ESTABLISHMENT OF A HUMAN B CELL LINE THAT RESPONDS SPECIFICALLY TO B CELL GROWTH FACTOR

A human B cell line （3D5） that responds specifically to B cell growth factor （BCGF） hasbeen developed by a sequence of Staphylococcus aureus Cowen I activation,EB virus im-mortalization,and cloning.Proliferative response to PHA-stimulated T cell supernatant（PHA-T-Sup） and nonresponsiveness to rIL-2 stimulation were factors used to screen positivecells.Phenotype analysis with a flow cytometer indicated that:1) 3D5 is a B cell line:100% of the cells were positive for B1 marker and 59% were positive for sIg,while T3and Mo 1 were negative:2) 3D5 is an activated B cell line:both Tac and 4F2 markersof activated （but not of resting） B cells were 100% positive:3) 3D5 expresses high molecularweight BCGF （HMW-BCGF） receptor-associated epitope BA5.3D5 cells proliferated inresponse to cpBCGF stimulation in a dose-dependent manner.HMW-BCGF also induced3D5 cells to proliferate.Interestingly.no proliferation could be detected in the presenceof rIL-2,rIL-4,or rIFN-r.The data show that 3D5 cells are specifically BCGF-responsiveB cells.Using 3D5 cells as target,BCGF activity was detected in crude BCGF preparationsedimented by 85% （NH4）2SO4 and chromatographed in a DEAE-Sephadex A-25 column fromPHA-T-Sup.T24 cell supernatant with B cell differentiation factor （BCDF） activity couldnot induce 3D5 cells to differentiate into immunoglobulin-secreting cells.

Effects of 530 nm monochromatic light on basic fibroblast growth factor and transforming growth factor-β1 expression in Müller cells

AIMTo expose rat retinal Müller cells to 530 nm monochromatic light and investigate the influence of varying light illumination times on basic fibroblast growth factor (bFGF) and transforming growth factor-β1 (TGF-β1) expression.METHODSThree groups of rat retinal Müller cells cultured in vitro under a 530 nm monochromatic light were divided into 6, 12 and 24h experimental groups, while cells incubated under dark conditions served as the control group. The bFGF and TGF-β1 mRNA expression, protein levels and fluorescence intensity of the Müller cells were analyzed.RESULTSThe bFGF mRNA expression and protein levels were significantly upregulated in Müller cells in all three experimental groups compared with the control group (P<0.05), while that of TGF-β1 was downregulated (P<0.05). Also, bFGF expression was positively correlated, but TGF-β1 expression was negatively correlated with illumination time. The largest changes for both cytokines were seen in the 24h group. The changes in bFGF and TGF-β1 fluorescence intensity were highest in the 24h group, and significant differences were observed among the experimental groups (P<0.05).CONCLUSIONThe expressions of bFGF and TGF-β1 changed in a time-dependent manner in Müller cells exposed to 530 nm monochromatic light with 250 lx illumination intensity. Müller cells might play a role in the development of myopia by increasing bFGF expression or decreasing TGF-β1 expression. Changes in cytokine expression in retinal Müller cells may affect monochromatic light-induced myopia.

Vascular endothelial growth factor B inhibits insulin secretion in MIN6 cells and reduces Ca2+ and cyclic adenosine monophosphate levels through PI3K/AKT pathway

BACKGROUND Type 2 diabetes(T2 D) is characterized by insufficient insulin secretion caused by defective pancreatic β-cell function or insulin resistance,resulting in an increase in blood glucose.However,the mechanism involved in this lack of insulin secretion is unclear.The level of vascular endothelial growth factor B(VEGF-B) is significantly increased in T2 D patients.The inactivation of VEGF-B could restore insulin sensitivity in db/db mice by reducing fatty acid accumulation.It is speculated that VEGF-B is related to pancreatic β-cell dysfunction and is an important factor affecting β-cell secretion of insulin.As an in vitro model of normal pancreatic β-cells,the MIN6 cell line can be used to analyze the mechanism of insulin secretion and related biological effects.AIM To study the role of VEGF-B in the insulin secretion signaling pathway in MIN6 cells and explore the effect of VEGF-B on blood glucose regulation.METHODS The MIN6 mouse pancreatic islet β-cell line was used as the model system.By administering exogenous VEGF-B protein or knocking down VEGF-B expression in MIN6 cells,we examined the effects of VEGF-B on insulin secretion,Ca2+ and cyclic adenosine monophosphate(cAMP) levels,and the insulin secretion signaling pathway.RESULTS Exogenous VEGF-B inhibited the secretion of insulin and simultaneously reduced the levels of Ca2+ and cAMP in MIN6 cells.Exogenous VEGF-B also reduced the expression of phospholipase C gamma 1(PLCγ1),phosphatidylinositol 3-kinase(PI3 K),serine/threonine kinase(AKT),and other proteins in the insulin secretion pathway.Upon knockdown of VEGF-B,MIN6 cells exhibited increased insulin secretion and Ca2+ and cAMP levels and upregulated expression of PLCγ1,PI3 K,AKT,and other proteins.CONCLUSION VEGF-B can regulate insulin secretion by modulating the levels of Ca2+ and cAMP.VEGF-B involvement in insulin secretion is related to the expression of PLCγ1,PI3 K,AKT,and other signaling proteins.These results provide theoretical support and an experimental basis for the study of VEGF-B in the pathogenesis of T2 D.

Effect of NF-κB p65 antisense oligodeoxynucleotide on transdifferentiation of normal human lens epithelial cells induced by transforming growth factor-β2

AIM：To study the inhibition of nuclear factor kappa-B p65（NF-κB p65）antisense oligodeoxynucleotide（ASODN）on transdifferentiation of normal human lens epithelial cells induced by transforming growth factor-β2（TGF-β2）.·M ETHODS：NF-κBp65ASODNand NF-κBp65missense oligodeoxynucleotide（MSODN）were designed and synthesized.Human lens epithelial cell line（HLE B-3）cells were prepared for study and divided into 7 groups.Control group was HLE B-3 cells cultured in dulbecco’s modified eagle medium（DMEM）.T1,T2,and T3 group were HLE B-3 cells cultured in DMEM with 10 ng/m L TGF-β2 for 6h,12h,24h respectively.A＋T group was HLE B-3 cells cultured with 10 ng/m L TGF-β2for 24h after transfected by NF-κB p65 ASODN for 24h.M＋T group was HLE B-3 cells cultured with 10 ng/m L TGF-β2 for 24h after transfected by NF-κB p65 MSODN for 24h.The negative control group was HLE B-3 cells cultured with 10 ng/m L TGF-β2 for 24h after cultured with transfer agent（Hi Per Fect）for 24h.Cell morphology was observed at different time points using an inverted microscope.The expression of NF-κB p65 m RNA was detected with reverse transcription-polymerase chain reaction（RT-PCR）,and the expression ofα-smooth muscle actin（α-SMA）protein was assayed with ELISA.·RESULTS：With the TGF-β2 stimulation prolongation,the expression of NF-κB p65 m RNA and a-SMA protein increased in T1,T2,T3 groups compared with the control group,and the difference was statistically significant（〈0.05）.NF-κB p65 ASODN lowered the expression of NF-κB p65 m RNA andα-SMA protein induced by TGF-β2.NF-κB p65 MSODN and Hi Per Fect did not lower the expression of NF-κB p65 m RNA andα-SMA protein induced by TGF-β2.The difference between control group and A＋T group was not statistically significant（〉0.05）,but the difference among A＋T group and other groups was statistically significant（〈0.05）.·CONCLUSION：NF-κB p65 ASODN could lower the expression of NF-κB p65 m RNA andα-SMA protein induced by TGF-β2,and antagonized TGF-β2-induced transdifferentiation of HLE B-3.NF-κB p65ASODN could be used as a new biological therapeutic target of posterior capsular opacification.

Inhibition of EGFR attenuates EGF-induced activation of retinal pigment epithelium cell via EGFR/AKT signaling pathway

AIM:To explore the effect of epidermal growth factor receptor(EGFR)inhibition by erlotinib and EGFR siRNA on epidermal growth factor(EGF)-induced activation of retinal pigment epithelium(RPE)cells.METHODS:Human RPE cell line(ARPE-19 cells)was activated by 100 ng/mL EGF.Erlotinib and EGFR siRNA were used to intervene EGF treatment.Cellular viability,proliferation,and migration were detected by methyl thiazolyl tetrazolium(MTT)assay,bromodeoxyuridine(BrdU)staining assay and wound healing assay,respectively.EGFR/protein kinase B(AKT)pathway proteins and N-cadherin,α-smooth muscle actin(α-SMA),and vimentin were tested by Western blot assay.EGFR was also determined by immunofluorescence staining.RESULTS:EGF treatment for 24h induced a significant increase of ARPE-19 cells’viability,proliferation and migration,phosphorylation of EGFR/AKT proteins,and decreased total EGFR expression.Erlotinib suppressed ARPE-19 cells’viability,proliferation and migration through down regulating total EGFR and AKT protein expressions.Erlotinib also inhibited EGF-induced an increase of proliferative and migrative ability in ARPE-19 cells and clearly suppressed EGF-induced EGFR/AKT proteins phosphorylation and decreased expression of N-cadherin,α-SMA,and vimentin proteins.Similarly,EGFR inhibition by EGFR siRNA significantly affected EGF-induced an increase of cell proliferation,viability,and migration,phosphorylation of EGFR/AKT proteins,and up-regulation of N-cadherin,α-SMA,and vimentin proteins.CONCLUSION:Erlotinib and EGFR-knockdown suppress EGF-induced cell viability,proliferation,and migration via EGFR/AKT pathway in RPE cells.EGFR inhibition may be a possible therapeutic approach for proliferative vitreoretinopathy(PVR).

Effects of hypoxia-inducible factor-1α silencing on the proliferation of CBRH-7919 hepatoma cells

AIM:To study the effects of hypoxia-inducible factor1α(HIF-1α) silencing on the proliferation of hypoxic CBRH-7919 rat hepatoma cells.METHODS:The CBRH-7919 rat hepatoma cell line was used in this study and the hypoxic model was constructed using CoCl2.The HIF-1α-specific RNAi sequences were designed according to the gene coding sequence of rat HIF-1α obtained from GeneBank.The secondary structure of the HIF-1α gene sequence was analyzed using RNA draw software.The small interfering RNA(siRNA) transfection mixture was produced by mixing the siRNA and Lipofectamine2000TM,and transfected into the hypoxic hepatoma cells.Real time reverse transcription-polymerase chain reaction(RTPCR) and Western blotting assay were used to detect the expression levels of mRNA and protein.HIF-1α and vascular endothelial growth factor(VEGF) mRNA was determined using real time RT-PCR;the protein expression levels of AKT,p-AKT,p21 and cyclinD1 were determined using Western blotting.The proliferation of hepatoma cells was observed using the methyl thiazolyl tetrazolium(MTT) assay and the bromodeoxyuridine(BrdU) incorporation cell proliferation assay.RESULTS:Under induced hypoxia,the viability of the hepatoma cells reached a minimum at 800 μmol/L CoCl2;the viability of the cells was relatively high at CoCl2 concentrations between 100 μmol/L and 200 μmol/L.Under hypoxia,the mRNA and protein expression levels of HIF-1α and VEGF were significantly higher than that of hepatoma cells that were cultured in normaxia.HIF-1α-specific RNAi sequences were successfully transfected into hepatoma cells.The transfection of specific siRNAs significantly inhibited the mRNA and protein expression levels of HIF-1α and VEGF,along with the protein expression levels of p-AKT and cyclinD1;the protein expression of p21 was significantly increased,and there was no significant difference in the expression of AKT.The MTT assay showed that the amount of hepatoma cells in S phase in the siRNA transfection group was obviously smaller than that in the control group;in the siRNA transfection group,the amount of hepatoma cells in G1 phase was more than that in the control group.The BrdU incorporation assay showed that the number of BrdU positive hepatoma cells in the siRNA transfection group was less than that in the control group.The data of the MTT assay and BrdU incorporation assay suggested that HIF-1α silencing using siRNAs significantly inhibited the proliferation of hepatoma cells.CONCLUSION:Hypoxia increases the expression of HIF-1α,and HIF-1α silencing significantly inhibits the proliferation of hypoxic CBRH-7919 rat hepatoma cells.

Effects of transforming growth factor β2 and connective tissue growth factor on induction of epithelial mesenchymal transition and extracellular matrix synthesis in human lens epithelial cells

AIM:To Investigate the effects of transforming growth factorβ2(TGF-β2)and connective tissue growth factor(CTGF)on transdifferentiation of human lens epithelial cells(HLECs)cultured in vitro and synthesis of extracellular matrix(ECM).METHODS:HLECs were treated with TGF-β2(0,0.5,1.0,5,10μg/L)and CTGF(0,15,30,60,100μg/L)for different times(0,24,48,72h)in vitro and the expression ofα-smooth muscle actin(α-SMA),the main component of the extracellular matrix typeⅠcollagen(Col-1)and fibronectin(Fn)were measured by using real-time polymerase chain reaction(PCR)and western-blot.RESULTS:TGF-β2 and CTGF significantly increased expression ofα-SMA mRNA and protein(P【0.05,P【0.001),Fn mRNA and protein(P【0.001),Col-1 mRNA and protein(P【0.001).TGF-β2 could induce HLECs expression of CTGF mRNA and protein in dosedependent manner(P【0.05,P【0.001).TGF-β2 and CTGF could induce HLECs to expressα-SMA,Fn and Col-1 in time-dependent manner.Each time of TGF-β2and CTGF induced HELCs expression ofα-SMA,Fn,Col-1 mRNA and protein was significant increase compared with control(P【0.05,P【0.001).CONCLUSION:TGF-β2 and CTGF could induce HLECs epithelial mesenchymal transition and ECM synthesis.

Influence of baicalin on the expression of receptor activator of nuclear factor-κB ligand and osteoprotegerin in human periodontal ligament cells

Objective To study the effect of baicalin on the expression of receptor activator of nuclear factor-κB ligand(RANKL)and osteoprotegerin(OPG)in cultured human periodontal ligament(HPDL)cells.Methods Small interfering RNA(siRNA)eukaryotic expression vector targeted transforming growth factor βⅡ receptor(TGF-β RⅡ)was constructed and transfected into T cells.HPDL cells with T cells transfected with siRNA or not were placed in the culture medium that had been added with lipopolysaccharide(LPS)and baicalin.The obtained solution was divided into six groups according to the components(group Ⅰ:HPDL cells+LPS+T cells transfected with siRNA1+baicalin;group Ⅱ:HPDL cells+LPS+T cells transfected with siRNA1;group Ⅲ:HPDL cells+LPS+T cells+baicalin;group Ⅳ:HPDL cells+LPS+T cells;group Ⅴ:HPDL cells+baicalin;group Ⅵ:HPDL cells)and was cultured for 48 hours.RT-PCR was used to observe the effect of baicalin on the expression of OPG-RANKL in HPDL cells.Results The ratio of RANKL/OPG in group Ⅰ was lower than that in group Ⅱ(P<0.01)and higher than that in group Ⅲ(P<0.01);The ratio of RANKL/OPG in group Ⅲ was lower than that in group Ⅳ(P<0.01);the ratio of RANKL/OPG in group Ⅳ was higher than that in group Ⅵ(P<0.01);the ratio of RANKL/OPG in group Ⅴ was lower than that in group Ⅵ(P<0.05).Conclusion ① Baicalin could decrease the ratio of RANKL/OPG in HPDL cells.② The TGF-β signaling transduction plays an important role in the effect of baicalin on the RANKL/OPG ratio in HPDL cells.③ Baicalin acts not only through TGF-β to regulate RANKL/OPG in HPDL cells,but also through other pathways.

PCNA、Bcl-2及EGFR在喉癌组织中的表达及与临床病理特征、生存的关系

目的探讨增殖细胞核抗原(PCNA)、B淋巴细胞瘤-2(Bcl-2)及表皮生长因子受体(EGFR)在喉癌组织中的表达及与临床病理特征、生存的关系。方法选取2017年3月—2020年1月在苏州大学附属第一医院因喉癌行手术治疗的92例患者的喉癌组织及对应癌旁组织标本。检测癌组织与癌旁组织PCNA mRNA、Bcl-2mRNA、EGFR mRNA相对表达量,多元线性回归分析其癌组织表达与临床病理特征的关系。随访3年,采用Kaplain-Maier曲线分析不同PCNA、Bcl-2、EGFR表达水平患者生存情况差异。结果癌组织PCNA mRNA、Bcl-2 mRNA、EGFR mRNA相对表达量高于癌旁组织(P<0.05)。不同年龄、肿瘤部位患者PCNA mRNA、Bcl-2 mRNA、EGFR mRNA相对表达量比较,差异无统计学意义(P>0.05);低分化,临床分期Ⅲ、Ⅳ期及淋巴结转移患者PCNA mRNA、Bcl-2 mRNA、EGFR mRNA相对表达量分别高于中、高分化,临床分期Ⅰ、Ⅱ期,无淋巴结转移患者(P<0.05)。多元线性回归分析结果显示,肿瘤分化程度、临床分期、淋巴结转移是喉癌组织PCNA mRNA、Bcl-2 mRNA、EGFR mRNA表达的影响因素。Kaplain-Maier曲线分析结果显示,PCNA mRNA高表达患者3年无进展生存率、总生存率分别为59.57%和70.21%,低于低表达患者的80.00%和88.89%(P<0.05);Bcl-2 mRNA高表达患者3年无进展生存率、总生存率分别为60.78%和70.59%,低于低表达患者的80.49%和90.24%(P<0.05);EGFR mRNA高表达患者3年无进展生存率、总生存率分别为59.09%和70.45%,低于低表达患者的79.17%、87.50%(P<0.05)。结论喉癌组织PCNA、Bcl-2、EGFR呈高表达,且其高表达状态与肿瘤分期高、分化程度低、淋巴结转移有关,PCNA、Bcl-2、EGFR表达水平可在一定程度上反映患者预后。

Small interfering RNA targeting PGC-1α inhibits VEGF expression and tube formation in human retinal vascular endothelial cells

AIMTo determine whether small interfering RNA (siRNA) of PGC-1α could inhibit vascular endothelial growth factor (VEGF) expression and tube formation in human retinal vascular endothelial cells (hRVECs).METHODShRVECs transfected with peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) siRNA were incubated for 24h and then placed into a normoxic (20%, O2) or hypoxic (1%, O2) environment for another 16h. PGC-1α mRNA and protein levels were detected by real-time PCR and Western blot. VEGF mRNA and protein levels were detected by real-time PCR and ELISA. Cell proliferation was evaluated by BrdU incorporation assay. Forty-eight hours after siRNA transfection, hRVECs were planted into Matrigel-coated plates and cultured under normoxic (20%, O2) or hypoxic (1%, O2) conditions for another 48h. The tube formation of hRVECs was observed under an optical microscope and quantified by counting the number of branch points and calculating the total tube length.RESULTSPGC-1α mRNA and protein levels were significantly reduced by PGC-1α siRNA, and VEGF mRNA and protein levels also decreased significantly. The percentage of BrdU-labeled cells in siPGC-1α groups were significantly decreased compared with control siRNA groups under normoxia and hypoxia in cell proliferation assay. In the tube formation assay, PGC-1α siRNA treated cells formed significantly fewer tubes.CONCLUSIONBlocking PGC-1α expression can inhibit VEGF expression in hRVECs and inhibit their ability to form tubes under both normoxic and hypoxic conditions.

Protein tyrosine phosphatase 1B regulates migration of ARPE-19 cells through EGFR/ERK signaling pathway

AIMTo evaluate whether protein tyrosine phosphatase 1B (PTP1B) contributed to initiate human retinal pigment epithelium cells (A)-19 migration and investigate the signaling pathways involved in this process.METHODSARPE-19 cells were cultured and treated with the siRNA-PTP1B. Expression of PTP1B was confirmed by quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR). AG1478 [a selective inhibitor of epidermal growth factor receptor (EGFR)] and PD98059 (a specific inhibitor of the activation of mitogen-activated protein kinase) were used to help to determine the PTP1B signaling mechanism. Western blot analysis verified expression of EGFR and extracellular signal-regulated kinase (ERK) in ARPE-19 cells. The effect of siRNA-PTP1B on cell differentiation was confirmed by immunostaining for &#x003b1;-smooth muscle actin (&#x003b1;-SMA) and qRT-PCR. Cell migration ability was analyzed by transwell chamber assay.RESULTSThe mRNA levels of PTP1B were reduced by siRNA-PTP1B as determined by qRT-PCR assay. SiRNA-PTP1B activated EGFR and ERK phosphorylation. &#x003b1;-SMA staining and qRT-PCR assay demonstrated that siRNA-PTP1B induced retinal pigment epithelium (RPE) cells to differentiate toward better contractility and motility. Transwell chamber assay proved that PTP1B inhibition improved migration activity of RPE cells. Treatment with AG1478 and PD98059 abolished siRNA-PTP1B-induced activation of EGFR and ERK, &#x003b1;-SMA expression and cell migration.CONCLUSIONPTP1B inhibition promoted myofibroblast differentiation and migration of ARPE-19 cells, and EGFR/ERK signaling pathway played important role in migration process.

EFFECT OF OXIDIZED-LDL ON NF-κB NUCLEAR TRANSLOCATION IN AORTIC SMOOTH MUSCLE CELLS ORIGINATED FROM RATS OF DIFFERENT AGES

Objective To investigate the molecular mechanism of atherosclerosis that related to age. Methods Immunohistochemistry staining and Western blot were adopted to determine the nuclear translocation of nuclear factor-kappa B (NF-κB) and expression of platelet-derived growth factor B (PDGF-B) in smooth muscle cells (SMCs) co-cultured with low density lipoprotein (LDL), oxidized LDL (ox-LDL), and ox-LDL+high density lipoprotein (HDL) originated from rats of 2 and 10 months old respectively. Fat stain was used to identify the lipid intake in SMCs. Results The optimal stimulation time of ox-LDL to SMCs was 12 hours. NF-κB intensity increased in most nuclei of SMCs that originated from rats of either 2 or 10 months old co-cultured with ox-LDL. The intensity of NF-κB and the amount of intracellular lipid taken in SMCs were more obvious in cells from 10-month-old rats than from the younger ones. Change of PDGF-B expression in SMCs was not remarkable in each group of rats. Conclusions The 10-month-old rats are more susceptive to ox-LDL than 2-month-old rats in activating nuclear transloca- tion of NF-κB. Maybe this is one of the important reasons contributing to the difference between the older and younger rats on the initiation and development of atherosclerosis lesion. Expression of PDGF-B is not associated with the activity of nuclear translocation of NF-κB.

α1-antitrypsin combined with bone marrow mesenchymal stem cells regulates retinopathy in diabetic rats via p38 MAPK/NF-κB signaling pathway

Objective:To investigate the effect ofα1-antitrypsin combined with bone marrow mesenchymal stem cells on retinopathy in diabetic rats and its mechanism.Methods:A model of diabetic retinopathy was established by intraperitoneal injection of streptozotocin.The 30 Wistar rats successfully modeled were randomly divided into a model group,a bone marrow mesenchymal stem cell group and a combined group(α1-antitrypsin combined with bone marrow Mesenchymal stem cells),the blood glucose and serum insulin levels of diabetic rats were measured 4 weeks after treatment.Enzyme-linked immunosorbent assay(ELISA)for measuring serum inflammatory factors IL-1β,IL-6 and TNF-α in rats.Observing the pathological morphology of rat retina under hematoxylin-eosin staining(HE).TUNEL staining to observe the apoptosis of rat retinal nerve cells.Immunohistochemical method to detect the expression level of CD45 in retinal tissue.Real-time fluorescence quantitative PCR was used to detect the expression of retinal vascular endothelial growth factor(VEGF),hypoxiainducible factor-1α(HIF-1α),and angiotensinⅡ(ANGⅡ)mRNA.Western blot was used to detect the expression of p38 MAPK/NF-κB signaling pathway-related proteins in the retinal tissue of each group of rats.Results:Compared with the control group,the rats in the model group had increased blood glucose,decreased insulin levels,increased serum IL-1β,IL-6,and TNF-α levels,and had obvious lesions in the retina.CD45 showed high expression in retinal tissue,VEGF,HIF-1α,ANGⅡ mRNA expression increased,p-p38,p-p65,p-IκBα protein expression increased(P<0.05).Compared with the model group,the bone marrow mesenchymal stem cell group and the combined group have decreased blood glucose,increased insulin levels,and decreased serum IL-1β,IL-6 and TNF-α levels.Retinopathy is improved,apoptosis of retinal nerve cells is reduced,CD45 expression in retinal tissue is reduced,VEGF,HIF-1α,ANGⅡ mRNA expression is decreased,and p-p38,p-p65,p-IκBα protein expression is decreased.Compared with the bone marrow mesenchymal stem cell group,the effect of the combined group was more obvious(P<0.05).Conclusion:α1-antitrypsin combined with bone marrow mesenchymal stem cell transplantation can improve the degree of retinopathy in diabetic rats.The mechanism may be related to the inhibition of p38 MAPK/NF-κB signaling pathway.

AP2M1抑制弥漫性大B细胞淋巴瘤细胞增殖和侵袭

目的探究接头相关蛋白质复合体2亚基μ1(AP2M1)对弥漫性大B细胞淋巴瘤(DLBCL)细胞增殖和侵袭的调控作用。方法将人弥漫性大B细胞淋巴瘤细胞系OCI-LY8分为对照组、NC-LV组、AP2M1-LV组。用Lipofectamine 2000进行细胞转染。四甲基偶氮唑盐(MTT)法检测细胞增殖,流式细胞仪检测细胞凋亡,Transwell小室法检测细胞迁移和侵袭。Western blot检测AP2M1、表皮生长因子受体(EGFR)、p-磷脂酰肌醇3激酶(PI3K)和p-蛋白质激酶B(AKT)蛋白表达。结果与对照组相比,AP2M1-shRNA组细胞中AP2M1的mRNA和蛋白相对表达量均降低(P<0.05),相对细胞活力升高(P<0.05),细胞凋亡率降低(P<0.05),迁移和侵袭细胞数量均升高(P<0.05);EGFR的蛋白相对表达量及PI3K和AKT的磷酸化水平均升高(P<0.05)。与对照组相比,AP2M1-LV组细胞中AP2M1的mRNA和蛋白相对表达量均升高(P<0.05),相对细胞活力降低(P<0.05),细胞凋亡率升高(P<0.05),迁移和侵袭细胞数量均降低(P<0.05),EGFR的蛋白相对表达量及PI3K和AKT的磷酸化水平均降低(P<0.05)。结论AP2M1的过表达部分通过抑制EGFR/PI3K/AKT信号通路来抑制DLBCL细胞的增殖和侵袭。

血清糖蛋白非转移性黑色素瘤蛋白B对表皮生长因子受体扩增伴随突变的非小细胞肺癌预后的预测作用

目的评估游离的糖蛋白非转移性黑色素瘤蛋白B(GPNMB)对表皮生长因子受体(EGFR)扩增伴随突变的非小细胞肺癌(NSCLC)患者的耐药和预后的预测价值。方法选取2018年3月至2019年9月在唐山市人民医院接受治疗的55例EGFR扩增伴随突变的NSCLC患者作为观察组,患者均应用EGFR酪氨酸激酶抑制剂(EGFR-TKI)作为一线治疗方案;随机选取同期体检中心67例健康人群血液样本作为对照组。比较2组游离GPNMB表达水平;采用t检验或χ2检验分析GPNMB表达和患者临床病理特征的相关性;结合临床疗效,评估其作为耐药标志物的价值。随访患者的无进展生存时间(PFS),并采用多因素Cox回归分析影响EGFR扩增伴随突变NSCLC患者生存的独立危险因素。结果与对照组相比,观察组游离GPNMB表达水平显著升高。观察组游离GPNMB水平和EGFR-TKI的临床疗效显著相关(P=0.016),GPNMB高表达患者的耐药程度更强,PFS也更短(P=0.032)。游离GPNMB高水平(HR=4.029,95%CI:1.942~8.358,P<0.001)是影响患者生存的独立危险因素。结论EGFR扩增伴随突变的NSCLC患者游离GPNMB表达水平显著上调;且其高表达与患者耐药程度的增强及不良预后显著相关,是影响患者生存的独立危险因素。

血清Bcl-2、IGF-1、CA19-9水平联合检测对食管癌的诊断价值

目的:分析血清B细胞淋巴瘤-2(Bcl-2)、胰岛素样生长因子-1(IGF-1)、糖类抗原19-9(CA19-9)水平联合检测对食管癌的诊断价值。方法:回顾性分析2020年1月至2023年8月该院收治的126例疑似食管癌患者的临床资料。所有患者均接受血清学检测和内镜下病理活检,比较良恶性患者及不同病理特征食管癌患者的血清Bcl-2、IGF-1、CA19-9水平。采用受试者工作特征(ROC)曲线分析血清Bcl-2、IGF-1、CA19-9水平单项及联合检测对食管癌的诊断价值。结果:126例疑似食管癌患者中,病理学检出恶性肿瘤(食管癌)70例,其中鳞癌54例,腺癌16例;良性病变56例,其中不典型增生37例,平滑肌瘤19例。恶性患者血清Bcl-2、IGF-1、CA19-9水平均高于良性患者,高分期、低分化、有淋巴结转移的食管癌患者血清Bcl-2、IGF-1、CA19-9水平高于低分期、中高分化、无淋巴结转移的患者,差异均有统计学意义(P<0.05);ROC曲线分析结果显示,血清Bcl-2、IGF-1、CA19-9水平单项及联合检测诊断食管癌的曲线下面积分别为0.802、0.836、0.767、0.918,联合检测的诊断价值高于单一指标检测。结论:血清Bcl-2、IGF-1、CA19-9水平联合检测诊断食管癌的价值高于单一指标检测。

NF-κB p50和VEGF在弥漫性大B细胞淋巴瘤组织中的表达及其临床意义

目的:探讨弥漫性大B细胞淋巴瘤(DLBCL)组织中NF-κB p50、VEGF和MVD的表达及其临床意义。方法:应用免疫组化SP法对62例DLBCL组织和10例反应增生淋巴结组织中NF-κB p50、VEGF及MVD进行检测。结果:62例DLBCL组织中NF-κBp50阳性表达率为63%(39/62),VEGF阳性表达率为68%(42/62),二者在反应增生淋巴结组织中都不表达。NF-κB p50的表达与病理类型和治疗反应有关(P<0.05)。VEGF和MVD的表达与IPI、LDH值、肿瘤分期、病理类型和治疗反应有关(P<0.05)。NF-κB p50,VEGF阳性表达组的MVD值明显高于阴性组。NF-κB p50的表达与VEGF的表达呈正相关(r=0.285,P=0.025)。结论:NF-κB p50对VEGF表达有正向调节作用,在DLBCL的发生和发展中起着重要的作用。

缺氧对血管内皮生长因子诱导牛冠状动脉内皮细胞脂蛋白通透性升高的调控及丹酚酸B的抑制作用

目的 为了了解缺氧对血管内皮生长因子(VEGF)增强血管内皮细胞通透性作用的影响及其与动脉粥样硬化的关系 ,考察了正常和缺氧状态下VEGF对体外培养牛冠状动脉内皮细胞 (BCEC)脂蛋白通透性的影响及丹酚酸B的抑制作用。方法 正常及缺氧条件下 ,将VEGF及丹酚酸B加入BCEC共孵育 ,用SN 695型液闪计数器测 [12 5I]低密度脂蛋白([12 5I]LDL)通过BCEC的百分率。结果 VEGF可显著增强BCEC对 [12 5I]LDL的通透性 ,这种增加具有浓度依赖性。缺氧 3h可促进VEGF所致的的通透性增加。丹酚酸B在正常和缺氧条件下均显著抑制VEGF诱导的BCEC通透性升高。结论 VEGF可增强血管内皮细胞的通透性 ,可能在动脉粥样硬化的形成和发展过程中起一定作用。

丹参酚酸B对转化生长因子β_1和血小板衍生生长因子-BB在肝星状细胞内信号传导的影响

目的探讨丹参酚酸B(SA-B)抗肝纤维化作用机制。方法分离大鼠肝星状细胞(HSC),于无包被塑料培养皿中原代培养7天,继以10-6mmol/L SA-B孵育,再加10ng/ml的转化生长因子β1(TGF-β1)或血小板衍生生长因子-BB(PDGF-BB)刺激。以Western blot法观察SA-B对TGF-β1或PDGF-BB刺激的HSC内ERK蛋白及其磷酸化表达的影响及对TGF-β1或PDGF-BB刺激的HSC表面TGF-β1Ⅰ型受体(TβRⅠ)、Ⅱ型受体(TβRⅡ)或PDGF受体β(PDGFR-β)表达的影响。结果SA-B可抑制原代正常培养9天的HSC及TGF-β1刺激的HSC中ERK1/2的磷酸化;对正常培养的HSC和TGF-β1刺激的HSC表面TβRⅠ和TβRⅡ的表达无明显影响。SA-B抑制原代正常培养9天的HSC和经PDGF-BB刺激的HSC中PDGFR-β的表达,但对PDGF-BB刺激的HSC中ERK1/2磷酸化没有明显作用。结论SA-B抑制TGF-β1诱导的HSC内ERK信号传导通路。这一抑制作用与HSC上TβR的表达无关,也与PDGF-BB在HSC内的ERK信号传导通路无关。SA-B对PDGF-BB在HSC内的信号传导通路的抑制作用在于抑制了HSC上PDGF受体的表达。

弥漫大B细胞淋巴瘤患者外周血CD14^+单核细胞TF、VEGF表达的临床意义

目的:探讨弥漫大B细胞淋巴瘤(DLBCL)患者外周血CD14^+单核细胞组织因子(TF)和血管内皮生长因子(VEGF)表达的临床意义。方法:应用流式细胞术检测41例DLBCL患者(病例组)化疗前、4个疗程化疗后以及20例健康体检者(对照组)外周血CD14^+单核细胞TF和VEGF的表达水平,分析TF和VEGF表达与国际预后指数(IPI)、短期疗效以及生存的关系。结果:病例组外周血CD14^+单核细胞TF、VEGF的表达水平显著高于对照组(P <0.01),2者表达呈正相关(r=0.755,P <0.01)。不同性别、年龄、亚型病例组CD14^+单核细胞TF、VEGF表达水平均无明显差异(P> 0.05)。具有预后危险因素患者CD14^+单核细胞TF、VEGF表达水平与不具有预后危险因素者比较均明显增高(P <0.05)。随着IPI指数的增高,病例组CD14^+单核细胞TF和VEGF表达均有递增趋势(P <0.01)。缓解组化疗前CD14^+单核细胞TF、VEGF表达水平低于未缓解组(P均<0.01);化疗后缓解组CD14^+单核细胞TF、VEGF表达水平均低于化疗前水平(P <0.01),未缓解组TF和VEGF表达水平较化疗前变化不大(P> 0.05)。TF和VEGF低表达组生存状况均优于TF、VEGF高表达组(P <0.05)。结论:DLBCL患者外周血CD14^+单核细胞高表达TF和VEGF,与国际预后指数、疗效、生存相关,对评估疗效、预后具有参考意义。