Expression of matrix metalloproteinase-2 and tissue inhibitor of metalloproteinase-1 in hepatic stellate cells during rat hepatic fibrosis and its intervention by IL-10

AIM: To investigate the expression of matrix metallopr-oteinase-2 and tissue inhibitor of metalloproteinase-1 in hepatic fibrosis and the antifibrogenic role of exogenous interleukin-10 (IL-10). METHODS: Hepatic fibrosis was induced by CCI4 administration and 60 male Sprague-Dawley rats were randomly divided into normal control group (group N, 8 rats), CCI4-induced group (group C, 28 rats) and IL-10-treated group (group I, 24 rats). At the beginning of the 7th and 11th wk, rats in each group were routinely perfused with pronase E and type IV collagenase through portal vein catheter and the suspension was centrifuged by 11% Nycodenz density gradient to isolate hepatic stellate cells (HSCs). RT-PCR was used to analyze mRNA of MMP-2 and TIMP-1 from freshly isolated cells. Densitometric data were standardized with β-actin signals. Immunocytochemistry was performed to detect MMP-2 and TIMP-1 expression in HSC cultured for 72 h. RESULTS: Compared to group N in the 7th wk, MMP-2 and TIMP-1 mRNA increased in group C (P= 0.001/0.001) and group I (P= 0.001/0.009). The level of MMP-2 and TIMP-1 mRNA in group I was significantly lower than that in group C (P= 0.001/0.001). In the 11th wk, MMP-2 mRNA in group I was still lower than that in group C (P = 0.005), but both dropped compared with that in the 7th week (P = 0.001/0.004). TIMP-1 mRNA in group I was still lower than that in group C (P= 0.001), and increased in group C (P= 0.001) while decreased in group I (P = 0.042) compared with that in the 7th wk. Same results were found by immunocytochemistry. CONCLUSION: Expression of MMP-2 and TIMP-1 is increased in hepatic fibrosis. IL-10 exhibits an antifibrogenic effect by suppressing MMP-2 and TIMP-1 expression.

Multifactor Regulation on Expressions of MMP-9 and TIMP-1 in Endometrial Stromal Cells

Objective To investigate the regulatory effect of multifactor on the matrix metalloproteinases-9 （MMP-9） and the tissue inhibitor of metalloproteinase-1 （TIMP-1） in endometrial stromal cells. Methods The endometrial stromal cells separated from the proliferative endometrial tissues were incubated with medium alone, 17-β estradiol （E2,10^-8 mol/L）, medroxyprogesterone acetate （MPA, 10^-6 mol/L）, E2（10^-8 mol/L）＋MPA （10^-6 mol/L）, E2 （10^-8 mol/L）＋MPA （10^-6 mol/L）＋RU486 （10^-5 mol/L） or HB-EGF （10 ng/ml） for 48 h respectively. The expressions of MMP-9 and TIMP-1 were detected by in situ hybridization, immunocytochemistry, reverse transcriptase-polymerase chain reaction （RT-PCR） and Western blotting. Results Compared with control group [mRNA, 0. 729 ± 0. 090 （MMP-9） and 1.056± 0.154 （TIMP-1）; protein, 0.545 ±0.086 （MMP-9） and 0.745 ±0.154 （TIMP-1）], expressions of MMP-9 and TIMP-1 in E2 alone, progestin alone or E2 combined with progestin group were respectively：mRNA, 0.413 ± 0.069, 0.402 ± 0.073 and 0.407 ± 0.039; 0.487 ± 0.093, 0.503 ± 0.093 and 0.468 ± 0.075：protein, 0.294 ± 0.076, 0.331 ±0.064 and 0.265 ±0.049; 0.425 ±0.085, 0.397 ±0.065 and 0.435 ± 0.099. RU486 weakened the expression level of down-regulation, while HB-EGF elevated the level of MMP-9 and TIMP-1 after 48 h treatment （mRNA, 0.955 ± 0.068 and 1.396 ± 0.238; protein, 0. 780 ± 0.109 and 0.985 ± 0.165）. Conclusions 1） Both E2 and progestin can down-regulate the expressions of MMP-9 and TIMP-1 in endometrial stromal cells, but RU486 can inhibit the effect. 2） HB-EGF can elevate the level of MMP-9 and TIMP-1. 3） E2, progestin and HB-EGF have effect on the ratio of MM-P/TIMP-1.

Contrary Regulation of TIMP-1 and MMP-9 by Hepatocyte Growth Factor Antibody after Lung Injury

Objective To study the influence of hepatocyte growth factor (HGF) antibody on the lung expression level of matrix metalloproteinases-9 (MMP-9) and tissue inhibitor of metalloproteinase-1 (TIMP-1). Methods Thirty male Wistar rats were randomly divided into 3 groups: control group, model group, and intervention group. Endotoxin was intratracheally infused in the model and intervention groups. HGF antibody was injected in the rats of the intervention group from day 1 to day 14, while the same volume of saline was injected in the control group. The rats were sacrificed on day 28 after endotoxin treatment. The amounts of MMP-9 mRNA and TIMP-1 mRNA were measured by reverse transcription-polymerase chain reaction, and protein expression levels of MMP-9 and TIMP-1 were measured by immunohistochemistry. Results In the model group, both mRNA and protein expression levels of TIMP-1 were significantly increased, the same as MMP-9. In the intervention group, the increase of TIMP-1 was remarkably reduced compared with the model group, while the mRNA and protein expression levels of MMP-9 were still increased. Conclusion HGF activity may accelerate the repair of lung injury through contrary regulating the expression levels of TIMP-1 and MMP-9.

Effects of Activating Blood Circulation to Resolve Blood Stasis and Glucosamine Capsules Therapy on the Expression of Matrix Metalloproteinase in Post-traumatic Knee Osteoarthritis

[Objectives] To observe the clinical effects of activating blood circulation to resolve blood stasis and glucosamine hydrochloride capsules therapy on post-traumatic knee osteoarthritis treatment,to measure the expression of matrix metalloproteinase in post-traumatic knee osteoarthritis patients before and after treatment,and to explore its relevant molecular mechanisms. [Methods]A total of 60 patients with posttraumatic knee osteoarthritis in early stage in Affiliated Hospital of Shannxi University of Chinese Medicine from January 2015 to December2017 were selected and randomly divided into the control group( n = 30) and the observation group( n = 30). The control group was given oral administration of glucosamine hydrochloride capsules for 6 courses of the treatment. The observation group was using the activating blood circulation to resolve blood stasis therapy for 6 courses of treatment. The recovery of traumatic knee osteoarthritis and the symptom of traditional Chinese medicine( TCM) in 2 groups was compared before and after treatment. The symptoms and the quantitative assessment rating scale of knee osteoarthritis,the pain index of knee and the TCM symptom scores were recorded in both groups. And the synovial fluid and serum of patients were collected to detect the expression of matrix metalloproteinase. [Results]The symptoms and the quantitative assessment rating scale of knee osteoarthritis,the pain index of knee and TCM symptom scores of the 2 groups in the 1 st week,the 12 th week and the 24 th week after the treatment were significantly lower than those before the treatment,and the difference was statistically significant( P < 0. 05). Two therapeutic methods had certain therapeutic effects on the recovery from traumatic knee osteoarthritis,and could significantly improve the TCM syndrome. There were better therapeutic effects on oral decoction of traditional Chinese medicine than oral glycosaminoglycans hydrochloride capsules on treating traumatic knee osteogenesis( P < 0. 05). The expression level of MMP3 was decreased significantly in both groups( P <0. 001),while the expression level of MMP13 had no significant change in both groups( P >0. 05). The expression level of MMP1 was decreased significantly in the observation group( P < 0. 001),but there was no significant change in the control group after treatment( P >0. 05). [Conclusions]Two treatment methods have been effective in this clinical study. We found that the expression level of MMP3 was decreased significantly after treatment in 2 therapies,which can be clinically applied.

Role of Matrix Metalloproteinase-1(MMP-1)/Protease-activated Receptor-1(PAR-1) Signaling Pathway in the Cervical Cancer Invasion

Objective To investigate the role of matrix metalloproteinase-1 （MMP-1）/protease- activated receptor-1 （PAR-l） signaling in the cervical cancer invasion. Methods RT-PCR was used to test the mRNA level of MMP-1 in 59 cases of cervical squamous cell cancer. Cell invasion was evaluated by Transwell assay to explore the effect of adding human recombinant MMP-1 （rhMMP-1） and PARI-siRNA on cervical cancer invasion. Results In cervical cancer tissues, more MMP-1 expression was observed than that in the normal cervical tissues, and its expression correlated with cancer status. Human recombinant MMP-1 （rhMMP-1） could promote Hela cell invasion, and its number of invasive cell correlated with the concentration of rhMMP-1. Disrupting the expression of PAR-1 reduced the MMP-1 promoting-effect on Hela cell invasion, but had no effect on non-MMP-1 invasive action. Conclusion The MMP-1/PAR-1 signaling pathway is involved in cervical cancer invasion. Therefore, blocking PAR-1 may represent a new therapeutic option for metastatic cervical cancer.

Expression of matrix metalloproteinase-1 mRNA in peripheral blood mononuclear cells of systemic lupus erythematosus patients and its relationship with atherosclerosis

Background Matrix metalloproteinase-1 （MMP-1） plays an important role in atherosclerosis. This study was to examine expression of MMP-1 mRNA in peripheral blood mononuclear cells （PBMCs） of patients with systemic lupus erythematosus （SLE）, and to explore its relationship with atherosclerosis in SLE. Methods Fluorescent quantitative reverse transcription polymerase chain reaction （RT-PCR） was used to examine the expression of MMP-1 mRNA in PBMCs in 80 SLE patients, including 39 prone to atherosclerosis （Group A） and 41 unprone to atherosclerosis （Group B）. Meanwhile, 30 patients who were free of cardiovascular diseases and 30 healthy individuals were selected as disease and normal control group （Groups C and D）. The changes of MMP-1 gene expression were analyzed by differences of cycle threshold （ACt）, with the following formula： ACt = Cttarget gene - Ctreference gene. Results The expression level of MMP-1 mRNA in Group A was significantly higher than that of group B （ACt=8.64±2.43 vs ACt=12.09±2.26, t=6.588, P 〈0.01）. The expression level of MMP-1 mRNA of SLE patients was significantly higher than that of Group C （ACt=10.41±2.90 vs ACt=12.29±2.51, t=3.135, P 〈0.01） and Group D （ACt=10.41±2.90 vs ACt=12.48±1.69, t=3.675, P 〈0.01）. Conclusions In comparison to disease and control group, expression of MMP-1 mRNA in PBMCs of SLE patients was significantly elevated, and significant difference of MMP-1 mRNA expression was also found between SLE patients prone and unprone to atherosclerosis, indicating that expression of MMP-1 mRNA may be correlated with the pathogenesis and activity of atherosclerosis in SLE.

活动期类风湿关节炎患者血清基质金属蛋白酶-3及其组织抑制剂-1水平的检测和意义

目的研究活动期类风湿关节炎(rheum atoid arthritis,RA)患者血清基质金属蛋白酶-3(m atrix m etalloproteinase3,M M P-3)、金属蛋白酶组织抑制剂-1(tissue inhibitors ofm etalloproteinase1,TIM P-1)的水平及其相关影响因素,探讨M M P-3及TIM P-1在R A的作用机制。方法选择41例初诊活动期RA患者和30名正常健康志愿者,以酶联免疫吸附试验(ELISA)分别检测血清M M P-3及TIM P-1水平,计算M M P-3/TIM P-1,同时测定关节功能、X线、关节肿胀数(SJC)、血沉(ESR)、类风湿因子(R F)、C反应蛋白(CR P)等相关实验室指标。结果活动期R A患者血清M M P-3、TIM P-1明显增高(P<0.01),且以M M P-3增高更为显著,M M P-3/TIM P-1较正常组亦增高(P<0.05)。不同关节功能分级时,上述指标差异无统计学意义(P>0.05),而不同X线分期时,各指标差异有统计学意义(P<0.01或P<0.05)。M M P-3、M M P-3/TIM P-1与SJC(P<0.01)、CRP(P<0.01)、ESR(P<0.05)呈正相关,二者与年龄、病程、晨僵时间、RF无明显相关性(P>0.05)。结论M M P-3、TIM P-1在RA血清中高水平存在,二者比例失衡导致RA发生。M M P-3、M M P-3/TIM P-1的高低可作为反映病情活动及预后的指标,阻断M M P-3高水平有可能成为治疗RA的新途径之一。

脑血管

高血压丘脑出血破入脑室的分型和治疗;中脑周围蛛网膜下腔出血32例临床分析及长期随访研究;89例颅内动脉瘸栓塞术后血管造影随访报告；ICAM-1、MMP-2基因在颈动脉内膜切除术后早期再狭窄中的动态表达；Matrix电解弹簧圈栓塞治疗颅内动脉瘤；

Regulatory Action of Charred Gossamer Urocteae on the Functions of Mouse Oral Fibroblasts

Objective： To explore the influence of charred Gossamer urocteae （CGU） on the functions of primary cultured mouse oral fibroblasts and reveal its mechanism in wound healing. Methods： CGU was extracted with different solvents and ethanol extract （EE）, ethyl acetate fraction （EF）, n-butanol fraction （BF） and aqueous fraction （AF） were obtained. The effects of different fractions on the proliferation, matrix metaUoproteinase-2,9 （MMP-2,9） activities, synthesis of collagen and tissue inhibitor of metalloproteinase 1 （TIMP-1） in the mouse oral fibroblasts were determined by MTT, gelatin zymography, chloramine-T method, and enzyme-linked immunosorbent assay （ELISA） respectively. Results： EE, EF and BF at high concentrations could significantly inhibit proliferation of fibroblasts （P〈0.05 or P〈0.01）, and at low concentrations EF and BF could promote proliferation of fibroblasts, and BF and AF could significantly inhibit collagen synthesis （P〈0.05 or P〈0.01）. EE, EF and AF at high concentrations could significantly increase the MMP-9 activity, and BF and AF could significantly inhibit synthesis of TIMP-1. Conclusion： CGU at high concentrations can inhibit the proliferations of fibroblasts and synthesis of collagen, and in healing of wound, CGU at high concentrations possibly has the functions of anti-fibrosis and anti-scar, and the mechanism to promote degradation of collagen is possibly related to the increase in MMP-9 activity and the inhibition of TIMP-1 synthesis.