Objective:Verrucous epidermal nevus(VEN),seborrheic keratosis(SK),verruca plana(VP),verruca vulgaris(VV),and nevus sebaceous(NS)are common verrucous proliferative skin diseases with similar clinical appearances,often ...Objective:Verrucous epidermal nevus(VEN),seborrheic keratosis(SK),verruca plana(VP),verruca vulgaris(VV),and nevus sebaceous(NS)are common verrucous proliferative skin diseases with similar clinical appearances,often posing diagnostic challenges.Dermoscopy and reflectance confocal microscopy(RCM)can aid in their differentiation,yet their specific features under these tools have not been systematically described.This study aims to summarize and analyze the dermoscopic and RCM features of VEN,SK,VP,VV,and NS.Methods:A total of 121 patients with histopathologically confirmed verrucous proliferative skin diseases were enrolled.Dermoscopy and RCM imaging was used to observe and analyze the microscopic features of these conditions.Results:Under dermoscopy,the 5 diseases displayed distinct characteristics:VEN typically showed gyriform structures;SK was characterized by gyriform structures,comedo-like openings,and milia-like cysts;VP and VV featured dotted vessels and frogspawn-like structures;NS presented as brownish-yellow globules.RCM revealed shared features such as hyperkeratosis and acanthosis across all 5 diseases.Specific features included gyriform structures and elongated rete ridges in VEN;pseudocysts and gyriform structures in SK;evenly distributed ring-like structures in VP;vacuolated cells and papillomatous proliferation in VV;and frogspawn-like structures in NS.Conclusion:These 5 verrucous proliferative skin conditions exhibit distinguishable features under both dermoscopy and RCM.The combination of these 2 noninvasive imaging modalities holds significant clinical value for the differential diagnosis of verrucous proliferative skin diseases.展开更多
Objective:Ovarian cancer(OC)ranks among the leading causes of mortality among the female cancers worldwide.Numerous studies have explored the development and progression of OC at multiple genetic regulatory levels.How...Objective:Ovarian cancer(OC)ranks among the leading causes of mortality among the female cancers worldwide.Numerous studies have explored the development and progression of OC at multiple genetic regulatory levels.However,relatively few studies have explored the impact of post-translational modifications(PTM)on OC progression,which is essential for uncovering new therapeutic targets.This study aimed to systematically identify the key PTM types involved in OCprogression,and to explore and evaluate their translational potential as therapeutic targets.Methods:First,we utilized multiple general PTM antibodies to compare gross PTM levels between normal ovarian and OC tissues from clinical females.After identifying lactylation as the PTM with the most significant differences,we selected representative samples for label-free mass spectrometry to identify specific lactylation sites.Next,we transfected A2780(OC)cells with either wild-type(WT)or mutant(K192A[Q])poly(ADP-ribose)polymerase 1(PARP1)conjugated to enhanced green fluorescent protein(EGFP)with a StrepⅡpeptide tag and assessed various cellular indexes related to cell proliferation(clonogenicity assay),migration(scratch wound healing assay),and reactive oxygen species levels.Results:Pan-lactylation was significantly upregulated in clinical OC samples,with PARP1 lactylation at K192 being one of the most common modifications.The growth and migration of A2780 cells were markedly suppressed by overexpressing PARP1-WT but not mutant PARP1.Overexpressing PARP1 significantly downregulated the phosphorylation of extracellular signal-regulated kinases 1/2(ERK1/2).Conclusion:This study uncovered a novel PTM of PARP1 in OC,lactylation,and demonstrated that lactylation at K192 is crucial in regulating OC cell growth and migration via the ERK1/2 pathway.Further investigations are required to elucidate the broader functional implications of PARP1 lactylation and its therapeutic potential.展开更多
Objective To investigate the effect of mucin 1(MUC1)on the proliferation and apoptosis of nasopharyngeal carcinoma(NPC)and its regulatory mechanism.Methods The 60 NPC and paired para-cancer normal tissues were collect...Objective To investigate the effect of mucin 1(MUC1)on the proliferation and apoptosis of nasopharyngeal carcinoma(NPC)and its regulatory mechanism.Methods The 60 NPC and paired para-cancer normal tissues were collected from October 2020 to July 2021 in Quanzhou First Hospital.The expression of MUC1 was measured by real-time quantitative PCR(qPCR)in the patients with PNC.The 5-8F and HNE1 cells were transfected with siRNA control(si-control)or siRNA targeting MUC1(si-MUC1).Cell proliferation was analyzed by cell counting kit-8 and colony formation assay,and apoptosis was analyzed by flow cytometry analysis in the 5-8F and HNE1 cells.The qPCR and ELISA were executed to analyze the levels of TNF-αand IL-6.Western blot was performed to measure the expression of MUC1,NFкB and apoptosis-related proteins(Bax and Bcl-2).Results The expression of MUC1 was up-regulated in the NPC tissues,and NPC patients with the high MUC1 expression were inclined to EBV infection,growth and metastasis of NPC.Loss of MUC1 restrained malignant features,including the proliferation and apoptosis,downregulated the expression of p-IкB、p-P65 and Bcl-2 and upregulated the expression of Bax in the NPC cells.Conclusion Downregulation of MUC1 restrained biological characteristics of malignancy,including cell proliferation and apoptosis,by inactivating NF-κB signaling pathway in NPC.展开更多
Objective:IgA nephropathy(IgAN)is the most common primary glomerular disease in China,but its pathogenesis remains unclear.This study aims to explore the regulatory role of the mammalian target of rapamycin(mTOR)signa...Objective:IgA nephropathy(IgAN)is the most common primary glomerular disease in China,but its pathogenesis remains unclear.This study aims to explore the regulatory role of the mammalian target of rapamycin(mTOR)signaling pathway in autophagy and mesangial proliferation during renal injury in IgA.Methods:The activity of mTOR and autophagy was evaluated in kidney samples from IgAN patients and in an IgAN mouse model induced by oral bovine serum albumin and carbon tetrachloride(CCl4)injection.mTOR inhibitors(rapamycin)and activators[bpV(phen)]were administered to the IgAN mouse model to observe the effects of mTOR on autophagy and renal lesions.In human mesangial cells treated with polymeric IgA1(p IgA1)and mTOR modulators,the expression and distribution of cell cycle proteins were assessed,along with the effects of mTOR on mesangial cell proliferation and autophagy.Results:Increased mTOR activity and decreased autophagy were observed in kidney tissues from IgAN patients and the mouse model,as evidenced by elevated phosphorylated mTOR(p-mTOR)levels and reduced LC3 expression.In the IgAN mouse model,rapamycin inhibited mTOR,restored autophagy,reduced mesangial IgA deposition,alleviated mesangial cell proliferation,and decreased proteinuria(all P<0.05).In contrast,bpV(phen)activated mTOR,further suppressed autophagy,exacerbated kidney damage,and increased proteinuria(all P<0.05).In vitro,p-IgA1 induced mesangial cell proliferation and inhibited autophagy,effects that were reversed by rapamycin and aggravated by bpV(phen)(all P<0.05).mTOR regulated mesangial cell proliferation by altering cell cycle distribution,with rapamycin inducing G1 phase arrest and bpV(phen)promoting cell cycle progression.Additionally,cyclinD1 expression in renal cortex was up-regulated in the IgAN mouse model,further increased by bpV(phen),and reduced by rapamycin(all P<0.05).Conclusion:Inhibition of the mTOR signaling pathway enhances renal autophagy,reduces mesangial cell proliferation,and improves renal injury in IgAN.展开更多
OBJECTIVE Peroxisome proliferator activated receptor alpha(PPARα)is an important protective factor in neurovascular diseases such as ischemic stroke.Although PPARαexpression is higher in neurons than astrocytes and ...OBJECTIVE Peroxisome proliferator activated receptor alpha(PPARα)is an important protective factor in neurovascular diseases such as ischemic stroke.Although PPARαexpression is higher in neurons than astrocytes and microglia,the pathophysiological functions of neuronal specific-PPARαin isch⁃emic stroke remains unknown.Here,we report that neuronal PPARαdeficiency is a key factor of neuronal injury.PPARαexpression markedly decreased in neurons after ischemic stroke.METHODS AND RESULTS Neuronal-specific PPARαknockout(NCKO)exacerbates neuronal damage and brain ischemic injury.PPARαdefi⁃ciency disrupts axonal microtubule organization and mitochondrial transport by decreasing the expression of dynein light chain Tctex-type 1(Dynlt1),which is implicated in cytoprotective role with damaged neurons.Furthermore,resto⁃ration of Dynlt1 expression in neurons of NCKO mice rescue mitochondrial transport disorder,cognitive deficits and brain ischemic injury asso⁃ciated with PPARαdeletion.CONCLUSION These results reveal a critical role for neuronal PPARαin ischemic brain injury by modulating axonal mitochondrial transportation.展开更多
OBJECTIVE Platinum compounds such as cisplatin and carboplatin are frequently used as the first-line chemotherapy for the treatment of the head and neck squamous cell carcinoma(HNSCC).In the present study,we investiga...OBJECTIVE Platinum compounds such as cisplatin and carboplatin are frequently used as the first-line chemotherapy for the treatment of the head and neck squamous cell carcinoma(HNSCC).In the present study,we investigated whether garcinol,apolyisoprenylated benzophenone can chemosensitize HNSCC to cisplatin.METHODS The effect of garcinol and cisplatin on HNSCC was assessed by MTT,Western blotting,real time PCR,FACS,immunohistochemistry,DNA binding assay and xenograft mouse model.RESULTS We found that garcinol inhibited the viability of a panel of diverse HNSCC cell lines,enhanced the apoptotic effect of cisplatin,suppressed constitutive as well as cisplatin-induced NF-κB activation,and downregulated the expression of various oncogenic gene products(cyclin D1,Bcl-2,survivin and VEGF).In vivo study showed that administration of garcinol alone(0.5 mg·kg-1,ip five times/week)significantly suppressed the growth of the tumor,and this effect was further increased by cisplatin.Both the markers of proliferation index(Ki-67)and microvessel density(CD31)were downregulated in tumor tissues by the combination of cisplatin and garcinol.The pharmacokinetic results of garcinol indicated that good systemic exposure was achievable after ip administration of garcinol at 0.5and 2mg·kg-1 with mean peak concentration(cmax)of 1825.4 and 6635.7nmol·L-1 in the mouse serum,respectively.CONCLUSION Overall,our results suggest that garcinol can indeed potentiate the effects of cisplatin by negative regulation of various inflammatory and proliferative biomarkers.展开更多
OBJECTIVE To investigate the role and mechanism of G protein-coupled receptor kinase 2(GRK2)involving in hepatocel ular carcinoma(HCC)progression.METHODS Cel Counting Kit 8 and tumor colony formation assay were design...OBJECTIVE To investigate the role and mechanism of G protein-coupled receptor kinase 2(GRK2)involving in hepatocel ular carcinoma(HCC)progression.METHODS Cel Counting Kit 8 and tumor colony formation assay were designed to detect HCC cell proliferation,wound healing assay was to detect HCC migration.The correlation between GRK2 and early growth response-1(EGR1)were detected by RT-PCR and real-time PCR assays.Co-immunoprecipitation and Western blot assay were adopted to detect the relationship between GRK2and insulin-like growth factor 1 receptor(IGF-1R)signaling pathway.RESULTS In this study we find that GRK2plays an inhibition role in IGF1-induced HCC cell proliferation and migration.Overexpression of GRK2 causes a decrease in EGR1 expression,while knockdown of GRK2 leads to the dramatically increase in EGR1 expression in the treatment of IGF1.Through co-immunoprecipitation and Western blot assay,we confirm that GRK2can interact with IGF-1R and inhibiting IGF1-induced activation of IGF1R signaling pathway.Silencing EGR1attenuates GRK2 overexpression-caused inhibition of cell proliferation,tumor colony number and migrationactivity,while overexpressing of EGR1 restores the antiproliferative and migratory effect by GRK2 overexpression in HCCLM3 cells.CONCLUSION Taken together,these results suggest that GRK2 may inhibit IGF1-induced HCC cell growth and migration through down-regulation of EGR1 and indicate that enforced GRK2 may offer a potential therapeutic approach against HCC.展开更多
The biocompatibility evaluation of calcium phosphate based biomaterials is performed by tissue culture in vitro model. Three kinds of bioceramic materials which are potential to deal with bone trauma and/or conduct ti...The biocompatibility evaluation of calcium phosphate based biomaterials is performed by tissue culture in vitro model. Three kinds of bioceramic materials which are potential to deal with bone trauma and/or conduct tissue growth are recommodated. The biological research results show that human and animal osteoblast cells anchor the materials surface in two hours in culture. Confocal laser scanning microscopy (CLSM) demonstrated the normal cell distribution and proliferation on both of dense and porous biomaterials. Hydroxyapatite and tricalcium phosphate stimulate cell proliferation. However, DNA and protein synthesis were considerably limited and the apoptosis phenomenon would be present on the hydroxyapatite (HA) materials by adding Al, Mg elements. Several important methods of biocompatibility evaluation of implant materials are described and the related biological molecular techniques such as tissue culture, cell transfection, cellular DNA stain, and Lowry assay are involved in the present research.展开更多
Since the diccovery of neural stem cells(NSCs)in the embryonic and adult mammalian central nerous system,it provided novel ideas forneurogenesis as the potential of proliferation and differentiation of NSCs.One of the...Since the diccovery of neural stem cells(NSCs)in the embryonic and adult mammalian central nerous system,it provided novel ideas forneurogenesis as the potential of proliferation and differentiation of NSCs.One of the ways to promote the clinical application of neural stem cells(NSCs)is searching effective methods which regulate the proliferation and differentiation.This is also a problem urgently to be solved in medical field.Plenty of earlier studies have shown that traditional chinese medicine can promote the proliferation and differentiation of NSCs by regulating the related signaling pathway in vivo and in vitro.The reports of Chinese and foreign literatures on regulating the proliferation and differentiation of neural stem cells in recent ten years and their target and signaling pathways is analyzed in this review.The traditional chinese medicine regulate proliferation and differentiation of NSCs by the signaling pathways of Notch,PI3K/Akt,Wnt/β-catenin,and GFs.And,those signaling pathways have cross-talk in the regulation progress.Moreover,some traditional Chinese medicine,such as astragalus,has a variety of active ingredients to regulate proliferation and differentiation of NSCs through different signaling pathways.However,to accelerate the clinical application of neural stem cells,the studies aboutthe proliferation and differentiation of NSCs and Chinese medicine should be further deepened,the mechanism of multiple targets and the comprehensive regulation function of traditional Chinese medicine should be clarified.展开更多
The experiment was conducted to study the effects and possible mechanism of GLP-2 on proliferation,metabolism and apoptosis of cultured enterocytes from a 28-d weaned piglet injured by exposure to β-conglycinin.A cel...The experiment was conducted to study the effects and possible mechanism of GLP-2 on proliferation,metabolism and apoptosis of cultured enterocytes from a 28-d weaned piglet injured by exposure to β-conglycinin.A cell damage model was established to investigate cell proliferation, metabolism and apoptosis by exposing primary cell cultures of intestinal epithelial cells(IEC) to 1.2 and 2.4 mg/mL β-conglycinin.A 2×3 factorial experiment was then used to study the effect of different GLP-2 concentrations of(1×10<sup>-9</sup>,1×10<sup>-8</sup> and 1×10<sup>-7</sup>mol/L),in combination with the two concentrations ofβ-conglycinin.Cells exposed to the allergenβ-conglycinin had decreased(P【0.05) MTT OD;decreased (P【0.01) protein retention and total protein content of cells;increased(P【0.01) LDH and caspase-3 activities and decreased(P【0.05) Na<sup>+</sup>,K<sup>+</sup>-ATPase activity.When GLP-2 was used in combination withβ-conglycinin,MTT OD,protein retention,total protein content and Na<sup>+</sup>,K<sup>+</sup>-ATPase activity significantly increased(P【0.05);LDH activity gradually decreased(P【0.05 or P【0.01) and Caspase-3 activity significantly decreased(P【0.01) with increasing concentrations of GLP-2.The results indicated thatβ-conglycinin had adverse effects on proliferation and integrity of IEC in vitro.GLP-2 relieved or prevented the adverse effects ofβ-conglycinin on proliferation and integrity of IEC by regulating Na<sup>+</sup>,K<sup>+</sup>- ATPase and Caspase-3 activities,and consequently affecting cell metabolism.展开更多
The effects of FSH on the proliferation of sertoli cells of new born calves were studied in order to provide some data for theoretical research and practical use of spermatogenesis in vitro. Different concentrations o...The effects of FSH on the proliferation of sertoli cells of new born calves were studied in order to provide some data for theoretical research and practical use of spermatogenesis in vitro. Different concentrations of FSH （0, 0.01, 0.02, 0.04, and 0.08 IU· mL^-1） were taken to treat bovine sertoli cells in vitro culture, the number of sertoli cells and the expression of seven genes were determined at 6, 12 and 24 h after FSH treatments. FSH could significantly promote the proliferation of in vitro cultured sertoli cells. FSH had no significant effects on the expression of CDC25A and could significantly improve the expression of CDC25B. 0.04 IU· mL^-1 and 0.08 IU· mL^-1 FSH treatments decreased the expression of CDC25C at 12 h. 0.08 IU· mL^-1 FSH treatment decreased the expression of CDC25C at 24 h. 0.04 IU. mLI FSH could significantly decrease the expression of GSK-3β and improve the expression of β-catenin at 6, 12 and 24 h. 0.02, 0.04 and 0.08 IU· mL^-1 FSH treatments enhanced the expressions of CYCLIND1 and C-MYC. In conclusion, FSH promoted the proliferation of sertoli cells and 0.04 IU· mL^-1 FSH concentration could significantly promote the proliferation of in vitro cultured sertoli cells. FSH promoted the proliferation of sertoli cells by CDC25B and WNT/ β-eatenin and CDC25B might be the key regulator to the proliferating rate of sertoli cells of bovine calf.展开更多
Objective:Arterial stiffening occurs in the progression of natural aging and cardiovascular diseases.Vascular smooth muscle cells(VSMCs),the major components of vascular walls,which largely contribute to the pathophys...Objective:Arterial stiffening occurs in the progression of natural aging and cardiovascular diseases.Vascular smooth muscle cells(VSMCs),the major components of vascular walls,which largely contribute to the pathophysiological states of blood vessels,are influenced by environmental cues of blood vessels reciprocally as well.Consistently,the increased proliferation of VSMCs has been reported to be observed in stiffening blood vessel and on rigid substrates,the underlying mechanism of which remains not yet fully clarified.Our previous work has demonstrated that Ca2+-activated K+(IKCa)channel participates in the stiff substrate-induced vascular smooth muscle cell(VSMC)proliferation.In the present work,from the standpoint of calcium entry and extracellular regulated protein kinases 1 and 2(ERK 1/2)activation,we further investigated the underlying mechanisms by which IKCa channels functions in the process mentioned above.Methods Soft(0.21 MPa)and stiff(1.72 MPa)PDMS substrates where VSMCs were seeded after coated with fibronectin(FN),were fabricated through the blending of sylgard 184 gel and sylgard 527 gel.After that,intracellular calcium level of VSMCs was compared with or without the treatment of IKCa specific blocker,TRAM-34,using a calcium-sensitive dye,fluo 4-AM.1mM Ethylene glycol-bis(2-aminoethyl ether)-N,N,N’,N’-tetraacetic acid(EGTA)was added into the culture media for the removal of the extracellular calcium ions as well as cell counting kit-8(CCK-8)assay was applied,which is to explore the role that calcium ion entry played in proliferation process.The activation level of ERK 1/2 was described by the expression level of phospho-ERK 1/2 using western blotting with or without TRAM-34 treatment.The role of the activation of ERK1/2 in VSMC proliferation was examined by CCK-8 assay with or without the treatment of PD98095,an ERK1/2 inhibitor.Results Compared with soft substrate,stiff substrate caused an increase of intracellular calcium level,which was attenuated by IKCa blockade.In addition,compared with soft substrate,stiff substrate also caused an activation of ERK1/2,which was significantly suppressed by IKCa blockade.Furthermore,extracellular calcium ion reduction by adding EGTA significantly inhibited the stiff substrate-induced VSMC proliferation,which whereas had no effect on VSMC proliferation on soft substrate.Finally,ERK1/2 inhibition had similar inhibitory effect on stiff substrate-induced proliferation.Conclusions Stiff substrate causes an IKCa channel-mediated calcium entry and ERK1/2 activation,both of which play important roles in stiff substrate-induced VSMC proliferation.Combined the previous results that IKCa channel participated in stiff substrate-induced VSMC proliferation,our present work suggests that IKCa channel functioned in the proliferation process through mediating calcium entry and subsequent ERK1/2 activations.These findings provide a new insight into how substrate stiffness regulates VSMC proliferation,and additional considerations for vascular tissue engineering and vascular disease treatment.展开更多
Aim：Investigate the influence of IgD on T/B cell activation and construct hIgD-Fc-Ig fusion protein to competitive inhibition IgD binding with IgDR. Methods differences of mIgD and IgD-R level between different T/B w...Aim：Investigate the influence of IgD on T/B cell activation and construct hIgD-Fc-Ig fusion protein to competitive inhibition IgD binding with IgDR. Methods differences of mIgD and IgD-R level between different T/B was detected by ELISA. Human IgD-Fc-IgGI-Fc sequence T/B cells were sorted by magnetic cell sorting. The cell subtypes were detected by FCM. Serum IgD level was amplified by cross-PCR and then subcloned into PET28a（ ＋ ） empty vector. After prokaryotic expression through escherichia？ coli, we obtained the hIgD-Fc-Ig fu- sion protein by affinity chromatograph. Western blot was used to identify the hIgD-Fc-Ig fusion protein. Human pe- ripheral blood monouclear cells （PBMC） and fibroblast like synoviocytes （FLS） proliferation were detected using a cell counting kit-8 （ CCK-8 ）. Results The percentage of CD3 ＋/CD4 ＋ , CD3 ＋/IgD ＋ , CD3 ＋/CD4 ＋/IgD ＋ , CD3 ＋/IgD-R ＋ and CD3 ＋/CD4 ＋/IgD-R ＋ cells increased significantly in RA patients comparing to healthy people. IgD can stimulate PBMC proliferation. IgD （1, 3, 10, 30 μg ＂ ml^-1） stimulate PBMC proliferation significantly after 24 h. We obtained stable and active hlgD-Fc-Ig fusion protein. The hlgD-Fc-Ig fusion protein showed no effect on PBMC proliferation. But it could downregulate human IgD protein promoting proliferation effects in human PBMC. Conclusion This result suggests that IgD and IgDR play an important role on T/B cell activation in RA patients and the hIgD-Fc-Ig fusion protein may competitively inhibit IgD＇s function and may play an therapeutic role in autoimmune diseases.展开更多
Objective Mechanical stretch regulates mesenchymal stem cell(MSC)function,which much more emphasis has been placed its prolonged effect on lineage differentiation,especially osteogenic differentiation.In contrast,ther...Objective Mechanical stretch regulates mesenchymal stem cell(MSC)function,which much more emphasis has been placed its prolonged effect on lineage differentiation,especially osteogenic differentiation.In contrast,there are few reports about its short term effect on MSC proliferation.In the present study,effects of short-term mechanical stretch on the proliferation and osteogenic differentiation of mesenchymal stem cell proliferation were investigated.In addition,the stretchinfluenced expression of transient receptor potential cation channel,subfamily C,member 1(TRPC1)was also investigated due to its mechanosensitivity and positive correlation with MSC proliferation.Methods MSCs,harvested from rat bone marrow,were seeded on collagen l-coated silicone chamber and exposed to mechanical stretch with various magnitude(0%,5%,10%and 15%)or various duration(2 h,6 h,12 h and 24 h).Cell proliferation was examined by cell counting kit-8(CCK-8)assay and cell cycle analysis.The gene and protein expression of two makers for osteogenic differentiation,collagen I and Cbfα1,and TRPC1 were determined by RT-PCR and western blotting,respectively.BMSC were harvested,and total RNA was isolated with Trizol reagent.A 2μg portion of total RNA was synthesized to cDNA according to the manufacturer s instructions.cDNA was used as a template for each PCR amplification.BMSC were solubilized in RIPA lysis buffer on ice for 30 min.Phenylmethane sulfonyl fluoride(PMSF)was added to avoid proteolysis.Equal portions of the cell lysates were separated on 10%sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE)and transferred to polyvinylidene fluoride(PVDF)membranes.The membranes were incubated with primary antibodies to TRPC1 and GAPDH at4℃overnight to identify the specific proteins.The PVDF membranes were washed with TBST three times and incubated with a horseradish peroxidase(HRP)-conjugated secondary antibody.Immunoreactive bands were visualized using an enhanced chemiluminescent(ECL)system.Results The OD value for the three stretch cases(5%-15%)was increased^1.4-fold compared with that for control(0%).There is no significantly difference among the three stretch cases.The percentage of cells for three stretch cases were more in the S phase but less in the G0/G1 phase compared to those for control.The cell cycle distribution still had no significant difference among the three stretch cases.In addition,the stretch application for 24 h didn’t affect the gene or protein level for collagen I and Cbfα1 compared with those of control.Application of 10%stretch for 2 h didn’t affect TRPC1 gene or protein expression,but that application for 6-24 h significantly up-regulated TRPCl gene and protein level.That increase exhibited a stretch duration-independent manner.Conclusions Short-term mechanical stretch promoted MSC proliferation in a magnitude-independent manner,whereas had no effect on its oesteogenic differentiation.Paralleled to which,TRPC1 was up-regulated by stretch,implying that TRPCl may be implicated in that proliferation courses.Future work is still needed to confirm whether TRPC1 participates in that stretch-induced MSC proliferation using RPC1 blockade or knockout.展开更多
OBJECTIVE This study aimed to investigate the influence of IgD on T/B cell activation and construct h IgD-Fc-Igfusion protein to competitive inhibition IgD binding with IgDR.METHODS T/B cells were sorted by magnetic c...OBJECTIVE This study aimed to investigate the influence of IgD on T/B cell activation and construct h IgD-Fc-Igfusion protein to competitive inhibition IgD binding with IgDR.METHODS T/B cells were sorted by magnetic cell sorting.The differences of m IgD and IgD-R level between different T/B cell subtypes were detected by FCM.Serum IgD level was detected by ELISA.Human IgD-Fc-IgG1-Fc sequence was amplified by cross-PCR and then subcloned into PET28 a(+) empty vector.After prokaryotic expression through escherichia coli,we obtained the h IgD-Fc-Igfusion protein by affinity chromatograph.Western blot was used to identify the h IgD-Fc-Igfusion protein.Human peripheral blood monouclear cells(PBMC) and fibroblast like synoviocytes(FLS) proliferation were detected using a cell counting kit-8(CCK-8).RESULTS The percentage of CD3^+/CD4^+,CD3^+/IgD^+,CD3^+/CD4^+/IgD^+,CD3^+/IgD-R+and CD3^+/CD4^+/IgD-R+cells increased significantly in RA patients comparing to healthy people.IgD can stimulate PBMC proliferation.IgD(1,3,10,30 μg·mL^(-1)) stimulate PBMC proliferation significantly after 24 h.We obtained stable and active h IgD-Fc-Igfusion protein.The h IgD-Fc-Igfusion protein showed no effect on PBMC proliferation.But it could downregulate human IgD protein promoting proliferation effects in human PBMC.CONCLUSION This result suggests that IgD and IgDR play an important role on T/B cell activation in RA patients and the h IgD-Fc-Igfusion protein may competitively inhibit IgD′s function and may play an therapeutic role in autoimmune diseases.展开更多
OBJECTIVE Cervical cancer is the third most malignant tumor in the world.Farnesoid X receptor(FXR) is a member of nuclear receptor superfamily.It is highly expressed in liver,kidney and small intestine,while it showed...OBJECTIVE Cervical cancer is the third most malignant tumor in the world.Farnesoid X receptor(FXR) is a member of nuclear receptor superfamily.It is highly expressed in liver,kidney and small intestine,while it showed low expression level in other tissues.It not only plays an important role in the metabolism of bile acids and sugars,but also in the production of chronic inflammation in the early stage of cancer,the proliferation and migration of tumor.Compared with the normal tissue,the expression of FXR in most tumor tissues decreased.But there is no correlation between cervical cancer and FXR.So we aimed to find out the relationship between FXR and cervical cancer.METHODS A clinical study using q PCR,western blot and immunohistochemistry detected the expression of FXR in tumor tissues and normal tissues of clinical patients.FXR was activated by agonists or over-expressed by lentivirus.MTT,clone formation and flow cytometry were used to detect the relationship between FXR and proliferation of cervical cell lines.Tumor growth ability of FXR was detected by nude mice tumorigenicity.The interaction between FXR and CDKN2A-p14^(ARF)-MDM2-p53 pathway was detected by q PCR,Western blot and immunohistochemistry.RESULTS FXR was decreased in cancer tissues compared to normal control.Activation of FXR by agonist or constitutively-over-expression of FXR inhibited cervical cell proliferation.Over-expressed FXR attenuated Caski,Hela and Siha xenograft tumor growth in nude mice compared with control.Over-expression of FXR caused G1 cell-cycle arresting and up-regulated CDKN2A-p14^(ARF)-MDM2-p53 pathway.CONCLUSION FXR inhibits cervical cancer cell proliferation and cervical tumorigenicity which is related to CDKN2A-p14^(ARF)-MDM2-p53 pathway.Activation or overexpression of FXR may be a potential target for the treatment of cervical cancer.展开更多
OBJECTIVE To investigate the inhibitory effects of Spatholobus Suberectus Column Extract(SSCE)on estrogen receptor positive(ER+)breast cancer cel MCF-7and its possible molecular mechanism.METHODS MCF-7cells were cultu...OBJECTIVE To investigate the inhibitory effects of Spatholobus Suberectus Column Extract(SSCE)on estrogen receptor positive(ER+)breast cancer cel MCF-7and its possible molecular mechanism.METHODS MCF-7cells were cultured without estrogen and with 17-β-estrogen(10-8mol·L-1),respectively,then treated with SSCE(0,40,80,160,320μg·m L-1).MTT assay was employed to evaluate cell viability.Flow cytometry assays were performed to underlying apoptosis and detecting cel cycle of MCF-7 cells treated with SSCE(0,80,160,320μg·mL-1).Wound healing assays was conducted to detect the migration ability.Dual luciferase reporter system was used to detect the activity of p-ERα,p-ERβpresented in intra-nuclear estrogen response element(ERE).Western blotting assay was employed to identify the expression of protein such as Bax,Bcl-2,p-ERα,p-ERβ,ERK1/2,p-ERK1/2,AKT,p-AKT,m TOR,p-m TOR,PI3K,p-PI3K.RESULTS It showed that SSCE(80,160and 320μg·mL-1)significantly decreased the viability of MCF-7.SSCE also triggered apoptosis,arrested cell cycle at G0/G1phase,inhibited migration.Dual luciferase reporter system showed that SSCE suppressed intra-nuclear p-ER activity,Western blotting analysis confirmed that SSCE did repress the expression of phosphorylated-ER alpha(p-ERα),ERK1/2,p-ERK1/2,AKT,p-AKT,pmT OR,PI3K,p-PI3K,which indicate that SSCE suppress MAPK PI3K/AKT signal pathway.CONCLUSION Our result showed that SSCE cause ER+MCF-7 cells apoptosis,G0/G1phase arresting,migration decreasing,via hypo-active of ER,suppress MAPK PI3K/AKT pathway.展开更多
OBJECTIVE To investigate the effect of gap junctions on the anti-tumor function induced by mi R-34a in glioma U87 cells.METHODS 1.Transfection(miR-34a mimics were transfected into glioma cells to upregulate their expr...OBJECTIVE To investigate the effect of gap junctions on the anti-tumor function induced by mi R-34a in glioma U87 cells.METHODS 1.Transfection(miR-34a mimics were transfected into glioma cells to upregulate their expression);2.Co-culture assay(U87cells were transfected with mi R-34a co-cultured with U87 cells that was transfected PCMV-eG FP plasmid);3.Flow cytometry analysis(select the e GFP labed U87 cells);4.RNA isolation and real-time PCR;5.CCK-8 assay;6.Western blotting.RESULTS Mi R-34a mimics transfered between the U87 cells.Parachute assay showed that GJ inhibition(CBX and 18-α-GA)can decrease mi R-34a expression than co-culture group.RA and galanglin enhanced mi R-34a expression than co-culture group.Mi R-34a relative expression reduced after co-culture,while gap junctions composed of Cx43 were down-regulated by sh RNA.Transfected with mi R-34a mimics reduced the survival of U87 cells in a dose-dependent manner.To more specifically establish the role of GJIC in mi R-34a induced growth inhibition of U87 cells,si RNA was used to knockdown the expression of Cx43,the dominant connexin expressed in U87 cells.CCK-8 assay showed that siR NAs have no effect on cell growth,but they could aggravate the growth inhibition of miR-34a to U87 cels.CONCLUSION Gap junctions enhance the antiproliferative effect of miR NA-34a in glioma cells.展开更多
基金supported by the Project of Health Committee of Hunan Province(D202304128868),China.
文摘Objective:Verrucous epidermal nevus(VEN),seborrheic keratosis(SK),verruca plana(VP),verruca vulgaris(VV),and nevus sebaceous(NS)are common verrucous proliferative skin diseases with similar clinical appearances,often posing diagnostic challenges.Dermoscopy and reflectance confocal microscopy(RCM)can aid in their differentiation,yet their specific features under these tools have not been systematically described.This study aims to summarize and analyze the dermoscopic and RCM features of VEN,SK,VP,VV,and NS.Methods:A total of 121 patients with histopathologically confirmed verrucous proliferative skin diseases were enrolled.Dermoscopy and RCM imaging was used to observe and analyze the microscopic features of these conditions.Results:Under dermoscopy,the 5 diseases displayed distinct characteristics:VEN typically showed gyriform structures;SK was characterized by gyriform structures,comedo-like openings,and milia-like cysts;VP and VV featured dotted vessels and frogspawn-like structures;NS presented as brownish-yellow globules.RCM revealed shared features such as hyperkeratosis and acanthosis across all 5 diseases.Specific features included gyriform structures and elongated rete ridges in VEN;pseudocysts and gyriform structures in SK;evenly distributed ring-like structures in VP;vacuolated cells and papillomatous proliferation in VV;and frogspawn-like structures in NS.Conclusion:These 5 verrucous proliferative skin conditions exhibit distinguishable features under both dermoscopy and RCM.The combination of these 2 noninvasive imaging modalities holds significant clinical value for the differential diagnosis of verrucous proliferative skin diseases.
文摘Objective:Ovarian cancer(OC)ranks among the leading causes of mortality among the female cancers worldwide.Numerous studies have explored the development and progression of OC at multiple genetic regulatory levels.However,relatively few studies have explored the impact of post-translational modifications(PTM)on OC progression,which is essential for uncovering new therapeutic targets.This study aimed to systematically identify the key PTM types involved in OCprogression,and to explore and evaluate their translational potential as therapeutic targets.Methods:First,we utilized multiple general PTM antibodies to compare gross PTM levels between normal ovarian and OC tissues from clinical females.After identifying lactylation as the PTM with the most significant differences,we selected representative samples for label-free mass spectrometry to identify specific lactylation sites.Next,we transfected A2780(OC)cells with either wild-type(WT)or mutant(K192A[Q])poly(ADP-ribose)polymerase 1(PARP1)conjugated to enhanced green fluorescent protein(EGFP)with a StrepⅡpeptide tag and assessed various cellular indexes related to cell proliferation(clonogenicity assay),migration(scratch wound healing assay),and reactive oxygen species levels.Results:Pan-lactylation was significantly upregulated in clinical OC samples,with PARP1 lactylation at K192 being one of the most common modifications.The growth and migration of A2780 cells were markedly suppressed by overexpressing PARP1-WT but not mutant PARP1.Overexpressing PARP1 significantly downregulated the phosphorylation of extracellular signal-regulated kinases 1/2(ERK1/2).Conclusion:This study uncovered a novel PTM of PARP1 in OC,lactylation,and demonstrated that lactylation at K192 is crucial in regulating OC cell growth and migration via the ERK1/2 pathway.Further investigations are required to elucidate the broader functional implications of PARP1 lactylation and its therapeutic potential.
文摘Objective To investigate the effect of mucin 1(MUC1)on the proliferation and apoptosis of nasopharyngeal carcinoma(NPC)and its regulatory mechanism.Methods The 60 NPC and paired para-cancer normal tissues were collected from October 2020 to July 2021 in Quanzhou First Hospital.The expression of MUC1 was measured by real-time quantitative PCR(qPCR)in the patients with PNC.The 5-8F and HNE1 cells were transfected with siRNA control(si-control)or siRNA targeting MUC1(si-MUC1).Cell proliferation was analyzed by cell counting kit-8 and colony formation assay,and apoptosis was analyzed by flow cytometry analysis in the 5-8F and HNE1 cells.The qPCR and ELISA were executed to analyze the levels of TNF-αand IL-6.Western blot was performed to measure the expression of MUC1,NFкB and apoptosis-related proteins(Bax and Bcl-2).Results The expression of MUC1 was up-regulated in the NPC tissues,and NPC patients with the high MUC1 expression were inclined to EBV infection,growth and metastasis of NPC.Loss of MUC1 restrained malignant features,including the proliferation and apoptosis,downregulated the expression of p-IкB、p-P65 and Bcl-2 and upregulated the expression of Bax in the NPC cells.Conclusion Downregulation of MUC1 restrained biological characteristics of malignancy,including cell proliferation and apoptosis,by inactivating NF-κB signaling pathway in NPC.
基金supported by the Key Funding Project of Hunan Provincial Health Commission(A202303057091)the Nephrology Medical Development Research Fund Project(20220316ZN),China.
文摘Objective:IgA nephropathy(IgAN)is the most common primary glomerular disease in China,but its pathogenesis remains unclear.This study aims to explore the regulatory role of the mammalian target of rapamycin(mTOR)signaling pathway in autophagy and mesangial proliferation during renal injury in IgA.Methods:The activity of mTOR and autophagy was evaluated in kidney samples from IgAN patients and in an IgAN mouse model induced by oral bovine serum albumin and carbon tetrachloride(CCl4)injection.mTOR inhibitors(rapamycin)and activators[bpV(phen)]were administered to the IgAN mouse model to observe the effects of mTOR on autophagy and renal lesions.In human mesangial cells treated with polymeric IgA1(p IgA1)and mTOR modulators,the expression and distribution of cell cycle proteins were assessed,along with the effects of mTOR on mesangial cell proliferation and autophagy.Results:Increased mTOR activity and decreased autophagy were observed in kidney tissues from IgAN patients and the mouse model,as evidenced by elevated phosphorylated mTOR(p-mTOR)levels and reduced LC3 expression.In the IgAN mouse model,rapamycin inhibited mTOR,restored autophagy,reduced mesangial IgA deposition,alleviated mesangial cell proliferation,and decreased proteinuria(all P<0.05).In contrast,bpV(phen)activated mTOR,further suppressed autophagy,exacerbated kidney damage,and increased proteinuria(all P<0.05).In vitro,p-IgA1 induced mesangial cell proliferation and inhibited autophagy,effects that were reversed by rapamycin and aggravated by bpV(phen)(all P<0.05).mTOR regulated mesangial cell proliferation by altering cell cycle distribution,with rapamycin inducing G1 phase arrest and bpV(phen)promoting cell cycle progression.Additionally,cyclinD1 expression in renal cortex was up-regulated in the IgAN mouse model,further increased by bpV(phen),and reduced by rapamycin(all P<0.05).Conclusion:Inhibition of the mTOR signaling pathway enhances renal autophagy,reduces mesangial cell proliferation,and improves renal injury in IgAN.
文摘OBJECTIVE Peroxisome proliferator activated receptor alpha(PPARα)is an important protective factor in neurovascular diseases such as ischemic stroke.Although PPARαexpression is higher in neurons than astrocytes and microglia,the pathophysiological functions of neuronal specific-PPARαin isch⁃emic stroke remains unknown.Here,we report that neuronal PPARαdeficiency is a key factor of neuronal injury.PPARαexpression markedly decreased in neurons after ischemic stroke.METHODS AND RESULTS Neuronal-specific PPARαknockout(NCKO)exacerbates neuronal damage and brain ischemic injury.PPARαdefi⁃ciency disrupts axonal microtubule organization and mitochondrial transport by decreasing the expression of dynein light chain Tctex-type 1(Dynlt1),which is implicated in cytoprotective role with damaged neurons.Furthermore,resto⁃ration of Dynlt1 expression in neurons of NCKO mice rescue mitochondrial transport disorder,cognitive deficits and brain ischemic injury asso⁃ciated with PPARαdeletion.CONCLUSION These results reveal a critical role for neuronal PPARαin ischemic brain injury by modulating axonal mitochondrial transportation.
基金The project supported by National Medical Research Council of Singapore and NUS Academic Research fund
文摘OBJECTIVE Platinum compounds such as cisplatin and carboplatin are frequently used as the first-line chemotherapy for the treatment of the head and neck squamous cell carcinoma(HNSCC).In the present study,we investigated whether garcinol,apolyisoprenylated benzophenone can chemosensitize HNSCC to cisplatin.METHODS The effect of garcinol and cisplatin on HNSCC was assessed by MTT,Western blotting,real time PCR,FACS,immunohistochemistry,DNA binding assay and xenograft mouse model.RESULTS We found that garcinol inhibited the viability of a panel of diverse HNSCC cell lines,enhanced the apoptotic effect of cisplatin,suppressed constitutive as well as cisplatin-induced NF-κB activation,and downregulated the expression of various oncogenic gene products(cyclin D1,Bcl-2,survivin and VEGF).In vivo study showed that administration of garcinol alone(0.5 mg·kg-1,ip five times/week)significantly suppressed the growth of the tumor,and this effect was further increased by cisplatin.Both the markers of proliferation index(Ki-67)and microvessel density(CD31)were downregulated in tumor tissues by the combination of cisplatin and garcinol.The pharmacokinetic results of garcinol indicated that good systemic exposure was achievable after ip administration of garcinol at 0.5and 2mg·kg-1 with mean peak concentration(cmax)of 1825.4 and 6635.7nmol·L-1 in the mouse serum,respectively.CONCLUSION Overall,our results suggest that garcinol can indeed potentiate the effects of cisplatin by negative regulation of various inflammatory and proliferative biomarkers.
基金The project supported by National Natural Science Foundation of China(81502123,81330081,81202596)Natural Science Foundation of Anhui Province(1308085QH130)+3 种基金Anhui Province Natural Science Foundation in University(KJ2014A119)Grants for Scientific Research of BSKY from Anhui Medical University(XJ201212)Specialized Research Fund for the Doctoral Program of Higher Education(20113420120006,20123420110003)Program for Tackling Key Problems in Science and Technology by Anhui Province(1301042098)
文摘OBJECTIVE To investigate the role and mechanism of G protein-coupled receptor kinase 2(GRK2)involving in hepatocel ular carcinoma(HCC)progression.METHODS Cel Counting Kit 8 and tumor colony formation assay were designed to detect HCC cell proliferation,wound healing assay was to detect HCC migration.The correlation between GRK2 and early growth response-1(EGR1)were detected by RT-PCR and real-time PCR assays.Co-immunoprecipitation and Western blot assay were adopted to detect the relationship between GRK2and insulin-like growth factor 1 receptor(IGF-1R)signaling pathway.RESULTS In this study we find that GRK2plays an inhibition role in IGF1-induced HCC cell proliferation and migration.Overexpression of GRK2 causes a decrease in EGR1 expression,while knockdown of GRK2 leads to the dramatically increase in EGR1 expression in the treatment of IGF1.Through co-immunoprecipitation and Western blot assay,we confirm that GRK2can interact with IGF-1R and inhibiting IGF1-induced activation of IGF1R signaling pathway.Silencing EGR1attenuates GRK2 overexpression-caused inhibition of cell proliferation,tumor colony number and migrationactivity,while overexpressing of EGR1 restores the antiproliferative and migratory effect by GRK2 overexpression in HCCLM3 cells.CONCLUSION Taken together,these results suggest that GRK2 may inhibit IGF1-induced HCC cell growth and migration through down-regulation of EGR1 and indicate that enforced GRK2 may offer a potential therapeutic approach against HCC.
文摘The biocompatibility evaluation of calcium phosphate based biomaterials is performed by tissue culture in vitro model. Three kinds of bioceramic materials which are potential to deal with bone trauma and/or conduct tissue growth are recommodated. The biological research results show that human and animal osteoblast cells anchor the materials surface in two hours in culture. Confocal laser scanning microscopy (CLSM) demonstrated the normal cell distribution and proliferation on both of dense and porous biomaterials. Hydroxyapatite and tricalcium phosphate stimulate cell proliferation. However, DNA and protein synthesis were considerably limited and the apoptosis phenomenon would be present on the hydroxyapatite (HA) materials by adding Al, Mg elements. Several important methods of biocompatibility evaluation of implant materials are described and the related biological molecular techniques such as tissue culture, cell transfection, cellular DNA stain, and Lowry assay are involved in the present research.
基金supported by National Natural Science Foundation of China(81473549)Fundamental Research Funds for Central Universities(XDJK2017E158)
文摘Since the diccovery of neural stem cells(NSCs)in the embryonic and adult mammalian central nerous system,it provided novel ideas forneurogenesis as the potential of proliferation and differentiation of NSCs.One of the ways to promote the clinical application of neural stem cells(NSCs)is searching effective methods which regulate the proliferation and differentiation.This is also a problem urgently to be solved in medical field.Plenty of earlier studies have shown that traditional chinese medicine can promote the proliferation and differentiation of NSCs by regulating the related signaling pathway in vivo and in vitro.The reports of Chinese and foreign literatures on regulating the proliferation and differentiation of neural stem cells in recent ten years and their target and signaling pathways is analyzed in this review.The traditional chinese medicine regulate proliferation and differentiation of NSCs by the signaling pathways of Notch,PI3K/Akt,Wnt/β-catenin,and GFs.And,those signaling pathways have cross-talk in the regulation progress.Moreover,some traditional Chinese medicine,such as astragalus,has a variety of active ingredients to regulate proliferation and differentiation of NSCs through different signaling pathways.However,to accelerate the clinical application of neural stem cells,the studies aboutthe proliferation and differentiation of NSCs and Chinese medicine should be further deepened,the mechanism of multiple targets and the comprehensive regulation function of traditional Chinese medicine should be clarified.
基金supported by Program for Changjiang Scholars and Innovative Reseach Team in University (IRTO 555)Applied Basic Research(045Y029-031) of Sichuan Province,People's Republic of China
文摘The experiment was conducted to study the effects and possible mechanism of GLP-2 on proliferation,metabolism and apoptosis of cultured enterocytes from a 28-d weaned piglet injured by exposure to β-conglycinin.A cell damage model was established to investigate cell proliferation, metabolism and apoptosis by exposing primary cell cultures of intestinal epithelial cells(IEC) to 1.2 and 2.4 mg/mL β-conglycinin.A 2×3 factorial experiment was then used to study the effect of different GLP-2 concentrations of(1×10<sup>-9</sup>,1×10<sup>-8</sup> and 1×10<sup>-7</sup>mol/L),in combination with the two concentrations ofβ-conglycinin.Cells exposed to the allergenβ-conglycinin had decreased(P【0.05) MTT OD;decreased (P【0.01) protein retention and total protein content of cells;increased(P【0.01) LDH and caspase-3 activities and decreased(P【0.05) Na<sup>+</sup>,K<sup>+</sup>-ATPase activity.When GLP-2 was used in combination withβ-conglycinin,MTT OD,protein retention,total protein content and Na<sup>+</sup>,K<sup>+</sup>-ATPase activity significantly increased(P【0.05);LDH activity gradually decreased(P【0.05 or P【0.01) and Caspase-3 activity significantly decreased(P【0.01) with increasing concentrations of GLP-2.The results indicated thatβ-conglycinin had adverse effects on proliferation and integrity of IEC in vitro.GLP-2 relieved or prevented the adverse effects ofβ-conglycinin on proliferation and integrity of IEC by regulating Na<sup>+</sup>,K<sup>+</sup>- ATPase and Caspase-3 activities,and consequently affecting cell metabolism.
基金Supported by the National International Scientific and Technological Cooperation Project(2011DFA30760-2-1)Fund of Key Lab.of Northeast Agricultural University,Harbin,China(GXZDSYS-2012-07)
文摘The effects of FSH on the proliferation of sertoli cells of new born calves were studied in order to provide some data for theoretical research and practical use of spermatogenesis in vitro. Different concentrations of FSH （0, 0.01, 0.02, 0.04, and 0.08 IU· mL^-1） were taken to treat bovine sertoli cells in vitro culture, the number of sertoli cells and the expression of seven genes were determined at 6, 12 and 24 h after FSH treatments. FSH could significantly promote the proliferation of in vitro cultured sertoli cells. FSH had no significant effects on the expression of CDC25A and could significantly improve the expression of CDC25B. 0.04 IU· mL^-1 and 0.08 IU· mL^-1 FSH treatments decreased the expression of CDC25C at 12 h. 0.08 IU· mL^-1 FSH treatment decreased the expression of CDC25C at 24 h. 0.04 IU. mLI FSH could significantly decrease the expression of GSK-3β and improve the expression of β-catenin at 6, 12 and 24 h. 0.02, 0.04 and 0.08 IU· mL^-1 FSH treatments enhanced the expressions of CYCLIND1 and C-MYC. In conclusion, FSH promoted the proliferation of sertoli cells and 0.04 IU· mL^-1 FSH concentration could significantly promote the proliferation of in vitro cultured sertoli cells. FSH promoted the proliferation of sertoli cells by CDC25B and WNT/ β-eatenin and CDC25B might be the key regulator to the proliferating rate of sertoli cells of bovine calf.
基金supported by the National Natural Science Foundation of China ( 11872010)
文摘Objective:Arterial stiffening occurs in the progression of natural aging and cardiovascular diseases.Vascular smooth muscle cells(VSMCs),the major components of vascular walls,which largely contribute to the pathophysiological states of blood vessels,are influenced by environmental cues of blood vessels reciprocally as well.Consistently,the increased proliferation of VSMCs has been reported to be observed in stiffening blood vessel and on rigid substrates,the underlying mechanism of which remains not yet fully clarified.Our previous work has demonstrated that Ca2+-activated K+(IKCa)channel participates in the stiff substrate-induced vascular smooth muscle cell(VSMC)proliferation.In the present work,from the standpoint of calcium entry and extracellular regulated protein kinases 1 and 2(ERK 1/2)activation,we further investigated the underlying mechanisms by which IKCa channels functions in the process mentioned above.Methods Soft(0.21 MPa)and stiff(1.72 MPa)PDMS substrates where VSMCs were seeded after coated with fibronectin(FN),were fabricated through the blending of sylgard 184 gel and sylgard 527 gel.After that,intracellular calcium level of VSMCs was compared with or without the treatment of IKCa specific blocker,TRAM-34,using a calcium-sensitive dye,fluo 4-AM.1mM Ethylene glycol-bis(2-aminoethyl ether)-N,N,N’,N’-tetraacetic acid(EGTA)was added into the culture media for the removal of the extracellular calcium ions as well as cell counting kit-8(CCK-8)assay was applied,which is to explore the role that calcium ion entry played in proliferation process.The activation level of ERK 1/2 was described by the expression level of phospho-ERK 1/2 using western blotting with or without TRAM-34 treatment.The role of the activation of ERK1/2 in VSMC proliferation was examined by CCK-8 assay with or without the treatment of PD98095,an ERK1/2 inhibitor.Results Compared with soft substrate,stiff substrate caused an increase of intracellular calcium level,which was attenuated by IKCa blockade.In addition,compared with soft substrate,stiff substrate also caused an activation of ERK1/2,which was significantly suppressed by IKCa blockade.Furthermore,extracellular calcium ion reduction by adding EGTA significantly inhibited the stiff substrate-induced VSMC proliferation,which whereas had no effect on VSMC proliferation on soft substrate.Finally,ERK1/2 inhibition had similar inhibitory effect on stiff substrate-induced proliferation.Conclusions Stiff substrate causes an IKCa channel-mediated calcium entry and ERK1/2 activation,both of which play important roles in stiff substrate-induced VSMC proliferation.Combined the previous results that IKCa channel participated in stiff substrate-induced VSMC proliferation,our present work suggests that IKCa channel functioned in the proliferation process through mediating calcium entry and subsequent ERK1/2 activations.These findings provide a new insight into how substrate stiffness regulates VSMC proliferation,and additional considerations for vascular tissue engineering and vascular disease treatment.
文摘Aim：Investigate the influence of IgD on T/B cell activation and construct hIgD-Fc-Ig fusion protein to competitive inhibition IgD binding with IgDR. Methods differences of mIgD and IgD-R level between different T/B was detected by ELISA. Human IgD-Fc-IgGI-Fc sequence T/B cells were sorted by magnetic cell sorting. The cell subtypes were detected by FCM. Serum IgD level was amplified by cross-PCR and then subcloned into PET28a（ ＋ ） empty vector. After prokaryotic expression through escherichia？ coli, we obtained the hIgD-Fc-Ig fu- sion protein by affinity chromatograph. Western blot was used to identify the hIgD-Fc-Ig fusion protein. Human pe- ripheral blood monouclear cells （PBMC） and fibroblast like synoviocytes （FLS） proliferation were detected using a cell counting kit-8 （ CCK-8 ）. Results The percentage of CD3 ＋/CD4 ＋ , CD3 ＋/IgD ＋ , CD3 ＋/CD4 ＋/IgD ＋ , CD3 ＋/IgD-R ＋ and CD3 ＋/CD4 ＋/IgD-R ＋ cells increased significantly in RA patients comparing to healthy people. IgD can stimulate PBMC proliferation. IgD （1, 3, 10, 30 μg ＂ ml^-1） stimulate PBMC proliferation significantly after 24 h. We obtained stable and active hlgD-Fc-Ig fusion protein. The hlgD-Fc-Ig fusion protein showed no effect on PBMC proliferation. But it could downregulate human IgD protein promoting proliferation effects in human PBMC. Conclusion This result suggests that IgD and IgDR play an important role on T/B cell activation in RA patients and the hIgD-Fc-Ig fusion protein may competitively inhibit IgD＇s function and may play an therapeutic role in autoimmune diseases.
基金supported by the National Natural Science Foundation of China ( 11872010)
文摘Objective Mechanical stretch regulates mesenchymal stem cell(MSC)function,which much more emphasis has been placed its prolonged effect on lineage differentiation,especially osteogenic differentiation.In contrast,there are few reports about its short term effect on MSC proliferation.In the present study,effects of short-term mechanical stretch on the proliferation and osteogenic differentiation of mesenchymal stem cell proliferation were investigated.In addition,the stretchinfluenced expression of transient receptor potential cation channel,subfamily C,member 1(TRPC1)was also investigated due to its mechanosensitivity and positive correlation with MSC proliferation.Methods MSCs,harvested from rat bone marrow,were seeded on collagen l-coated silicone chamber and exposed to mechanical stretch with various magnitude(0%,5%,10%and 15%)or various duration(2 h,6 h,12 h and 24 h).Cell proliferation was examined by cell counting kit-8(CCK-8)assay and cell cycle analysis.The gene and protein expression of two makers for osteogenic differentiation,collagen I and Cbfα1,and TRPC1 were determined by RT-PCR and western blotting,respectively.BMSC were harvested,and total RNA was isolated with Trizol reagent.A 2μg portion of total RNA was synthesized to cDNA according to the manufacturer s instructions.cDNA was used as a template for each PCR amplification.BMSC were solubilized in RIPA lysis buffer on ice for 30 min.Phenylmethane sulfonyl fluoride(PMSF)was added to avoid proteolysis.Equal portions of the cell lysates were separated on 10%sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE)and transferred to polyvinylidene fluoride(PVDF)membranes.The membranes were incubated with primary antibodies to TRPC1 and GAPDH at4℃overnight to identify the specific proteins.The PVDF membranes were washed with TBST three times and incubated with a horseradish peroxidase(HRP)-conjugated secondary antibody.Immunoreactive bands were visualized using an enhanced chemiluminescent(ECL)system.Results The OD value for the three stretch cases(5%-15%)was increased^1.4-fold compared with that for control(0%).There is no significantly difference among the three stretch cases.The percentage of cells for three stretch cases were more in the S phase but less in the G0/G1 phase compared to those for control.The cell cycle distribution still had no significant difference among the three stretch cases.In addition,the stretch application for 24 h didn’t affect the gene or protein level for collagen I and Cbfα1 compared with those of control.Application of 10%stretch for 2 h didn’t affect TRPC1 gene or protein expression,but that application for 6-24 h significantly up-regulated TRPCl gene and protein level.That increase exhibited a stretch duration-independent manner.Conclusions Short-term mechanical stretch promoted MSC proliferation in a magnitude-independent manner,whereas had no effect on its oesteogenic differentiation.Paralleled to which,TRPC1 was up-regulated by stretch,implying that TRPCl may be implicated in that proliferation courses.Future work is still needed to confirm whether TRPC1 participates in that stretch-induced MSC proliferation using RPC1 blockade or knockout.
基金supported by National Natural Science Foundation of China(81330081,81202596)Specialized Research Fund for the Doctoral Program of Higher Education(20123420110003)+1 种基金Program for Tackling Key Problems in Science and Technology by Anhui Province(1301042098)China Postdoctoral Science Foundation(2013M540508)
文摘OBJECTIVE This study aimed to investigate the influence of IgD on T/B cell activation and construct h IgD-Fc-Igfusion protein to competitive inhibition IgD binding with IgDR.METHODS T/B cells were sorted by magnetic cell sorting.The differences of m IgD and IgD-R level between different T/B cell subtypes were detected by FCM.Serum IgD level was detected by ELISA.Human IgD-Fc-IgG1-Fc sequence was amplified by cross-PCR and then subcloned into PET28 a(+) empty vector.After prokaryotic expression through escherichia coli,we obtained the h IgD-Fc-Igfusion protein by affinity chromatograph.Western blot was used to identify the h IgD-Fc-Igfusion protein.Human peripheral blood monouclear cells(PBMC) and fibroblast like synoviocytes(FLS) proliferation were detected using a cell counting kit-8(CCK-8).RESULTS The percentage of CD3^+/CD4^+,CD3^+/IgD^+,CD3^+/CD4^+/IgD^+,CD3^+/IgD-R+and CD3^+/CD4^+/IgD-R+cells increased significantly in RA patients comparing to healthy people.IgD can stimulate PBMC proliferation.IgD(1,3,10,30 μg·mL^(-1)) stimulate PBMC proliferation significantly after 24 h.We obtained stable and active h IgD-Fc-Igfusion protein.The h IgD-Fc-Igfusion protein showed no effect on PBMC proliferation.But it could downregulate human IgD protein promoting proliferation effects in human PBMC.CONCLUSION This result suggests that IgD and IgDR play an important role on T/B cell activation in RA patients and the h IgD-Fc-Igfusion protein may competitively inhibit IgD′s function and may play an therapeutic role in autoimmune diseases.
基金supported by Medical Science and Technology Research Foundation of Guangdong Province(2015120104622949)
文摘OBJECTIVE Cervical cancer is the third most malignant tumor in the world.Farnesoid X receptor(FXR) is a member of nuclear receptor superfamily.It is highly expressed in liver,kidney and small intestine,while it showed low expression level in other tissues.It not only plays an important role in the metabolism of bile acids and sugars,but also in the production of chronic inflammation in the early stage of cancer,the proliferation and migration of tumor.Compared with the normal tissue,the expression of FXR in most tumor tissues decreased.But there is no correlation between cervical cancer and FXR.So we aimed to find out the relationship between FXR and cervical cancer.METHODS A clinical study using q PCR,western blot and immunohistochemistry detected the expression of FXR in tumor tissues and normal tissues of clinical patients.FXR was activated by agonists or over-expressed by lentivirus.MTT,clone formation and flow cytometry were used to detect the relationship between FXR and proliferation of cervical cell lines.Tumor growth ability of FXR was detected by nude mice tumorigenicity.The interaction between FXR and CDKN2A-p14^(ARF)-MDM2-p53 pathway was detected by q PCR,Western blot and immunohistochemistry.RESULTS FXR was decreased in cancer tissues compared to normal control.Activation of FXR by agonist or constitutively-over-expression of FXR inhibited cervical cell proliferation.Over-expressed FXR attenuated Caski,Hela and Siha xenograft tumor growth in nude mice compared with control.Over-expression of FXR caused G1 cell-cycle arresting and up-regulated CDKN2A-p14^(ARF)-MDM2-p53 pathway.CONCLUSION FXR inhibits cervical cancer cell proliferation and cervical tumorigenicity which is related to CDKN2A-p14^(ARF)-MDM2-p53 pathway.Activation or overexpression of FXR may be a potential target for the treatment of cervical cancer.
基金The project supported by Beijing Natural Science Foundation(7122083)Beijing Science and Technology Projec(tD161100005116005)
文摘OBJECTIVE To investigate the inhibitory effects of Spatholobus Suberectus Column Extract(SSCE)on estrogen receptor positive(ER+)breast cancer cel MCF-7and its possible molecular mechanism.METHODS MCF-7cells were cultured without estrogen and with 17-β-estrogen(10-8mol·L-1),respectively,then treated with SSCE(0,40,80,160,320μg·m L-1).MTT assay was employed to evaluate cell viability.Flow cytometry assays were performed to underlying apoptosis and detecting cel cycle of MCF-7 cells treated with SSCE(0,80,160,320μg·mL-1).Wound healing assays was conducted to detect the migration ability.Dual luciferase reporter system was used to detect the activity of p-ERα,p-ERβpresented in intra-nuclear estrogen response element(ERE).Western blotting assay was employed to identify the expression of protein such as Bax,Bcl-2,p-ERα,p-ERβ,ERK1/2,p-ERK1/2,AKT,p-AKT,m TOR,p-m TOR,PI3K,p-PI3K.RESULTS It showed that SSCE(80,160and 320μg·mL-1)significantly decreased the viability of MCF-7.SSCE also triggered apoptosis,arrested cell cycle at G0/G1phase,inhibited migration.Dual luciferase reporter system showed that SSCE suppressed intra-nuclear p-ER activity,Western blotting analysis confirmed that SSCE did repress the expression of phosphorylated-ER alpha(p-ERα),ERK1/2,p-ERK1/2,AKT,p-AKT,pmT OR,PI3K,p-PI3K,which indicate that SSCE suppress MAPK PI3K/AKT signal pathway.CONCLUSION Our result showed that SSCE cause ER+MCF-7 cells apoptosis,G0/G1phase arresting,migration decreasing,via hypo-active of ER,suppress MAPK PI3K/AKT pathway.
基金The project supported by National Natural Science Foundation of China(81473234)
文摘OBJECTIVE To investigate the effect of gap junctions on the anti-tumor function induced by mi R-34a in glioma U87 cells.METHODS 1.Transfection(miR-34a mimics were transfected into glioma cells to upregulate their expression);2.Co-culture assay(U87cells were transfected with mi R-34a co-cultured with U87 cells that was transfected PCMV-eG FP plasmid);3.Flow cytometry analysis(select the e GFP labed U87 cells);4.RNA isolation and real-time PCR;5.CCK-8 assay;6.Western blotting.RESULTS Mi R-34a mimics transfered between the U87 cells.Parachute assay showed that GJ inhibition(CBX and 18-α-GA)can decrease mi R-34a expression than co-culture group.RA and galanglin enhanced mi R-34a expression than co-culture group.Mi R-34a relative expression reduced after co-culture,while gap junctions composed of Cx43 were down-regulated by sh RNA.Transfected with mi R-34a mimics reduced the survival of U87 cells in a dose-dependent manner.To more specifically establish the role of GJIC in mi R-34a induced growth inhibition of U87 cells,si RNA was used to knockdown the expression of Cx43,the dominant connexin expressed in U87 cells.CCK-8 assay showed that siR NAs have no effect on cell growth,but they could aggravate the growth inhibition of miR-34a to U87 cels.CONCLUSION Gap junctions enhance the antiproliferative effect of miR NA-34a in glioma cells.
