BACKGROUND Gastric cancer(GC)is one of the most common malignant tumors.Osteopontin(OPN)is thought to be closely related to the occurrence,metastasis and prognosis of many types of tumors.AIM To investigate the effect...BACKGROUND Gastric cancer(GC)is one of the most common malignant tumors.Osteopontin(OPN)is thought to be closely related to the occurrence,metastasis and prognosis of many types of tumors.AIM To investigate the effects of OPN on the proliferation,invasion and migration of GC cells and its possible mechanism.METHODS The mRNA and protein expression of OPN in the GC cells were analyzed by realtime quantitative-reverse transcription polymerase chain reaction and western blotting,and observe the effect of varying degree expression OPN on the proliferation and other behaviors of GC.Next,the effects of OPN knockdown on GC cells migration and invasion were examined.The short hairpin RNA(shRNA)and negative control shRNA targeting OPN-shRNA were transfected into the cells according to the manufacturer’s instructions.Non transfected cells were classified as control in the identical transfecting process.24 h after RNA transfection cell proliferation activity was detected by 3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-diphenytetrazoliumromide assay,and cell invasiveness and migration were detected by Trans well assay.Meanwhile,the expression of protein kinase B(AKT),matrix metalloproteinase 2(MMP-2)and vascular endothelial growth factor(VEGF)in the human GC cell lines was detected by reverse transcription polymerase chain reaction and western blotting.RESULTS The results of this study revealed that OPN mRNA and protein expression levels were highly expressed in SGC-7901 cells.OPN knockdown by specific shRNA noticeably reduced the capabilities of proliferation,invasion and migration of SGC-7901 cells.Moreover,in the experiments of investigating the underlying mechanism,results showed that OPN knockdown could down-regulated the expression of MMP-2 and VEGF,it also decreased the phosphorylation of AKT.Meanwhile,the protein expression levels of MMP-2,VEGF and phosphorylated AKT was noticeable lower than that in control group in the GC cells after they were added to phosphatidylinositol-3-kinase(PI3K)inhibitor(LY294002).CONCLUSION These results suggested that OPN though PI3K/AKT/mammalian target of rapamycin signal pathway to upregulate MMP-2 and VEGF expression,which contribute SGC-7901 cells to proliferation,invasion and migration.Thus,our results demonstrate that OPN may serve as a novel prognostic biomarkers as well as a potential therapeutic targets for GC.展开更多
目的研究鼻咽癌相关基因osteopontin SNP rs2728127和rs34527305对鼻咽癌易感性的影响。方法采用多重单碱基延伸PCR技术,对264例广西百色地区鼻咽癌患者和264例健康对照者的osteopontin基因SNP rs2728127和rs34527305进行基因分型,比较...目的研究鼻咽癌相关基因osteopontin SNP rs2728127和rs34527305对鼻咽癌易感性的影响。方法采用多重单碱基延伸PCR技术,对264例广西百色地区鼻咽癌患者和264例健康对照者的osteopontin基因SNP rs2728127和rs34527305进行基因分型,比较广西百色地区人群rs2728127和rs34527305基因型和等位基因型频率与其他地区人群的差异,分析rs2728127和rs34527305对鼻咽癌易感性的影响。结果广西百色地区人群osteopontin基因SNP rs2728127和rs34527305基因型和等位基因型频率与云南西双版纳地区人群差异无统计学意义(P>0.05),但是与北京、越南胡志明、日本东京[除rs34527305等位基因型频率与北京相比较差异无统计学意义(P>0.05)外]、秘鲁利马和芬兰地区人群相比差异有统计学意义(P<0.05)。对照组和病例组rs2728127和rs34527305基因型和等位基因型频率分布差异无统计学意义(P>0.05)。但是携带GA单倍型的个体相比携带AA单倍型个体罹患鼻咽癌的风险更低(OR=0.69,95%CI=0.48~0.96,P=0.038)。结论在广西百色地区人群中,osteopontin基因SNP rs2728127和rs34527305基因型和等位基因型可能与鼻咽癌的易感性无关,但是其GA单倍型可能降低鼻咽癌的风险。展开更多
AIM: To investigate the relationship between osteopontin plasma concentrations and the severity of portal hypertension and to assess osteopontin prognostic value.METHODS: A cohort of 154 patients with confirmed liver ...AIM: To investigate the relationship between osteopontin plasma concentrations and the severity of portal hypertension and to assess osteopontin prognostic value.METHODS: A cohort of 154 patients with confirmed liver cirrhosis(112 ethylic, 108 men, age 34-72 years)were enrolled in the study. Hepatic venous pressure gradient(HVPG) measurement and laboratory and ultrasound examinations were carried out for all patients. HVPG was measured using a standard catheterization method with the balloon wedge technique. Osteopontin was measured using the enzyme-linked immunosorbent assay(ELISA) method in plasma. Patients were followed up with a specific focus on mortality. The control group consisted of 137 healthy age- and sex- matched individuals.RESULTS: The mean value of HVPG was 16.18 ± 5.6 mm Hg. Compared to controls, the plasma levels of osteopontin in cirrhotic patients were significantly higher(P < 0.001). The plasma levels of osteopontin were positively related to HVPG(P = 0.0022, r = 0.25) and differed among the individual Child-Pugh groups of patients. The cut-off value of 80 ng/m L osteopontin distinguished patients with significant portal hypertension(HVPG above 10 mm Hg) at 75% sensitivity and 63% specificity. The mean follow-up of patients was 3.7 ± 2.6 years. The probability of cumulative survival was 39% for patients with HVPG > 10 mm Hg and 65% for those with HVPG ≤ 10 mm Hg(P = 0.0086, odds ratio(OR), 2.92, 95% confidence interval(CI): 1.09-7.76). Osteopontin showed a similar prognostic value to HVPG. Patients with osteopontin values above 80 ng/m L had significantly lower cumulative survival compared to those with osteopontin ≤ 80 ng/m L(37% vs 56%, P = 0.00035; OR = 2.23, 95%CI: 1.06-4.68).CONCLUSION: Osteopontin is a non-invasive parameter of portal hypertension that distinguishes patients with clinically significant portal hypertension. It is a strong prognostic factor for survival.展开更多
AIM:To investigate the effects of osteopontin(OPN)gene expression knockdown on colon cancer Lovo cells in vitro.METHODS:Four candidate small interfering RNA(siRNA)constructs targeting the OPN gene and a scrambled cont...AIM:To investigate the effects of osteopontin(OPN)gene expression knockdown on colon cancer Lovo cells in vitro.METHODS:Four candidate small interfering RNA(siRNA)constructs targeting the OPN gene and a scrambled control sequence(NC-siRNA)were synthesized and inserted into a pGPU6/GFP/Neo expression vector.After confirmation by restriction enzyme digestion and DNA sequencing,the recombinant plasmids were subsequently transfected into a human colon cancer cell line(Lovo)using a liposome transfection method.Stably transfected cells were maintained with G418 selection and referred to as Lovo-OPN-1,-2,-3,-4,and Lovo-NC cells.Knockdown efficiency of each of the four siRNA constructs was determined by realtime reverse transcription polymerase chain reaction assays and western blotting,and the construct with the most effective silencing was used for subsequent experiments.Cell proliferation,adhesion,and Matrigel invasion assays were performed to analyze the effects of OPN knockdown in stably transfected Lovo cells.The levels of four angiogenic factors,namely vascular endothelial growth factor(VEGF),matrix metalloproteinase(MMP)-2,MMP-9 and urokinase plasminogen activator were detected by enzyme-linked immunosorbent assays(ELISA).RESULTS:Recombinant vectors containing OPNspecific and scrambled siRNA sequences were successfully constructed and stably transfected into Lovo cells.Compared with the control Lovo and Lovo-NC cells,the levels of OPN mRNA and protein expression in LovoOPN-1,-2,-3,and-4 were significantly reduced(all P<0.05),with the most efficient reduction observed in Lovo-OPN-4 cells(P<0.05).Relative to untransfected Lovo cells,OPN mRNA expression levels in Lovo-NC and Lovo-OPN-4 cells were 1.008±0.067 and 0.160±0.023,respectively.The relative OPN protein expression levels in Lovo,Lovo-NC,and Lovo-OPN-4 cells were 3.024±0.211,2.974±0.630,and 0.121±0.008,respectively.Moreover,transfection with the scrambled sequence had no effect on the expression of OPN.After24,48,72,and 96 h of cultivation,absorption values at 450 nm to assess proliferation of Lovo-OPN-4 cells were 0.210±0.017,0.247±0.024,0.314±0.037,and 0.359±0.043,respectively,which were significantly lower than those of Lovo(0.244±0.031,0.313±0.024,0.513±0.048 and 0.783±0.051)and LovoNC cells(0.241±0.029,0.309±0.022,0.563±0.023,and 0.735±0.067)(all P<0.05).The absorption values at 595 nm,which were measured in a cell adhesion assay,showed that adhesion of Lovo-OPN-4 cells(0.215±0.036)was significantly decreased compared to Lovo(0.490±0.037)and Lovo-NC cells(0.462±0.043)(P<0.05).The number of invasive Lovo-OPN-4 cells(16.1±1.9)was also significantly decreased compared to Lovo(49.9±5.4)and Lovo-NC cells(48.8±4.5)(P<0.05).ELISA assays showed significant reductions in Lovo-OPN-4 cells compared to Lovo and Lovo-NC cells with regard to the expression of VEGF(1687.85±167.84 ng/L vs 2348.54±143.80 ng/L and 2284.39±138.62 ng/L,respectively),MMP-2(2966.07±177.36μg/L vs 4084.74±349.54μg/L and 4011.41±424.48μg/L,respectively),MMP-9(3782.89±300.64μg/L vs5062.90±303.02μg/L and 4986.38±300.75μg/L,respectively)and uPA(1152.69±120.79μg/L vs1380.90±147.25μg/L and 1449.80±189.92μg/L,respectively)(all P<0.05).CONCLUSION:Knockdown of OPN gene expression suppresses colon cancer cell growth,adherence,invasion,and expression of angiogenic factors.展开更多
AIM: To analyze plasma osteopontin levels and liver stiffness using transient elastography in postoperative biliary atresia (BA) children compared with healthy controls. METHODS: Thirty children with postoperative BA ...AIM: To analyze plasma osteopontin levels and liver stiffness using transient elastography in postoperative biliary atresia (BA) children compared with healthy controls. METHODS: Thirty children with postoperative BA and 10 normal controls were enrolled. The patients were categorized into two groups according to their jaundicestatus. Plasma levels of osteopontin were determined using commercially available enzyme-linked immunosorbent assay. Liver stiffness was measured by using transient elastography (Fibroscan). Ten validated Fibroscan measurements were performed in each patient and control with the result expressed in kilopascals (kPa). RESULTS: Plasma osteopontin was significantly elevated in BA children compared with that of healthy controls (47.0 ± 56.4 ng/mL vs 15.1 ± 15.0 ng/mL, P = 0.01). The liver stiffness measurement was markedly elevated in the patients with BA compared with that of controls (26.9 ± 24.6 kPa vs 3.9 ± 0.7 kPa, P = 0.001). Subgroup analysis showed that the BA patients with jaundice had more pronounced plasma osteopontin levels than those without jaundice (87.1 ± 61.6 ng/mL vs 11.9 ± 6.1 ng/mL, P = 0.001). Furthermore, the mean liver stiffness was significantly greater in the jaundiced BA patients compared with non-jaundiced patients (47.7 ± 21.8 kPa vs 8.7 ± 3.0 kPa, P = 0.001). Additionally, plasma osteopontin was positively related to serum total bilirubin (r = 0.64, P < 0.001). There was also a correlation between plasma osteopontin and liver stiffness values (r = 0.60, P < 0.001). CONCLUSION: High plasma osteopontin positively correlated with degree of hepatic fibrosis and could be used as a biochemical parameter reflecting disease severity in postoperative BA children.展开更多
Objective:To explore the significance of osteopontin and nuclear factorκB(NF-κB) expression in patients with knee osteoarthritis.Methods:RT-PCR and enzyme-linked immunosorbent assay were used to measure the Osteopon...Objective:To explore the significance of osteopontin and nuclear factorκB(NF-κB) expression in patients with knee osteoarthritis.Methods:RT-PCR and enzyme-linked immunosorbent assay were used to measure the Osteopontin(OPN) and NF-κB concentration of knee joint synovial fluid of patients with knee osteoarthritis and trauma fractures,and analyze the relationship between the expressiones of them.Results:OPN and NF-κB expression at the mRNA and protein levels of patients with knee osteoarthritis were significantly higher than the control group, the result showed statistical significance(P【0.05).There was a positive correlation between the OPN levels in synovial fluid of patients with knee osteoarthritis and NF-κB expression levels (P【0.05).Conclusions:The high expression of OPN and NF-κB are closely related to occurrence and development of knee osteoarthritis.展开更多
AIM: To analyze osteopontin (OPN) expression in vitreous and proliferative retinal membranes of patients with proliferative vitreous retinopathy (PVR). METHODS: A total of 54 vitreous fluid samples were obtained betwe...AIM: To analyze osteopontin (OPN) expression in vitreous and proliferative retinal membranes of patients with proliferative vitreous retinopathy (PVR). METHODS: A total of 54 vitreous fluid samples were obtained between 2009 and 2010, which contained 45 with PVR (group A) and 9 without PVR (group B). Enzyme-linked immunosorbent assay was applied to quantify the OPN concentrations in vitreous fluid. Four samples of proliferative retinal membrane were also obtained at the time of vitrectomy, and their contents of OPN were measured by Real-time RT-PCR. RESULTS: The OPN levels in the vitreous fluid were 778.48±62.06ng/mL in group A and 452.99±32.52ng/mL in group B. The vitreous OPN levels in group A were significantly higher than those in group B and to rise by time in the early stages of PVR. The average OPN levels in the proliferative retinal membranes (F =0.14) were also higher than those in the retinal pigment cells (F =0) using Real-time RT-PCR. CONCLUSION: The high vitreous and proliferative retinal membrane OPN levels in PVR suggest that OPN might promote the development of PVR. The vitreous OPN concentrations are rising by the time in the early phases of PVR.展开更多
Osteopontin(OPN)is an extracellular matrix protein with a diverse range of functions,including roles in cell adhesion,migration,and immunomodulation,which are associated with the modulation of neuroinflammation in the...Osteopontin(OPN)is an extracellular matrix protein with a diverse range of functions,including roles in cell adhesion,migration,and immunomodulation,which are associated with the modulation of neuroinflammation in the central nervous system.The present study was performed to evaluate the involvement of OPN in the eyes of an experimental autoimmune uveoretinitis(EAU)model.The EAU model was developed by immunization of Lewis rats with interphotoreceptor retinoid-binding protein.The results showed the OPN level was remarkably upregulated in the eye of EAU rats on day 9 post-immunization.The level of CD44,a ligand of OPN,was increased in the ciliary body of EAU rats.Furthermore,OPN was also detected in the ciliary body and activated microglia/macrophages in the EAU retina.The results suggest that OPN was significantly upregulated in the eyes of EAU rats,and that it may be useful as an early biomarker of ocular autoimmune diseases.All animal experiments were approved by the Institutional Animal Care and Use Committee of Jeju National University(approval No.2020-0012)on March 11,2020.展开更多
The influence of short hairpin RNA(shRNA)-mediated osteopontin(OPN)gene silencing on the proliferation and invasion of human renal cancer ACHN cells was investigated.Four types of OPN shRNA recombinant plasmids were c...The influence of short hairpin RNA(shRNA)-mediated osteopontin(OPN)gene silencing on the proliferation and invasion of human renal cancer ACHN cells was investigated.Four types of OPN shRNA recombinant plasmids were constructed and RT-PCR assays were used to screen the most highly functional shRNA recombinant plasmids,which were transferred into the cultured ACHN cells by LipofectamineTM 2000.The cells transfected by shRNA expression vectors(ACHN/OPN)were visualized under an inverted microscope and screened by G418.Untreated cells(ACHN)and cells transfected by mock vectors(ACHN/Vect)were used as control groups.The expression levels of OPN mRNA and protein were detected by real-time PCR and Western blot respectively.The cell cycle and ratios of apoptotic cells were assessed by flow cytometry.MTT method was used for drawing the growth curve and observing cell proliferation in vitro.The abilities of migration and invasion in three groups were measured by Transwell chamber test.The expression levels of matrix metalloproteinase(MMP)-2 and MMP-9 in three groups were examined by Western blot.Our results showed that the recombinant plasmid could be successfully transferred into ACHN cells by LipofectamineTM 2000.Compared with untreated cells,the expression levels of OPN mRNA and protein in ACHN/OPN cells were decreased by 59.68% and 76.42%,respectively(P<0.05),ACHN/OPN cells were blocked in S phase and apoptotic ratio increased significantly(P<0.05),however,no significant differences were found between ACHN/Vect and ACHN.Recombinant plasmid significantly attenuated expression levels of MMP-2 and MMP-9 proteins and suppressed the proliferation,migration,and invasion of ACHN cells.This study suggested that OPN may play an important role in the growth and invasion of human renal cancer ACHN cells,and these processes are correlated with the activations of MMP-2 and MMP-9.Our data provided preliminary experimental evidence for the feasibility of RNA interference technology in gene therapy of human renal cancer.展开更多
Objectives To observe the effects of perindopril on left ventricular remodeling and myocardial osteopontin expression in rats with myocardial infarction. Methods In this study male adult SD rats were randomly divided ...Objectives To observe the effects of perindopril on left ventricular remodeling and myocardial osteopontin expression in rats with myocardial infarction. Methods In this study male adult SD rats were randomly divided into 3 groups: sham-operation group, MI-saline group and MI-perindopril group. Left anterior descending artery was ligated to generate myocardial infarction. Perindopril (2 mg/kg body weight/day) was administered from the next day of MI. Four weeks later, left ventricular diameter (LVEDD and LVESD) and left ventricular ejection fraction was estimated with echocardiography, LVSP, LVEDP and±dp/dtmax was detected with hemodynamic measurement, cardiomyocyte diameter and interstitial fibrosis infiltration were evaluated with histological methods, and myocardium osteopontin protein expression level was detected with western blot. Results ①Compared with the sham-operation group, all rats with MI developed significant systolic and diastolic dysfunction, as was indicated by decreased LVEF, LVSP and±dp/dtmax, as well as increased LVEDP. ②Rats with MI showed significantly dilated left ventricles and higher ventricular weight / body weight ratio, significantly increased cardiomyocyte diameter and marked interstitial fibrosis in the non-infarction area. ③Perindopril treatment partly prevented cardiac dysfunction and left ventricular remodeling as indicated by the parameters mentioned above. ④No osteopontin protein was detected in myocardium of sham-operation rats. In rats with MI, high level osteopontin protein expression was significantly inhibited by perindopril treatment. Conclusions In rats with MI, perindopril treatment significantly prevented left ventricular remodeling and myocardium osteopontin protein expression.展开更多
Osteopontin (OPN) is a protein found at higher concentrations in the seminal plasma of bulls with above average fertility. Polymorphisms have been reported within the OPN gene promoter that can affect production of th...Osteopontin (OPN) is a protein found at higher concentrations in the seminal plasma of bulls with above average fertility. Polymorphisms have been reported within the OPN gene promoter that can affect production of this protein and thus, affect fertility. Therefore, Angus (n = 5) and Angus x Gelbvieh (Balancer, n = 14) and Angus x Brahman (n = 15) bulls were evaluated for presence of single nucleotide polymorphisms (SNP) in the Bos taurus OPN gene (GenBank: AY878328.1) promoter region, and their possible effects on bull semen quality as evaluated by computer-assisted semen analysis (CASA). Semen was collected by electroejaculation 6 to 9 times from each bull, and each semen collection was evaluated by CASA for motile, progressive and rapid sperm within 5 mins of ejaculation. The bulls were genotyped for reported single nucleotide polymorphisms (SNP) in the promoter region of the OPN gene through amplification of two 700 base pair (bp) DNA fragments and sequencing of the resulting PCR products. Seven SNP sites were identified, at bp 3379, 3490, 3492, 5075, 5205, 5209, and 5263 of the OPN gene. The SNP identified at bp 5205, 5209 and 5263 had not been previously reported. Individual SNP sites were evaluated as the main effect on CASA sperm motility variables in a SAS MIXED model for repeated measures. A thymine to guanine substitution at bp 3379 was associated with increased (P ≤ 0.02) percentage of motile, progressive and rapid sperm in Angus x Brahman bulls, and tended (P ≤ 0.10) to increase the same sperm motility parameters in Angus, and Angus x Gelbvieh bulls. The percentages of motile, progressive and rapid sperm were similar (P ≥ 0.05) among genotypes for the other 6 SNP identified. These results suggest that identification and genotyping of polymorphisms within the promoter region of the bovine OPN gene may be useful for selecting bulls with improved sperm motility parameters.展开更多
Objective:To analyze mRNA and protein expression of osteopontin(OPN)in human endome-trium throughout menstrual cycle.Methods:Immunohistochemieal method was used to determine the level and the location ofOPN in normal ...Objective:To analyze mRNA and protein expression of osteopontin(OPN)in human endome-trium throughout menstrual cycle.Methods:Immunohistochemieal method was used to determine the level and the location ofOPN in normal cycling endometrium of 50 women.Western Blot analysis was also used to detectthe existence of OPN at proliferative and secretory phases in six samples.The levels of OPNmRNA in 18 samples were measured by RT-PCR.Results:The OPN mainly expressed in human endometrial glandular epithelium but not instromal cells.The expression was highest at mid and late secretory phases and menstruous phaseas well.No expression was found in early and mid proliferative endometria.Two subtypes ofOPN proteins(40 kDa-75 kDa)were detected by Western blot analysis in homogenized endome-tria at secretory phase.The level of OPN mRNA in secretory phase endometrium was significantlyhigher than that in proliferative phase.Conclusions:OPN was only found in endometrial glandular epithelial cells,and the OPN andits mRNA showed a pronounced cycle-dependent expression in human endometrium,higher ex-pression at mid and late secretory phases and menstruous phase.It's probably involved in embry-onic implantation and endometrial shedding.展开更多
文摘BACKGROUND Gastric cancer(GC)is one of the most common malignant tumors.Osteopontin(OPN)is thought to be closely related to the occurrence,metastasis and prognosis of many types of tumors.AIM To investigate the effects of OPN on the proliferation,invasion and migration of GC cells and its possible mechanism.METHODS The mRNA and protein expression of OPN in the GC cells were analyzed by realtime quantitative-reverse transcription polymerase chain reaction and western blotting,and observe the effect of varying degree expression OPN on the proliferation and other behaviors of GC.Next,the effects of OPN knockdown on GC cells migration and invasion were examined.The short hairpin RNA(shRNA)and negative control shRNA targeting OPN-shRNA were transfected into the cells according to the manufacturer’s instructions.Non transfected cells were classified as control in the identical transfecting process.24 h after RNA transfection cell proliferation activity was detected by 3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-diphenytetrazoliumromide assay,and cell invasiveness and migration were detected by Trans well assay.Meanwhile,the expression of protein kinase B(AKT),matrix metalloproteinase 2(MMP-2)and vascular endothelial growth factor(VEGF)in the human GC cell lines was detected by reverse transcription polymerase chain reaction and western blotting.RESULTS The results of this study revealed that OPN mRNA and protein expression levels were highly expressed in SGC-7901 cells.OPN knockdown by specific shRNA noticeably reduced the capabilities of proliferation,invasion and migration of SGC-7901 cells.Moreover,in the experiments of investigating the underlying mechanism,results showed that OPN knockdown could down-regulated the expression of MMP-2 and VEGF,it also decreased the phosphorylation of AKT.Meanwhile,the protein expression levels of MMP-2,VEGF and phosphorylated AKT was noticeable lower than that in control group in the GC cells after they were added to phosphatidylinositol-3-kinase(PI3K)inhibitor(LY294002).CONCLUSION These results suggested that OPN though PI3K/AKT/mammalian target of rapamycin signal pathway to upregulate MMP-2 and VEGF expression,which contribute SGC-7901 cells to proliferation,invasion and migration.Thus,our results demonstrate that OPN may serve as a novel prognostic biomarkers as well as a potential therapeutic targets for GC.
基金Supported by The Internal Grant Agency of the Czech Ministry of Health(http://iga.mzcr.cz/public Web/),No.NT 12290/4the Charles University in Prague(http://www.cuni.cz/UKEN-1.html),No.SVV 260156/2015the Czech Ministry of Health(http://mzcr.cz),No.MZCR-RVO VFN64165
文摘AIM: To investigate the relationship between osteopontin plasma concentrations and the severity of portal hypertension and to assess osteopontin prognostic value.METHODS: A cohort of 154 patients with confirmed liver cirrhosis(112 ethylic, 108 men, age 34-72 years)were enrolled in the study. Hepatic venous pressure gradient(HVPG) measurement and laboratory and ultrasound examinations were carried out for all patients. HVPG was measured using a standard catheterization method with the balloon wedge technique. Osteopontin was measured using the enzyme-linked immunosorbent assay(ELISA) method in plasma. Patients were followed up with a specific focus on mortality. The control group consisted of 137 healthy age- and sex- matched individuals.RESULTS: The mean value of HVPG was 16.18 ± 5.6 mm Hg. Compared to controls, the plasma levels of osteopontin in cirrhotic patients were significantly higher(P < 0.001). The plasma levels of osteopontin were positively related to HVPG(P = 0.0022, r = 0.25) and differed among the individual Child-Pugh groups of patients. The cut-off value of 80 ng/m L osteopontin distinguished patients with significant portal hypertension(HVPG above 10 mm Hg) at 75% sensitivity and 63% specificity. The mean follow-up of patients was 3.7 ± 2.6 years. The probability of cumulative survival was 39% for patients with HVPG > 10 mm Hg and 65% for those with HVPG ≤ 10 mm Hg(P = 0.0086, odds ratio(OR), 2.92, 95% confidence interval(CI): 1.09-7.76). Osteopontin showed a similar prognostic value to HVPG. Patients with osteopontin values above 80 ng/m L had significantly lower cumulative survival compared to those with osteopontin ≤ 80 ng/m L(37% vs 56%, P = 0.00035; OR = 2.23, 95%CI: 1.06-4.68).CONCLUSION: Osteopontin is a non-invasive parameter of portal hypertension that distinguishes patients with clinically significant portal hypertension. It is a strong prognostic factor for survival.
基金Supported by Grants from the National Natural Science Foundation of China,No.81260364,No.81270472 and No.81070310the"Chunhui"Program of Ministry of Education in China,No.Z2012007the Natural Science Foundation of Inner Mongolian Autonomous Region,No.2012MS1123 and No.2013MS1132
文摘AIM:To investigate the effects of osteopontin(OPN)gene expression knockdown on colon cancer Lovo cells in vitro.METHODS:Four candidate small interfering RNA(siRNA)constructs targeting the OPN gene and a scrambled control sequence(NC-siRNA)were synthesized and inserted into a pGPU6/GFP/Neo expression vector.After confirmation by restriction enzyme digestion and DNA sequencing,the recombinant plasmids were subsequently transfected into a human colon cancer cell line(Lovo)using a liposome transfection method.Stably transfected cells were maintained with G418 selection and referred to as Lovo-OPN-1,-2,-3,-4,and Lovo-NC cells.Knockdown efficiency of each of the four siRNA constructs was determined by realtime reverse transcription polymerase chain reaction assays and western blotting,and the construct with the most effective silencing was used for subsequent experiments.Cell proliferation,adhesion,and Matrigel invasion assays were performed to analyze the effects of OPN knockdown in stably transfected Lovo cells.The levels of four angiogenic factors,namely vascular endothelial growth factor(VEGF),matrix metalloproteinase(MMP)-2,MMP-9 and urokinase plasminogen activator were detected by enzyme-linked immunosorbent assays(ELISA).RESULTS:Recombinant vectors containing OPNspecific and scrambled siRNA sequences were successfully constructed and stably transfected into Lovo cells.Compared with the control Lovo and Lovo-NC cells,the levels of OPN mRNA and protein expression in LovoOPN-1,-2,-3,and-4 were significantly reduced(all P<0.05),with the most efficient reduction observed in Lovo-OPN-4 cells(P<0.05).Relative to untransfected Lovo cells,OPN mRNA expression levels in Lovo-NC and Lovo-OPN-4 cells were 1.008±0.067 and 0.160±0.023,respectively.The relative OPN protein expression levels in Lovo,Lovo-NC,and Lovo-OPN-4 cells were 3.024±0.211,2.974±0.630,and 0.121±0.008,respectively.Moreover,transfection with the scrambled sequence had no effect on the expression of OPN.After24,48,72,and 96 h of cultivation,absorption values at 450 nm to assess proliferation of Lovo-OPN-4 cells were 0.210±0.017,0.247±0.024,0.314±0.037,and 0.359±0.043,respectively,which were significantly lower than those of Lovo(0.244±0.031,0.313±0.024,0.513±0.048 and 0.783±0.051)and LovoNC cells(0.241±0.029,0.309±0.022,0.563±0.023,and 0.735±0.067)(all P<0.05).The absorption values at 595 nm,which were measured in a cell adhesion assay,showed that adhesion of Lovo-OPN-4 cells(0.215±0.036)was significantly decreased compared to Lovo(0.490±0.037)and Lovo-NC cells(0.462±0.043)(P<0.05).The number of invasive Lovo-OPN-4 cells(16.1±1.9)was also significantly decreased compared to Lovo(49.9±5.4)and Lovo-NC cells(48.8±4.5)(P<0.05).ELISA assays showed significant reductions in Lovo-OPN-4 cells compared to Lovo and Lovo-NC cells with regard to the expression of VEGF(1687.85±167.84 ng/L vs 2348.54±143.80 ng/L and 2284.39±138.62 ng/L,respectively),MMP-2(2966.07±177.36μg/L vs 4084.74±349.54μg/L and 4011.41±424.48μg/L,respectively),MMP-9(3782.89±300.64μg/L vs5062.90±303.02μg/L and 4986.38±300.75μg/L,respectively)and uPA(1152.69±120.79μg/L vs1380.90±147.25μg/L and 1449.80±189.92μg/L,respectively)(all P<0.05).CONCLUSION:Knockdown of OPN gene expression suppresses colon cancer cell growth,adherence,invasion,and expression of angiogenic factors.
基金Supported by Ratchadapiseksompotch Fund, Faculty of Medicine, Chulalongkorn University, Thailand Research Fund, and the Commission on Higher Education
文摘AIM: To analyze plasma osteopontin levels and liver stiffness using transient elastography in postoperative biliary atresia (BA) children compared with healthy controls. METHODS: Thirty children with postoperative BA and 10 normal controls were enrolled. The patients were categorized into two groups according to their jaundicestatus. Plasma levels of osteopontin were determined using commercially available enzyme-linked immunosorbent assay. Liver stiffness was measured by using transient elastography (Fibroscan). Ten validated Fibroscan measurements were performed in each patient and control with the result expressed in kilopascals (kPa). RESULTS: Plasma osteopontin was significantly elevated in BA children compared with that of healthy controls (47.0 ± 56.4 ng/mL vs 15.1 ± 15.0 ng/mL, P = 0.01). The liver stiffness measurement was markedly elevated in the patients with BA compared with that of controls (26.9 ± 24.6 kPa vs 3.9 ± 0.7 kPa, P = 0.001). Subgroup analysis showed that the BA patients with jaundice had more pronounced plasma osteopontin levels than those without jaundice (87.1 ± 61.6 ng/mL vs 11.9 ± 6.1 ng/mL, P = 0.001). Furthermore, the mean liver stiffness was significantly greater in the jaundiced BA patients compared with non-jaundiced patients (47.7 ± 21.8 kPa vs 8.7 ± 3.0 kPa, P = 0.001). Additionally, plasma osteopontin was positively related to serum total bilirubin (r = 0.64, P < 0.001). There was also a correlation between plasma osteopontin and liver stiffness values (r = 0.60, P < 0.001). CONCLUSION: High plasma osteopontin positively correlated with degree of hepatic fibrosis and could be used as a biochemical parameter reflecting disease severity in postoperative BA children.
文摘Objective:To explore the significance of osteopontin and nuclear factorκB(NF-κB) expression in patients with knee osteoarthritis.Methods:RT-PCR and enzyme-linked immunosorbent assay were used to measure the Osteopontin(OPN) and NF-κB concentration of knee joint synovial fluid of patients with knee osteoarthritis and trauma fractures,and analyze the relationship between the expressiones of them.Results:OPN and NF-κB expression at the mRNA and protein levels of patients with knee osteoarthritis were significantly higher than the control group, the result showed statistical significance(P【0.05).There was a positive correlation between the OPN levels in synovial fluid of patients with knee osteoarthritis and NF-κB expression levels (P【0.05).Conclusions:The high expression of OPN and NF-κB are closely related to occurrence and development of knee osteoarthritis.
基金National Natural Science Foundation of China(No. 30973257and 81070743)Research Found of Jiangsu Health Department, China (No. H200908)
文摘AIM: To analyze osteopontin (OPN) expression in vitreous and proliferative retinal membranes of patients with proliferative vitreous retinopathy (PVR). METHODS: A total of 54 vitreous fluid samples were obtained between 2009 and 2010, which contained 45 with PVR (group A) and 9 without PVR (group B). Enzyme-linked immunosorbent assay was applied to quantify the OPN concentrations in vitreous fluid. Four samples of proliferative retinal membrane were also obtained at the time of vitrectomy, and their contents of OPN were measured by Real-time RT-PCR. RESULTS: The OPN levels in the vitreous fluid were 778.48±62.06ng/mL in group A and 452.99±32.52ng/mL in group B. The vitreous OPN levels in group A were significantly higher than those in group B and to rise by time in the early stages of PVR. The average OPN levels in the proliferative retinal membranes (F =0.14) were also higher than those in the retinal pigment cells (F =0) using Real-time RT-PCR. CONCLUSION: The high vitreous and proliferative retinal membrane OPN levels in PVR suggest that OPN might promote the development of PVR. The vitreous OPN concentrations are rising by the time in the early phases of PVR.
基金supported by the National Research Foundation of Korea,No.NRF-2019R1A2C1087753(to TS)。
文摘Osteopontin(OPN)is an extracellular matrix protein with a diverse range of functions,including roles in cell adhesion,migration,and immunomodulation,which are associated with the modulation of neuroinflammation in the central nervous system.The present study was performed to evaluate the involvement of OPN in the eyes of an experimental autoimmune uveoretinitis(EAU)model.The EAU model was developed by immunization of Lewis rats with interphotoreceptor retinoid-binding protein.The results showed the OPN level was remarkably upregulated in the eye of EAU rats on day 9 post-immunization.The level of CD44,a ligand of OPN,was increased in the ciliary body of EAU rats.Furthermore,OPN was also detected in the ciliary body and activated microglia/macrophages in the EAU retina.The results suggest that OPN was significantly upregulated in the eyes of EAU rats,and that it may be useful as an early biomarker of ocular autoimmune diseases.All animal experiments were approved by the Institutional Animal Care and Use Committee of Jeju National University(approval No.2020-0012)on March 11,2020.
基金supported by a grant from the Major State Basic Research Development Program of China(973 Program)(No.2002CB513100)
文摘The influence of short hairpin RNA(shRNA)-mediated osteopontin(OPN)gene silencing on the proliferation and invasion of human renal cancer ACHN cells was investigated.Four types of OPN shRNA recombinant plasmids were constructed and RT-PCR assays were used to screen the most highly functional shRNA recombinant plasmids,which were transferred into the cultured ACHN cells by LipofectamineTM 2000.The cells transfected by shRNA expression vectors(ACHN/OPN)were visualized under an inverted microscope and screened by G418.Untreated cells(ACHN)and cells transfected by mock vectors(ACHN/Vect)were used as control groups.The expression levels of OPN mRNA and protein were detected by real-time PCR and Western blot respectively.The cell cycle and ratios of apoptotic cells were assessed by flow cytometry.MTT method was used for drawing the growth curve and observing cell proliferation in vitro.The abilities of migration and invasion in three groups were measured by Transwell chamber test.The expression levels of matrix metalloproteinase(MMP)-2 and MMP-9 in three groups were examined by Western blot.Our results showed that the recombinant plasmid could be successfully transferred into ACHN cells by LipofectamineTM 2000.Compared with untreated cells,the expression levels of OPN mRNA and protein in ACHN/OPN cells were decreased by 59.68% and 76.42%,respectively(P<0.05),ACHN/OPN cells were blocked in S phase and apoptotic ratio increased significantly(P<0.05),however,no significant differences were found between ACHN/Vect and ACHN.Recombinant plasmid significantly attenuated expression levels of MMP-2 and MMP-9 proteins and suppressed the proliferation,migration,and invasion of ACHN cells.This study suggested that OPN may play an important role in the growth and invasion of human renal cancer ACHN cells,and these processes are correlated with the activations of MMP-2 and MMP-9.Our data provided preliminary experimental evidence for the feasibility of RNA interference technology in gene therapy of human renal cancer.
文摘Objectives To observe the effects of perindopril on left ventricular remodeling and myocardial osteopontin expression in rats with myocardial infarction. Methods In this study male adult SD rats were randomly divided into 3 groups: sham-operation group, MI-saline group and MI-perindopril group. Left anterior descending artery was ligated to generate myocardial infarction. Perindopril (2 mg/kg body weight/day) was administered from the next day of MI. Four weeks later, left ventricular diameter (LVEDD and LVESD) and left ventricular ejection fraction was estimated with echocardiography, LVSP, LVEDP and±dp/dtmax was detected with hemodynamic measurement, cardiomyocyte diameter and interstitial fibrosis infiltration were evaluated with histological methods, and myocardium osteopontin protein expression level was detected with western blot. Results ①Compared with the sham-operation group, all rats with MI developed significant systolic and diastolic dysfunction, as was indicated by decreased LVEF, LVSP and±dp/dtmax, as well as increased LVEDP. ②Rats with MI showed significantly dilated left ventricles and higher ventricular weight / body weight ratio, significantly increased cardiomyocyte diameter and marked interstitial fibrosis in the non-infarction area. ③Perindopril treatment partly prevented cardiac dysfunction and left ventricular remodeling as indicated by the parameters mentioned above. ④No osteopontin protein was detected in myocardium of sham-operation rats. In rats with MI, high level osteopontin protein expression was significantly inhibited by perindopril treatment. Conclusions In rats with MI, perindopril treatment significantly prevented left ventricular remodeling and myocardium osteopontin protein expression.
文摘Osteopontin (OPN) is a protein found at higher concentrations in the seminal plasma of bulls with above average fertility. Polymorphisms have been reported within the OPN gene promoter that can affect production of this protein and thus, affect fertility. Therefore, Angus (n = 5) and Angus x Gelbvieh (Balancer, n = 14) and Angus x Brahman (n = 15) bulls were evaluated for presence of single nucleotide polymorphisms (SNP) in the Bos taurus OPN gene (GenBank: AY878328.1) promoter region, and their possible effects on bull semen quality as evaluated by computer-assisted semen analysis (CASA). Semen was collected by electroejaculation 6 to 9 times from each bull, and each semen collection was evaluated by CASA for motile, progressive and rapid sperm within 5 mins of ejaculation. The bulls were genotyped for reported single nucleotide polymorphisms (SNP) in the promoter region of the OPN gene through amplification of two 700 base pair (bp) DNA fragments and sequencing of the resulting PCR products. Seven SNP sites were identified, at bp 3379, 3490, 3492, 5075, 5205, 5209, and 5263 of the OPN gene. The SNP identified at bp 5205, 5209 and 5263 had not been previously reported. Individual SNP sites were evaluated as the main effect on CASA sperm motility variables in a SAS MIXED model for repeated measures. A thymine to guanine substitution at bp 3379 was associated with increased (P ≤ 0.02) percentage of motile, progressive and rapid sperm in Angus x Brahman bulls, and tended (P ≤ 0.10) to increase the same sperm motility parameters in Angus, and Angus x Gelbvieh bulls. The percentages of motile, progressive and rapid sperm were similar (P ≥ 0.05) among genotypes for the other 6 SNP identified. These results suggest that identification and genotyping of polymorphisms within the promoter region of the bovine OPN gene may be useful for selecting bulls with improved sperm motility parameters.
文摘Objective:To analyze mRNA and protein expression of osteopontin(OPN)in human endome-trium throughout menstrual cycle.Methods:Immunohistochemieal method was used to determine the level and the location ofOPN in normal cycling endometrium of 50 women.Western Blot analysis was also used to detectthe existence of OPN at proliferative and secretory phases in six samples.The levels of OPNmRNA in 18 samples were measured by RT-PCR.Results:The OPN mainly expressed in human endometrial glandular epithelium but not instromal cells.The expression was highest at mid and late secretory phases and menstruous phaseas well.No expression was found in early and mid proliferative endometria.Two subtypes ofOPN proteins(40 kDa-75 kDa)were detected by Western blot analysis in homogenized endome-tria at secretory phase.The level of OPN mRNA in secretory phase endometrium was significantlyhigher than that in proliferative phase.Conclusions:OPN was only found in endometrial glandular epithelial cells,and the OPN andits mRNA showed a pronounced cycle-dependent expression in human endometrium,higher ex-pression at mid and late secretory phases and menstruous phase.It's probably involved in embry-onic implantation and endometrial shedding.
