EFFECTS OF CEREALS FROM KASHIN－BECK DISEASE ENDEMIC AREA ON FIBRILLOGENESIS IN VITRO OF CARTILAGE COLLAGEN TYPE Ⅱ IN RATS

In this feeding trial rats were fed on diets of cereals from Kashin-Beck Disease (KBD) endemic area, Se-supplemented cereals from the above area, and cereals from Non-KBD endemic area. The purpose of this paper was to investigate the effects of cereals from KBD endemic area and Se on the formation kinetics of cartilage type Ⅱ collagen fibril, the stability and ultrastructure of fibrils formed in vitro. The results indicated that low-selenium cereals from KBD endemic area resulted in a decelerated rate and extant of forming the type Ⅱ collagen fibril, the fibril stability reduced, fibril diameters diminished, and fibril banding periods increased or decreased in vitro. Sesupplemented cereals from KBD endemic area partially rectified the pathologic changes mentioned above. These data are important for further studying the etiology and pathology of KBD.

TGF-β_1、BMP-2和TypeⅡ Collagen在黄韧带中的表达及意义

目的研究TGF-1β、BM P-2和typeⅡco llagen在退行性腰椎滑脱(degenerative lum bar spondy lo listhes is,DLS)和腰椎间盘突出症(lum bar d isc hern iation,LDH)黄韧带中的表达及其意义。方法37例手术切除的腰椎椎板间部黄韧带标本分为3组,第1组为退行性腰椎滑脱组(DLS)10例;第2组为腰椎间盘突出症组(LDH)17例,第3组为正常对照组10例,其中7例取自腰椎骨折手术病人,3例取自意外死亡者。应用EnV is ion二步免疫组化的方法检测其TGF-1β、BM P-2和typeⅡco llagen的表达情况,普通光镜观察,计算出各标本的表达阳性率和表达强度,数据以x-±s标准差及表达强度表示,结果分别用Spss统计软件和R id it进行分析。结果TGF-1β、BM P-2和typeⅡco llagen的阳性表达产物见于成纤维细胞、成软骨细胞和软骨细胞中,而Ⅱ型胶原染色还可同时见于基质。TGF-1β、BM P-2和typeⅡco llagen在DLS组中的表达明显高于LDH组和正常组(P<0.01或P<0.05),Ⅱ型胶原基质染色明显深于LDH组和对照组。LDH组的TGF-1β和typeⅡco llagen的表达阳性率和表达强度与正常组之间差异无显著性(P>0.05),而BM P-2的表达阳性率和表达强度在LDH组与正常组之间具有统计学意义(P<0.01)。结论黄韧带所受到的异常机械牵张力可以增加TGF-1β在黄韧带细胞中的合成,而TGF-1β则促进退行性腰椎滑脱黄韧带中的Ⅱ型胶原合成,导致黄韧带的退变和肥厚。BM P-2在退变黄韧带中的表达异常增高,可能与黄韧带的软骨化倾向有关。

The Effects of Tetrandrine (TT) and Polyvinylpyridine-N-Oxide (PVNO) on Gene Expression of Type Ⅰand Type ⅢCollagens during Experimental Silicosis

In the screening tests of drugs for silicosis in our laboratory, we found that TT, a type of alkaloid isolated from Stephania tetrandra, could inhibit the development of experimental silicosis of rats and the synthesis of collagen in rat lung. Chest X-rays of silicotic patients treaied with TT for 1-3 years showed obvious changes. The silicotic nodules became smallel and shadows became clearer. PVNO was proved to have anti-silicotic effect on animal and clinically. This presentation reports the effect of them on collagen mRNA.Dot blot results showed that 1 (Ⅰ) and 1 (Ⅲ) mRNA levels increased significantly at 60 and 120 days after the rats were exposed to silica dust. The mRNA levels went down at 1 and 3 months after treated by TT and PVNO. In situ hybridization observation revealed that the silver grains of Type Ⅰand Type Ⅲ collagen were scattered within the fibroblasts in cellular nodules and in thickened interstitium of silicosis tissue. The amounts of mRNA silver grains decreased in the lung tissue treated by TT and PVNO. It was suggested that TT and PVNO may inhibil the gene expression of collagen during silicosis

Is type Ⅰ alpha 2 collagen gene responsible for intracranial aneurysm in Northeast China?

In this study, we investigated whether a single nucleotide polymorphism （rs42524 G 〉 C） in the type I alpha 2 collagen gene was associated with sporadic ruptured intracranial aneurysm or its clinical characteristics in patients from Northeast China. Genotyping of the rs42524 G 〉 C polymorphism was carried out using a polymerase chain reaction-restriction fragment length polymorphism assay. The data showed that the frequency of the rs42524 GC ＋ CC genotype was significantly higher than the GG genotype among intracranial aneurysm patients whose Hunt and Hess grading scale was 〉 3. In addition, the rs42524 G 〉 C genotype was found to have a statistically significant association with intracranial aneurysm risk. These findings indicate that the type I alpha 2 collagen gene gene may be involved in a predisposition to intracranial aneurysm in the Northeast Chinese population. Crucially, the rs42524 C allele may be an important risk factor for increased severity of the condition in patients with ruptured intracranial aneurysms.

Autosomal dominant osteopetrosis typeⅡresulting from a de novo mutation in the CLCN7 gene:A case report

BACKGROUND Osteopetrosis is a family of extremely rare diseases caused by failure of osteoclasts and impaired bone resorption. Among them, autosomal dominant osteopetrosis type Ⅱ(ADO Ⅱ), related to the chloride channel 7(CLCN7) gene, is the most frequent form of osteopetrosis. In this study, we report a de novo mutation of CLCN7 in a patient without the family history of ADO Ⅱ.CASE SUMMARY A 5-year-old Chinese boy with ADO Ⅱ was found to have a de novo mutation in the CLCN7 gene [c.746 C>T(p.P249 L)]. Typical clinical manifestations, including thickening of the cortex of spinal bones and long bones, non-traumatic fracture of the femoral neck, and femoral head necrosis, were found in this patient. The patient is the first reported case of ADO Ⅱ with the missense mutation c.746 C>T(p.P249 L) of the CLCN7 gene reported in China. We also review the available literature on ADO Ⅱ-related CLCN7 mutations, including baseline patient clinical features, special clinical significance, and common mutations.CONCLUSION Our report will enrich the understanding of mutations in ADO Ⅱ patients. The possibility of a de novo mutation should be considered in individuals who have no family history of osteopetrosis.

The effects of radiofrequency hyperthermia on type Ⅱ collagen formation in the osteoarthritic knee

Objective: To explore the effect of radiofrequency hyperthermia on type Ⅱ collagen expression in a rabbit model of osteoarthri-tis (OA). Methods: Experimental model of knee OA was replicated in the right hind limbs of 54 male rabbits by using modified Hulth modeling method. The rabbits were randomly divided into Model group, Lugua Polypeptide group and Radiofrequency Hyper-thermia group. After modeling, Lugua Polypeptide group was given intramuscular injection of Lugua polypeptide;Radiofre-quency Hyperthermia group was treated with radiofrequency hyperthermia;Model group was not given any special treatment. On the 7th, 13th and 19th day after radiofrequency hyperthermia, six experimental rabbits were chosen from each group and sacrificed to take out medial femoral condyle cartilages in the right hind limbs. Modified Mankins rating was applied to the morphological evaluation. Meanwhile, quantitative real-time PCR was used to detect the content of type Ⅱ collagen in cartilage tissues of medial femoral condyle. Results: At each time point after treatment, Mankins scores were decreased in all the 3 groups, with that of Model group sig-nificantly higher than those of the other two groups (Model group > Lugua Polypeptide group > Radiofrequency Hyperthermia group). The contents of type Ⅱ collagen in cartilage tissues were increased in all the 3 groups, with that of Radiofrequency Hyperthermia group significantly higher than those of the other two group (Model group < Lugua Polypeptide group < Ra-diofrequency Hyperthermia group). The difference between groups was of statistical significance (p < .05). For Radiofrequency Hyperthermia group, Mankins scores were decreased gradually as the treatment time went by, with the content of type Ⅱ colla-gen in cartilage tissues increased. The difference between time points was of statistical significance (p < .05). Conclusions: Radiofrequency hyperthermia is superior to Lugua polypeptide in the treatment of knee OA, at least in rabbits. Its therapeutic mechanism may be related to the significant increase in type Ⅱ collagen in cartilages.

Type XIX collagen: a promising biomarker from the basement membranes

Among collagen members in the collagen superfamily,type XIX collagen has raised increasing interest in relation to its structural and biological roles.Type XIX collagen is a Fibril-Associated Collagen with Interrupted Triple helices member,one main subclass of collagens in this superfamily.This collagen contains a triple helix composed of three polypeptide segments aligned in parallel and it is associated with the basement membrane zone in different tissues.The molecular structure of type XIX collagen consists of five collagenous domains,COL1 to COL5,interrupted by six non-collagenous domains,NCI to NC6.The most relevant domain by which this collagen exerts its biological roles is NCI domain that can be cleavage enzymatically to release matricryptins,exerting anti-tumor and anti-angiogenic effect in murine and human models of cancer.Under physiological conditions,type XIX collagen expression decreases after birth in different tissues although it is necessary to keep its basal levels,mainly in skeletal muscle and hippocampal and telencephalic interneurons in brain.Notwithstanding,in amyotrophic lateral sclerosis,altered transcript expression levels show a novel biological effect of this collagen beyond its structural role in basement membranes and its anti-tumor and anti-angiogenic properties.Type XIX collagen can exert a compensatory effect to ameliorate the disease progression under neurodegenerative conditions specific to amyotrophic lateral sclerosis in transgenic SOD1 G93 A mice and amyotrophic lateral sclerosis patients.This novel biological role highlights its nature as prognostic biomarker of disease progression in and as promising therapeutic target,paving the way to a more precise prognosis of amyotrophic lateral sclerosis.

阿克他利对Ⅱ型胶原性关节炎小鼠的治疗作用及部分机制研究

目的 研究阿克他利 (Acta)对小鼠Ⅱ型胶原性关节炎 (CIA)的治疗作用。方法 采用Ⅱ型胶原 (CⅡ )乳剂皮内注射诱导的小鼠CIA模型。在此基础上 ,检测小鼠足肿胀、免疫功能的改变 ,以及对CⅡ的迟发性变态反应 (DTH)和血清中抗CⅡ抗体的测定 ,同时进行了病理组织学的检查。结果 Acta(10 ,30 ,90mg·kg-1)ig对CⅡ诱导的小鼠足肿胀有明显的抑制作用 ;体外研究发现 ,Acta(10 ,30 ,90mg·kg-1)能使CIA小鼠过高的ConA增殖反应和IL 2的产生恢复至接近正常 ,同时对CIA小鼠过高的IL 1产生有明显的抑制作用 ;Acta(10 ,30 ,90mg·kg-1)ig可以明显减轻CⅡ诱发迟发性变态反应 (DTH) ,但Acta对CIA小鼠体内的抗体的产生无显著影响。病理学检查表明 ,Acta(10 ,30 ,90mg·kg-1)ig可以减轻CIA小鼠的骨膜增生和软骨破坏。结论 Acta对CIA小鼠具有治疗作用 ,该作用可能是通过细胞免疫调节实现的。

Ⅱ型胶原蛋白与弗氏完全佐剂大鼠关节炎模型的建立和比较

目的对Ⅱ型胶原蛋白（ＣⅡ－Ａ）和弗氏完全佐剂（Ａ－Ａ）大鼠关节炎模型在大体外观和足部组织病理学切片等方面进行观察比较。方法分别采用Ⅱ型胶原蛋白和弗氏完全佐剂诱导建立大鼠关节炎模型，利用排水法对大鼠足部体积进行测定，并将大鼠后足进行组织病理学切片观察。结果从大体外观和足部病理切片上两种大鼠模型均显示出有明显的病变，但ＣⅡ－Ａ大鼠与Ａ－Ａ大鼠比较，滑膜增生及软骨破坏等继发性病变特征更为明显，关节炎持续时间也较长，更接近于人的类风湿性关节炎。结论ＣⅡ－Ａ大鼠模型与Ａ－Ａ相比是研究ＲＡ较为理想的动物模型。

丹参酮Ⅱ_A对AngⅡ诱导的心肌成纤维细胞增殖及Ⅰ型胶原合成的影响

目的:通过观察丹参酮ⅡA(TSN)对AngⅡ诱导的心肌成纤维细胞(CFs)增殖及Ⅰ型胶原合成的影响,探讨其抗心肌纤维化的作用机制。方法:差速贴壁法提取原代CFs,采用四氮唑盐(MTT)比色法测定细胞数目,ELISA法检测Ⅰ型胶原合成,RT-PCR法半定量检测Ⅰ胶原mRNA表达。结果:①AngⅡ组OD值高于空白对照组,两者比较有显著性差异(P<0.01);AngⅡ+TSN组OD值低于AngⅡ组,两者比较有显著性差异(P<0.01)。②AngⅡ组有促进心肌成纤维细胞分泌Ⅰ型胶原的作用,与空白对照组比较,有显著性差异(P<0.01);AngⅡ+TSN组Ⅰ型胶原含量低于AngⅡ组,两者比较,有显著性差异(P<0.01)。③AngⅡ组Ⅰ型胶原mRNA表达高于空白对照组,两者比较有显著性差异(P<0.05);AngⅡ+TSN组Ⅰ型胶原mRNA表达低于AngⅡ组,两者比较有显著性差异(P<0.05)。结论:AngⅡ可直接诱导CFs的增殖,促进其分泌Ⅰ型胶原,并能显著增加Ⅰ型胶原mRNA的表达;TSN对AngⅡ诱导的CFs增殖及Ⅰ型胶原分泌增加,及Ⅰ型胶原mRNA表达增强均有抑制作用。提示:TSN抗心肌纤维化作用的产生可能与丹参酮ⅡA抑制心脏局部的RAS系统有关。

鸡Ⅱ型胶原对小鼠胶原性关节炎的治疗作用

目的 研究鸡Ⅱ型胶原 (CⅡ )对小鼠胶原性关节炎(CIA)的治疗作用。方法 在建立小鼠CIA模型的基础上 ,观测关节积分变化 ,同时检测脾淋巴细胞增殖反应 ,白细胞介素 1(IL 1)和IL 2。结果 CⅡ 5、10、2 0 μg·kg-1ig× 7d能明显减轻CIA小鼠的关节炎症 ;抑制CIA小鼠过高的ConA诱导的脾细胞增殖反应及小鼠腹腔巨噬细胞IL 1的过度产生 ,降低脾细胞IL 2的产生 ;同时病理检查亦发现CⅡ治疗用药可以减轻CIA小鼠的骨膜增生和炎细胞浸润。结论 CⅡ对CIA具有治疗作用 ,其机制与免疫功能调节有关。

可溶性猪软骨Ⅱ型胶原蛋白的提取与鉴定

目的从猪膝关节软骨中提取纯化Ⅱ型胶原蛋白,并对其进行鉴定。方法选择猪膝关节软骨为提取原料,用盐酸胍去除蛋白多糖、胃蛋白酶两步消化、氯化钠盐析、超纯水透析、离心浓缩等方法提取纯化Ⅱ型胶原蛋白;采用SDS-PAGE、氨基酸成分分析对提取的Ⅱ型胶原蛋白进行鉴定;利用冻干的方法计算提取的浓度。结果 SDS-PAGE电泳结果显示提取的Ⅱ型胶原蛋白分子量约120 kD;氨基酸成分分析显示甘氨酸、脯氨酸和丙氨酸含量最高,并且可以检出羟脯氨酸、羟赖氨酸,符合Ⅱ型胶原特征;该法提取的Ⅱ型胶原蛋白浓度为66 mg/mL。结论从猪关节软骨提取的Ⅱ型胶原蛋白纯度高,方法简便,提取的胶原蛋白浓度更高。

小鼠Ⅱ型胶原性关节炎模型的制备

目的 制备Ⅱ型胶原诱发的小鼠关节炎模型。方法 利用Ⅱ型胶原乳剂皮内注射。结果 与对照组比较，Ⅱ型胶原性关节组小鼠的足爪肿胀和关节评分明显增加，并且Ⅱ型胶原诱发的迟发性变态反应呈阳性，血清中也检测出抗Ⅱ型胶原的抗体。结论 Ⅱ型胶原乳剂皮内注射可制备关节炎小鼠模型。

骨关节炎软骨中Ⅰ型和Ⅱ型胶原的分布

目的 :研究骨关节炎软骨中Ⅰ型和Ⅱ型胶原的分布。方法 :从正常关节软骨和骨关节炎软骨上取样本做切片。所有样本行HE、蕃红 0染色及Ⅰ型和Ⅱ型胶原免疫组化。结果 :骨关节炎软骨中Ⅱ型胶原免疫组化染色不均匀。Ⅰ型胶原染色 ,在表层和中层的部分区域有不规则着色。纤维样组织中 ,Ⅰ型胶原免疫组化呈阳性 ,Ⅱ型胶原免疫组化不着色。结论 :骨关节软骨基质中Ⅱ型胶原和蛋白聚糖的破坏增强与软骨细胞对其合成增强同时存在 ,软骨修复的过程中 ,部分软骨细胞发生去分化 。

类风湿性关节炎患者血清抗Ⅱ型胶原抗体的检测及意义

目的 探讨类风湿性关节炎 (RA)患者血清抗Ⅱ型胶原 (CⅡ )抗体阳性的意义。方法 分别以人Ⅱ型胶原蛋白(HCⅡ )及牛Ⅱ型胶原蛋白CⅡ (BCⅡ )作为包被抗原进行间接ELISA法 ,检测 30例RA患者 ,对照组 (30例非RA的风湿病患者和 34例健康人 )血清抗CⅡ抗体。结果 以HCⅡ及BCⅡ作为包被抗原检测RA患者血清抗CⅡ抗体的阳性率分别约为 30 .0 %和 33.3% ;对照组血清中抗体阳性率均约为 1.6 % ;二者差异极其显著 (P <0 .0 1)。抗CⅡ抗体阳性的RA患者RF的阳性率高于抗CⅡ抗体阴性的RA患者。与HCⅡ和BCⅡ均发生阳性反应的血清 8例 ,均为早期RA患者。结论 抗CⅡ抗体可作为判断RA患者病情的辅助手段之一 。

猪软骨Ⅱ型胶原蛋白的提取纯化与鉴定

目的:从猪关节软骨中提取纯化Ⅱ型胶原蛋白,并对其进行鉴定。方法:选择猪关节软骨为提取原料,用盐酸胍去除蛋白多糖、胃蛋白酶消化、氯化钠盐析、DE22纤维树脂处理等步骤,提取纯化Ⅱ型胶原蛋白。采用SDS-PAGE、吸收光谱分析、氨基酸成分分析等方法对Ⅱ型胶原蛋白进行鉴定。结果:4 mol/L盐酸胍能有效去除蛋白多糖;分步加入胃蛋白酶的限制性酶解结果满意;氯化钠盐析浓度为2.4 mol/L时最佳。SDS-PAGE电泳结果显示提取纯化的Ⅱ型胶原蛋白与S igm a公司的产品一致。Ⅱ型胶原最大吸收峰为230 nm,氨基酸成分分析显示甘氨酸、脯氨酸和丙氨酸含量最高。结论:实验结果提示从猪关节软骨提取的Ⅱ型胶原蛋白纯度高,符合II型胶原的特征。提取材料广泛、实验条件简便,结果可靠。

Ⅱ型胶原蛋白的结构、功能及其最新研究进展

Ⅱ型胶原蛋白是动物体内透明软骨的主要成分,具有良好的生物相容性、可降解性、可促进细胞生长和再分化等生物学特性,可作为人体组织工程材料,也可应用于食品、日化、包装等领域。利用基因工程技术生产重组Ⅱ型胶原蛋白是该领域的研究热点。综述了Ⅱ型胶原蛋白的结构、功能及应用,阐述了基因工程技术生产重组Ⅱ型胶原蛋白的最新研究进展,对未来羟化胶原蛋白在大肠杆菌中的大规模生产进行了展望,以期为Ⅱ型胶原蛋白的进一步开发和同行研究提供指导和帮助。

髁突骨上组织改建中Ⅰ、Ⅱ型胶原的免疫组织化学研究

目的 :研究髁突骨上组织改建中Ⅰ、Ⅱ型胶原的变化特征 ,增加对颞颌关节改建的进一步认识。方法 :用免疫组织化学的方法检测拔除成年家兔左侧下颌磨牙后 2周、1月和 3月时Ⅰ、Ⅱ型胶原分布及含量的变化。结果 :术后 2周Ⅰ、Ⅱ型胶原的表达均明显减少 ,1月和 3月其表达有所回升 ,并且Ⅰ型胶原出现不均衡分布 ;拔牙侧与非拔牙侧相比 ,前者Ⅰ型胶原的表达较后者稍强 ,而Ⅱ型胶原的表达则较弱。结论 :成年家兔颞颌关节的改建能力有限 ,超过一定限度时 ,细胞合成及分泌Ⅰ、Ⅱ型胶原的能力发生改变 ,使纤维软骨的抗压性和弹性下降。

复元胶囊对大鼠膝骨关节炎软骨Ⅰ、Ⅱ型胶原及蛋白多糖的影响

目的观察复元胶囊对大鼠膝骨关节炎(OA)软骨Ⅰ、Ⅱ型胶原及蛋白多糖的影响,探讨其保护关节软骨的作用机理。方法采用Hulth法建立OA模型,随机分为6组:正常组、模型组、四环素组及复元胶囊低、中、高剂量组。各药物组分别给不同剂量药物灌胃每天1次,持续12周。免疫组化检测软骨Ⅰ、Ⅱ型胶原表达,甲苯胺蓝染色软骨蛋白多糖含量。结果复元胶囊各组软骨Ⅱ型胶原及蛋白多糖显著高于模型组(P<0.01),而Ⅰ型胶原低于模型组(P<0.05)。结论复元胶囊能增加大鼠膝骨关节炎软骨中Ⅱ型胶原表达及蛋白多糖水平,同时降低Ⅰ型胶原表达,对维持正常胶原表型、保护关节软骨有一定的作用。

骨桥蛋白干预体外培养人膝骨关节炎软骨细胞蛋白多糖和Ⅱ型胶原的表达

背景:骨桥蛋白基因和蛋白表达与骨关节炎的病变程度高度相关。目的:进一步验证骨桥蛋白对体外培养人膝骨关节炎软骨细胞蛋白多糖和Ⅱ型胶原mR NA表达的影响。方法:体外培养人膝骨关节炎软骨细胞。取第1代人膝骨关节炎软骨细胞,分为空白对照组、0.1 mg/L骨桥蛋白组和1 mg/L骨桥蛋白组。骨桥蛋白组加入重组骨桥蛋白0.1 mg/L或1 mg/L干预48 h;空白对照组未予任何干预继续培养48 h。用Real-time PCR测量蛋白多糖与Ⅱ型胶原的mR NA表达量。结果与结论:经0.1 mg/L与1 mg/L两种质量浓度骨桥蛋白干预后,人膝骨关节炎软骨细胞蛋白多糖与Ⅱ型胶原mR NA表达水平显著上升(P<0.05),1 mg/L骨桥蛋白组人膝骨关节炎软骨细胞蛋白多糖与Ⅱ型胶原mR NA表达水平高于0.1 mg/L骨桥蛋白组(P<0.05),人膝骨关节炎软骨细胞蛋白多糖mR NA表达水平与骨桥蛋白干预质量浓度呈正相关(r=0.751,P<0.01)。人膝骨关节炎软骨细胞Ⅱ型胶原mR NA表达水平与骨桥蛋白干预浓度呈正相关(r=0.676,P<0.01)。结果提示骨桥蛋白上调人膝骨关节炎软骨细胞蛋白多糖与Ⅱ型胶原mR NA的表达,并具有浓度依赖性。