茵陈水提物对胆红素诱导神经母瘤细胞凋亡实验研究

目的采用分子生物学技术方法探讨胆红素诱导神经母瘤细胞(SH-SY5Y)凋亡和茵陈水提物(Yinchengextract,YCE)的保护机制。方法噻唑蓝(MTT)观察茵陈提取物对SH-S5Y5细胞增殖影响,Western blotting观察NF-KB蛋白核质分布,免疫荧光显微镜观察JC-1探针标记线粒体电位变化,流式细胞仪Annexin-FITC/PI观察细胞凋亡,荧光定量PCR观察IL-1β、TNF-α变化。结果茵陈提取物浓度的时间依赖性提高胆红素对细胞增殖的抑制作用(p<0.05)。胆红素促进NF-KB核转移,茵陈提取物抑制NF-KB核转移。免疫荧光显微镜观察到胆红素处理组与正常组对比线粒体膜电位明显降低(p<0.05)表现早期凋亡特性。茵陈提取物抑制线粒体膜电位降低(p<0.05)。流式细胞Annexin-FITC/PI结果显示,胆红素诱导细胞凋亡,YCE保护胆红素诱导的凋亡。荧光定量PCR结果显示茵陈提取物抑制NF-KB转录因子调控的炎症相关蛋白IL-1β、TNF-α基因的表达(p<0.05)。结论胆红素明显诱导SH-S5Y5细胞凋亡,YCE能有效抑制NK-KB激活和抑制线粒体膜电位变化,呈现多靶点抑制胆红素诱导的细胞凋亡。     

喜树碱诱导鼻咽癌细胞凋亡的线粒体机制探讨

目的 :了解喜树碱 (CPT)诱导鼻咽癌细胞凋亡是否与线粒体膜电位改变有关。方法 :用 2 μmol/ L 的CPT 处理 CNE- 2 Z细胞 ,经流式细胞仪、Hoechst332 5 8/ PI双重染色鉴定凋亡细胞 ,同时用罗丹明 12 3(Rhodamine12 3)染色后 ,经流式细胞仪检测线粒体膜电位。结果 :CPT可明显降低线粒体膜电位 ,2 μm ol/ L 的CPT处理 CNE- 2 Z细胞 2 4h后 ,罗丹明 12 3摄取量低的细胞明显增加 ,达 (19.0± 3.0 ) % ,与 DMSO对照组的(7.0± 1.4) %相比 ,差异有显著性意义 (P <0 .0 5 )。结论 :降低线粒体膜电位可能是 CPT诱导鼻咽癌细胞凋亡的机制之一。     

LMP1诱导鼻咽癌细胞发生TRAIL抵抗的体外实验

 目的探讨EBV膜潜伏蛋白1(latent membrane protein 1,LMP1)基因过表达后对肿瘤坏死因子相关的凋亡诱导配体(TNF related apoptosis inducing ligand,TRAIL)凋亡诱导作用和凋亡信号传导的影响。方法利用脂质体介导的pGL6 LMP1上调LMP1低表达的TRAIL抵抗性鼻咽癌细胞CNE 1中LMP1的表达;通过MTT比色法、流式细胞术观察LMP1过表达后对CNE 1细胞TRAIL敏感性的影响;流式细胞术、Western blot、线粒体膜电位检测观察LMP1过表达后对死亡受体表达、细胞内外凋亡信号通路激活、线粒体膜电位改变的影响。结果死亡受体荧光标记后流式细胞术检测发现CNE 1和CNE 1 LMP1之间细胞膜蛋白死亡受体4（death receptor 4,DR4）,死亡受体5（death receptor,DR5）表达差异无统计学意义（P>0.05）。Western blot结果显示TRAIL(100 ng/ml)分别作用2、4、6、12 h后,CNE 1细胞中caspase 8 p43/p41,p18亚单位蛋白和caspase 3 p17,p10亚单位蛋白表达明显高于CNE 1 LMP1细胞,而Bcl 2相互作用域死亡激动蛋白的截短形式（truncated form of Bid,tBid）和caspase 9 p35亚单位蛋白表达无明显改变,且两个细胞株间也无明显区别。JC 1线粒体膜电位检测法结果显示TRAIL干预后,线粒体膜电位无明显改变,且两个细胞株间也无明显区别。结论LMP1过表达抑制TRAIL对鼻咽癌细胞的凋亡诱导作用和其细胞外凋亡信号的传导,提示LMP1是通过抑制细胞外信号通路激活诱导鼻咽癌细胞发生TRAIL抵抗。     

美洛昔康对人喉癌Hep-2细胞株作用的研究

目的探讨美洛昔康对喉癌细胞株Hep-2凋亡作用和对体外培养喉癌Hep-2细胞周期的影响及作用机制。方法应用肿瘤细胞培养技术,随机分实验组和空白对照组,培养不同时间后,用流式细胞仪检测美洛昔康对Hep-2细胞凋亡发生率及对细胞周期的影响,同时检测美洛昔康作用Hep-2细胞后细胞线粒体跨膜电位的变化。结果流式细胞仪分析显示美洛昔康呈浓度依赖性诱导Hep-2细胞凋亡,且实验组G1期细胞数明显增加,与对照组比较差异有统计学意义（P〈0.05）;实验组Hep-2细胞线粒体跨膜电位明显下降,与对照组比较差异有统计学意义（P〈0.05）。结论美洛昔康具有诱导Hep-2细胞凋亡及抑制细胞分化的作用,其机制可能与触发了线粒体凋亡途径有关。     

姜黄素诱导鼻咽癌NCE细胞凋亡机制的研究

目的探讨姜黄素诱导人鼻咽癌NCE细胞凋亡的分子机制。方法通过吖啶橙-嗅化乙锭复合染色、原位末端脱氧核糖核苷酸转移酶分析法检测、电镜和DNA片段化分析法分析姜黄素对NCE细胞凋亡的影响，并应用流式细胞术、Western Blot、RT—PCR方法探讨姜黄素对细胞线粒体膜电位、半胱氨酸蛋白酶-3（cysteine proteases-3，caspase-3）酶活性、胞质CytC、Fas mRNA及蛋白表达的影响。结果终浓度100μmol／L的姜黄素可诱导NCE细胞凋亡；与姜黄素共孵育12、24、48h，凋亡细胞的阳性率分别为25．6％、40．3％、54．5％。姜黄素处理后NCE细胞线粒体膜电位下降，12、24、48h的线粒体膜电位下降的阳性率分别是26．8％、42．3％、68．2％。caspase-3酶活性增强，12、24、48h表达caspase-3酶活性细胞的阳性率分别是80．5％、100％、100％。胞质CytC含量显著增强，Fas mRNA及蛋白表达水平上升，Fas蛋白阳性率由33．6％上升到89．9％。结论姜黄素可能通过线粒体途径与死亡受体途径诱导鼻咽癌NCE细胞凋亡。     

线粒体参与荛花酚诱导人鼻咽癌CNE细胞凋亡机制

目的探讨荛花酚诱导人鼻咽癌CNE细胞(简称CNE细胞)凋亡的效应,初步探索其与线粒体相互作用的机制。方法以不同浓度的荛花酚作用于CNE细胞,MTT检测分析荛花酚对细胞增殖的影响,Annexin V/PI双标后流式细胞仪检测凋亡情况,Western blot检测凋亡相关蛋白(caspase-3、caspase-9、PARP、细胞色素C、Bax、Bcl-2)的表达。JC-1染色检测线粒体膜电位,DCFH-DA染色检测活性氧(reactive oxygen species,ROS)生成。结果荛花酚以时间和剂量依赖性方式抑制CNE细胞的增殖,荛花酚处理诱导CNE细胞发生明显凋亡,Bax/Bcl-2比值增加,细胞色素C从线粒体释放到胞浆,caspase-3和caspase-9活化,PARP发生剪切。荛花酚还可诱导CNE细胞线粒体膜电位下降,线粒体膜通透转运孔(PTP)开放,ROS含量升高。结论荛花酚可显著诱导CNE细胞的凋亡,机理与影响线粒体功能密切相关。     

新生豚鼠高胆红素耳蜗核谷氨酸、γ-氨基丁酸含量表达

目的观察高胆红素模型新生豚鼠耳蜗核谷氨酸(glutamicacid,Glu)和γ-氨基丁酸(gamma-aminobutyricacid,GABA)含量的变化,探讨Glu和GABA在高胆红素损伤模型低位听觉中枢耳蜗核中的作用和意义。方法选择正常新生豚鼠30只随机分为三组:对照组、低胆红素(100μg/g)处理组和高胆红素(200μg/g)处理组,8h后处死动物取材处理标本免疫荧光观察各组耳蜗核Glu和GABA含量,Western blotting(线粒体凋亡相关蛋白检测)观察细胞质细胞色素C含量变化。结果与对照组相比,高浓度和低浓度胆红素处理组耳蜗核中GABA的含量水平明显下降,差异有显著性(p<0.05),而Glu的含量水平则明显上升(p<0.01);Western blotting显示细胞色素C在胆红素高低浓度处理组表达明显升高。结论新生豚鼠高胆红素模型Glu和GABA含量的动态变化反映了神经元的活动状况,提示二者在胆红素耳神经毒性中可能起重要作用。细胞质细胞色素C增高说明高胆红素可能介导神经细胞早期凋亡。     

抗癌活性肽对鼻咽癌细胞周期的影响

目的探讨抗癌活性肽(anti-cancer bioactive peptide,ACBP)肝肽和科脾肽对人鼻咽癌细胞株CNE体外增殖抑制作用及细胞周期的影响。方法将人鼻咽癌CNE细胞与不同浓度的ACBP进行体外培养,采用四甲基偶氮唑蓝法测定不同浓度的ACBP对CNE细胞的生长抑制率;光镜下观察形态学变化;流式细胞仪分析细胞周期与凋亡率。结果质量浓度5．0-20．0μg／ml的ACBP均能抑制CNE细胞的生长,且随浓度及作用时间延长抑制率上升,20．0μg／ml ACBP作用72 h后,肝肽组的抑制率为63．0％,科脾肽组抑制率为46．7％。光镜下可观察到ACBP作用后的细胞凋亡现象。流式细胞仪结果显示:5．0μg/ml肝肽和科脾肽作用CNE细胞24 h的早期凋亡率较高,分别为(11．8±0．3)％和(8．1±0．2)％,分别与对照组比较,差异均有统计学意义(t值分别为42．535和47．300, P值均为0．000);20．0μg／ml的肝肽和科脾肽作用CNE细胞,随作用时间延长S期细胞构成比有明显上升。结论肝肽和科脾肽对CNE细胞均有抑制增殖的作用,其抗鼻咽癌细胞增殖作用机制可能为诱导肿瘤细胞凋亡,调控肿瘤细胞周期。     

高胆红素血症对听神经病的致病机制

听神经病是儿童期耳聋的重要原因之一,新生儿高胆红素血症是导致听神经病的最重要因素,当血清胆红素浓度达到一定水平时可引起新生儿听觉障碍、神经肌肉不协调、智能发育障碍等严重后遗症,甚至可危及生命,因此高胆红素血症已经成为引起小儿听觉损伤的重要病因之一。本文通过对相关文献的分析,就高胆红素血症与听神经病之间可能存在的联系做一综述。     

胆红素的神经毒性及耳毒性机制

胆红素脑病是高胆红素血症的主要并发症,它反映了胆红素对神经细胞的毒性作用,这种毒性作用主要累及部位包括基底神经节、耳蜗核及动眼神经核团.听觉系统对胆红素具有高度敏感性.胆红素对神经细胞的损伤机制尚未完全明确,很可能是以在膜水平上千扰膜通透性和功能开始的,继而导致细胞能量代谢紊乱,细胞内Ca2+增多,最终导致细胞凋亡或坏死.对胆红素神经毒性机制更深入的探索在保护中枢系统方面具有重要意义.     

The expression of vascular endothelial growth factor (VEGF) and VEGF‐C in early laryngeal cancer: relationship with radioresistance

The expression of vascular endothelial growth factor (VEGF) and VEGF‐C in early laryngeal cancer: relationship with radioresistance Angiogenesis is essential for tumour growth and invasion. Vascular endothelial growth factor (VEGF) is a prime mediator of tumour angiogenesis. VEGF‐C is a closely related protein that effects lymphatic endothelial cells and may be important in the process of lymphatic metastasis. The purpose of this study was to evaluate the expression of these cytokines in patients with T1 and T2a glottic, squamous cell carcinoma, in comparison with normal epithelial control tissue, to ascertain any association with radioresistance. Twenty‐two tumours treated by radiotherapy (13 radiosensitive, nine radioresistant) and seven normal control tissues were studied. The minimum follow‐up was 2 years after radiotherapy. Expression of VEGF and VEGF‐C was evaluated by immunohistochemistry of formalin‐fixed, paraffin‐embedded biopsy specimens. Analysis was carried out using a quantitative computer image analyser. Both VEGF and VEGF‐C were detectable in tumour and normal control specimens. There was increased expression in tumour specimens of both VEGF (P = 0.03) and VEGF‐C (P < 0.001). In addition, the expression of VEGF‐C was associated with tumours of higher histological grade (P = 0.021). There was, however, no difference in VEGF and VEGF‐C expression between radioresistant and radiosensitive tumours. The expression of VEGF and VEGF‐C is increased in early laryngeal squamous cell carcinoma (SCC). However, measuring the expression of these proteins cannot predict radioresistance in this tumour group.     

Treatment of Maxillary Sinus Carcinoma: Clinical Results Using the Kitasato Modality

The conventional therapeutic regimen for maxillary sinus carcinoma consists of dissection of the maxilla, full-dose irradiation and extensive chemotherapy. However, the results obtained with this treatment are often poor. Even when patients recover, their quality of life is significantly reduced as a result of deformity of facial structures and swallowing and articulation dysfunctions. A retrospective analysis of 68 patients with maxillary sinus carcinoma treated with the Kitasato modality between 1975 and 1999 was conducted. All patients underwent pergingival maxillary sinus surgery combined with pre- and postoperative irradiation therapy with standardized total doses of 16 Gy; the postoperative irradiation was given in combination with regional intra-arterial infusion chemotherapy administered via the superficial temporal artery. All visible tumor lesions were removed where possible in order to preserve or facilitate cellular immunity after surgery. The cumulative 5-year survival rates were 85.7% for Stage II patients, 88.1% for Stage III, 76.6% for Stage IVA and 75.0% for Stage IVB.     

Tongue Base Reduction with Temperature-controlled Radiofrequency Volumetric Tissue Reduction for Treatment of Obstructive Sleep Apnea Syndrome

In recent years a considerable effort has been made to establish the use of different surgical techniques for the treatment of obstructive sleep apnea syndrome (OSAS). Nevertheless, treatment of hypopharyngeal obstruction due to tongue base hypertrophy remains in many ways an unsolved problem. The aim of this study was to evaluate the safety and efficacy of tongue base reduction with temperature-controlled radiofrequency volumetric tissue reduction in the treatment of OSAS. Twenty patients with OSAS and tongue base hypertrophy were treated with radiofrequency tissue ablation. An intensified treatment protocol was used, delivering 2,800 J per treatment session under local anesthesia. Two nights of polysomnography testing were performed before and after treatment. Daytime sleepiness, snoring and postoperative morbidity were assessed using questionnaires. Mean respiratory disturbance index (RDI) was reduced from 32.1 to 24.9/h after a mean of 3.4 treatment sessions. Six patients (33%) were cured after the procedure (reduction in RDI of &#83 50% and a postoperative RDI of <15/h) and ten (55%) showed an improvement of >20% in their RDI. Daytime sleepiness and snoring improved significantly. Peri- and postoperative morbidity was low; one severe complication occurred (tongue base abscess). We were able to achieve similar cure and responder rates to those reported in a recently published pilot study but with a reduced number of treatment sessions. We believe that this technique may improve patient acceptance and have beneficial cost implications.     

Cervical Reflex Induced by Click Stimuli in Cats

We studied click-evoked potentials in the anterior horn of the spinal cord in 17 cats. A concentric needle electrode was inserted into the anterior horn of the spinal cord at levels C3-C6. Potentials evoked with 105 dB SPL clicks were recorded with a peak latency of 4.89-5.10 ms only at the C3 level. These responses were observed 45-60 dB SPL above the auditory brainstem response (ABR) threshold, and no potentials were evoked by stimulation of the contralateral ear. Average was performed 100 times with changes in stimulation frequency of 1-20 Hz. The amplitude of the potentials decreased with increasing stimulus frequency, but there were no changes in ABRs. The responses disappeared after destruction of the medial vestibulospinal tract at the obex level, but ABRs were still recorded. The spinal nucleus of the accessory nerves was located in the anterior horn of the spinal cord at levels C1-C6, and the sternocleidomastoid muscle motoneurons were found at levels C1-C3. The click-evoked potentials recorded in this study reflect responses of the spinal nucleus of accessory nerves through the vestibulospinal tract to click stimulation. The responses have the same characteristics as vestibular-evoked myogenic potentials that can be recorded using surface electrodes over the sternocleidomastoid muscles of humans.     

Lack of significant estrogen and progesterone receptor expression in nasal telangiectasias in hereditary hemorrhagic telangiectasia: an immunohistochemical analysis

Obstructive sleep apnea syndrome (OSAS) is characterized by snoring and apnea during sleep leading to decreased oxygen saturation and disturbed sleep, excessive daytime sleepiness and neuropsychological disturbances. This study investigates cognitive neuropsychological abilities in a group of 53 OSAS patients before and after treatment with uvulopalatopharyngoplasty. General intellectual ability, verbal learning and memory as well as executive functioning were measured at baseline and 6 months postoperatively. After surgery there were significant improvements in verbal learning and memory (mean change - 39, SD 57.3, p <0.001), recall (mean change - 24.3, SD 39.3, p <0.001) and executive functioning (as assessed by percentage of errors on the Wisconsin Card Sorting Test; mean change-9.1, SD 15.7, p <0.001). These improvements were in accordance with improvements in the degree of sleep apnea, the oxygen desaturation index (mean change -9.7, SD 15.9, p <0.001) and arterial minimum oxygen saturation (mean change 4.5%, SD 10.2%, p <0.01). Surgical treatment seems to improve verbal learning, memory and recall and executive functions in parallel with better oxygenation in OSAS.     

Measurement of chemosensory function

Although hundreds of thousands of patients seek medical help annually for disorders of taste and smell, relatively few medical practitioners quantitatively test their patients' chemosensory function, taking their complaints at face value. This is clearly not the approach paid to patients complaining of visual, hearing, or balance problems. Accurate chemosensory testing is essential to establish the nature, degree, and veracity of a patient's complaint, as well as to aid in counseling and in monitoring the effectiveness of treatment strategies and decisions. In many cases, patients perseverate on chemosensory loss that objective assessment demonstrates has resolved. In other cases, patients are malingering. Olfactory testing is critical for not only establishing the validity and degree of the chemosensory dysfunction, but for helping patients place their dysfunction into perspective relative to the function of their peer group. It is well established, for example, that olfactory dysfunction is the rule, rather than the exception, in members of the older population. Moreover, it is now apparent that such dysfunction can be an early sign of neurodegenerative diseases such as Alzheimer's and Parkinson's. Importantly, older anosmics are three times more likely to die over the course of an ensuring five-year period than their normosmic peers, a situation that may be averted in some cases by appropriate nutritional and safety counseling. This review provides the clinician, as well as the academic and industrial researcher, with an overview of the available means for accurately assessing smell and taste function, including up-to-date information and normative data for advances in this field.