【作者】 张彬彬；

【导师】 江新梅；

【作者基本信息】 吉林大学 ， 临床医学， 2011， 硕士

【摘要】 目的：分析Duchenne型肌营养不良的肌酸激酶随病程及发病年龄的变化规律；分析Duchenne型肌营养不良血清心肌酶的变化及早期心肌损害的情况；探讨Duchenne型肌营养不良临床病变程度、病理病变程度与免疫组织化学染色抗a-sarcoglycan、dysferlin、laminA/C、MHC-Ⅰ单克隆抗体的相关性。方法：采用回顾性研究方法,收集2008年10月～2010年10月期间,在吉林大学第一医院神经内科及儿科门诊、住院部就诊拟诊为Duchenne型肌营养不良的18例男性患者及2例女性携带者,行血清心肌酶检测、肌电图、肌肉超声、开放性骨骼肌活检、连续冰冻切片、组织化学染色、抗dystrophin-N,-C,-R单克隆抗体免疫组织化学染色,肌细胞膜dystrophin完全或近完全缺失确诊为DMD,并行抗a-sarcoglycan、dysferlin、laminA/C、MHC-Ⅰ单克隆抗体免疫组织化学染色,比较其结果,进行分析总结。结果：1、本组18例DMD患者临床资料,首发症状表现不一：以双下肢症状起病多见,共14例,占77.8%,心肌酶升高3例,占16.7%,转氨酶升高1例,占5.5%。18例均有不同程度的翼状肩胛、腓肠肌假性肥大、鸭步及Gower征,11例出现不同程度肌萎缩,占61.1%。11例近端肌力3～4级,占61.1%,7例近端肌力正常,占38.9%。11例膝反射减弱,占61.1%,12例足跟不能行走,占66.7%,3例足尖不能行走,占16.7%,不伴有肌肉握痛,病理征阴性。18例患者均无智力低下。14例行肌电图检查,均为肌源性损害。2例DMD女性携带者,均无明显阳性体征。2、血清肌酸激酶的变化规律,在病程上,18例DMD血清CK在发病初期随病程进展呈上升趋势,约在发病2～3年达到最高峰,均值为17577.78～16963.7U/L,随后随着病程持续进展,病变程度逐渐加重,血清CK整体呈逐渐下降趋势；发病年龄上,血清CK在2～5岁时达到峰值,均值为10838.25～12937U/L,其后随着年龄的增长,病变程度加重,肌肉坏死增多,血清CK呈下降趋势。3、本组18例DMD患者行血清心肌酶学检查,均有不同程度心肌酶升高,其中CK升高尤为显著,平均10331.16±8095.62U/L,CK-MB平均417.88±304.69U/LLDH平均1023.19±393.18U/L,HBD平均910.23±358.19U/L, AST平均217.96±108.12U/L。CK-MB/CK>6%者,7例。4、18例DMD患者行肌肉超声者6例,均有不同程度皮下脂肪增厚,双侧肌束对称性变细,约0.3～0.5mm左右,肌束膜回声增强及肌束连续性中断或消失,肌束纹理模糊不清,呈弥漫性细腻样脂肪回声。5、组织化学染色,光镜下HE切片,肌萎缩纤维直径10～70gm之间,大小不等,萎缩肌纤维呈圆形,镶嵌状分布,可见多数不透明纤维,散在坏死及再生肌纤维,部分切片肌膜核增多,核内移,肌纤维间质增宽,肌间结缔组织不同程度增生。6、18例DMD患者抗dystrophin-N,-R,-C单克隆抗体免疫组织化学染色,肌纤维膜dystrophin呈完全或近完全缺失,其中,14例α-SG不同程度表达欠缺,4例表达正常,18例抗dysferlin、laminA/C均正常表达,6例MHC-Ⅰ表达上调,12例MHC-Ⅰ表达正常。结论：1、DMD患者血清CK值显著升高,可达正常值的数十至数百倍,在病程2～3年、发病年龄2-5岁时,血清CK达到峰值,以后随病程进展和年龄增长逐年呈下降趋势。2、CK-MB/CK>6%对判定早期心肌受损有临床意义,可作为早期检测DMD心肌损害的指标之一。3、肌肉超声检查目前正逐渐成为研究肌肉疾病一种新的有效手段。4、肌纤维的变性病变与增生病变共存是DMD患者肌肉病理特征性的改变,Opaque纤维是DMD的特征性变化之一,但不具有特异性。随病情进展,肌纤维逐渐被脂肪结缔组织代替,临床症状越重,这种替代越重。5、DMD患者抗dystrophin-N,-R,-C呈完全或近完全缺失,同时部分伴有a-sarcoglycan欠缺,说明dystrophin缺失会影响a-sarcoglycan的表达,推测sarcoglycans参与了肌纤维的变性坏死过程。6、DMD患者抗dysferlin、laminA/C均表达正常,推测其合成与装配与dystrophin无直接相关性,同时也进一步证明dysferlin蛋白与肌营养不良糖蛋白复合体(DGC)无关。MHC-Ⅰ在部分DMD骨骼肌细胞膜表达上调,推测可能与肌细胞变性、坏死后继发炎症反应有关,确切机制有待于进一步研究。更多还原

【Abstract】 Objective:To find the relationship between serum creatine kinase level in DMD patients with the course of the disease and age of onset; Analyze serum myocardial enzymes changes and early myocardial injury in DMD patients; Explore correlation between extent of clinical damage and degree of pathological lesions, immunohistochemical staining resistance a-sarcoglycan, dysferlin, laminA/C, MHC-I monoclonal antibodies.Methods:Using a retrospective study method, departments of Neurology and Pediatrics in the first hospital of jilin university, out-patient and in-patient cases, collection of 18 cases of male patients fitting DMD and 2 cases of female carriers from Oct.2008 to Oct.2010 period. The patients underwent serum myocardial enzymes detection, electromyography, and muscle ultrasound, skeletal muscle biopsy, continuous freezing in liquid nitrogen,histochemistry dyeing, all cases chosen in the study having loss of immunohistochemical staining to N-terminal, Rod and C-terminal of dystrophin portion of muscle cell membrane, parallel resistance ofα-sarcoglycan, dysferlin, laminA/C, MHC-I monoclonal antibody immunohisto-chemical, and then compared, analyzed and summarized the results.Results:1. Initial complains are different, with bilateral lower limbs weakness accounting for majority (14 cases,77.8%), asymptomatic myocardial enzymes elevation in 3 cases (16.7%), transaminase elevation in 1 case (5.5%); different degrees of scapular winging, pseudo hypertrophy of calf muscles, Gower sign,11 cases of different degree muscle wasting (61.1%).11 cases of proximal muscle strength grade 3-4 (61.1%).7 cases proximal muscle normal (38.9%).11 cases knee flection is affected (61.1%) and 12 cases cannot walk on ankle (66.7%),3 cases cannot walk on toes (16.7%), All denied muscle pain and Babinski negative on examination. All patients were not mental retarded.14 patients showed myopathic changes on EMG, the two female carriers had no obvious positive signs.2. Serum creatine kinase level varied, during the course, all show ascending trend coming on earlier with progression, about coming on 2-3 years, range between (17577.78-16963.7U/L), as disease progress, pathological damages are progressive, serum CK whole show downtrend gradually; from onset ages view point, serum Ck reached peak in 2-5 years old, with range (10838.25-12937U/L). With age, the pathological damages are aggravating, muscle necrosis increased, serum CK decreased.3. serum myocardial enzyme level done in all of our cases, results showed varying degree of increase but CK elevation was particularly remarkable, the average increase (10331.16±8095.62) U/L, CK-MB(417.88±304.69) U/L, LDH (1023.19±393.18) U/L, HBD (910.23±358.19) U/L, AST (217.96±108.12) U/L. CK-MB/CK>6%,7 cases.4. Only in 6 cases muscle ultrasound were done, to have the varying degree subcutaneous fat thickening, bilateral symmetric muscle wasting, about 0.3-0.5mm or so, muscle bundle membrane echogenicity increased and its continuity interrupted or disappeared, its texture was also unclear, diffuse exquisite sample fat echo.5. Histochemical staining, H/E slides, atrophied muscle fiber diameter between 10-70μm sizes, round in shape in cross section,majority are opaque fibers, scattered muscle necrosis and regeneration, some slides showed increasing in membranous nuclei, internal nuclei,fibromuscular stroma widened, muscle connective tissue hyperplasia of different level.6. Immunohistochemical staining with dystrophin-N,-C,-R monoclonal antibody,muscle membrane dystrophin are totally or almost totally absent,14 casesα-SG staining showed different degree of deficiency,4 cases express normal, all are dysferlin and laminA/C normally expressed,6 cases MHC-Ⅰdown-regulated, 12 cases MHC-Ⅰnormally expressed.Conclusions:1. DMD patients serum CK values significantly increased, can reach dozens to hundreds times, this research from course and onset ages, two aspects of the changes in serum CK, analyze and obtain during the course of the 2-3 years, age 2-5 years old, serum CK peak, later decline as the disease progress and age year by year.2. CK-MB/CK>6% to determine early myocardial damaged clinically meaningful and can be used as early detection DMD myocardial injury indexes.3. Muscle ultrasonic examination are gradually becoming an effective means for new research of muscle diseases.4. Muscle fiber co-existence degenerative, hyperplasic, Opaque fiber lesions is DMD characteristic changes, but has no specificity. With progression, muscle fibers quantity gradually decrease, to the end almost disappear, replaced by fat and connective tissue, the more serious clinical symptoms, the more increase in fat & C.T.5. Absence of dystrophin, reduction in a-sarcoglycan, explains that dystrophin missing affect a-sarcoglycan expression, presumably, the sarcoglycans participated in the muscle degeneration necrosis process.6. Normal expression of dysferlin and laminA/C, presumably, indicates that their synthesis and assembly has no direct correlation with dystrophin, also further prove that dysferlin and DGC are not related. MHC-Ⅰup regulation indicates DMD skeletal muscle over-active, it is postulated that inflammatory reaction secondary to muscle cells degeneration and necrosis, exact mechanism is unknown, needs for further researches.更多还原