ENAM gene polymorphisms associated with dental anomalies in individuals with cleft lip and palate

Aim: This study aimed to investigate the occurrence of enamelin gene (ENAM) single nucleotide polymorphisms (SNP) and ENAM polymorphism association with dental anomalies (DA) in individuals with unilateral or bilateral cleft lip and palate (CLP). Methods: Saliva samples were collected from 147 individuals aged between 6 and 15 years-old, both genders, and divided into 4 groups: Group 1 (G1) - CLP and DA; Group 2 (G2) - CLP without DA; Group 3 (G3) - without CLP with DA; Group 4 (G4) - without CLP and DA. The genomic DNA was extracted from saliva samples and the following ENAM SNPs markers were genotyped: rs3796703, rs3796704, rs3796705, rs7671281, rs2609428, and rs35951442. Fisher exact and Pearson's Chi-square tests statistically analyzed the results (α=5%). Results: Individuals without CLP with DA (Group 3 - 19.2%) showed statistically higher prevalence of SNP rs2609428 heterozygotes (p=0.006) than individuals with CLP and DA (Group 1 - 0%). Individuals without CLP (10%) exhibited statistically higher prevalence of mutated heterozygotes/homozygous (p=0.028) than in individuals with CLP (1.3%). Conclusion: SNP rs2609428 marker of ENAM gene may be associated with dental anomalies in individuals without cleft lip and palate

Evaluation of the fibulin 5 gene polymorphism as a factor related to the occurrence of pelvic organ prolapse

SUMMARY OBJECTIVE Pelvic organ prolapse (POP) is a very frequent situation in our population that may lead to a significant decrease in patients' quality of life. Currently, we are looking for predictive factors for the development of POPs; thus, this study seeks to evaluate whether the Fibulin 5 polymorphism (FBLN5) is associated with the occurrence of POP. METHODS This is a cohort study with postmenopausal women who were divided into groups by POP stage: POP stages 0 and I (control group) and POP stages III and IV (case group). Subsequently, analyses of genetic polymorphisms of FBLN5 were performed using the Restriction Fragment Length Polymorphism (RFLP) technique. RESULTS A total of 292 women were included in the study. Pregnancy, parity and vaginal delivery in the patients, as well as in data described in the literature, were related to the occurrence of POP in the univariate analysis. However, after binary logistic regression, home birth and age remained independent risk factors for POP. We found no association between the FBLN5 polymorphism and the occurrence of POP (p = 0.371). CONCLUSION There was no association between the FBLN5 polymorphism and the occurrence of POP in Brazilian women.

RESUMO OBJETIVOS O prolapso de órgãos pélvicos (POP) é uma situação muito frequente em nossa população que pode levar a uma diminuição significativa da qualidade de vida dos pacientes. Atualmente, buscam-se fatores preditivos para o desenvolvimento de POPs e, assim, este estudo correlaciona um polimorfismo de Fibulina 5 (FBLN5) com a ocorrência da doença. MÉTODOS Estudo de coorte com mulheres na pós-menopausa, divididas por grupos pelos estádios 0 e I do POP (grupo controle) e POP III e IV (grupo caso). Posteriormente, análises do polimorfismo genético de FBLN5 foram realizadas utilizando a técnica de Polimorfismo de Comprimento de Fragmentos de Restrição (RFLP). RESULTADOS Um total de 292 mulheres foi incluído no estudo. Gestação, paridade e parto vaginal, como bem descritos na literatura, foram relacionados à ocorrência de POPs na análise univariada. No entanto, após a regressão logística binária, o parto domiciliar e a idade permaneceram como fatores de risco independentes para os POPs. Não encontramos associação deste polimorfismo FBLN5 com a ocorrência de POP (p=0,371). CONCLUSÃO Não houve associação deste polimorfismo FBLN5 com a ocorrência de POPs em mulheres brasileiras.

Expression of Mindin, a pattern recognition molecule, in a mouse model of acute liver injury induced by carbon tetrachloride / 临床肝胆病杂志

ObjectiveTo investigate the dynamic expression of Mindin protein in mice with acute liver injury induced by carbon tetrachloride (CCl4) and its mechanism of action. MethodsA total of 48 male C57B1/6 mice were selected and divided into experimental group (n=40) and control group (n=8). The mice in the experimental group were given intraperitoneal injection of CCl4 olive oil to induce acute liver injury, and the liver tissue was collected for pathological observation at 6, 12, 24, 48, and 72 hours after modeling. The mice in the control group were given intraperitoneal injection of olive oil. Western blot was used to measure the protein expression of Mindin and its change trend, and quantitative real-time PCR was used to measure the mRNA expression of Mindin. The t-test was used for comparison of continuous data between two groups. ResultsAbnormal liver structure was observed after liver injury was induced by CCl4, with the most significant pathological change at 48 hours. There was a low protein expression level of Mindin within 12-72 hours after the injection of CCl4. The mRNA expression of Mindin reached the lowest level at 12 hours after CCl4 injection and there was a significant difference between the experimental group and the control group at this time point (0.183±0.105 vs 1.023±0.247, t=8.841, P＜0.01); then there was a gradual increase in the mRNA expression of Mindin, and at 48 and 72 hours, the mRNA expression of Mindin in the experimental group was more than 2 times that in the control group (48 hours: 2.548±0.775 vs 1.023±0.247, t=5.428, P＜0.01; 72 hours: 2.699±0995 vs 1.023±0.247, t=4.621, P＜0.01). ConclusionThere is a significant change in Mindin protein during the process of acute liver injury induced by CCl4 in mice, which is characterized by the downregulation of protein expression and the regulation of post-transcriptional mRNA level, suggesting that Mindin may play an important role in the process of acute liver injury induced by CCl4.

Direct-acting antivirals-based therapy decreases hepatic fibrosis serum biomarker microfibrillar-associated protein 4 in hepatitis C patients

BACKGROUND/AIMS: An estimated 80 million people worldwide are infected with viremic hepatitis C virus (HCV). Even after eradication of HCV with direct acting antivirals (DAAs), hepatic fibrosis remains a risk factor for hepatocarcinogenesis. Recently, we confirmed the applicability of microfibrillar-associated protein 4 (MFAP4) as a serum biomarker for the assessment of hepatic fibrosis. The aim of the present study was to assess the usefulness of MFAP4 as a biomarker of liver fibrosis after HCV eliminating therapy with DAAs. METHODS: MFAP4 was measured using an immunoassay in 50 hepatitis C patients at baseline (BL), the end-of-therapy (EoT), and the 12-week follow-up (FU) visit. Changes in MFAP4 from BL to FU and their association with laboratory parameters including alanine aminotransferase (ALT), aspartate aminotransferase (AST), platelets, the AST to platelet ratio index (APRI), fibrosis-4 score (FIB-4), and albumin were analyzed. RESULTS: MFAP4 serum levels were representative of the severity of hepatic fibrosis at BL and correlated well with laboratory parameters, especially APRI (Spearman correlation, R²=0.80). Laboratory parameters decreased significantly from BL to EoT. MFAP4 serum levels were found to decrease from BL and EoT to FU with high statistical significance (Wilcoxon p<0.001 for both). CONCLUSIONS: Our findings indicate that viral eradication resulted in reduced MFAP4 serum levels, presumably representing a decrease in hepatic fibrogenesis or fibrosis. Hence, MFAP4 may be a useful tool for risk assessment in hepatitis C patients with advanced fibrosis after eradication of the virus.

CCN3 inhibits deposition of extracellular matrix of glomerular mesangial cells via promoting the expression of microRNA-29a / 中华肾脏病杂志

Objective To investigate the effects of nephroblastoma over-expressed protein (CCN3) on the formation of extracellular matrix (ECM) induced by transforming growth factor-β1 (TGF-β1) in human mesangial cells (HMCs) and its underlying signal transduction mechanism related with microRNA-29(miRNA-29).Methods HMCs were pretreated with different doses of exogenous CCN3 (5 μg/L,50 μg/L and 500 μg/L) or transfected with pcDNA3.1(+)-CCN3 before exposed to TGF-β1(2 μg/L),to observe the expression of fibronectin (FN),type I collagen (COL I) and miRNA-29a,b and c.The mimics or inhibitor of the miRNA-29a were transfected into HMCs to analyze whether miRNA-29a affect CCN3.The expressions of FN mRNA,COL I mRNA and miRNA-29 family were detected by real time PCR.The protein expressions of FN and COL I were detected by Western blotting and cell immunofluorescence.Results (1) Compared with the normal control group,the expressions of FN and COL I were up-regulated in TGF-β1 group,while the expressions of miRNA-29a,b,c were down-regulated in TGF-β1 group (all P ＜ 0.05).(2) Compared with the TGF-β1 group,the expressions of FN and COL I were decreased when pretreated with the different doses of exogenous of CCN3 or transfected with pcDNA3.1(+)-CCN3 (all P ＜ 0.05).Meanwhile,the expression of miRNA-29a was significantly increased when pretreated with 50 μg/L and 500 μg/L CCN3 or transfected with pcDNA3.1(+)-CCN3 (all P ＜ 0.05);whereas miRNA-29b and c had no statistical difference (all P ＞ 0.05).(3) Compared with TGF-β1+CCN3 group,the expressions of FN and COL I were decreased in CCN3+TGF-β1+miRNA-29a mimics group (all P ＜ 0.05),whereas the expressions of FN and COL I in CCN3+TGF-β1+miRNA-29a inhibitors group were increased (all P ＜ 0.05).Conclusions CCN3 reduces the TGF-β1-induced production of ECM by the up-regulation of miRNA-29a.

Effect of thymosin β4 on transforming growth factor-β/connective tissue growth factor of renal tubular interstitial fibrosis rats / 中华肾脏病杂志

Objective To investigate the influence of thymosin beta 4 (Tβ4) with two different dosages on the expression of transforming growth factor beta (TGF-β) and connective tissue growth factor (CTGF) in rats with renal tubular interstitial fibrosis,and to further estimate the changes of renal tubular interstitial lesions.Methods Rat models of renal tubular interstitial fibrosis were established by unilateral ureteral occlusion (UUO).The male SD rats were randomly divided into 4 groups and 15 rats in each group:sham group,model group,treatment group with 1 mg/L Tβ4 and treatment group with 5 mg/L T34.Rats in sham group and model group were poured into the same amount of saline.The renal function and renal pathological changes were observed after the second week.The mRNA and protein expression of TGF-β and CTGF in renal tissues was tested by in-situ hybridization and Western blotting.Results Compared with that in sham group,the expression of TGF-β mRNA and its protein,CTGF mRNA and its protein was significantly higher in model group (all P ＜ 0.01).Compared with rats of model group,Tβ4 treatment rats had lower mRNA and protein expression of TGF-β and CTGF (all P ＜ 0.01),and the expression in treatment group with 5 mg/L Tβ4 was lower than that in treatment group with 1 mg/L Tβ4 (P ＜ 0.05).And the expression of TGF-β mRNA was positively correlated with CTGF mRNA expression (r=0.697,P ＜ 0.01).The 24 h total urinary protein and the area of renal tubular interstitial lesion in model group were significantly more than those in sham group,and also more than those in Tβ4 treatment group (all P ＜ 0.05).Tβ4 treatment attenuated kidney damage,and the effects in treatment group with 5 mg/L Tβ4 were better than those in treatment group with 1 mg/L Tβ4.No difference in serum creatinine and blood urea nitrogen was observed among 4 groups (all P ＞ 0.05).Conclusions Tβ4 treatment can inhibit the renal TGF-β and CTGF expression of rats with tubular interstitial fibrosis in a dose-dependent manner,and play a protective role in kidney.

Effects of cartilage oligomeric matrix protein on BMP-2 induced cell differentiation of mesenchymal stem cells / 中华骨科杂志

Objective To investigate the effect of overexpression of cartilage oligomeric matrix protein (COMP) on BMP-2 induced cell osteogenic and chondrogenic differentiation of mesenchymal stem cells (MSCs).Methods MSCs,transfected with plasmid DNA encoding recombinant human COMP,were induced to differentiate into osteocytes and chondrocytes by BMP-2.Realtime PCR of osteogenic related markers (Col1a1,RUNX2,OPN,BGP) and chondrogenic related markers (Col2a1,SOX9,Aggrecan) were performed to evaluate the process of cell differentiation.ALP staining,Alizarin red S staining for osteogenic differentiation and alcian blue staining for chandrogenic differentiation were conducted to evaluate the tendency of cell differentiation.Results Real-time PCR assay presented the significantly higher (P＜0.05) COMP expression of MSCs when COMP gene was transfected into cells.The expression level of OPN was significantly (P＜0.05) down-regulated at all the time points in experimental group compared with that in control group.A final significant (P＜0.05) up-regulation of expression appeared in experimental group at the late stage of induction (day 7,14) compared with that in control group,even though a decrease (P＜0.05) expression of Col1a1,RUNX2 and BGP in experimental group occurred at the early stage of induction (day 3).The expression of Aggrecan and Col2a1 in experimental group was up-regulated (P＜0.05) at different time points compared with that in control group.And a significant higher (P＜0.05) expression of SOX9 in experimental group only appeared at day 7 compared with that in control group.ALP staining and Alizarin red S staining were weakened while alcian blue staining was enhanced.Conclusion COMP may inhibit BMP-2 induced osteogenic differentiation and promote BMP-2 induced chondrogenic differentiation,which may provide new insight for cartilage tissue engineering.

Clinical investigation and mutation analysis of ECM1 gene in a family with lipoid proteinosis / 中华皮肤科杂志

Objective To report a family with lipoid proteinosis (LP) from Shandong province and to analyze mutations in the extracellular matrix protein 1 (ECM1) gene in this family.Methods Eight members in a threegeneration family with LP were clinically investigated,and two patients were identified to suffer from LP,including the proband (Ⅲ 1) and her mother (Ⅱ 2).Both of the patients presented with papules on the palpebral margin,short and thick lingual frenum,and hoarseness.Indirect laryngoscopy showed infiltrating and thickening of the vocal cord.Pathological examination of lesions on the palpebral margin and laryngeal mucosa revealed deposits of hyaline-like material in the dermis,which was strongly positive for periodic acid-Schiff (PAS) staining and resistant to diastase digestion.The pathological diagnosis was LP.Blood samples were collected from all the family members and 100 ethnically matched,unrelated and unaffected Chinese human controls followed by DNA extraction.PCR and sequencing were performed to detect the ECM1 gene,and nested PCR followed by agarose gel electrophoresis to analyze mutations in the coding region of the ECM1 gene.Results Both of the two patients were compound heterozygotes.Three missense mutations,incluing p.P169T,p.A44T and p.R392W,were found in the ECM1 gene of the affected mother,with p.P169T in one allele and p.A44T as well as p.R392W in the other.The girl patient inheried the missence mutation p.P169T from her mother and a synonymous mutation c.879G ＞ A from her father (Ⅱ 1).Nested PCR showed that the c.978G ＞ A mutation generated a splice-acceptor site AG,which leaded to a splicing defect.Conclusion A novel synonymous splice-acceptor site mutation c.879G ＞ A in the ECM1 gene is identified in the family with LP.

The relationship of Slit2 and bone marrow mesenchymal stem cells with the angiogenesis / 中国组织工程研究

BACKGROUND:Bone marrow mesenchyme stem cells are important non-hematopoietic stem cells in the bone marrow, which can stimulate angiogenesis. While, Slit can also stimulate angiogenesis, as many studies have proved. OBJECTIVE:To review the biological functions, clinical application and effects of bone marrow mesenchymal stem cells and Slit2 on promoting angiogenesis. METHODS:A computer-based online research of CNKI and PubMed databases was performed to col ect articles published between 1980 and 2014 with the keywords“MSCs”and“Slit2”in Chinese and English. There were 436 articles after the initial survey. Final y, 65 articles were included according inclusion and exclusion criteria. RESULTS AND CONCLUSION:Both bone marrow mesenchymal stem cells and Slit2 play an important role in promoting angiogenesis, but the relevance of bone marrow mesenchymal stem cells and Slit2 is stil controversial. If assuming that bone marrow mesenchymal stem cells secrete Slit2, more researches should be done to reveal whether bone marrow mesenchymal stem cells promoting angiogenesis is relevant to Slit2 and through which signaling pathway Slit2/Robo functions to adjust bone marrow mesenchymal stem cells thus to promote angiogenesis. If relevant, the transplantation of the Slit2 and bone marrow mesenchymal stem cells wil be a promising treatment of cerebral infarction and other central nervous injuries.

Fisetin attenuates diabetic nephropathy by regulating transcriptional coactivator p300 and matrix metalloproteinase-2 / 中华内分泌代谢杂志

Rat models of diabetes were established by injecting streptozocin intraperitoneally.According to random number table,three groups were divided:normal group,diabetic group,and fisetin-treated group.After 24 weeks,all rats were sacrificed.Biochemical parameters of blood and urine samples were tested.The pathological changes were observed by paraffin sections staining with HE.The expression of extracellular matrix proteins was analyzed via PAS and Masson staining.Location of p300 protein expression was analyzed by immunohistochemistry.The protein expressions of p300 and MMP-2 were determined by Western blotting.The mRNA expressions of MMP-2 were analyzed via real-time PCR.The biochemical parameters and kidney pathological images in fisetin-treated group were better than those in diabetic group.The expression of extraeellular matrix proteins was lower than that in diabetic group.Immunohistochemistry analysis showed that among three groups the expression of p300 was mainly in glomeruli,and was also expressed in cell nucleus and cytoplasm and the coloration of fisetin-treated group was weakened as compared with diabetic group.Western blotting analysis showed that the expression of p300 protein in fisetin-treated group was lower than that in diabetic group(P＜O.05).The expressions of MMP-2 mRNA and MMP-2 protein were higher than those in diabetic group (P ＜ 0.05).It is suggested that fisetin may attenuate diabetes associated abnormalities in the kidney of rats,owing probably to inhibiting the expressions of p300 and enhancing the expressions of MMP-2.

Cysteinyl Cathepsins: Multifunctional Enzymes in Cardiovascular Disease / 전남의대학술지

Until recently, the role of lysosomal cysteine protease cathepsins in intracellular protein degradation was believed to be mainly restricted to scavenging. However, recent studies have revealed nontraditional roles for cysteine protease cathepsins in the extracellular space during the development and progression of cardiovascular disease. Although the precise mechanisms are unknown, data from animal studies suggest that members of the cathepsin family, like other extracellular proteases, contribute to extracellular matrix protein remodeling and interstitial matrix degradation, as well as to cell signaling and cell apoptosis in heart disease. Inflammatory cytokines and hormones regulate the expression and secretion of cathepsins in cultured cardiovascular cells and macrophages. Serum levels of cathepsins L, S, and K and their endogenous inhibitor cystatin C may be useful predictive biomarkers in patients with coronary artery disease and cardiac disease. Furthermore, in vivo pharmacological intervention with a synthetic cathepsin inhibitor and cardiovascular drugs (including statins and angiotensin II type 1 receptor antagonists) has the potential for pharmacologic targeting of cathepsins in cardiovascular disease. This review focuses on cathepsin biology (structure, synthesis, processing, activation, secretion, activity regulation, and function) and the involvement of cysteinyl cathepsins in the pathogenesis of several heart and vessel diseases, especially with respect to their potential application as diagnostic and prognostic markers and drug targets to prevent inappropriate proteolysis in cardiovascular disease.

Cysteinyl Cathepsins: Multifunctional Enzymes in Cardiovascular Disease / 전남의대학술지

Until recently, the role of lysosomal cysteine protease cathepsins in intracellular protein degradation was believed to be mainly restricted to scavenging. However, recent studies have revealed nontraditional roles for cysteine protease cathepsins in the extracellular space during the development and progression of cardiovascular disease. Although the precise mechanisms are unknown, data from animal studies suggest that members of the cathepsin family, like other extracellular proteases, contribute to extracellular matrix protein remodeling and interstitial matrix degradation, as well as to cell signaling and cell apoptosis in heart disease. Inflammatory cytokines and hormones regulate the expression and secretion of cathepsins in cultured cardiovascular cells and macrophages. Serum levels of cathepsins L, S, and K and their endogenous inhibitor cystatin C may be useful predictive biomarkers in patients with coronary artery disease and cardiac disease. Furthermore, in vivo pharmacological intervention with a synthetic cathepsin inhibitor and cardiovascular drugs (including statins and angiotensin II type 1 receptor antagonists) has the potential for pharmacologic targeting of cathepsins in cardiovascular disease. This review focuses on cathepsin biology (structure, synthesis, processing, activation, secretion, activity regulation, and function) and the involvement of cysteinyl cathepsins in the pathogenesis of several heart and vessel diseases, especially with respect to their potential application as diagnostic and prognostic markers and drug targets to prevent inappropriate proteolysis in cardiovascular disease.

Effect of estrogen on the expression of matrix GLA protein in ovariectomized SD rats / 中华妇产科杂志

Objective To investigate the effect of estrogen on expression of matrix GLA protein (MGP)in ovariectomized Sprague-Dawley(SD)rats and the role of estrogen in improving postmenopausal osteoporosis.Methods Thirty-six SD female rats were allocated into 3 groups randomly,every 12 rats in ovariectomized group(OVX group),estrogen group(E group)and control group(sham group).Rats in OVX and E group all underwent bilateral ovariectomy,those rats in E group were given by estradiol benzoate intramuscularly after 3 weeks of ovariectomy.Rats in sham group underwent bilateral lipectomy near the ovary.All rats were kept the urine and the serum every three weeks and were sacrificed after 15 weeks.The pathology changes of uterus,lumbar vertebral bones were observed by immunohistochemistry.Bone mineral density(BMD)of lumbar vertebra of rats was determined by dual energy X ray absorptiometry(DEXA).The content of MGP in serum and urine was determined by ELISA.Expression of underearboxylated matrix GLA Protein(MGP)was detected by immunohistochemistry.Relative quantification of MGP mRNA expression in lumbar vertebra bone was detected by Fluorescent real-time quantitative polymerase chain reaction.Results(1)After 15 weeks of ovariectomized,the endometrium of uterus and lumbar vertebra exhibit remarkable pathologic changes in OVX group.The serum estrogen of(454±66)pmol/L in OVX group were lower than in(527 ±77)pmol/L in sham group and(556 ±80)pmol/L in E group significantly (P ＜ 0.05).The BMD of lumbar vertebra of(0.263 ± 0.030)g/cm2 in OVX group were lower than (0.295 ±0.024)g/cm2 in sham group and(0.279 ±0.024)g/cm2 in E group significantly(P ＜0.01).(2)The serum MPG protein in OVX group and E group showed decreased trends after ovariectomized,which were(104 ±64)ng/L in OVX group and(134 ±6)ng/L in E group at 9 weeks,which reached statistical difference(P ＜ 0.05).However,MGP in urine in sham group did not exhibit significant difference after 15 weeks of surgery(P ＞0.05).The MGP in urine of E group showed increased trends after 12 weeks of surgery,which reached(110.0 ±3.4)ng/L at 15 weeks,in the mean time,it was found that(86.5 ±2.5)ng/L of MGP in urine in OVX group,which showed significant difference(P ＜ 0.05).(3)MGP could be observed in lumbar vertebra in OVX group by immunochemistry staining.In the other two groups,the expression of MGP was not dominant.(4)Relative quantification of MGP mRNA expression in lumbar vertebra was defined as 1 in OVX group,when compared with 0.289 ±0.260 in E group and 0.103 ±0.098 in sham group,the difference showed statistically significant(P ＜ 0.01).Conclusion Estrogen could increase the expression of MGP mRNA and protein in ovariectomized rats and might play an important role in improving postmenopausal osteoporosis.

The prognostic significance of preoperative serum levels of extracellular matrix protein 1 in patients with hepatocellular carcinoma / 中华普通外科杂志

Objective To evaluate the prtognostic significance of preoperative serum extracellular matrix protein 1 ( ECM1 ) levels in patients with hepatocellular carcinoma (HCC).Methods Preoperative serum levels of ECM1 were detected by enzyme-linked immunosorbent assay (ELISA) in 117 HCC patients and 53 healthy volunteers.Corrrlations to the clinicopathological characteristics and patients survival were analyzed.Results ECM1 were detected in all the samples of 117 HCC patients and 53 healthy volunteers,the median serum ECM1 level in HCC patients was significantly higher than that in healthy volunteers ( 166.39 vs 108.06 pg/ml,Z =- 7.805,P ＜ 0.001 ).Median serum ECM 1 levels were significantly higher in patients with invasive phenotypes,such as larger tumor size (Z =- 3.454,P =0.001 ),multiple nodule ( Z =- 2.201,P =0.028 ),vascular invasion ( Z =- 4.685,P ＜ 0.001 ),and advanced TN M stage ( Z =-4.610,P ＜ 0.001 ).Patients with lower serum ECM1 level (≤ 180 pg/ml ) have significantly better overall and disease free survival thanthose with higher levels ( ＞ 180 pg/ml).By Cox proportional-hazard model multivariate analysis,high serum ECM1 level ( ＞ 180 pg/ml) was an independent factor for OS and DFS in HCC patients.Conclusions Serum ECM1 levels are significantly correhted to the invasive phenotypes and survival rate.Serum ECM1 level could be used as a predictive marker for HCC recurrence and prognostic factor of HCC patients after surgery.

Effects of vascular endothelial cell growth factor on apoptosis of rat intervertebral disc cells cultured in vitro / 中华创伤杂志

Objective To observe the effect of vascular endothelial cell growth factor (VEGF)on apoptosis and matrix metabolism of rat intervertebral.disc cells cultured in vitro. Methods Culture system of rat intervertebral disc cells was established to culture the monolayer intervertebral disc cells in vitro.The well-grown intervertebral disc cells of the second generation were chosen and anti-Fas antibody was applied to induce their apoptosis.Then,VEGF at different concentrations ( 10,100,1 000 μg/L)were used to affect their apoptosis and metabolism.The apoptosis of the cells stained with Propidium Iodide (PI) was detected by using flow cytometry.The levels of hydroxyproline and proteoglycan in the culture medium were detected by using chloramines T method and DMB chromometry method,respectively.Results ( 1 ) The apoptotic ratio of the intervertebral disc cells affected by different concentrations of VEGF (10,100,1 000 μg/L) was (87.62 ±11.06)％,(53.30 ±9.23)％ and (16.75 ±4.21)％ respectively,with significant differences in comparison with the control group (P ＜ 0.01 ).(2)The contents of hydroxyproline [ (6.71 ±0.33) μg/L,(9.12 ±0.41 ) μg/L,( 11.58 ±0.12) μg/L] and proteoglycan [ (23.21 ± 2.87) μg/L,( 32.45 ± 5.23 ) μg/L,(37.18 ± 3.22) μg/] in the culture medium raised with the increase of VEGF concentration ( 10,100,1 000 μg/L),with statistical differences compared with those of the control group (P ＜0.01 ).The contents of hydroxyproline and proteoglycan were positively correlated with the concentration of VEGF (ra=0.972,P＜0.01;rb =0.907,P＜0.01). Conclusion VEGF can inhibit the apoptosis of the rat intervertebral disc cells cultured in vitro and promote collagen and proteoglycan synthesis of the cell matrix.

Osteopontin expression and its possible functions in the aortic disorders and coronary artery disease / A expressão da osteopontina e as suas funções possíveis nas desordens aórticas e doença arterial coronariana

BACKGROUND: Osteopontin (OPN) has been verified to be closely associated with oncogenesis and remodeling processes. But this cytokine was rarely assessed in the presence of aortopathies, especially acute aortic dissection. The aim of the present study was to evaluate the expressions of OPN by way of molecular biological approaches so as to offer a better understanding of the possible mechanisms of the aortopathies. METHODS: Consecutive patients with type A acute aortic dissection (20 patients), aortic aneurysm (nine patients) or coronary artery disease (21 patients) referred to this hospital for surgical operations were enrolled into this study. Blood samples of the surgical patients after systematic heparinization, and control fast morning blood samples drawn from 21 young healthy volunteers who had no evidence of any healthy problems were investigated for enzyme linked immunosorbent assay (ELISA). The surgical specimens of the aortic tissues collected from the surgical patients during the operations were obtained for quantitative realtime reverse transcription polymerase chain reaction (RT-PCR) for OPN mRNA, western blot assay for OPN protein, and for immunohistochemical staining of OPN. Ascending aortic tissues from the autopsies of the healthy individuals dying of accident were obtained as controls of immunohistochemistry. RESULTS: By quantitative RT-PCR, the expressions of OPN mRNA were all upregulated in all three surgical groups. The quantitative results did not reveal any intergroup differences. Western blot assay revealed that OPN was positive with similar intensities of expressions in all three surgical groups. Quantitative western blot analyses of OPN expressions did not show any significance between groups. The OPN expressions by ELISA in the aortic tissue were 3.09311 ± 1.65737, 3.40414 ± 1.15095, and 1.68243 ± 0.31119 pg/mg protein in the aortic dissection, aortic aneurysm, and coronary artery disease groups, respectively. The OPN level of the patients with coronary artery disease was much lower than those with aortic dissection (P = 0.033) or with aortic aneurysm (P = 0.019). By unparametric tests, there were significant differences in the aortic OPN contents among aortic dissection, aortic aneurysm and coronary artery disease groups (P < 0.01). A significant direct correlation was present between plasma OPN concentration and the time interval from the onset to surgery of aortic dissection (Y = 0.1420X + 2.4838, r² = 0.5623, r = 0.750, P = 0.032). By immunohistochemistry, OPN was expressed in the aortic cells: in the intima, it was weaker in all three surgical groups in comparison with the healthy control; in the media, it was weak in the aortic dissection, intense positive in aortic aneurysm, focal positive in the coronary artery disease, but evenly positive in the healthy control groups; and in the adventitia, it was positive in the aortic dissection, coronary artery disease and healthy control groups, but weak positive in the aortic aneurysm group. CONCLUSION: These data may provide evidences that OPN may play a role in the pathogenesis of aortopathies including aortic dissection, aortic aneurysm, and coronary artery disease. OPN might be of potential perspective as a clinically diagnostic tool in the evaluations of the complex remodeling process incorporating vascular injury and repair.

OBJETIVOS: A osteopontina (OPN) está estreitamente associada com os processos de oncogênese e remodelação. Entretanto, essa citocina era raramente avaliada na presença de aortopatias, especialmente na dissecção aórtica aguda. O objetivo do presente estudo foi avaliar a expressão de OPN por meio de abordagens moleculares biológicas, de modo a oferecer uma melhor compreensão dos possíveis mecanismos das aortopatias. MÉTODOS: Pacientes consecutivos com um tipo de dissecção aguda da aorta (20 pacientes), aneurisma da aorta (nove pacientes) ou doença arterial coronária (21 pacientes) foram incluídos neste estudo. As amostras de sangue depois da heparinização sistemática e de 21 voluntários jovens e saudáveis não apontaram nenhuma evidência de qualquer problema ao serem investigados por ensaio imunoenzimático (ELISA). Os espécimes cirúrgicos dos tecidos aórtica coletados dos pacientes durante as operações foram obtidos para a reação de transcrição reversa quantitativa em tempo real em cadeia da polimerase (RT-PCR) para OPN mRNA, técnica de Western blot para a proteína OPN, e imunohistoquímica de OPN. Amostras da aorta de indivíduos saudáveis que morreram de acidente foram obtidos para controle imunohistoquímico. RESULTADOS: Com uso do RT-PCR quantitativo, as expressões de OPN mRNA foram suprarreguladas em todos os três grupos cirúrgicos. Os resultados quantitativos não revelaram quaisquer diferenças intergrupais. Western blot revelou que OPN foi positiva com intensidade semelhante de expressões em todos os três grupos. As análises quantitativas Western blot de expressões OPN não apresentaram significâncias entre os grupos. As expressões OPN medidas pelo teste ELISA no tecido aórtico foram 3,09311 ± 1,65737, 3,40414 ± 1,15095 e 1,68243 ± 0,31119 pg/mg de proteína na dissecção de aorta, aneurisma da aorta, e grupos de doença arterial coronariana, respectivamente. O nível de OPN dos pacientes com doença arterial coronariana foi muito menor do que aqueles com dissecção aórtica (P = 0,033) ou com aneurisma da aorta (P = 0,019). Testes não-paramétricos apontaram diferenças significativas nos teores de aorta OPN entre dissecção aórtica, aneurisma da aorta e grupos com doença arterial coronariana (P <0,01). Uma correlação direta significativa estava presente entre a concentração plasmática OPN e o intervalo de tempo entre o início da cirurgia de dissecção de aorta (Y = 2,4838 + 0,1420X, r² = 0,5623, r = 0,750, P = 0,032). Pela imunohistoquímica, a OPN foi expressa em células aórticas: na íntima, foi fraca em todos os três grupos cirúrgicos em comparação ao grupo saudável; na média, era fraca na dissecção aórtica, positiva intensa no aneurisma de aorta, focal positivo na doença arterial coronariana, mas igualmente positiva no grupo controle; e na adventícia, positiva para a dissecção da aorta, doença arterial coronariana e grupos de controle saudáveis, mas fraca positiva no grupo de aneurisma da aorta. CONCLUSÃO: Estes dados fornecem evidências de que a OPN pode desempenhar um papel na patogênese da aortopatias, incluindo dissecção aórtica, aneurisma da aorta R e doença arterial coronariana. OPN tem perspectiva potencial como ferramenta de diagnóstico clínico nas avaliações do processo de remodelação complexa, incluindo lesão vascular e de reparação.

Effects of intra-articular hyaluronan injection on the expression of cartilage oligomeric matrix protein in synovium of strenuous running rats / 中国医师杂志

Objective To investigate the effects of intra-articular hyaluronan(HA)injection on the expression of cartilage oligomeric matrix protein(COMP)in synovium of strenuous running rats,and investigate the possibility of predicting the effectiveness of HA based on COMP in synovium.Methods 36 healthy male Wistar rats were randomly divided into control group,strenuous running group and strenuous running group and HA injection group.Strenuous running group and HA injection group were intra- articularly injected with HA once a week for 5 consecutive weeks.The histological changes of synovium of knee joint was examined by H.E.staining and immunohistochemical expression of COMP in three groups after 6 weeks' strenuous running.Results Synovial inflammation was less severe in strenuous running and HA injection group than strenuous running group(t =7.15,P ＜0.01).The immunohistochemical expression of COMP in rats'synovium of knee joint in strenuous running and HA injection group was significantly lower than that in rats'synovium in strenuous running group(t = 6.30,P ＜ 0.01).Conclusions Intra- articular HA injection suppressed synovitis,and the expression of COMP in synovium could be used to predict the effectiveness of HA.

Expression of extracellular matrix protein 1 in hepatocellular carcinoma is correlated with prognosis / 中华普通外科杂志

Objective To examine the expression of extracellular matrix protein 1 ( ECM1 ) in hepatocellular carcinoma (HCC) and its correlation with prognosis and recurrence of HCC.Methods Immunohistochemistry was used to detect expression of ECM1 in cancerous tissues from 77 HCC patients. The correlation between ECM1 expression and clinicopathologic features and prognosis were analyzed. Results ECM1 is mainly expressed in the cytoplasm of liver cells. The positive rate of ECM1 expression in HCC tissues were 74. 0% (57/77), and the expression level was significantly correlated with vascular invasion (χ2 =6.523, P =0.011 ) and TNM stage (χ2 =6.989, P =0.030). No significant correlation was found between the expression of ECM1 and patient's gender, age, AFP level of plasm, tumor size, number of nodules, and tumor differentiation. Patients with positive ECM1 expression have significantly poorer overall survival (OS) and disease-free survival (DFS) after curative resection than those with negative ECM1 expression (P =0. 016). The Cox multivariate analysis demonstrated that among the factors analyzed, ECM1 expression is an independent prognostic factors for OS and DFS in HCC patients after a curative resection (RR=3.721, P=0.002; RR=2.323, P=0.031). Conclusions Positive ECM1 is correlated with postoperative metastasis and invasion of HCC and poor prognosis.

Comparison of the effect of ibuprofen and glucosamine on synoviocyte proliferation and cartilage oligomeric matrix protein expression in knee osteoarthritis of human / 中华风湿病学杂志

Objective To compare the effect of ibuprofen and glucosamine on synoviocyte proliferation and cartilage oligomeric matrix protein (COMP) expression in human knee osteoarthritis. Methods Human synoviocytes were isolated from synovium (earlier stage and late stage of OA) by tissue culture and were cocultured with ibuprofen and glucosamine. The concentration of COMP was determined by MTS/PMS method and hCOMP kit. Two-tailed t-test was used for statistical analysis. Results The observation time of tissue culture was determined at 5～7 day by the MTS/PMS method. The A values of glucosamine [ late stage group (0.054±0.021), early stage group (0.777±0.034)] were less than the normal serum control group (P＜0.05).Both ibuprofen [late stage group (35.4±1.9), early stage group (46.0±2.2)] and glucosamine [late stagegroup (36.6±1.3), early stage group (48.8±1.3) ] could decrease the concentration of COMP in synoviocyte secretion in vitro (P＜0.05). Conclusion Glucosamine can inhibit the synoviocyte proliferation of human knee osteoarthritis (both early stage and late stage) in vitro. Both ibuprofen and glucosamine can inhibit the COMP secretion of synoviocyte in vitro.

Expression and Significance of CD147 and MMP9 Protein Expression in Lung Metastasis Model of Breast Cancer in TAⅡ Inbred Mice / 天津医药

Objective: To study the expressions of CD147and MMP9 in the lung metastasis model of breast cancer in BCML-TAII99 mice and their correlation with tumor growth and metastasis thereof. Methods: The breast cancer model with lung metastasis was established in BCML-TAII99 mice. The immunohistochemical staining method(SP method) was used to analyze the expressions of CD147 and MMP9 in mice lung metastasis tissue. The relationship between the tumor growth and the metastasis was analyzed. Results: The positive rates of CD147 and MMP9 were 63.33%（19/30）and 53.33%（16/30）respectively in mouse models. There was significant difference in positive rate of expressions between the metastasis group and the non-metastasis group（χ~2= 6.238， P ＝ 0．013； χ~2＝ ＝5．129， P ＝ 0．024). There was a statistically significant positive correlation between the expression of CD147 and MMP9（r = 0．786， P ＝ 0.025). Conclusion: The expression of CD147 may play a crucial role in infiltration and metastasis of breast cancer by inducing the expression of MMP9.