Objective To investigate the role and molecular mechanism of exosomal miR-224-5p in colorectal cancer(CRC).Methods The miR-224-5p expression in CRC patient tissues and cell-derived exosomes was measured by laser captu...Objective To investigate the role and molecular mechanism of exosomal miR-224-5p in colorectal cancer(CRC).Methods The miR-224-5p expression in CRC patient tissues and cell-derived exosomes was measured by laser capture microdissection and qRT-PCR,respectively.Dual-luciferase reporter gene assay was used to determine the target gene of miR-224-5p.The protein expressions of p53 and unc-51 like kinase 2(ULK2)in CRC cells were detected by western blot.Flow cytometry was used to detect cell cycle and apoptosis.Cell proliferation was measured by CCK8 and EdU assay.Results The miR-224-5p expression was upregulated in CRC tissues and increased progressively with the rise of CRC stage.CRC cells secreted extracellular miR-224-5p mainly in an exosome-dependent manner,and then miR-224-5p could be transferred to surrounding tumor cells to regulate cell proliferation in the form of autocrine or paracrine.Moreover,ULK2 was characterized as a direct target of miR-224-5p and was downregulated in CRC tissues.Interestingly,ULK2 inhibited CRC cell proliferation in a p53-dependent manner.Furthermore,exosome-derived miR-224-5p partially reversed the proliferation regulation of ULK2 on CRC cells.Conclusion Our findings demonstrate that exosome-transmitted miR-224-5p promotes p53-dependent cell proliferation by targeting ULK2 in CRC,which may offer promising targets for CRC prevention and therapy.展开更多
Prolife ration of neural stem cells is crucial for promoting neuronal regeneration and repairing cerebral infarction damage.Transcranial magnetic stimulation(TMS)has recently emerged as a tool for inducing endogenous ...Prolife ration of neural stem cells is crucial for promoting neuronal regeneration and repairing cerebral infarction damage.Transcranial magnetic stimulation(TMS)has recently emerged as a tool for inducing endogenous neural stem cell regeneration,but its underlying mechanisms remain unclea r In this study,we found that repetitive TMS effectively promotes the proliferation of oxygen-glucose deprived neural stem cells.Additionally,repetitive TMS reduced the volume of cerebral infa rction in a rat model of ischemic stro ke caused by middle cerebral artery occlusion,im p roved rat cognitive function,and promoted the proliferation of neural stem cells in the ischemic penumbra.RNA-sequencing found that repetitive TMS activated the Wnt signaling pathway in the ischemic penumbra of rats with cerebral ischemia.Furthermore,PCR analysis revealed that repetitive TMS promoted AKT phosphorylation,leading to an increase in mRNA levels of cell cycle-related proteins such as Cdk2 and Cdk4.This effect was also associated with activation of the glycogen synthase kinase 3β/β-catenin signaling pathway,which ultimately promotes the prolife ration of neural stem cells.Subsequently,we validated the effect of repetitive TMS on AKT phosphorylation.We found that repetitive TMS promoted Ca2+influx into neural stem cells by activating the P2 calcium channel/calmodulin pathway,thereby promoting AKT phosphorylation and activating the glycogen synthase kinase 3β/β-catenin pathway.These findings indicate that repetitive TMS can promote the proliferation of endogenous neural stem cells through a Ca2+influx-dependent phosphorylated AKT/glycogen synthase kinase 3β/β-catenin signaling pathway.This study has produced pioneering res ults on the intrinsic mechanism of repetitive TMS to promote neural function recove ry after ischemic stro ke.These results provide a stro ng scientific foundation for the clinical application of repetitive TMS.Moreover,repetitive TMS treatment may not only be an efficient and potential approach to support neurogenesis for further therapeutic applications,but also provide an effective platform for the expansion of neural stem cells.展开更多
BACKGROUND Gastrointestinal stromal tumor(GIST)is a common neoplasm with high rates of recurrence and metastasis,and its therapeutic efficacy is still not ideal.There is an unmet need to find new molecular therapeutic...BACKGROUND Gastrointestinal stromal tumor(GIST)is a common neoplasm with high rates of recurrence and metastasis,and its therapeutic efficacy is still not ideal.There is an unmet need to find new molecular therapeutic targets for GIST.TATA-boxbinding protein-associated factor 15(TAF15)contributes to the progress of various tumors,while the role and molecular mechanism of TAF15 in GIST progression are still unknown.AIM To explore new molecular therapeutic targets for GIST and understand the biological role and underlying mechanisms of TAF15 in GIST progression.METHODS Proteomic analysis was performed to explore the differentially expressed proteins in GIST.Western blotting and immunohistochemical analysis were used to verify the expression level of TAF15 in GIST tissues and cell lines.Cell counting kit-8,colony formation,wound-healing and transwell assay were executed to detect the ability of TAF15 on cell proliferation,migration and invasion.A xenograft mouse model was applied to explore the role of TAF15 in the progression of GIST.Western blotting was used to detect the phosphorylation level and total level of RAF1,MEK and ERK1/2.RESULTS A total of 1669 proteins were identified as differentially expressed proteins with 762 upregulated and 907 downregulated in GIST.TAF15 was selected for the further study because of its important role in cell proliferation and migration.TAF15 was significantly over expressed in GIST tissues and cell lines.Overexpression of TAF15 was associated with larger tumor size and higher risk stage of GIST.TAF15 knockdown significantly inhibited the cell proliferation and migration of GIST in vitro and suppressed tumor growth in vivo.Moreover,the inhibition of TAF15 expression significantly decreased the phosphorylation level of RAF1,MEK and ERK1/2 in GIST cells and xenograft tissues,while the total RAF1,MEK and ERK1/2 had no significant change.CONCLUSION TAF15 is over expressed in GIST tissues and cell lines.Overexpression of TAF15 was associated with a poor prognosis of GIST patients.TAF15 promotes cell proliferation and migration in GIST via the activation of the RAF1/MEK/ERK signaling pathway.Thus,TAF15 is expected to be a novel latent molecular biomarker or therapeutic target of GIST.展开更多
In mammals,microRNAs(miRNAs)play key roles in multiple biological processes by regulating the expression of target genes.Studies have found that the levels of miR-370-5p expression differ significantly in the skins of...In mammals,microRNAs(miRNAs)play key roles in multiple biological processes by regulating the expression of target genes.Studies have found that the levels of miR-370-5p expression differ significantly in the skins of sheep with different hair colors;however,its function remains unclear.In this study,we investigated the roles of miR-370-5p in sheep melanocytes and found that the overexpression of miR-370-5p significantly inhibited cell proliferation(P<0.01),tyrosinase activity(P=0.001)and significantly reduced(P<0.001)melanin production.Functional prediction revealed that the 3′-untranslated region(UTR)of MAP3K8 has a putative miR-370-5p binding site,and the interaction between these two molecules was confirmed using luciferase reporter assays.In situ hybridization assays revealed that MAP3K8 is expressed in the cytoplasm of melanocytes.The results of quantitative RT-PCR and Western blotting analyses revealed that overexpression of miR-370-5p in melanocytes significantly inhibits(P<0.01)MAP3K8 expression via direct targeting of its 3′UTR.Inhibition of MAP3K8 expression by siRNA-MAP3K8 transfection induced a significant inhibition(P<0.01)of melanocyte proliferation and significant reduction(P<0.001)in melanin production,which is consistent with our observations for miR-370-5p.Target gene rescue experiments indicated that the expression of MAP3K8 in melanocytes co-transfected with miR-370-5p and MAP3K8-cDNA(containing sites for the targeted binding to miR-370-5p)was significantly rescued(P≤0.001),which subsequently promoted significant increases in cell proliferation(P<0.001)and melanin production(P<0.01).Collectively,these findings indicate that miR-370-5p plays a functional role in inhibiting sheep melanocyte proliferation and melanogenesis by downregulating the expression of MAP3K8.展开更多
Follicle-stimulating hormone(FSH),an important hypothalamic-pituitary-gonadal axis(HPG)hormone,is secreted by the pituitary gland.This study confirms that FSH is expressed in chicken follicles at different stages,and ...Follicle-stimulating hormone(FSH),an important hypothalamic-pituitary-gonadal axis(HPG)hormone,is secreted by the pituitary gland.This study confirms that FSH is expressed in chicken follicles at different stages,and positive FSHβ mRNA signals were stronger(P<0.05)in granulosa cells than in oocytes.The 369 bp coding sequence of FSHβ in ovaries is 100%identical to that in the pituitary gland.The experiment in vitro revealed that the ovary possessed FSH secretory capacity.Further,FSHβ mRNA was significantly upregulated(P<0.05)in follicles and significantly higher(P<0.05)than that in the pituitary gland by approximately 2–23 times with the development.The number of granulosa cells decreased significantly(P<0.05)in the cells with siRNA treatment,confirming that the ovarian FSH could promote granulosa cell proliferation.This view was supported by cell cycle analysis and CCND2 and CCNE2 expression.Further research indicated that no difference(P>0.05)was observed between the number of granulosa cells treated with FSHβ siRNA and in exogenous FSH.However,the number of granulosa cells without FSHβ siRNA transfection was significantly higher(P<0.05)for exogenous FSH.This finding suggests that the proliferative effect of exogenous FSH on ovarian granulosa cells depend on endogenous FSH.This study demonstrated that the FSH gene was expressed in chicken follicles and promoted ovarian granulosa cell proliferation,which enriched the theory on HPG axis.展开更多
BACKGROUND Colorectal cancer(CRC)is a worldwide problem,which has been associated with changes in diet and lifestyle pattern.As a result of colonic fermentation of dietary fibres,short chain free fatty acids are gener...BACKGROUND Colorectal cancer(CRC)is a worldwide problem,which has been associated with changes in diet and lifestyle pattern.As a result of colonic fermentation of dietary fibres,short chain free fatty acids are generated which activate free fatty acid receptors(FFAR)2 and 3.FFAR2 and FFAR3 genes are abundantly expressed in colonic epithelium and play an important role in the metabolic homeostasis of colonic epithelial cells.Earlier studies point to the involvement of FFAR2 in colorectal carcinogenesis.AIM To understand the role of short chain FFARs in CRC.METHODS Transcriptome analysis console software was used to analyse microarray data from CRC patients and cell lines.We employed short-hairpin RNA mediated down regulation of FFAR2 and FFAR3 genes,which was validated using quantitative real time polymerase chain reaction.Assays for glucose uptake and cyclic adenosine monophosphate(cAMP)generation was done along with immunofluorescence studies to study the effects of FFAR2/FFAR3 knockdown.For measuring cell proliferation,we employed real time electrical impedancebased assay available from xCELLigence.RESULTS Microarray data analysis of CRC patient samples showed a significant down regulation of FFAR2 gene expression.This prompted us to study the FFAR2 in CRC.Since,FFAR3 shares significant structural and functional homology with FFAR2,we knocked down both these receptors in CRC cell line HCT 116.These modified cell lines exhibited higher proliferation rate and were found to have increased glucose uptake as well as increased level of glucose transporter 1.Since,FFAR2 and FFAR3 signal through G protein subunit(Gαi),knockdown of these receptors was associated with increased cAMP.Inhibition of protein kinase A(PKA)did not alter the growth and proliferation of these cells indicating a mechanism independent of cAMP/PKA pathway.CONCLUSION Our results suggest role of FFAR2/FFAR3 genes in increased proliferation of colon cancer cells via enhanced glucose uptake and exclude the role of PKA mediated cAMP signalling.Alternate pathways could be involved that would ultimately result in increased cell proliferation as a result of down regulated FFAR2/FFAR3 genes.This study paves the way to understand the mechanism of action of short chain FFARs in CRC.展开更多
The present study aimed to clarify the role of SENEX in malignant cell proliferation in diffuse large B-cell lymphoma(DLBCL).22 DLBCL patients(6 newly diagnosed cases,7 cases at complete remission,and 9 relapsed cases...The present study aimed to clarify the role of SENEX in malignant cell proliferation in diffuse large B-cell lymphoma(DLBCL).22 DLBCL patients(6 newly diagnosed cases,7 cases at complete remission,and 9 relapsed cases)were included in the study.Our results indicated that both SENEX gene and protein were significantly increased in peripheral blood mononuclear cells(PBMCs)and tumor cells of relapsed DLBCL patients,accompanied by overexpression of p21,p16,and phosphorylated retinoblastoma(Rb).Silencing the SENEX gene in a DLBCL cell line caused a significant decrease in cell proliferation and inhibited cell cycle progression in the G1 phase.Phosphorylated Rb and E2F1 were also decreased,and activation of the Rb/E2F1 pathway was obviously suppressed.To conclude,the SENEX gene promotes proliferation in PBMC and tumor cells of DLBCL patients by activating the Rb/E2F1 pathway,in a manner suggesting that increased SENEX expression affects the relapse of DLBCL and may serve as an important target for DLBCL therapy.展开更多
This study investigated cold plasmas for multiple biological applications. Our previous work has found dielectric barrier discharge plasma improves chicken sperm quality. The number of Sertoli cells(SCs) decides sperm...This study investigated cold plasmas for multiple biological applications. Our previous work has found dielectric barrier discharge plasma improves chicken sperm quality. The number of Sertoli cells(SCs) decides spermatogenesis. However, whether cold plasma can regulate SC proliferation remains unclear. This study explored the effects of cold plasma on immature chicken SC proliferation and the regulation mechanism. Results showed that cold plasma exposure at 2.4 W for 30 s twice with an interval of 6 h produced(P<0.05) the maximum SC viability, cell growth, and cell cycle progression. SC proliferation-promoting effect of cold plasma treatment was regulated by increasing(P<0.05) the adenosine triphosphate production and the respiratory enzyme activity in the mitochondria. This process was potentially mediated by the adenosine monophosphate-activated protein kinase(AMPK)–mammalian target of rapamycin(m TOR) signaling pathway, which was regulated by the micro RNA(mi RNA) targeting regulation directly and by the intracellular reactive oxygen species homeostasis indirectly. The cold plasma treatment increased(P<0.01) the mi R-7450-5 p expression and led to a decreased(P<0.01) AMPKα1 level. On the other hand, mi R-100-5 p expression was reduced(P<0.05) and led to an increased(P<0.05) m TOR level in SCs. A single-stranded synthetic mi R-7450-5 p antagomir and a double-stranded synthetic mi R-100-5 p agomir reduced(P<0.05) the SC proliferation. However, this could be ameliorated(P<0.05) by the cold plasma treatment. Our findings suggest that appropriate cold plasma treatment provides a safe strategy to improve SC proliferation, which is beneficial to elevating male chicken reproductive capacity.展开更多
Chemotherapy may cause cellular oxidative stress to bone marrow.Oxidative damage of bone marrow hematopoietic microenvironment is closely related to chronic myelosuppression after chemotherapeutic treatment.Angelica s...Chemotherapy may cause cellular oxidative stress to bone marrow.Oxidative damage of bone marrow hematopoietic microenvironment is closely related to chronic myelosuppression after chemotherapeutic treatment.Angelica sinensis polysaccharides(ASP)are major effective ingredients of traditional Chinese medicine Angelica with multi-target anti-oxidative stress features.In the current study,we investigated the protective roles and mechanisms of ASP on chemotherapy-induced bone marrow stromal cell(BMSC)damage.The human bone marrow stromal cell line HS-5 cells were divided into control group,5-FU group,5-FU+ASP group,and 5-FU+LiCl group to investigate the mechanism of ASP to alleviate 5-FU-induced BMSC proliferation inhibition.The results showed that 5-FU inhibits the growth of HS-5 cells in a time and dose-dependent manner;however,ASP partially counteracted the 5-FU-induced decrease in cell viability,whereas Wnt signaling inhibitor Dkk1 antagonized the effect of ASP on HS-5 cells.ASP reversed the decrease in total cytoplasmicβ-catenin,p-GSK-3β,and CyclinD1 following 5-FU treatment and modulated nuclear expression ofβ-catenin,Lef-1,and C-myc proteins.Furthermore,ASP also enhanced the antioxidant capacity of cells and reduced 5-FU-induced oxidative stress,attenuated FoxO1 expression,thus weakened its downstream apoptosis-related proteins and G0/G1 checkpoint-associated p27^(Kip1) expression to alleviate 5-FU-induced apoptosis and to promote cell cycle progression.All the results above suggest that the protective role of ASP in 5-FU-treated BMSCs proliferation for the chemotherapy may be related to its activating Wnt/β-catenin signaling and keeping homeostasis betweenβ-catenin and FoxO1 under oxidative stress.The study provides a potential therapeutic strategy for alleviating chemotherapeutic damage on BMSCs.展开更多
AIM:To explore the effect of the Andrographis paniculata(A.paniculata)polysaccharide on the proliferation and apoptosis of human retinoblastoma(RB)Y79 cells and its mechanism.METHODS:The refined A.paniculata polysacch...AIM:To explore the effect of the Andrographis paniculata(A.paniculata)polysaccharide on the proliferation and apoptosis of human retinoblastoma(RB)Y79 cells and its mechanism.METHODS:The refined A.paniculata polysaccharide was obtained using techniques such as water extraction,ethanol precipitation,and decompression concentration.The inhibition effect of the A.paniculata polysaccharide on the proliferation of Y79 cells was detected by cell proliferation assay.Flow cytometr y was used for the detection of cell apoptosis rate and cycle change.Realtime qunatitative polymerase chain reaction(RT q PCR)and Western blotting were used to detect the expression of cell apoptosis signal pathway-related factors(caspase-3,caspase-8,and caspase-9)and cell cycle signal pathwayrelated factors(CDK1 and cyclin B1)at the transcriptional and translational levels.RESULTS:Infrared and ultraviolet spectrum scanning showed that the extracted drug was a polysaccharide withhigh purity.After being treated with different concentrations of A.paniculata polysaccharide for different periods of time,the Y79 cells showed different degrees of proliferation inhibition.Flow cytometric observations showed that the cell apoptosis rate and the proportion of cells blocked in the G2/M phase were significantly increased after A.paniculata polysaccharide treatment.Further analysis revealed that the m RNA and protein expression of caspase-3,caspase-8,and caspase-9 in the A.paniculata polysaccharide treatment groups increased significantly compared with that in the control groups,while the expression of CDK1 and cyclin B1 decreased significantly.CONCLUSION:The A.paniculata polysaccharide could inhibit the proliferation and induce apoptosis of Y79 cells.Its possible mechanism is via the upregulation of caspase-3,caspase-8,and caspase-9 expression in the cell apoptotic signaling pathway and the downregulation of CDK1 and cyclin B1 expression in the cell cycle signaling pathway.展开更多
In order to explore the role of forkhead box protein O1(FoxO1)in the lipid metabolism and cell proliferation,goose primary hepatocytes were isolated and incubated with insulin or PI3K-Akt-mTOR pathway dual inhibitor N...In order to explore the role of forkhead box protein O1(FoxO1)in the lipid metabolism and cell proliferation,goose primary hepatocytes were isolated and incubated with insulin or PI3K-Akt-mTOR pathway dual inhibitor NVPBEZ235,and then transfected with FoxO1 interference plasmid.The related parameters of lipid metabolism and cell proliferation were measured.The results firstly showed that FoxO1 interference increased the intracellular TG and lipids concentration(P<0.05);and increased the proliferative index(PI),cell DNA synthesis,protein expression of Cyclin D1 in goose primary hepatocytes(P<0.05).Secondly,the co-treatment of insulin and FoxO1 interference increased the mRNA level and protein content of Cyclin D1(P<0.05);however,there was no significant difference between the insulin treatment and the co-treatment of insulin and miR-FoxO1 interference in the intracellular TG and lipids concentration and PI(P>0.05).Lastly,the decrease of intracellular TG and lipids concentration and PI induced by NVP-BEZ235 was up-regulated by FoxO1 interference significantly(P<0.05).In summary,FoxO1 could regulate the lipids metabolism and cell proliferation mediated by PI3K-Akt-mTOR signaling pathway in goose primary hepatocytes.Further investigations are required to highlight the potential role of FoxO1 in the lipid metabolism and cell proliferation mediated by insulin in goose primary hepatocyte.展开更多
BACKGROUND Dietary zinc deficiency has been shown to be associated with the development of esophageal cancer in humans,but the exact mechanism of action is not known AIM To observe the effects of dietary zinc deficien...BACKGROUND Dietary zinc deficiency has been shown to be associated with the development of esophageal cancer in humans,but the exact mechanism of action is not known AIM To observe the effects of dietary zinc deficiency on esophageal squamous cell proliferation.METHODS Thirty C57BL/6 mice were randomly divided into three groups:A zinc-sufficient(ZS)group,zinc-deficient(ZD)group,and zinc-replenished(ZR)group.For weeks 1–10,zinc levels in the mice diets were 30.66–30.89 mg/kg in the ZS group and 0.66–0.89 mg/kg in the ZD and ZR groups.During weeks 10–12,the ZR group was switched to the ZS diet;the other two groups had no changes in their diets.Changes in body weight,serum,and esophageal tissue zinc concentrations were assessed as well as differences in the expression of proliferating cell nuclear antigen(PCNA),mitogen-activated protein kinase p38(p38MAPK),nuclear factor kappa B(NF-κB)p105,NF-κB p65,and cyclooxygenase(COX)-2 proteins in the esophageal mucosa.RESULTS The body weight and zinc concentration in the serum and esophageal mucosa were significantly lower in the ZD and ZR groups than in the ZS group(P<0.05).In ZD mice,there was a marked proliferation of basal cells in the esophageal mucosa,resulting in a disturbance in the arrangement of basal cells in layers 2–4,a thickening of the squamous layer,and a significant increase in the expression of the above-mentioned five proteins involved in proliferation and inflammation in the esophageal mucosa.Two weeks after switching to the ZS diet,the serum zinc concentration in the ZR group increased,and the expression of PCNA,NF-κB p105,and COX-2 decreased,but the concentration of zinc in the esophageal mucosa and the structure of the esophageal mucosa did not display any significant changes CONCLUSION The ZD diet decreased the growth rate and promoted the proliferation of esophageal squamous cells in mice.The mechanism of proliferation was related to the induced overexpression of COX-2,P38,PCNA,and NF-κB(p105 and p65),and the ZR diet reduced the expression of PCNA,NF-κB p105,and COX-2,thereby reversing this process.展开更多
Background Normally, few immunocompetent cell are present in the endolymphatic sac (ES). During an active immune response in the inner ear, large amount of inflammatory cells, including immunocompetent cells, are seen...Background Normally, few immunocompetent cell are present in the endolymphatic sac (ES). During an active immune response in the inner ear, large amount of inflammatory cells, including immunocompetent cells, are seen in the ES. The current study aimed at assessing cellular proliferation within the ES during induced immune response in the inner ear. Methods Fifteen healthy, female SD rats were sensitized systemically with keyhole limpet hemocyanin (KLH), followed by local inoculation in the cochlea through basal turn fenestration with the same antigen. On Days 3, 7 and 14 following inoculation, the animal was sacrificed after intraperitoneal administration of 5-bromo-2'-deoxyuridine (BrdUrd), and the temporal bone harvested. Following decalcification, infiltration by BrdUrd- and IgG-positive cells in the ES was studied on frozen sections with H & E and immunohistochemical staining. Results During the secondary immune response in the inner ear against T-dependent antigens, there is increased cellular proliferation in the ES. The proliferated cells may differentiate into immunocompetent cells at the same location. Conclusions These findings indicate that the ES plays an important role in immune response of inner ear.展开更多
Taking grass carps with the initial weight of about 20g as the research object,the basic feeds of grass carps were added with0.0%,0.1%,0.3%,0.6%,0.8%,and 1% of nano-sustained release sodium butyrate to prepare 6 types...Taking grass carps with the initial weight of about 20g as the research object,the basic feeds of grass carps were added with0.0%,0.1%,0.3%,0.6%,0.8%,and 1% of nano-sustained release sodium butyrate to prepare 6 types of experimental feeds with equal nitrogen and energy.The effects of different concentrations of nano-sustained release sodium butyrate were surveyed on growth and intestinal cell proliferation of grass carps.The experiment was carried out in cages with 50 carps per cage,and each treatment was repeated 3 times for60 days.Experimental results indicated that the addition of nano-sustained release sodium butyrate significantly promoted the growth of grass carps and significantly increased the ratio of intestinal villus to crypt depth.When the addition of nano-sustained release sodium butyrate was0.6%,the weight increase rate,specific growth rate,fullness and intestinal villus height of grass carps were the highest,which was significantly higher than that of the control group(P < 0.05).The study results indicated that addition of appropriate amount of nano-sustained release sodium butyrate can promote the growth of grass carps through increasing the intestinal villus height,and the suitable addition dosage was0.6%.展开更多
It was previously found that the electric charge of water determines its ability to interact with other substances,including biologically significant ones.It is shown here that the electric charge of water can also de...It was previously found that the electric charge of water determines its ability to interact with other substances,including biologically significant ones.It is shown here that the electric charge of water can also determine its ability to penetrate and accumulate in living cells.In particular,it has been shown that the high penetrating ability of positively charged water determines both its active penetration into cells and accumulation in them,which creates favourable conditions for cell proliferation.At the same time,it has been shown that the low penetrating ability of negatively charged water determines its ability to slow down cell proliferation.It also discusses how medics can obtain and use water at different charges.展开更多
Objective:To investigate the mechanism of long non-coding RNA SNHG7 and its regulatory effect on the proliferation,migration,and epithelial-mesenchymal transition of cholangiocarcinoma cells.Methods:A total of 20 pair...Objective:To investigate the mechanism of long non-coding RNA SNHG7 and its regulatory effect on the proliferation,migration,and epithelial-mesenchymal transition of cholangiocarcinoma cells.Methods:A total of 20 pairs of cholangiocarcinoma and adjacent non-tumor bile duct tissues were collected from patients with cholangiocarcinoma who underwent surgery in the Affiliated Hospital of Hebei University(Hebei,China).Cholangiocarcinoma cell lines CCLP-1,QBC939,RBE,and HCC-9810 as well as normal human biliary epithelial cell line HIBEC were purchased for cell culture.We performed cell transfection,quantitative real-time polymerase chain reaction(qRT-PCR)to detect gene expression,Cell Counting Kit-8(CCK-8)experiment to determine cell proliferation ability,scratch test to determine cell migration ability,and Transwell test to detect cell invasion ability.Results:The expression of lncRNA SNHG7 in cholangiocarcinoma cell lines CCLP-1,QBC939,RBE,and HCCC-9810 was 3.21±1.01,3.03±1.02,2.98±1.21,and 3.12±1.14,respectively,while its expression in normal cell line HIBEC was 3.21±1.21;the expression of lncRNA SNHG7 in CCLP-1 was the highest;compared with HIBEC,the p values were all less than 0.05,indicating that the difference was statistically significant.The expression of miR-520f-3p in CCLP-1,QBC939,RBE,and HCCC-9810 was 1.45±0.75,1.55±0.71,1.54±0.73,and 1.61±0.72,respectively,while its expression in normal cell line HIBEC was 3.21±1.21;the expression of miR-520f-3p in CCLP-1 was the lowest,and compared with HIBEC,the p values were all less than 0.05,indicating that the difference was statistically significant.In qRT-PCR,the expression of lncRNA SNHG7 of si-NC(3.21±1.11)was significantly higher than that of si-SNHG7(1.14±0.76),and the p value was less than 0.05,indicating that the difference was statistically significant.In the CCK-8 experiment,the proliferation ability of CCLP-1 cells of the si-NC group at 24 h,48 h,and 72 h was 0.61±0.59,0.75±0.68,and 1.36±0.71,respectively;the proliferation ability of CCLP-1 cells of the si-SNHG7 group at 24 h,48 h,and 72 h was 0.51±0.64,0.59±0.59,and 0.63±0.61,respectively;there was a significant decrease in the proliferation ability,and the p value was less than 0.05,indicating a statistically significant difference.After 24 h of scratch treatment,compared with the si-NC group,the migration ability of CCLP-1 cells of the si-SNHG20 group was reduced(t=6.356,P=0.026).The results of Transwell test showed that the cell invasion ability of CCLP-1 in the si-SNHG20 group was significantly reduced compared with the si-NC group(t=7.845,P=0.032).Conclusion:Exploring the gene expression mechanism in relation to the occurrence and development of cholangiocarcinoma is beneficial to future clinical work in terms of diagnosis,treatment,and prognosis.The knockdown of lncRNA SNHG7 can effectively inhibit the proliferation,migration,and invasion of cholangiocarcinoma.展开更多
To clone human arresten gene and investigate biological activity of the recombinant protein.Methods Human arresten gene was obtained from the plasmid pGEMArr and subcloned into the BamHⅠ and Pst Ⅰ restriction sites ...To clone human arresten gene and investigate biological activity of the recombinant protein.Methods Human arresten gene was obtained from the plasmid pGEMArr and subcloned into the BamHⅠ and Pst Ⅰ restriction sites of prokaryotic expression vector pRSET containing T7 promoter.The recombinant plasmid pRSETAN was subsequently transformed into the strain E.coli BL21(DE3),and the target gene was expressed under induction of IPTG.The expressed protein was extracted,purified by Ni 2+ chelation affinity chromatography and refoled.The effect of the recombinant protein on proliferation of human umbilical vein endothelial cells (HUVECs) was also analyzed with the MTT assay.Results Endonuclease digesting and DNA sequencing confirmed that the arresten gene was correctly inserted into the expression vector.The recombinant protein was hightly expressed in the form of inclusion body in the host bacteria after induction.SDS-PAGE analysis revealed that the recombinant protein with a molecular weight of 26×103 amounted to 27% of the total bacterial proteins.The purity of the expected protein could reach over 96% through affinity chromatography.After renaturation,the recombinant protein could reach over 96% through affinity chromatography.After renaturation,the recombinant protein could significantly suppress proliferation of human umbilical vein endothelia cells(HUVECs) induced by vascular endothelial growth factor(VEGF).Conclusion Human arresten gene was successfully cloned into the expression vector pRSET and expressed at high level in Escherichia coli.Purified and refolded arresten protein could effectively inhibit proliferation of vascular endothelia cells.2 refs.展开更多
The regulation of spermatogonial proliferation and apoptosis is of great significance for maintaining spermatogenesis.The single-cell RNA sequencing(scRNA-seq)analysis of the testis was performed to identify genes upr...The regulation of spermatogonial proliferation and apoptosis is of great significance for maintaining spermatogenesis.The single-cell RNA sequencing(scRNA-seq)analysis of the testis was performed to identify genes upregulated in spermatogonia.Using scRNAseq analysis,we identified the spermatogonia upregulated gene origin recognition complex subunit 6(Orc6),which is involved in DNA replication and cell cycle regulation;its protein expression in the human and mouse testis was detected by western blot and immunofluorescence.To explore the potential function of Orc6 in spermatogonia,the C18-4 cell line was transfected with control or Orc6 siRNA.Subsequently,5-ethynyl-2-deoxyuridine(EdU)and terminal deoxynucleotidyl transferase dUTP nick end labeling(TUNEL)assays,flow cytometry,and western blot were used to evaluate its effects on proliferation and apoptosis.It was revealed that ORC6 could promote proliferation and inhibit apoptosis of C18-4 cells.Bulk RNA sequencing and bioinformatics analysis indicated that Orc6 was involved in the activation of wingless/integrated(Wnt)/β-catenin signaling.Western blot revealed that the expression ofβ-catenin protein and its phosphorylation(Ser675)were significantly decreased when silencing the expression of ORC6.Our findings indicated that Orc6 was upregulated in spermatogonia,whereby it regulated proliferation and apoptosis by activating Wnt/β-catenin signaling.展开更多
Human-specific insertions play important roles in human phenotypes and diseases.Here we reported a 446-bp insertion(Insert-446)in intron 11 of the TBC1D8B gene,located on chromosome X,and traced its origin to a portio...Human-specific insertions play important roles in human phenotypes and diseases.Here we reported a 446-bp insertion(Insert-446)in intron 11 of the TBC1D8B gene,located on chromosome X,and traced its origin to a portion of intron 6 of the EBF1 gene on chromosome 5.Interestingly,Insert-446 was present in the human Neanderthal and Denisovans genomes,and was fixed in humans after human-chimpanzee divergence.We have demonstrated that Insert-446 acts as an enhancer through binding transcript factors that promotes a higher expression of human TBC1D8B gene as compared with orthologs in macaques.In addition,over-expression TBC1D8B promoted cell proliferation and migration through“a dual finger”catalytic mechanism(Arg538 and Gln573)in the TBC domain in vitro and knockdown of TBC1D8B attenuated tumorigenesis in vivo.Knockout of Insert-446 prevented cell proliferation and migration in cancer and normal cells.Our results reveal that the human-specific Insert-446 promotes cell proliferation and migration by upregulating the expression of TBC1D8B gene.These findings provide a significant insight into the effects of human-specific insertions on evolution.展开更多
基金supported by the National Natural Science Foundation of China[Grant Number:81972803]。
文摘Objective To investigate the role and molecular mechanism of exosomal miR-224-5p in colorectal cancer(CRC).Methods The miR-224-5p expression in CRC patient tissues and cell-derived exosomes was measured by laser capture microdissection and qRT-PCR,respectively.Dual-luciferase reporter gene assay was used to determine the target gene of miR-224-5p.The protein expressions of p53 and unc-51 like kinase 2(ULK2)in CRC cells were detected by western blot.Flow cytometry was used to detect cell cycle and apoptosis.Cell proliferation was measured by CCK8 and EdU assay.Results The miR-224-5p expression was upregulated in CRC tissues and increased progressively with the rise of CRC stage.CRC cells secreted extracellular miR-224-5p mainly in an exosome-dependent manner,and then miR-224-5p could be transferred to surrounding tumor cells to regulate cell proliferation in the form of autocrine or paracrine.Moreover,ULK2 was characterized as a direct target of miR-224-5p and was downregulated in CRC tissues.Interestingly,ULK2 inhibited CRC cell proliferation in a p53-dependent manner.Furthermore,exosome-derived miR-224-5p partially reversed the proliferation regulation of ULK2 on CRC cells.Conclusion Our findings demonstrate that exosome-transmitted miR-224-5p promotes p53-dependent cell proliferation by targeting ULK2 in CRC,which may offer promising targets for CRC prevention and therapy.
基金supported by the National Natural Science Foundation of China,Nos.81672261(to XH),81972151(to HZ),82372568(to JL)the Natural Science Foundation of Guangdong Province,Nos.2019A1515011106(to HZ),2023A1515030080(to JL)。
文摘Prolife ration of neural stem cells is crucial for promoting neuronal regeneration and repairing cerebral infarction damage.Transcranial magnetic stimulation(TMS)has recently emerged as a tool for inducing endogenous neural stem cell regeneration,but its underlying mechanisms remain unclea r In this study,we found that repetitive TMS effectively promotes the proliferation of oxygen-glucose deprived neural stem cells.Additionally,repetitive TMS reduced the volume of cerebral infa rction in a rat model of ischemic stro ke caused by middle cerebral artery occlusion,im p roved rat cognitive function,and promoted the proliferation of neural stem cells in the ischemic penumbra.RNA-sequencing found that repetitive TMS activated the Wnt signaling pathway in the ischemic penumbra of rats with cerebral ischemia.Furthermore,PCR analysis revealed that repetitive TMS promoted AKT phosphorylation,leading to an increase in mRNA levels of cell cycle-related proteins such as Cdk2 and Cdk4.This effect was also associated with activation of the glycogen synthase kinase 3β/β-catenin signaling pathway,which ultimately promotes the prolife ration of neural stem cells.Subsequently,we validated the effect of repetitive TMS on AKT phosphorylation.We found that repetitive TMS promoted Ca2+influx into neural stem cells by activating the P2 calcium channel/calmodulin pathway,thereby promoting AKT phosphorylation and activating the glycogen synthase kinase 3β/β-catenin pathway.These findings indicate that repetitive TMS can promote the proliferation of endogenous neural stem cells through a Ca2+influx-dependent phosphorylated AKT/glycogen synthase kinase 3β/β-catenin signaling pathway.This study has produced pioneering res ults on the intrinsic mechanism of repetitive TMS to promote neural function recove ry after ischemic stro ke.These results provide a stro ng scientific foundation for the clinical application of repetitive TMS.Moreover,repetitive TMS treatment may not only be an efficient and potential approach to support neurogenesis for further therapeutic applications,but also provide an effective platform for the expansion of neural stem cells.
基金Supported by National Natural Science Foundation of China,No.81870453.
文摘BACKGROUND Gastrointestinal stromal tumor(GIST)is a common neoplasm with high rates of recurrence and metastasis,and its therapeutic efficacy is still not ideal.There is an unmet need to find new molecular therapeutic targets for GIST.TATA-boxbinding protein-associated factor 15(TAF15)contributes to the progress of various tumors,while the role and molecular mechanism of TAF15 in GIST progression are still unknown.AIM To explore new molecular therapeutic targets for GIST and understand the biological role and underlying mechanisms of TAF15 in GIST progression.METHODS Proteomic analysis was performed to explore the differentially expressed proteins in GIST.Western blotting and immunohistochemical analysis were used to verify the expression level of TAF15 in GIST tissues and cell lines.Cell counting kit-8,colony formation,wound-healing and transwell assay were executed to detect the ability of TAF15 on cell proliferation,migration and invasion.A xenograft mouse model was applied to explore the role of TAF15 in the progression of GIST.Western blotting was used to detect the phosphorylation level and total level of RAF1,MEK and ERK1/2.RESULTS A total of 1669 proteins were identified as differentially expressed proteins with 762 upregulated and 907 downregulated in GIST.TAF15 was selected for the further study because of its important role in cell proliferation and migration.TAF15 was significantly over expressed in GIST tissues and cell lines.Overexpression of TAF15 was associated with larger tumor size and higher risk stage of GIST.TAF15 knockdown significantly inhibited the cell proliferation and migration of GIST in vitro and suppressed tumor growth in vivo.Moreover,the inhibition of TAF15 expression significantly decreased the phosphorylation level of RAF1,MEK and ERK1/2 in GIST cells and xenograft tissues,while the total RAF1,MEK and ERK1/2 had no significant change.CONCLUSION TAF15 is over expressed in GIST tissues and cell lines.Overexpression of TAF15 was associated with a poor prognosis of GIST patients.TAF15 promotes cell proliferation and migration in GIST via the activation of the RAF1/MEK/ERK signaling pathway.Thus,TAF15 is expected to be a novel latent molecular biomarker or therapeutic target of GIST.
基金funded by the Natural Science Foundation of Anhui Province,China(2008085QC158)the University Natural Science Research Project of Anhui Province(KJ2019A0165)。
文摘In mammals,microRNAs(miRNAs)play key roles in multiple biological processes by regulating the expression of target genes.Studies have found that the levels of miR-370-5p expression differ significantly in the skins of sheep with different hair colors;however,its function remains unclear.In this study,we investigated the roles of miR-370-5p in sheep melanocytes and found that the overexpression of miR-370-5p significantly inhibited cell proliferation(P<0.01),tyrosinase activity(P=0.001)and significantly reduced(P<0.001)melanin production.Functional prediction revealed that the 3′-untranslated region(UTR)of MAP3K8 has a putative miR-370-5p binding site,and the interaction between these two molecules was confirmed using luciferase reporter assays.In situ hybridization assays revealed that MAP3K8 is expressed in the cytoplasm of melanocytes.The results of quantitative RT-PCR and Western blotting analyses revealed that overexpression of miR-370-5p in melanocytes significantly inhibits(P<0.01)MAP3K8 expression via direct targeting of its 3′UTR.Inhibition of MAP3K8 expression by siRNA-MAP3K8 transfection induced a significant inhibition(P<0.01)of melanocyte proliferation and significant reduction(P<0.001)in melanin production,which is consistent with our observations for miR-370-5p.Target gene rescue experiments indicated that the expression of MAP3K8 in melanocytes co-transfected with miR-370-5p and MAP3K8-cDNA(containing sites for the targeted binding to miR-370-5p)was significantly rescued(P≤0.001),which subsequently promoted significant increases in cell proliferation(P<0.001)and melanin production(P<0.01).Collectively,these findings indicate that miR-370-5p plays a functional role in inhibiting sheep melanocyte proliferation and melanogenesis by downregulating the expression of MAP3K8.
基金This work was funded by the National Natural Science Foundation of China(31802057)the Second Batch of Special Grant from China Postdoctoral Science Foundation(2020TQ0252)+1 种基金the Postdoctoral Research Funding Project of Jiangsu Province,China in 2020(2020Z213)the Open Project Program of Joint International Research Laboratory of Agriculture and Agri-Product Safety,the Ministry of Education of China(JILAR-KF202017).
文摘Follicle-stimulating hormone(FSH),an important hypothalamic-pituitary-gonadal axis(HPG)hormone,is secreted by the pituitary gland.This study confirms that FSH is expressed in chicken follicles at different stages,and positive FSHβ mRNA signals were stronger(P<0.05)in granulosa cells than in oocytes.The 369 bp coding sequence of FSHβ in ovaries is 100%identical to that in the pituitary gland.The experiment in vitro revealed that the ovary possessed FSH secretory capacity.Further,FSHβ mRNA was significantly upregulated(P<0.05)in follicles and significantly higher(P<0.05)than that in the pituitary gland by approximately 2–23 times with the development.The number of granulosa cells decreased significantly(P<0.05)in the cells with siRNA treatment,confirming that the ovarian FSH could promote granulosa cell proliferation.This view was supported by cell cycle analysis and CCND2 and CCNE2 expression.Further research indicated that no difference(P>0.05)was observed between the number of granulosa cells treated with FSHβ siRNA and in exogenous FSH.However,the number of granulosa cells without FSHβ siRNA transfection was significantly higher(P<0.05)for exogenous FSH.This finding suggests that the proliferative effect of exogenous FSH on ovarian granulosa cells depend on endogenous FSH.This study demonstrated that the FSH gene was expressed in chicken follicles and promoted ovarian granulosa cell proliferation,which enriched the theory on HPG axis.
文摘BACKGROUND Colorectal cancer(CRC)is a worldwide problem,which has been associated with changes in diet and lifestyle pattern.As a result of colonic fermentation of dietary fibres,short chain free fatty acids are generated which activate free fatty acid receptors(FFAR)2 and 3.FFAR2 and FFAR3 genes are abundantly expressed in colonic epithelium and play an important role in the metabolic homeostasis of colonic epithelial cells.Earlier studies point to the involvement of FFAR2 in colorectal carcinogenesis.AIM To understand the role of short chain FFARs in CRC.METHODS Transcriptome analysis console software was used to analyse microarray data from CRC patients and cell lines.We employed short-hairpin RNA mediated down regulation of FFAR2 and FFAR3 genes,which was validated using quantitative real time polymerase chain reaction.Assays for glucose uptake and cyclic adenosine monophosphate(cAMP)generation was done along with immunofluorescence studies to study the effects of FFAR2/FFAR3 knockdown.For measuring cell proliferation,we employed real time electrical impedancebased assay available from xCELLigence.RESULTS Microarray data analysis of CRC patient samples showed a significant down regulation of FFAR2 gene expression.This prompted us to study the FFAR2 in CRC.Since,FFAR3 shares significant structural and functional homology with FFAR2,we knocked down both these receptors in CRC cell line HCT 116.These modified cell lines exhibited higher proliferation rate and were found to have increased glucose uptake as well as increased level of glucose transporter 1.Since,FFAR2 and FFAR3 signal through G protein subunit(Gαi),knockdown of these receptors was associated with increased cAMP.Inhibition of protein kinase A(PKA)did not alter the growth and proliferation of these cells indicating a mechanism independent of cAMP/PKA pathway.CONCLUSION Our results suggest role of FFAR2/FFAR3 genes in increased proliferation of colon cancer cells via enhanced glucose uptake and exclude the role of PKA mediated cAMP signalling.Alternate pathways could be involved that would ultimately result in increased cell proliferation as a result of down regulated FFAR2/FFAR3 genes.This study paves the way to understand the mechanism of action of short chain FFARs in CRC.
基金supported by the National Natural Science Foundation of China(Grant No.81670179)the Research Fund Project of Anhui Medical University(No.2018xkj026)the National Natural Science Foundation Incubation Project of the Second Hospital of Anhui Medical University(No.2019GQFY11).
文摘The present study aimed to clarify the role of SENEX in malignant cell proliferation in diffuse large B-cell lymphoma(DLBCL).22 DLBCL patients(6 newly diagnosed cases,7 cases at complete remission,and 9 relapsed cases)were included in the study.Our results indicated that both SENEX gene and protein were significantly increased in peripheral blood mononuclear cells(PBMCs)and tumor cells of relapsed DLBCL patients,accompanied by overexpression of p21,p16,and phosphorylated retinoblastoma(Rb).Silencing the SENEX gene in a DLBCL cell line caused a significant decrease in cell proliferation and inhibited cell cycle progression in the G1 phase.Phosphorylated Rb and E2F1 were also decreased,and activation of the Rb/E2F1 pathway was obviously suppressed.To conclude,the SENEX gene promotes proliferation in PBMC and tumor cells of DLBCL patients by activating the Rb/E2F1 pathway,in a manner suggesting that increased SENEX expression affects the relapse of DLBCL and may serve as an important target for DLBCL therapy.
基金supported by the National Natural Science Foundation of China(31902338)the Natural Science Foundation of Chongqing,China(cstc2019jcyjmsxm X0056)the Innovative Project of Chongqing Returned Overseas Person Entrepreneurship and Innovation Plan,China(cx2020057)。
文摘This study investigated cold plasmas for multiple biological applications. Our previous work has found dielectric barrier discharge plasma improves chicken sperm quality. The number of Sertoli cells(SCs) decides spermatogenesis. However, whether cold plasma can regulate SC proliferation remains unclear. This study explored the effects of cold plasma on immature chicken SC proliferation and the regulation mechanism. Results showed that cold plasma exposure at 2.4 W for 30 s twice with an interval of 6 h produced(P<0.05) the maximum SC viability, cell growth, and cell cycle progression. SC proliferation-promoting effect of cold plasma treatment was regulated by increasing(P<0.05) the adenosine triphosphate production and the respiratory enzyme activity in the mitochondria. This process was potentially mediated by the adenosine monophosphate-activated protein kinase(AMPK)–mammalian target of rapamycin(m TOR) signaling pathway, which was regulated by the micro RNA(mi RNA) targeting regulation directly and by the intracellular reactive oxygen species homeostasis indirectly. The cold plasma treatment increased(P<0.01) the mi R-7450-5 p expression and led to a decreased(P<0.01) AMPKα1 level. On the other hand, mi R-100-5 p expression was reduced(P<0.05) and led to an increased(P<0.05) m TOR level in SCs. A single-stranded synthetic mi R-7450-5 p antagomir and a double-stranded synthetic mi R-100-5 p agomir reduced(P<0.05) the SC proliferation. However, this could be ameliorated(P<0.05) by the cold plasma treatment. Our findings suggest that appropriate cold plasma treatment provides a safe strategy to improve SC proliferation, which is beneficial to elevating male chicken reproductive capacity.
基金supported by the National Natural Science Foundation of China(Grant No.81873103)the Foundation and Frontier Research Project of Chongqing Science and Technology Commission(Grant No.cstc2014jcyjA10001).
文摘Chemotherapy may cause cellular oxidative stress to bone marrow.Oxidative damage of bone marrow hematopoietic microenvironment is closely related to chronic myelosuppression after chemotherapeutic treatment.Angelica sinensis polysaccharides(ASP)are major effective ingredients of traditional Chinese medicine Angelica with multi-target anti-oxidative stress features.In the current study,we investigated the protective roles and mechanisms of ASP on chemotherapy-induced bone marrow stromal cell(BMSC)damage.The human bone marrow stromal cell line HS-5 cells were divided into control group,5-FU group,5-FU+ASP group,and 5-FU+LiCl group to investigate the mechanism of ASP to alleviate 5-FU-induced BMSC proliferation inhibition.The results showed that 5-FU inhibits the growth of HS-5 cells in a time and dose-dependent manner;however,ASP partially counteracted the 5-FU-induced decrease in cell viability,whereas Wnt signaling inhibitor Dkk1 antagonized the effect of ASP on HS-5 cells.ASP reversed the decrease in total cytoplasmicβ-catenin,p-GSK-3β,and CyclinD1 following 5-FU treatment and modulated nuclear expression ofβ-catenin,Lef-1,and C-myc proteins.Furthermore,ASP also enhanced the antioxidant capacity of cells and reduced 5-FU-induced oxidative stress,attenuated FoxO1 expression,thus weakened its downstream apoptosis-related proteins and G0/G1 checkpoint-associated p27^(Kip1) expression to alleviate 5-FU-induced apoptosis and to promote cell cycle progression.All the results above suggest that the protective role of ASP in 5-FU-treated BMSCs proliferation for the chemotherapy may be related to its activating Wnt/β-catenin signaling and keeping homeostasis betweenβ-catenin and FoxO1 under oxidative stress.The study provides a potential therapeutic strategy for alleviating chemotherapeutic damage on BMSCs.
文摘AIM:To explore the effect of the Andrographis paniculata(A.paniculata)polysaccharide on the proliferation and apoptosis of human retinoblastoma(RB)Y79 cells and its mechanism.METHODS:The refined A.paniculata polysaccharide was obtained using techniques such as water extraction,ethanol precipitation,and decompression concentration.The inhibition effect of the A.paniculata polysaccharide on the proliferation of Y79 cells was detected by cell proliferation assay.Flow cytometr y was used for the detection of cell apoptosis rate and cycle change.Realtime qunatitative polymerase chain reaction(RT q PCR)and Western blotting were used to detect the expression of cell apoptosis signal pathway-related factors(caspase-3,caspase-8,and caspase-9)and cell cycle signal pathwayrelated factors(CDK1 and cyclin B1)at the transcriptional and translational levels.RESULTS:Infrared and ultraviolet spectrum scanning showed that the extracted drug was a polysaccharide withhigh purity.After being treated with different concentrations of A.paniculata polysaccharide for different periods of time,the Y79 cells showed different degrees of proliferation inhibition.Flow cytometric observations showed that the cell apoptosis rate and the proportion of cells blocked in the G2/M phase were significantly increased after A.paniculata polysaccharide treatment.Further analysis revealed that the m RNA and protein expression of caspase-3,caspase-8,and caspase-9 in the A.paniculata polysaccharide treatment groups increased significantly compared with that in the control groups,while the expression of CDK1 and cyclin B1 decreased significantly.CONCLUSION:The A.paniculata polysaccharide could inhibit the proliferation and induce apoptosis of Y79 cells.Its possible mechanism is via the upregulation of caspase-3,caspase-8,and caspase-9 expression in the cell apoptotic signaling pathway and the downregulation of CDK1 and cyclin B1 expression in the cell cycle signaling pathway.
基金supported by the National Natural Science Funds of China(No.31672413)the National Waterfowl Industrial Technology System(No.CARS-43-6).
文摘In order to explore the role of forkhead box protein O1(FoxO1)in the lipid metabolism and cell proliferation,goose primary hepatocytes were isolated and incubated with insulin or PI3K-Akt-mTOR pathway dual inhibitor NVPBEZ235,and then transfected with FoxO1 interference plasmid.The related parameters of lipid metabolism and cell proliferation were measured.The results firstly showed that FoxO1 interference increased the intracellular TG and lipids concentration(P<0.05);and increased the proliferative index(PI),cell DNA synthesis,protein expression of Cyclin D1 in goose primary hepatocytes(P<0.05).Secondly,the co-treatment of insulin and FoxO1 interference increased the mRNA level and protein content of Cyclin D1(P<0.05);however,there was no significant difference between the insulin treatment and the co-treatment of insulin and miR-FoxO1 interference in the intracellular TG and lipids concentration and PI(P>0.05).Lastly,the decrease of intracellular TG and lipids concentration and PI induced by NVP-BEZ235 was up-regulated by FoxO1 interference significantly(P<0.05).In summary,FoxO1 could regulate the lipids metabolism and cell proliferation mediated by PI3K-Akt-mTOR signaling pathway in goose primary hepatocytes.Further investigations are required to highlight the potential role of FoxO1 in the lipid metabolism and cell proliferation mediated by insulin in goose primary hepatocyte.
文摘BACKGROUND Dietary zinc deficiency has been shown to be associated with the development of esophageal cancer in humans,but the exact mechanism of action is not known AIM To observe the effects of dietary zinc deficiency on esophageal squamous cell proliferation.METHODS Thirty C57BL/6 mice were randomly divided into three groups:A zinc-sufficient(ZS)group,zinc-deficient(ZD)group,and zinc-replenished(ZR)group.For weeks 1–10,zinc levels in the mice diets were 30.66–30.89 mg/kg in the ZS group and 0.66–0.89 mg/kg in the ZD and ZR groups.During weeks 10–12,the ZR group was switched to the ZS diet;the other two groups had no changes in their diets.Changes in body weight,serum,and esophageal tissue zinc concentrations were assessed as well as differences in the expression of proliferating cell nuclear antigen(PCNA),mitogen-activated protein kinase p38(p38MAPK),nuclear factor kappa B(NF-κB)p105,NF-κB p65,and cyclooxygenase(COX)-2 proteins in the esophageal mucosa.RESULTS The body weight and zinc concentration in the serum and esophageal mucosa were significantly lower in the ZD and ZR groups than in the ZS group(P<0.05).In ZD mice,there was a marked proliferation of basal cells in the esophageal mucosa,resulting in a disturbance in the arrangement of basal cells in layers 2–4,a thickening of the squamous layer,and a significant increase in the expression of the above-mentioned five proteins involved in proliferation and inflammation in the esophageal mucosa.Two weeks after switching to the ZS diet,the serum zinc concentration in the ZR group increased,and the expression of PCNA,NF-κB p105,and COX-2 decreased,but the concentration of zinc in the esophageal mucosa and the structure of the esophageal mucosa did not display any significant changes CONCLUSION The ZD diet decreased the growth rate and promoted the proliferation of esophageal squamous cells in mice.The mechanism of proliferation was related to the induced overexpression of COX-2,P38,PCNA,and NF-κB(p105 and p65),and the ZR diet reduced the expression of PCNA,NF-κB p105,and COX-2,thereby reversing this process.
文摘Background Normally, few immunocompetent cell are present in the endolymphatic sac (ES). During an active immune response in the inner ear, large amount of inflammatory cells, including immunocompetent cells, are seen in the ES. The current study aimed at assessing cellular proliferation within the ES during induced immune response in the inner ear. Methods Fifteen healthy, female SD rats were sensitized systemically with keyhole limpet hemocyanin (KLH), followed by local inoculation in the cochlea through basal turn fenestration with the same antigen. On Days 3, 7 and 14 following inoculation, the animal was sacrificed after intraperitoneal administration of 5-bromo-2'-deoxyuridine (BrdUrd), and the temporal bone harvested. Following decalcification, infiltration by BrdUrd- and IgG-positive cells in the ES was studied on frozen sections with H & E and immunohistochemical staining. Results During the secondary immune response in the inner ear against T-dependent antigens, there is increased cellular proliferation in the ES. The proliferated cells may differentiate into immunocompetent cells at the same location. Conclusions These findings indicate that the ES plays an important role in immune response of inner ear.
基金Supported by Science and Technology Planning Project of Changsha City(k1407023-31)
文摘Taking grass carps with the initial weight of about 20g as the research object,the basic feeds of grass carps were added with0.0%,0.1%,0.3%,0.6%,0.8%,and 1% of nano-sustained release sodium butyrate to prepare 6 types of experimental feeds with equal nitrogen and energy.The effects of different concentrations of nano-sustained release sodium butyrate were surveyed on growth and intestinal cell proliferation of grass carps.The experiment was carried out in cages with 50 carps per cage,and each treatment was repeated 3 times for60 days.Experimental results indicated that the addition of nano-sustained release sodium butyrate significantly promoted the growth of grass carps and significantly increased the ratio of intestinal villus to crypt depth.When the addition of nano-sustained release sodium butyrate was0.6%,the weight increase rate,specific growth rate,fullness and intestinal villus height of grass carps were the highest,which was significantly higher than that of the control group(P < 0.05).The study results indicated that addition of appropriate amount of nano-sustained release sodium butyrate can promote the growth of grass carps through increasing the intestinal villus height,and the suitable addition dosage was0.6%.
文摘It was previously found that the electric charge of water determines its ability to interact with other substances,including biologically significant ones.It is shown here that the electric charge of water can also determine its ability to penetrate and accumulate in living cells.In particular,it has been shown that the high penetrating ability of positively charged water determines both its active penetration into cells and accumulation in them,which creates favourable conditions for cell proliferation.At the same time,it has been shown that the low penetrating ability of negatively charged water determines its ability to slow down cell proliferation.It also discusses how medics can obtain and use water at different charges.
文摘Objective:To investigate the mechanism of long non-coding RNA SNHG7 and its regulatory effect on the proliferation,migration,and epithelial-mesenchymal transition of cholangiocarcinoma cells.Methods:A total of 20 pairs of cholangiocarcinoma and adjacent non-tumor bile duct tissues were collected from patients with cholangiocarcinoma who underwent surgery in the Affiliated Hospital of Hebei University(Hebei,China).Cholangiocarcinoma cell lines CCLP-1,QBC939,RBE,and HCC-9810 as well as normal human biliary epithelial cell line HIBEC were purchased for cell culture.We performed cell transfection,quantitative real-time polymerase chain reaction(qRT-PCR)to detect gene expression,Cell Counting Kit-8(CCK-8)experiment to determine cell proliferation ability,scratch test to determine cell migration ability,and Transwell test to detect cell invasion ability.Results:The expression of lncRNA SNHG7 in cholangiocarcinoma cell lines CCLP-1,QBC939,RBE,and HCCC-9810 was 3.21±1.01,3.03±1.02,2.98±1.21,and 3.12±1.14,respectively,while its expression in normal cell line HIBEC was 3.21±1.21;the expression of lncRNA SNHG7 in CCLP-1 was the highest;compared with HIBEC,the p values were all less than 0.05,indicating that the difference was statistically significant.The expression of miR-520f-3p in CCLP-1,QBC939,RBE,and HCCC-9810 was 1.45±0.75,1.55±0.71,1.54±0.73,and 1.61±0.72,respectively,while its expression in normal cell line HIBEC was 3.21±1.21;the expression of miR-520f-3p in CCLP-1 was the lowest,and compared with HIBEC,the p values were all less than 0.05,indicating that the difference was statistically significant.In qRT-PCR,the expression of lncRNA SNHG7 of si-NC(3.21±1.11)was significantly higher than that of si-SNHG7(1.14±0.76),and the p value was less than 0.05,indicating that the difference was statistically significant.In the CCK-8 experiment,the proliferation ability of CCLP-1 cells of the si-NC group at 24 h,48 h,and 72 h was 0.61±0.59,0.75±0.68,and 1.36±0.71,respectively;the proliferation ability of CCLP-1 cells of the si-SNHG7 group at 24 h,48 h,and 72 h was 0.51±0.64,0.59±0.59,and 0.63±0.61,respectively;there was a significant decrease in the proliferation ability,and the p value was less than 0.05,indicating a statistically significant difference.After 24 h of scratch treatment,compared with the si-NC group,the migration ability of CCLP-1 cells of the si-SNHG20 group was reduced(t=6.356,P=0.026).The results of Transwell test showed that the cell invasion ability of CCLP-1 in the si-SNHG20 group was significantly reduced compared with the si-NC group(t=7.845,P=0.032).Conclusion:Exploring the gene expression mechanism in relation to the occurrence and development of cholangiocarcinoma is beneficial to future clinical work in terms of diagnosis,treatment,and prognosis.The knockdown of lncRNA SNHG7 can effectively inhibit the proliferation,migration,and invasion of cholangiocarcinoma.
文摘To clone human arresten gene and investigate biological activity of the recombinant protein.Methods Human arresten gene was obtained from the plasmid pGEMArr and subcloned into the BamHⅠ and Pst Ⅰ restriction sites of prokaryotic expression vector pRSET containing T7 promoter.The recombinant plasmid pRSETAN was subsequently transformed into the strain E.coli BL21(DE3),and the target gene was expressed under induction of IPTG.The expressed protein was extracted,purified by Ni 2+ chelation affinity chromatography and refoled.The effect of the recombinant protein on proliferation of human umbilical vein endothelial cells (HUVECs) was also analyzed with the MTT assay.Results Endonuclease digesting and DNA sequencing confirmed that the arresten gene was correctly inserted into the expression vector.The recombinant protein was hightly expressed in the form of inclusion body in the host bacteria after induction.SDS-PAGE analysis revealed that the recombinant protein with a molecular weight of 26×103 amounted to 27% of the total bacterial proteins.The purity of the expected protein could reach over 96% through affinity chromatography.After renaturation,the recombinant protein could reach over 96% through affinity chromatography.After renaturation,the recombinant protein could significantly suppress proliferation of human umbilical vein endothelia cells(HUVECs) induced by vascular endothelial growth factor(VEGF).Conclusion Human arresten gene was successfully cloned into the expression vector pRSET and expressed at high level in Escherichia coli.Purified and refolded arresten protein could effectively inhibit proliferation of vascular endothelia cells.2 refs.
基金This work was supported by the National Key Research and Development Program of China(No.2022YFC2702700)the National Natural Science Foundation of China(No.82171597)Clinical Research Plan of Shanghai Hospital Development Center(No.SHDC2020CR3077B).
文摘The regulation of spermatogonial proliferation and apoptosis is of great significance for maintaining spermatogenesis.The single-cell RNA sequencing(scRNA-seq)analysis of the testis was performed to identify genes upregulated in spermatogonia.Using scRNAseq analysis,we identified the spermatogonia upregulated gene origin recognition complex subunit 6(Orc6),which is involved in DNA replication and cell cycle regulation;its protein expression in the human and mouse testis was detected by western blot and immunofluorescence.To explore the potential function of Orc6 in spermatogonia,the C18-4 cell line was transfected with control or Orc6 siRNA.Subsequently,5-ethynyl-2-deoxyuridine(EdU)and terminal deoxynucleotidyl transferase dUTP nick end labeling(TUNEL)assays,flow cytometry,and western blot were used to evaluate its effects on proliferation and apoptosis.It was revealed that ORC6 could promote proliferation and inhibit apoptosis of C18-4 cells.Bulk RNA sequencing and bioinformatics analysis indicated that Orc6 was involved in the activation of wingless/integrated(Wnt)/β-catenin signaling.Western blot revealed that the expression ofβ-catenin protein and its phosphorylation(Ser675)were significantly decreased when silencing the expression of ORC6.Our findings indicated that Orc6 was upregulated in spermatogonia,whereby it regulated proliferation and apoptosis by activating Wnt/β-catenin signaling.
基金supported by Key Research and Development Program of Yunnan(202203AC100010)the National Natural Science Foundation of China(31760311,32160236,81830087,U2102203)+4 种基金the National Key Research and Development Program of China(2022YFC2601604,2018YFC2000400,2020YFA0112300)Spring City Plan:the Highlevel Talent Promotion and Training Project of Kunming(2022SCP001)the Yunnan Fundamental Research Projects(CY22624104,202101AS070050)the open project of State Key Laboratory of Genetic Resources and Evolution,Kunming Institute of Zoology,Chinese Academy of Sciences(GREKF17-01)Yunnan University's new round of"Double First-Class"Construction Project—For People’s Life and Health(CY22624104)。
文摘Human-specific insertions play important roles in human phenotypes and diseases.Here we reported a 446-bp insertion(Insert-446)in intron 11 of the TBC1D8B gene,located on chromosome X,and traced its origin to a portion of intron 6 of the EBF1 gene on chromosome 5.Interestingly,Insert-446 was present in the human Neanderthal and Denisovans genomes,and was fixed in humans after human-chimpanzee divergence.We have demonstrated that Insert-446 acts as an enhancer through binding transcript factors that promotes a higher expression of human TBC1D8B gene as compared with orthologs in macaques.In addition,over-expression TBC1D8B promoted cell proliferation and migration through“a dual finger”catalytic mechanism(Arg538 and Gln573)in the TBC domain in vitro and knockdown of TBC1D8B attenuated tumorigenesis in vivo.Knockout of Insert-446 prevented cell proliferation and migration in cancer and normal cells.Our results reveal that the human-specific Insert-446 promotes cell proliferation and migration by upregulating the expression of TBC1D8B gene.These findings provide a significant insight into the effects of human-specific insertions on evolution.
