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1.
Effect of purified human interleukin-1 on cartilage degradation 总被引:3,自引:0,他引:3
The effects of highly purified human monocyte-derived interleukin-1 (IL-1) on bovine nasal cartilage breakdown were investigated. Cartilage degradation was determined by quantifying the fraction of total proteoglycan released from cartilage during 8 days of culture. The response appeared to be chondrocyte-dependent, for IL-1 stimulated proteoglycan (PG) release from living but not from dead (frozen-thawed) cartilage. IL-1 action on living cartilage was heat labile and concentration dependent, with significant effect at 5 U/ml and maximal effect at 10-20 U/ml. Kinetic studies showed significant stimulation of PG release by 3 days of incubation with 10 U/ml IL-1. Studies in which IL-1 was removed on day 1 or day 4 showed that the cartilage-degrading effect of this monokine was reversible. Although IL-1 caused little change in the Sepharose CL-2B chromatographic profile of released PGs using an associative elution buffer, a significant shift to lower mol wt was observed under dissociative conditions. To probe the mechanism of IL-1 action, cartilage samples were incubated with IL-1 in the presence of the protein synthesis inhibitor, cycloheximide, or the lysosomal membrane-stabilizing steroid, hydrocortisone. Cycloheximide at 5-10 micrograms/ml completely blocked IL-1-induced breakdown. One the other hand, 3 x 10(-7) M hydrocortisone had little or no effect on IL-1 action. IL-1 was also shown to stimulate the degradation of human articular cartilage. 相似文献
2.
Biosynthesis of proteoglycan in bone and cartilage of parathyroid hormone-related protein knockout mice 总被引:1,自引:0,他引:1
Ogihara Y Suda N Hammond VE Senior PV Beck F Yanagishita M 《Journal of bone and mineral metabolism》2001,19(1):4-12
Proteoglycans are suggested to regulate cell adhesion, differentiation and mineralization of hard tissues. In vitro studies
have shown that many humoral and local factors regulate proteoglycan synthesis. Among them, parathyroid hormone (PTH) and
parathyroid hormone-related protein (PTHrP) have potent stimulating effects on proteoglycan synthesis. However, the exact
role of PTHrP on the biosynthesis and metabolism of proteoglycans during skeletal development is not clear. To clarify this
point, we examined bony and cartilaginous explants of newborn mice with disrupted PTHrP alleles. Ribs of homozygous PTHrP-knockout
mice and wild-type littermates were dissected into bony and cartilaginous regions and metabolically labeled with [35S]sulfate in culture. Radiolabeled proteoglycans were analyzed by column chromatography. The elution profiles of [35S]-labeled proteoglycan from cartilaginous explants did not differ between homozygous PTHrP-knockout mice and wild-type littermates.
However, the amount of labeled proteoglycan in homozygous PTHrP-knockout mice was only 4%–5% that of wild-type littermates.
In contrast with cartilaginous explants, the amount of labeled proteoglycans in bony explants did not differ between the two
genotypes. Interestingly, besides the common major peak (Kd = 0.10–0.16) observed in the bony explants of both genotypes, a minor peak (Kd = 0.42) was specifically present in homozygous PTHrP-knockout mice. This minor peak was earlier than that of free glycosaminoglycan
(GAG) chains, suggesting that the core protein, but not GAG chain, was cleaved in the bony explants of homozygous PTHrP. These
findings demonstrate a crucial and nonredundant role of PTHrP in the regulation of proteoglycan synthesis and metabolism during
skeletal development.
Received: February 21, 2000 / Accepted: June 16, 2000 相似文献
3.
目的探讨饰胶蛋白聚糖(DCN)与转化生长因子β1(TGF-β1)对大鼠肾系膜细胞(MsC)生长及相关信号途径的影响。方法经脂质体介导将DCN基因转染体外培养的大鼠肾MsC,经筛选和鉴定后,收集阳性细胞克隆的培养上清(DCN上清)。采用流式细胞仪检测经TGF-β1(2μg/L)和(或)DCN上清作用后MsC细胞周期的变化。采用Western印迹法分别检测两者单独或联合作用后,MsC磷酸化丝裂原活化蛋白激酶(p-MAPK)、磷酸化Smad2 (p-Smad2)和p21蛋白表达情况。采用免疫共沉淀方法检测阳性细胞克隆上清中DCN与TGF-β1的结合情况。结果TGF-β1或DCN上清单独作用时均可抑制MsC的增殖,G_2-M期细胞数分别为对照组的81%及86.5%,而两者共同作用时G_2-M期细胞数与对照组差异无统计学意义。经TGF-β1和DCN上清单独作用12 h后,p-ERK1/2表达为对照组的4.4、3.7倍和3.2、3.7倍;p-SAPK/JNK表达为对照组的2.0、1.5倍和1.9、1.5倍;而两者共同作用组p-ERK1/2和p-SAPK/JNK表达与对照组差异无统计学意义。TGF-β1单独作用12 h后,MsC p-Smad2的表达为对照组的2.6倍,而DCN单独作用组及2者联合作用组其表达与对照组差异无统计学意义。DCN单独作用12 h后p21蛋白的表达为对照组的2.6倍,而TGF-β1单独作用及2者联合作用组其表达与对照组差异无统计学意义。阳性细胞克隆上清中DCN可以直接与TGF-β1结合。结论TGF-β1与DCN单独作用均可抑制MsC增殖及激活MAPK信号途径,但2者共同作用时其作用相互抵消。TGF-β1激活Smad信号途径作用可被DCN阻断;DCN上调p21蛋白作用可被TGF-β1阻断。2者通过不同信号分子抑制细胞生长,其作用可能与两者相互结合有关。 相似文献
4.
Heterotopic ossification is a metabolically active process which shares several properties of orthotopic bone formation and, therefore, represents an excellent model for studying bone matrix components. Immunohistochemical methods were used to investigate the distribution pattern of the small proteoglycans decorin and proteoglycan-100 during different stages of heterotopic ossification of pressure sores of paraplegic patients. Decorin and proteoglycan-100 exhibited a substantially divergent distribution pattern. Decorin was detectable in the perivascular matrix of granulation tissue as well as in the stroma of heterotopic ossification. The ossification zone was stained most strongly. In contrast, proteoglycan-100 was predominantly detectable in fibroblasts and preosteoblasts in early areas of osteogenesis. In more mature forms of heterotopic ossification immunostaining was markedly reduced in osteoblasts and osteocytes and even absent in so-called bone-lining cells. However, at least some osteoclasts were strongly positive.These results suggest indicate that decorin and proteoglycan-100 are important components during the formal pathogenesis of heterotopic ossification. The expression of the small proteoglycans, especially of proteoglycan-100, correlates with different phases during heterotopic ossification, showing a maximum for proteoglycan-100 in matrix-forming cells in early phases of bone formation, but in osteoclasts in mature bone. 相似文献
5.
《Joint, bone, spine : revue du rhumatisme》2014,81(2):175-177
Erdheim-Chester disease is a rare non-langerhans systemic histiocytosis of unknown origin, associated with bone diseases and severe visceral complications. Therapies have been disappointing. A recombinant form of interleukin-1 receptor antagonist (anakinra) has been used in a few cases when usual treatment fails. We report a new case of successfully interleukin-1 receptor antagonist treatment in Erdheim-Chester disease. 相似文献
7.
目的 建立稳定表达丙型肝炎病毒(HCV)核心蛋白的原核表达系统,获得高产量的纯化核心蛋白.方法 以HCV株全长Cdna 序列为模板,PCR扩增获得核心蛋白基因,构建原核表达载体Pet-32a(+)-HCV-Core.转化大肠埃希菌DH5α,提取质粒,验证正确后,再次转化大肠埃希菌BL21中,IPTG诱导表达,SDS-PAGE、Western blot验证融合蛋白的表达.超声破碎表达菌,Ni+-亲和柱对表达蛋白进行纯化及柱上复性.结果 成功构建了原核表达载体Pet-32a(+)-HCV-Core,并表达了预期分子量大小的目的蛋白.结论 成功表达、纯化HCV核心融合蛋白,为进一步研究其生物学功能奠定了坚实的基础. 相似文献
8.
Kazuhiro Yoshimura Osamu Miyake Masao Tsujihata Toshiaki Yoshioka Seiji Yamaguchi Takuo Koide Shiro Takahara Akihiko Okuyama 《International journal of urology》1999,6(11):567-571
BACKGROUND: There are only a few papers reporting on the role of proteoglycan core protein in calcium oxalate stone formation. The present study was carried out to investigate the role of core protein of proteoglycan in human urine on calcium oxalate (CaOx) crystallization. METHODS: Proteoglycans were collected from whole human urine. The covalently bound glycosaminoglycans (GAG) of proteoglycans were then digested by GAG lyase. The inhibitory activity on CaOx crystal growth in vitro was measured before and after enzyme digestion of proteoglycans. Sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of the core protein of proteoglycans and the analysis of amino acid sequence were performed. RESULTS: The core protein showed significant inhibitory activity on CaOx crystal growth, which scarcely changed when compared with that of proteoglycans before enzyme digestion. The SDS-PAGE revealed that the core protein was a single unit with a molecular weight of 26 kDa and amino acid sequencing demonstrated high homology to interalpha-trypsin inhibitor (ITI) light chain (bikunin) with Kunitz inhibitor domain as a core protein. CONCLUSIONS: The results suggested that human urine contains proteoglycans and a major part of them is ITI light chain (bikunin). The Kunitz inhibitor domain, a core protein of bikunin, has significant inhibitory activity on CaOx crystallization without GAG bound covalently to the core protein. 相似文献
9.
目的:探讨针刺对骨关节炎(OA)软骨中炎性细胞因子的影响作用。方法:选用SD雌性大鼠40只,采用随机数字表,随机分为正常组、模型组、针刺组和对照组,各10只。采用单侧后肢跟腱切除法建立OA动物模型,切除左后肢跟腱,分别采用电针和扶他林乳剂对针刺组和对照组非手术侧(右后肢)进行治疗。采用免疫组化技术,观察各组关节软骨中白细胞介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)表达的特点,并比较各组间的差异。结果:模型组IL-1β和TNF-α的表达都较正常组上调(P〈0.01);模型组、针刺组和对照组3组比较,IL-1β和TNF-α的表达差异有统计学意义,针刺组和对照组的表达均较模型组下调(P〈0.01),两者在针刺组与对照组的表达差异无统计学意义(P〉0.05)。结论:针刺能下调OA软骨细胞IL-1β、TNF-α的表达,与扶他林乳剂作用相当,说明针刺对OA软骨具有一定的保护作用。 相似文献
10.
白细胞介素13对肾小球系膜细胞增殖及白细胞介素1β表达的影响 总被引:11,自引:0,他引:11
目的:探讨白细胞介素13(IL-13)对体外培养的大鼠系膜细胞的增殖及其产生白细胞介素1β(IL-1β)的影响。方法:用四甲基偶氮唑(MTT)法测定系膜细胞增殖,用逆转录聚合酶链反应(RT-PCR)及酶联免疫吸附(ELISA)法测定系膜细胞IL-1β mRNA表达IL-1β蛋白水平。结果IL-13在1.0,10,100ng/ml浓度范围呈剂量依赖性地抑制系膜细胞的增殖。含5%小牛血清(FCS)的RPMI 1640培养状态下的系膜细胞检测不出IL-1β,IL-13在抑制系膜细胞的增殖的相应浓度坷显著地抑制LPS诱导的系膜细胞IL-1β mRNAg表达及L-1β分必,IL-13在10ng/ml浓度时即可几乎完全抑制LPS诱导的系膜细胞IL-1β mRNA表达,结论:IL-13对体外培养的系膜细胞增殖及炎症效应具有抑制作用。 相似文献
11.
目的 探讨核心蛋白聚糖(DCN)对膀胱癌细胞生长的影响。方法 以膀胱癌T24细胞株为研究对象,采用MTT法检测不同浓度、不同时间的DCN对T24细胞存活率的作用,采用流式细胞术分析DCN对T24细胞周期及凋亡的影响,采用ELISA和Western blot法检测DCN对转化生长因子-β1(TGF-β1)和P21蛋白表达的影响。结果 与其他浓度相比,40、50 μg/mL DCN作用72 h时对T24细胞的抑制作用最强,差异有统计学意义(P<0.05),且G1期细胞达到最高值,S期细胞达最低值(P<0.001)。5、10、20、30、40、50 μg/mL DCN作用72 h后均能促进T24细胞凋亡,且在40 μg/mL时达到最大值(P<0.001);与0 μg/mL相比,5、10、20、30、40、50 μg/mL DCN作用72 h对TGF-β1表达均有抑制作用, 最明显的抑制作用浓度为40 μg/mL(P<0.001);与0 μg/mL相比,40 μg/mL DCN能促进P21蛋白上调(P<0.001)。结论 DCN在体外能够抑制膀胱癌T24细胞生长,诱导其凋亡,其可能的作用机制为下调TGF-β1及上调P2l蛋白表达。 相似文献
12.
目的 旨在研究白细胞介素(interleukin,IL)-10对高迁移率组蛋白l(high mobility group protein 1,HMGB1)释放的影响及其肺保护作用,进而探讨IL-10在治疗急性肺损伤中的可能机制. 方法 采用腹腔注射脂多糖(lipopolysaccharide,LPS)致大鼠脓毒症急性肺损伤模型,健康SPF级雄性SD大鼠72只,体重180 g~220 g,采用随机数字表法,随机分为对照组即磷酸盐缓冲液(phosphate buffered solution,PBS)组(P组,6只)、急性肺损伤组即LPS组(L组,30只)、PBS+IL-10组(PI组,6只)、LPS+IL-10组(LI组,30只).P组和PI组腹腔注射等体积PBS的同时分别经由气道滴注5 ml的PBS和IL-10,L组和LI组腹腔注射LPS的同时分别经由气道滴注等体积的PBS和IL-10.L组和LI组注射LPS后的4、8、16、24 h和48 h分别检测各组大鼠肺湿/干重比(wet/dry weight ratio,W/D)和支气管肺泡灌洗液(bronchoalveolar lavage fluid,BALF)中总蛋白浓度,酶联免疫吸附实验法检测BALF中炎性因子肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)和IL-6水平,苏木精-伊红(hematoxylin-eosin,HE)染色观察肺组织的病理变化,Western Blot分析肺组织中HMGB1表达水平. 结果 腹腔注射LPS后,L组大鼠肺组织W/D和BALF中总蛋白浓度分别为(5.68±0.12) mg/L和(254±105) mg/L,与P组比较,分别增加了12%和297%(P<0.05),BALF中TNF-α和IL-6水平明显增加(P<0.05),HE染色显示在注射LPS后4h肺组织的细胞浸润达到峰值随后减少,肺组织中HMGB1水平升高(P<0.05);使用IL-10后,LI组肺组织W/D和BALF中总蛋白浓度分别为(5.28±0.14) mg/L和(109±48) mg/L,与L组比较,分别降低了7%和57% (P<0.05),BALF中炎性因子水平降低(P<0.05),肺组织细胞浸润改善,HMGB1表达下调(P<0.05). 结论 IL-10对急性肺损伤具有保护作用,其机制可能为降低BALF中炎性因子的水平,下调晚期炎症介质HMGB1的表达,从而改善肺组织的细胞? 相似文献
13.
白细胞介素1对椎间盘细胞软骨特异性基因Sox9 mRNA的调节作用 总被引:3,自引:0,他引:3
目的探讨白细胞介素1(interleukin-1,IL-1)对人椎间盘细胞软骨特异性基因Sox9和Ⅱ型胶原基因表达的调节作用。方法应用RT—PCR技术检测IL-1对培养的椎间盘细胞中软骨特异性基因so西和Ⅱ型胶原基因mRNA表达的调节作用。结果在IL-1浓度为0.1ng/ml、1ng/ml和10ng/ml培养24h时,其对椎间盘细胞Sox9和Ⅱ型胶原基因mRNA可起到显著的负向调控作用;10ng/ml的IL-1随着培养时间的延长对椎间盘细胞中Sox9和Ⅱ型胶原基因mRNA出现显著的负向调控作用。结论IL-1可以按照剂量及时间依赖方式负向调节椎间盘细胞Sox9和Ⅱ型胶原基因的表达。 相似文献
14.
Elizabeth C. Arner 《Journal of orthopaedic research》1994,12(3):321-330
Rheumatoid arthritis and osteoarthritis are characterized by an early depletion of cartilage proteoglycans, which leads to a decrease in cartilage compressibility and, eventually, to a loss of joint function. Interleukin-1, which is thought to have a role in mediating this loss of proteoglycans in arthritis, induces an acute depletion of proteoglycans from articular cartilage following intra-articular injection in rabbits. As the structure and metabolism of proteoglycans are known to change with age, my laboratory investigated the effect of age on depletion and recovery of proteoglycans in response to interleukin-1 in the rabbit. Loss of cartilage proteoglycans induced by interleukin-1 was less servere in immature animals, increased until the age of sexual maturity, and then remained constant. The rate of recovery and compensatory overshoot in the rate of proteoglycan synthesis following challenge with interleukin-1 was more rapid in immature animals and may have been responsible for the quicker return of the cartilage proteoglycan content to control levels in younger animals. With multiple exposures to interleukin-1 at time intervals too short for recovery to occur, smaller amounts of interleukin-1 induced loss of proteoglycans, and the proteoglycan content and the rate of synthesis remained depressed longer after treatment had stopped. The decreased ability of mature cartilage to replace proteoglycans rapidly after exposure to cytokines would increase the probability of subsequent inflammatory episodes before recovery is complete; this may result in increased susceptibility of adult cartilage to proteoglycan depletion. 相似文献
15.
16.
Persistent bone-sparing effect of interleukin-1 receptor antagonist: A hypothesis on the role of IL-1 in ovariectomy-induced bone loss 总被引:1,自引:0,他引:1
R. B. Kimble R. Kitazawa J. L. Vannice R. Pacifici 《Calcified tissue international》1994,55(4):260-265
The recent finding that treatment with the interleukin-1 (IL-1) inhibitor, interleukin-1 receptor antagonist (IL-1ra) decreases bone loss and bone resorption in ovariectomized rats, strongly suggested that IL-1 mediates, at least in part, the effects of estrogen deficiency on bone resorption. Although in vitro studies have shown that IL-1 activates mature osteoclasts and stimulates osteoclastogenesis, the two main mechanisms by which estrogen deficiency stimulates bone resorption, it is still unclear whether IL-1 mediates both effects of estrogen deficiency in vivo. To investigate this matter, we have examined the changes in bone mineral density (BMD) which occur in ovariectomized rats after completion of 1 month of estrogen or IL-1ra treatment begun at the time of ovariectomy. Ovariectomy caused a marked decreased in BMD which was blocked by 17 estradiol and decreased by IL-1ra. Cessation of estrogen therapy was followed by a rapid induction of bone loss, indicating that estrogen blocks the activation and utilization of mature osteoclasts without depleting the bone microenvironment of osteoclast precursors and mature, inactive osteoclasts. In contrast, ovariectomized rats treated with IL-1ra maintained a stable bone density for the first 4 weeks after completion of the treatment. In these rats, bone loss resumed not earlier than 6 weeks after discontinuation of the IL-1ra treatment. Estrogen deficiency was necessary to unveil the bone-sparing effect of IL-1ra because in a control experiment in which rats were treated with IL-1ra for the 4 weeks before ovariectomy, BMD began to decrease immediately after ovariectomy. Based on these results we propose the hypothesis that in conditions of estrogen deficiency, the main effect of IL-1ra is to block the proliferation and differentiation of osteoclast precursors, an event that results in the depletion of mature, rapidly responsive osteoclasts. We also suggest that estrogen may have important direct effects on the regulation of osteoclast activity. 相似文献
17.
18.
白介素-1受体拮抗剂对椎间盘基质代谢的影响 总被引:1,自引:0,他引:1
目的:探讨白介素-1受体拮抗剂(IL-1Ra)对双后肢大鼠椎间盘基质代谢的影响。方法:建立双后肢大鼠模型36只,随机分为实验组和对照组,每组18只,实验组从造模当天即予腹腔注射IL-1R(a50ng/kg),隔天重复注射至处死,对照组18只不予任何处理。于造模后1、3、6个月每组分别处死6只大鼠,完整取出L4/5椎间盘固定、切片,予SABC法进行免疫组化染色,并使用图像分析系统对纤维环中Ⅱ型胶原染色进行灰度值扫描,同理取出L5/6椎间盘,使用间苯三酚法测量蛋白多糖含量。结果:术后第1个月,实验组椎间盘髓核中Ⅱ型胶原灰度值及蛋白多糖含量分别为83.67±7.97和2.376±0.161,对照组分别为87.25±8.70和2.297±0.101,两组间无显著性差异(P>0.05);第3个月时实验组Ⅱ型胶原染色的灰度值及蛋白多糖含量分别为107.42±6.50和2.093±0.131,对照组分别为145.91±7.42和1.039±0.092,两组间比较有非常显著性差异(P<0.01);第6个月时实验组Ⅱ型胶原染色的灰度值及蛋白多糖含量分别为129.83±4.03和1.796±0.065,对照组分别为203.76±5.98和0.654±0.048,两组间比较有非常显著性差异(P<0.01)。结论:IL-1Ra对双后肢大鼠椎间盘基质降解有明显的抑制作用。 相似文献
19.
目的:观察丙型肝炎病毒(HCV)的核心蛋白(core)对肝细胞甘油三酯(TG)合成的影响。方法将HCV 1b基因型core表达载体pcDNA3.1-myc/his(-)-core1b和HCV 3a基因型core表达载体pcDNA3.1-myc/his(-)-core3a分别转染至HepG2细胞,利用定量测定法检测细胞内TG含量,用实时荧光定量逆转录聚合酶链反应法(real time qPCR)、蛋白免疫印迹(Western blot)方法观察脂质合成相关基因肝X受体α(LXRα)、甾醇调节元件结合蛋白1c(SREBP1c)和脂肪酸合酶(FASN)的mRNA和蛋白的表达变化,并进行比较分析。结果 HepG2细胞转染两个基因型HCV core表达载体之后均能显著增加细胞内的TG含量(P<0.05),尤以基因型core3a组为差异更显著(P<0.001)。Real-time PCR结果显示,两个基因型core均上调LXRα的mRNA水平(P<0.05),基因型core3a组差异更为显著(P<0.01);FASN的mRNA水平只在仅在基因型core3a组上调(P<0.05),而基因型core1b组差异无统计学意义。Western blot结果显示,HCV core表达可以明显上调表达可以显著上调SREBP1c的蛋白水平,尤其基因型core3a组更为显著。对FASN蛋白水平,基因型core3a上调其表达,但基因型core1b组无明显变化。结论 HCV可能通过LXRα介导肝细胞内的脂质合成诱导肝脂肪变,有待进一步研究。 相似文献
20.
青少年特发性脊柱侧凸椎体软骨终板核心蛋白和细胞因子的变化 总被引:4,自引:0,他引:4
目的了解青少年特发性脊柱侧凸患者顶端椎凸、凹侧椎体软骨终板内核心蛋白、TGFβ1及bFGF的变化,探讨其在特发性脊柱侧凸发病中的作用。方法12例患者,男4例,女8例;年龄12~20岁,平均14.9岁;侧凸Cobb角43°~102°,平均65.1°。所有患者均经X线、CTM和(或)MR检查排除先天性、神经肌肉性等原因所导致的侧凸。对前路手术取下的顶椎和端椎的软骨终板进行S-P法免疫组化染色,染色结果采用北京航空航天大学图像中心医学彩色病理图像分析系统进行分析。结果经非参数Wilcoxon秩和检验,侧凸顶椎和端椎凹侧终板软骨细胞内TGFβ1表达的面密度和数密度高于凸侧及顶椎凸侧表达的数密度高于端椎均有统计学意义(P<0.05);侧凸顶椎和端椎凹侧bFGF表达的面密度和数密度高于凸侧亦有统计学意义(P<0.05);顶椎及端椎凸侧蛋白多糖核心蛋白的面密度和数密度高于凹侧,但无统计学意义(P>0.05)。TGFβ1及bFGF及核心蛋白在侧凸顶椎的表达较端椎表达略高,但无统计学意义(P>0.05)。结论青少年特发性脊柱侧凸患者凹侧TGFβ1、bFGF高表达和核心蛋白低表达,表明细胞因子和核心蛋白的表达异常可能是脊柱侧凸的发病原因或继发于侧凸的变化。 相似文献