射频电流对成纤维细胞增殖及胶原蛋白表达的影响

目的射频除皱技术在临床上的运用已经相当广泛,并取得了很好的疗效。但缺乏基础研究,以致对射频电流刺激成纤维细胞分泌胶原的机制缺乏了解,本研究旨在观察射频对皮肤成纤维细胞的作用机制,从而更好的推进射频技术的临床运用。方法采用组织块法原代培养成人皮肤成纤维细胞,建立传代细胞系,实验用第4~6代细胞。取对数生长期的成纤维细胞,分为对照组和射频电流组,建立无创性射频除皱技术的体外细胞实验模型,应用倒置相差显微镜观察细胞形态并计数,绘制细胞生长图表,采用酶联免疫法检测细胞分泌胶原蛋白的水平。结果经射频电流作用后细胞数量明显增加,并且在一定范围内随输出电压的增加而增多,同时促进细胞分泌及合成胶原蛋白。结论射频电流刺激人皮肤成纤维细胞的生长及增殖,刺激成纤维细胞分泌胶原,是一种有效的除皱紧肤的方法。     

川芎嗪和硫氮唑酮对照射后NIH/3 T3成纤维细胞增殖及bFGF表达的影响

目的　探讨川芎嗪及硫氮唑酮对照射后ＮＩＨ／３Ｔ３ 成纤维细胞增殖及ｂＦＧＦ表达的影响,为防治放射性肺纤维化的药物研究提供实验依据。方法　采用ＭＴＴ 法测定离体培养的ＮＩＨ／３Ｔ３ 成纤维细胞增殖程度并用免疫细胞化学ＡＢＣ 方法观察ｂＦＧＦ蛋白表达改变。结果　不同组ＭＴＴ法测得光密度值,对照组为０－７１ ±０－２２ ;照射组（Ⅰ） 为１－０３ ±０－２８ ;川芎嗪＋ 照射组为０－４７±０－１５ ;照射组（ Ⅱ） 为１－０５ ±０－２１;硫氮唑酮＋ 照射组为２－０２ ±０－２１;不同组ｂＦＧＦ 蛋白表达平均光密度：对照组为０－０２８±０－００５ ;照射组为０－０４４ ±０－００６ ;川芎嗪＋ 照射组为０－０２９ ±０－００１ ;硫氮唑酮＋ 照射组为０－０３６ ±０－００３。结论　川芎嗪明显抑制γ射线照射后ＮＩＨ／３Ｔ３ 成纤维细胞的增殖及ｂＦＧＦ蛋白表达,硫氮唑酮则不具有这一作用。     

病理性瘢痕与细胞因子相关性研究进展

瘢痕组织是皮肤创伤修复过程中的一种病理性产物,是成纤维细胞--胶原蛋白--细胞因子三者过度失衡,使细胞外基质(ECM)增生所致.烧伤、创伤、感染等均可造成瘢痕形成与增生,尤其是深Ⅱ度烧伤所引起的瘢痕是多年来困扰着临床的难题之一.为了探讨瘢痕形成过程中细胞因子作用机制、影响因素以及生物学调控等问题,对病理性瘢痕与细胞因子的相关性进行研究,可为临床预防和治疗病理性瘢痕提供新的思路.     

纤维黏连蛋白基因在翼状胬肉中表达的研究

 目的 以50例中国翼状胬肉患者为研究对象,探讨纤维黏连蛋白(fibronectin ,FN)表达水平的改变在翼状胬肉发生中的作用.方法 以正常眼结膜组织体为对照,应用半定量RT-PCR方法检测翼状胬肉组织中FN基因mRNA表达水平;应用Western印迹杂交检测FN蛋白表达的改变.结果 在50例样本中,40例FN蛋白在翼状胬肉中表达升高,其中包括39例FN mRNA表达水平上调.结论 翼状胬肉组织中FN mRNA和蛋白表达水平明显上调,提示其有可能在翼状胬肉的发生中发挥一定的作用.     

二乙基二巯基氨基甲酸钠对大鼠皮肤成纤维细胞增殖及胶原蛋白合成的影响

目的：研究二乙基二巯基氨基甲酸钠（ＤＴＣ） 对大鼠皮肤成纤维细胞增殖及胶原蛋白合成的影响。方法：采用［３Ｈ］ － 胸腺嘧啶核苷和［３Ｈ］ － 脯氨酸掺入法,分别测定成纤维细胞增殖和胶原蛋白合成。结果：ＤＴＣ低剂量时促进成纤维细胞增殖,高剂量时抑制成纤维细胞的增殖和胶原蛋白合成。结论：ＤＴＣ能抑制胶原蛋白的合成,对成纤维细胞的增殖具有双向调节作用     

乳腺癌超声征象与分子生物学表达的相关性研究

目的 :探讨乳腺癌超声征象与分子生物学表达的相关性,以提高超声对乳腺癌的诊断水平,为乳腺癌治疗及预后评估提供可靠的影像学信息。方法:随机抽取经手术病理证实为乳腺癌的患者50例,均行乳腺高频超声检查,重点观察肿块的形态、边缘、后方回声、微小钙化、内部血流及腋窝淋巴结;均采用免疫组化法检测ER、PR及Cerb B-2基因表达,并分析其与超声表现的关系。结果:12 cm乳腺癌微小钙化检出率最高,其次为边缘毛刺状;形态不规则在2 cm的肿瘤检出率较高。2肿块纵横比1患者的ER、PR阳性率为77.8%、61.1%,较纵横比≤1者高(P0.05);3边缘毛刺征患者的ER、PR阳性率分别为65.5%、55.2%,较无毛刺组高(P0.05);4周边高回声晕患者的ER和PR阳性率分别为73.3%、60.0%,均高于无高回声晕患者(P0.05);5内部无回声区患者的PR阳性率22.2%,低于未见无回声区患者(P0.05);6微小钙化患者的Cerb B-2阳性率83.3%,高于未见微钙化者(P0.05)。结论:乳腺癌超声表现与分子生物学指标的表达有一定的关系,肿瘤生物学特性影响乳腺癌的超声表现。超声检查可为乳腺癌的预后评估及临床治疗方式的选择提供影像学依据。     

胰腺癌表观扩散系数与纤维化含量、成纤维细胞活化蛋白表达相关性研究

目的：探讨高场磁共振扩散成像表观扩散系数（ADC ）值与纤维化含量、成纤维细胞活化蛋白（FA P ）表达评分的相关性。方法对18例经病理证实的胰腺癌患者进行3．0T MRI平扫及扩散加权成像（DWI）扫描,使用单一层面法选取感兴趣区（ROI）测量癌区ADC值,并对胰腺癌患者的蜡块进行Masson染色及FAP免疫组化染色,采用 Pearson相关分析检验ADC值与纤维化含量、FA P染色评分的相关性。结果胰腺癌区的ADC值与纤维化含量呈负相关（ r＝－0．459）,但无统计学意义（ P＝0．056）,中～高分化组ADC值与纤维化含量呈负相关（ r＝－0．564,P＝0．044）。ADC值与 FA P染色评分呈负相关（ r＝－0．479,P＝0．036）。结论胰腺癌的ADC值与其病理参数间呈负相关,DWI有望对胰腺癌的病理特征实现定量成像,有助于进一步了解其病理特征。     

纤连蛋白的分段表达及与乙肝表面抗原相互作用研究

目的:克隆纤连蛋白(fibronectin)各功能区域基因,在大肠杆菌中表达以筛选与HBsAg相互作用的区域.方法:将纤连蛋白分成7个片段,设计引物,利用RT-PCR从HepG2.2.15中扩增出相应的PCR产物,连接到表达载体pGEX4T-1中,用IPTG诱导目的蛋白在E.coli中表达,通过Western印迹分析对表达产物进行鉴定.融合蛋白纯化后制备蛋白芯片,与HBsAg杂交,筛选纤连蛋白中与HBsAg相互作用的区域.结果:经RT-PCR从HepG2.2.15中扩增到各结构域片段的cDNA,序列分析均正确;SDS-PAGE分析表明,各结构域片段的可溶性表达产物均占细菌可溶性蛋白的10%以上;Western印迹分析提示,诱导表达产物可与GST单抗发生特异性反应.经制备蛋白芯片并杂交,发现片段FN1、FN2信号最强.结论:成功地表达了纤连蛋白各片段,蛋白芯片杂交结果显示纤连蛋白可能是通过Ⅱ型肝素结合区及其羰基端(c端)与HBsAg相互作用.     

周围神经损伤后碱性成纤维细胞生长因子mRNA表达的实验研究

目的　研究大鼠坐骨神经钳夹伤后碱性成纤维细胞生长因子（ｂＦＧＦ）ｍＲＮＡ表达的变化规律。方法　１０２ 只Ｗｉｓｔａｒ 大鼠随机分为正常组６ 只，对照及实验组各４８ 只，用持针钳钳夹坐骨神经，于伤后４ 小时、１，３，７，１０，１４，２１，２８ 天采用逆转录－ 聚合酶链反应技术（ＲＴ－ ＰＣＲ） 结合聚丙烯酰胺凝胶电泳及ａｌｐｈａ－３２ｐ－ ｄＣＴＰ放射自显影，检测大鼠坐骨神经钳夹伤局部、腰４ ～６ 背根节及相应脊髓节段ｂＦＧＦｍＲＮＡ的动态变化。结果　实验组伤后４ 小时ｂＦＧＦｍＲＮＡ在损伤局部、脊髓及背根节的表达均开始增强，分别为１．４１±０．１０，１．７８±０ ．１０ 和２．１７±０．１２；伤后７ 天表达值最高，分别为２．７２±０．１４，３ ．６５±０．１７ 和３．９０ ±０．０６；伤后２８ 天表达值为１．２３ ±０．０６ ，１．４３±０ ．０５，１ ．７８±０．０３，降至正常及对照组水平（Ｐ＞ ０．０５）。结论　周围神经损伤后内源性ｂＦＧＦｍＲＮＡ表达增强有助于神经再生     

碱性成纤维细胞生长因子与血脑屏障的实验研究

血脑屏障（ＢＢＢ）能否让碱性成纤维细胞生长因子（ｂＦＧＦ）通过并发挥其神经营养因子作用其机制尚不明确。本文采用１２５Ｉ－标记ｂＦＧＦ经动物腹腔注射后２５ｍｉｎ检测实验动物全身各组织放射活度,计算各组织ｂＦＧＦ的含量。结果：脑ｂＦＧＦ含量在成年鼠组为（１７．３６±８．３０）ｐｇ／ｍｇ．新生鼠组为（３１．９６±１０．１１）ｐｇ／ｍｇ,新生鼠缺氧组为（５６．６８±１５．７９）ｐｇ／ｍｇ,各组相差具有显著意义（Ｐ＜０．０５～０．０１）;以同一时相血液ｂＦＧＦ含量为参照,成年鼠组脑ｂＦＧＦ含量占全血ｂＦＧＦ含量的（６．５１±３．８０）％,在新生鼠组为（１４．０９±３．１２）％,在新生鼠缺氧组为（２６．２４±１１．８３）％,各组相差具有显著意义（Ｐ＜０．０５～０．００５）。结果提示：ｂＦＧＦ可通过血脑屏障。新生鼠组及其在缺氧状态下应用ｂＦＧＦ,可使脑内ｂＦＧＦ含量较成年鼠组显著增多,故ｂＦＧＦ可能起保护作用。     

Is the increase in type III collagen of the patellar tendon graft after ligament reconstruction really caused by “ligamentization” of the graft?

To test the hypothesis that extrinsic cells that infiltrate the devitalized patellar tendon (PT) synthesize type III collagen even in the environmental milieu of the native PT, we conducted the present experimental study using the rat in situ frozen–thawed PTs. Tissue culture showed no cell outgrowth from the tendons immediately after the freeze–thaw treatment. Analysis by RT-PCR showed that the expression level of type III procollagen mRNA in the frozen–thawed tendon was significantly higher than that in the sham-operated tendon at 6 and 12 weeks. Immunohistological findings showed positive type III collagen staining around cells that had infiltrated the necrotized tendon at 3, 6, and 12 weeks. In addition, the elastic modulus of the in situ frozen–thawed tendon at 6 weeks was significantly less than that of the sham-operated tendon. The present study indicates that extrinsic cells that had infiltrated the devitalized PT synthesized type III collagen at least for 12 weeks even in the environmental milieu of the native PT. These findings raised the question whether the increase in type III collagen of the PT graft after ACL reconstruction is really caused by “ligamentization,” the adaptation of the PT graft to the ACL environment.     

电子射线照射大鼠胶原及自由基改变的研究

目的 探讨加速器电子射线局部照射SD大鼠致放射损伤后胶原表达的变化、自由基的改变及两者的相关性。方法 采用直线加速器局部照射制作SD大鼠的辐射损伤模型，观察MDA、NOS、SOD、GSH-PX及Ⅰ/Ⅲ型胶原mRNA表达及蛋白含量的改变。结果 照射后在一定剂量范围内MDA、NOS上升；SOD、GSH．PX则下降，血清中指标变化显著而皮肤中变化无明显规律性。受照射局部皮肤Ⅰ/Ⅲ型胶原mRNA表达及蛋白含量有明显改变，但其自由基的变化与胶原的变化无明显相关性。结论 辐射损伤后能在全身产生自由基，其对受照射局部皮肤组织中胶原表达改变的作用不大，在电子射线放射损伤中胶原可能起重要作用。     

夹层胶原"三明治"法冻存复苏肝细胞的实验研究

目的 研究胶原、表皮生长因子对新鲜分离及冻仔复苏后Sprague-Dawley（SD）大鼠肝细胞活率、形态和功能的影响,期望建立一种较为完善的肝细胞体外培养及冻存复苏体系,为肝细胞移植、生物人工肝及相关肝细胞研究获取充足有效的肝细胞来源提供必须的技术支持和理论依据。方法 采用胶原酶二步原位循环肝脏灌注法分离SD大鼠肝细胞,测定不同条件培养及冻存复苏后各组肝细胞的活性及功能。结果 双层胶原及双层胶味＋表皮生长因子组细胞活率、蛋白质含量及细胞功能均优于普通组（P〈0．05）。结论 双层胶原体外培养体系更接近于肝细胞体内生长环境,能有效的减少各种理化因素对肝细胞的损伤,有利于肝细胞在体外长期保存,是一种较为理想的培养及冻存复苏体系。     

Protection of mice against X-ray injuries by the post-irradiation administration of guanosine and inosine

Purpose:?To examine the radioprotective action of guanosine (Guo) and inosine (Ino) administered to mice after irradiation with X-rays.

Materials and methods:?Survival of mice exposed to lethal and sublethal doses of X-rays was studied. Peripheral blood cells were counted using a light microscope. The damage to bone marrow cells was assessed by micronucleus (MN) test. Damage and repair of DNA in blood leukocytes were estimated using the comet assay.

Results:?Mice injected intraperitoneally (i.p.) with Guo or Ino (～30 μg g^?1, i.e., ～0.6 mg per 20-g mouse) 15 min after acute whole-body irradiation with 7 Gy recovered from X-ray injury. On the 30th day after irradiation, 50 and 40% of mice injected with Guo and Ino, respectively, remained alive. The dose reduction factor (DRF) was 1.23 for Guo and 1.15 for Ino. The protective effect gradually decreased as the time interval between the irradiation and injection was increased to 3, 5, 8 h. Guo and Ino facilitated the restoration of peripheral blood cell counts. These compounds protected bone marrow cells from damage and normalized erythropoiesis. Guo and Ino contributed to a more rapid and complete repair of DNA in mouse leukocytes irradiated both in vitro and in vivo.

Conclusion:?Guo and Ino introduced shortly after irradiation reduce leukopenia and thrombocytopenia and offer promise as therapeutic agents for treatment of radiation injuries.     

STAT3与辐射敏感相关性的研究进展

放疗是肿瘤治疗的主要方法之一, 但肿瘤的辐射抵抗是影响放疗疗效的一个重要原因。信号转导和转录激活因子3(STAT3)是STAT家族的重要成员, 与肿瘤的发生、发展紧密相关。电离辐射可以对STAT3的表达水平产生影响, 同时STAT3的表达水平也可反作用于细胞, 通过对肿瘤细胞DNA损伤、自噬、凋亡、增殖、血管生成、迁移、侵袭、免疫抑制等的调节影响其辐射敏感性, 这预示着以STAT3为突破口的辐射敏感性研究将会为肿瘤临床治疗提供新策略。     

EPR investigation of the gamma-ray-irradiated natural and tanned collagen

Free radicals produced in natural and tanned collagen by gamma-ray irradiation within 1–15 kGy absorbed dose ranges were investigated by EPR spectroscopy. Tanned collagen was prepared using formaldehyde as well as aluminum basic salts [Al(OH)SO₄] tanning processes. Both natural and formaldehyde-tanned irradiated collagen show the same kind of EPR spectrum consisting of a single broad, slightly asymmetric line. Irradiated collagen tanned by aluminum basic salts process displayed a complex EPR spectrum consisting of a superposition of broad and narrow lines. A computer simulation of this spectrum allowed to evidence the presence of seven different kinds of paramagnetic centers, including those observed in the irradiated natural collagen. Corresponding Spin Hamiltonian parameters (g-factor, hyperfine splitting constant) as well as relative concentrations of these centers were calculated. Experimentally determined relative concentrations display a positive correlation with the absorbed dose described by a linear-type dependence. After three weeks of storage at room temperature, the concentration of some centers diminished by about 50%. The possible nature of these centers is discussed in connection with the local structure of the tanned collagen.     

腰椎间盘MRI信号强度与髓核胶原蛋白含量的相关性

目的研究腰椎间盘 MRI 信号强度与髓核内胶原含量之间的相关性。方法 31例腰椎间盘突出症患者,全部行经皮穿刺腰椎间盘摘除术(PLD),得到31个髓核样本,用改良羟脯氨酸测定法测定其内胶原蛋白含量;在 MR 矢状面 T_2WI 上测量腰椎间盘髓核及脑脊液的信号强度,计算出二者的比值。分别计算胶原含量与信号强度比值、腰椎间盘 T_2WI 信号强度之间的线性相关系数。结果髓核内胶原含量为(231.0±63.5)mg/g,信号强度比值为0.19±0.07,二者之间的 r 值为-0.61(P<0.01)。腰椎间盘 T_2WI 信号强度为80.5±31.7,胶原含量与其之间的 r 值为-0.53(P<0.01)。结论腰椎间盘 MR 信号强度与髓核内胶原含量之间存在负相关;胶原蛋白增多是导致退变腰椎间盘 T_2WI 信号强度降低的因素之一。     

Analysis of the immunohistochemical localization of collagen type III and V for the time-estimation of human skin wounds

Summary Collagen type III and V were visualized immunohistochemically in 79 surgically treated human skin wounds with a wound age between 8 h and 2.5 months. Network-like structures positively staining for collagen type III and associated with fibroblastic cells in the wound area were first detectable in a 2.5-day-old skin lesion and occurred regularly in wounds more than 5 days old. Collagen type V appeared first in the wound area after about 3 days, slightly later than collagen type III, and was detectable regularly in wounds with a survival time of 6 days or more. The immunohistochemical detection of collagen type III or type V thus indicates a wound age of at least 2–3 days. The lack of a positive reaction in a sufficient number of specimens indicates a wound age of less than 6 days. Even though both collagen types could also be detected in older wounds (wound age 2.5 months), further information for the time-estimation of older skin wounds cannot be given due to the observation that the time period during which reparative processes can be observed depends on the extent of the wound area.     

Immunohistochemical localization of collagen types I and VI in human skin wounds

Summary A total of 74 human skin wounds were investigated and collagen types I and VI were localized in the wound area by immunohistochemistry. Collagen type I appeared in the form of ramifying string-like structures after approximately 5–6 days, but positive reactions in the form of a spot-like staining around isolated fibroblasts also occurred in a skin wound aged 4 days. Collagen VI was detectable after a post-infliction interval of at least 3 days showing a strongly positive reacting network associated with fibroblasts in the wound area. Both collagens appeared almost constantly after a wound age of 6–7 clays and could also be found in wounds aged a few months. Therefore, although a positive reaction for collagen type I in the form of string-like and ramifying structures around wound fibroblasts indicates a wound age of at least 5–6 days, a spot-like positive staining for collagen type I cannot exclude a wound age of at least 4 days. A positive staining for collagen type VI represents a post-infliction time of 3 days or more. The almost constant appearance of these collagen types suggests that negative results in a sufficient number of specimens indicate a wound age of less than 6–7 days, but cannot completely exclude longer post-infliction intervals. Since collagen type I and VI are also found in the granulation/scar tissue of lesions with advanced wound age, the immunohistochemical analysis of these proteins provides no further information for an age determination of older skin wounds.