects and between chronic rhinosinusitis without nasal poiys （CRSsNP） and chronic rhinosinusitis with nasal polys （CRSwNP） in the regulation of EGF pathways and the regulating proliferative position of classic Ras/Raf/MEK/ERK pathways. Methods We evaluated the proliferation rates of ethmoidal mucosal cells before and after stimulation with EGF, epidermal growth factor receptor （EGFR） kinase inhibitor AG1478, and extracellular signal-regulated kinase 1/2 （ERKI/2）inhibitor PD98059 using MTT assays. We also analyzed the sinonasal epithelial cells collected from control subjects and patients with CRS subtypes CRSsNP and CRSwNP for the expression of ERK1/2, phosphorylated ERK1/2, P21, P15, and P27 using western blotting analyses. Results The proliferation rates of sinonasal epithelial cells before and after EGF stimulation were lower in CRS patients than in the controls. AG1478 or PD98059 inhibitor treatment of control epithelial cells did not result in a significant difference in proliferation. Although, AG1478 and PD98059,inhibited the proliferation of CRS cells, the degree of proliferation inhibition was markedly different in CRSsNP. AG1478 suppressed the proliferation of CRSwNP epithelial cells, whereas PD98059 had no effect. The ratio of ERK1/2 phosphorylation in CRS cells was lower than that of the control cells. Cyclin-dependent kinase inhibitors were highly expressed in CRS cells compared with that of control cells. ERK1/2 and P27 showed differential expression in CRSsNP and CRSwNP. Conclusions Differences existed in EGF pathways in CRS patients and normal subjects as well as in CRSsNP and CRSwNP. Classical Ras/Raf/MEK/ERK pathway may assume absolute superiority in control cells. Ras/Raf/MEK/ERK classical pathway and other pathways might be active at the same time to stimulate epithelial cell proliferation in CRSsNP. The function of Ras/Raf/MEK/ERK classical pathway was weaker in CRSwNP than in CRSsNP and when the classical pathway was blocked in CRSwNP, some other pathway could have completely compensated the proliferation induced by the Ras/Raf/MEK/ERK pathway.

THESEUS1 positively modulates plant defense responses against Botrytis cinerea through GUANINE EXCHANGE FACTOR4 signaling

The plant cell wall is an important interface for sensing pathogen attack and activating signaling pathways that promote plant immune responses.THESEUS1(THE1) acts as a sensor of cell wall integrity that controls cell elongation during plant growth.However, no specific role for THE1 in plant defense responses has been reported. Here, we found that THE1 interacts with GUANINE EXCHANGE FACTOR4(GEF4)and that both proteins play regulatory roles in plant resistance to the necrotrophic fungus Botrytis cinerea.Genetic analysis showed that THE1 and GEF4 function in the same genetic pathway to mediate plant defense responses. In addition, using transcriptome analysis, we identified various genes(such as defense-related,secondary metabolite-related, and transcription factor genes) that are likely downstream targets in the THE1-GEF4 signaling pathway. Our results suggest that THE1 functions as an upstream regulator of GEF4 signaling to positively regulate defense responses against B. cinerea in Arabidopsis.