伴有肌病的中性脂肪沉积病7例临床、病理与基因分析

发布时间：2018-05-08 13:11

本文选题：脂质贮积病 + 肌疾病　；参考：《山东大学》2017年硕士论文

【摘要】：研究背景与目的:伴有肌病的中性脂肪沉积病(neutral lipid storage disease with myopathy,NLSDM)是一种十分罕见的常染色体隐性遗传病,它是由编码脂肪甘油三酯酯酶(adipose triglyceride lipase,ATGL)的基因(又称PPNLA2基因)突变导致的。ATGL调控甘油三酯水解的第一步,它的突变会导致大量甘油三酯在不同的组织细胞中沉积,如骨骼肌细胞、心肌细胞、血液粒细胞等,造成的损害以骨骼肌损害为主,也可伴有心肌损害。NLSDM临床进展缓慢,主要表现为左右不对称的四肢近端或远端无力,目前无有效疗法。本研究拟对山东大学齐鲁医院神经肌肉病理实验室2006年1月～2016年11月间诊治的7例NLSDM患者进行随访,并对其临床、病理及基因突变特点进行回顾与分析。本研究的主要目的是总结NLSDM的临床、病理与基因突变特点,为NLSDM的诊断与治疗提供一定参考和帮助。对象和方法:研究对象为山东大学齐鲁医院神经肌肉病理实验室2006年1月～2016年11月诊治的7例NLSDM患者。7例患者均在齐鲁医院神经内科门诊进行详细的病史采集、内科查体和神经系统体格检查,并进行血清学、电生理学、影像学、肌肉组织活检和PNPLA2基因等辅助检查。患者的冰冻肌肉标本切片后均行组织化学染色(H-E、MGT、NADH-TR、SDH、COX、SDH/COX、PAS、ORO、ATPase)和免疫组织化学染色(Dystrophin-Rod,Dystrophin-C,Dystrophin-N,Dysferlin,α-sarcoglycan,β-sarcoglycan,γ-sarcoglycan,δ-sarcoglycan,Caveolin-3,MHC-1,CD3,CD4,CD8,CD20,CD68)。结果:7例患者中,女性2人,男性5人。患者起病年龄为22～45岁,平均年龄为36.7岁。患者就诊时年龄28～49岁,平均40.6岁。7例患者起病至诊断的时间间隔为1～7年,其中2例约为1年,3例超过6年,最长达7年,平均4年。7例患者均成年起病,起病隐匿,病情缓慢进展,为单侧或双侧左右不对称的肢体无力和肌肉萎缩,四肢远近端均可受累。7例患者就诊时主要临床表现为上肢抬举无力、拿捏物体困难、下蹲起立困难、行走姿势异常、上楼困难、行走无力、易绊倒、全身疲劳等。其中,5例患者就诊时肌无力现象累及双侧肢体(5/7),2例患者只累及一侧肢体(2/7)。4例患者就诊时同时累及上下肢(4/7),2例患者只累及上肢(2/7),1例患者只累及下肢(1/7)。3例患者就诊时肌无力症状近端明显(3/7),4例患者远端明显(4/7)。7例患者均出现腱反射出现减弱或消失(7/7)。除骨骼肌受累之外,7例患者就诊时其他系统受累情况不明显,7例患者均无心脏受累、高TG血症、肝肿大及听力下降。7例患者外周血ORO涂片均可见有Jordans小体。患者就诊时CK值在651～2419U/L,平均1527.9U/L。肌电图检查示:2例患者为正常肌电图;3例患者为肌源性损害肌电图;1例患者为神经源性损害肌电图,上下肢均可见强直电位;1例患者既有肌源性损害(以上肢为主),也有神经源性损害(以下肢为主)。7例患者均行肌肉活检,6例患者取肱二头肌,1例患者取腓肠肌。肌肉活检后行冰冻切片染色,结果均显示为肌源性损害病理改变。在H-E染色上7例患者均可见肌纤维大小明显不等,小纤维可为小圆形、小多边形、小长条型和小角形;7例患者肌纤维内均存在大量细小空泡或粗大空泡,两型纤维均可存在;5例患者可见内核纤维轻至中度增加,所占比例为10%～50%,达到50%者有3例;2例患者可见肥大及劈裂肌纤维;3例患者可见选择性Ⅰ型纤维萎缩;5例患者可见Ⅰ型纤维优势;其中2例患者可见选择性Ⅰ型纤维优势伴萎缩;1例患者可见Ⅱ型纤维优势;5例患者可见坏死伴吞噬肌纤维;4例患者可见再生肌纤维;2例患者可见少量核袋;4例患者可见散在炎细胞浸润;1例患者可见血管周围有炎细胞浸润;3例患者可见肌内膜明显增生,1例轻度增生。在MGT染色上,5例患者可见镶边空泡;1例患者肌内神经内有髓纤维减少。在SDH染色上,有3例患者酶活性普遍减低。在ORO染色上,7例患者肌纤维内脂滴含量均显著增多,肌膜下可见大的圆形或椭圆形脂滴,肌纤维内脂滴较为细小。在免疫组织化学染色上,7例患者中有1例MHC-1呈弱阳性表达,1例血管周围炎细胞CD3呈阳性表达,5例坏死伴吞噬肌纤维CD68呈阳性表达。7例患者均行PNPLA2基因检测。其中6例患者为PNPLA2基因纯合子突变,分别为IVS6+1GT 剪切突变、c.467del C 碱基缺失突变(2 例)、c.475_478dupCTCC,p.(Gln160Profs*19)移码突变、c.6dupT,p.(Pro3fs)移码突变、c.434GA,p.(Ser145Asn)错义突变;1例患者为IVS6+2TC剪切突变和c.749AC(Q250P)错义突变的复合杂合突变。其中c.434GA,p.(Ser145Asn)错义突变和c.6dupT,p.(Pro3fs)移码突变未见文献报道,通过生物信息学软件分析,推测均有致病性。本组7例患者中有6例患者至少携带1个无正常功能蛋白表达的严重突变。本研究中7例NLSDM患者服用强的松、复合维生素B、维生素B1、维生素B2、维生素B3、维生素B6、维生素B12、维生素E、辅酶Q10、左旋肉碱、薄芝糖肽等进行治疗,肌无力和肌萎缩症状均未得到缓解和改善。7例患者目前均已自行停药。截至最后一次随访(2017年3月),7例患者的肌无力和肌萎缩症状均在缓慢进展,左右侧不对称。随访时有2例出现双侧听力下降,2例出现耳鸣,7例均无明显心脏受累、糖尿病及高TG血症。结论:1.本组病例均成年起病,起病隐匿,临床主要表现为左右不对称的进行性四肢肌无力和肌萎缩,肢体远近端均可受累,半数以上远端受累为主。肌无力和肌萎缩症状缓慢进展。2.本组病例除骨骼肌受累之外,其他系统受累不明显。3.Jordans小体可为临床快速筛查NLSDM提供依据,结合肌活检中两型肌纤维内大量脂滴沉积,可病理诊断为NLSDM。本组病例中半数以上会伴有镶边空泡、坏死伴吞噬及再生肌纤维和炎性细胞浸润。4.本组病例均存在PNPLA2基因突变,包括6例纯合突变,1例复合杂合突变。有6例患者至少携带1个无正常功能蛋白表达的严重突变。c.434GA错义突变和c.6dupT移码突变国内外迄今未有报道。5.本病尚缺乏特异性治疗。
[Abstract]:Background and objective: neutral lipid storage disease with myopathy (NLSDM) is a very rare autosomal recessive hereditary disease, which is controlled by the mutation of the gene encoding the triglyceride esterase (adipose triglyceride lipase, ATGL) of the fat (adipose triglyceride lipase, ATGL), also known as the PPNLA2 gene). The first step of ester hydrolysis, its mutation can cause a large number of triglycerides to be deposited in different tissue cells, such as skeletal muscle cells, cardiac myocytes, and hematogenous cells, causing damage mainly in skeletal muscle damage, but also associated with myocardial damage.NLSDM clinical progress slowly, mainly manifested by the near or distal weakness of the left and right extremities. 7 patients with NLSDM from January 2006 to November 2016 in the neuromuscular pathology laboratory of Qilu Hospital of Shandong University were followed up and the clinical, pathological and gene mutation characteristics were reviewed and analyzed. The main purpose of this study was to summarize the clinical, pathological and gene mutation characteristics of NLSDM, NLSD The diagnosis and treatment of M provided some reference and help. Object and methods: 7 patients with NLSDM from January 2006 to November 2016, the neuromuscular pathology laboratory of Qilu Hospital of Shandong University, were collected in the Department of Neurology Department of the Department of Neurology for detailed medical history collection, medical examination and nervous system physical examination. Serology, electrophysiology, imaging, muscle tissue biopsy, and PNPLA2 genes were examined. The frozen muscle specimens of the patients were stained with histochemical staining (H-E, MGT, NADH-TR, SDH, COX, SDH/COX, PAS, ORO, ATPase) and immunohistochemical staining (Dystrophin-Rod, Dystrophin-C, alpha, alpha, beta, etc.) Lycan, gamma -sarcoglycan, Delta -sarcoglycan, Caveolin-3, MHC-1, CD3, CD4, CD8, CD20, CD68). Results: of 7 patients, 2 women and 5 men. The onset age was 22~45 years and the average age was 36.7 years. The age of the patients was 28~49 years old, and the average age of 40.6 years of.7 was 1~7 years, 2 cases were 1 years, 3 cases were about 1 years. More than 6 years, the maximum of 7 years, the average 4 years of.7 patients were all onset, the onset of disease, slow progress, unilateral or bilateral asymmetry of limb weakness and muscle atrophy, and the extremities of the extremities can be affected by.7 patients in the main clinical manifestations of the upper limb lifting, difficulty in holding objects, squatting difficulties, walking posture Often, it was difficult to go upstairs, unable to walk, easy to stumble, and fatigue. Among them, 5 patients were involved in bilateral limbs (5/7), 2 cases involved only one side limb (2/7).4 patients involved upper and lower limbs (4/7) while 2 cases were only involved in the upper limb (2/7), and 1 patients were only involved in the.3 cases of lower limbs (1/7) patients with myasthenia. The proximal end of the form was obvious (3/7). 4 patients with far distal (4/7).7 cases showed a weakening or disappearance of the tendon reflex (7/7). In addition to the skeletal muscle involvement, 7 patients were not involved in other systemic involvement, 7 cases had no heart involvement, high TG, hepatomegaly and hearing loss in the peripheral blood ORO smears in the peripheral blood were all Jordans small. The CK value of the patients was from 651 to 2419U/L, and the average 1527.9U/L. electromyography examination showed that 2 patients were normal EMG, 3 patients were myogenic electromyography, 1 patients were neurogenic damage electromyography, and the upper and lower extremities were both tetanic potential; 1 patients had both muscular damage (mainly upper limbs) and neurogenic damage (lower limbs with lower extremity). .7 patients underwent muscle biopsy in all cases. 6 patients took the biceps brachii muscle and 1 patients took the gastrocnemius muscle. After the muscle biopsy, the frozen section staining was performed. All the results showed the pathological changes of myogenic damage. In 7 cases of H-E staining, the size of the muscle fibers was obviously different, and the small fibers could be small round, small polygon, small strip type and small angle shape; 7 cases were found. There were a large number of fine vacuoles or large vacuoles in the muscle fibers of the patients, and two types of fibers could exist in the 5 patients. The proportion of the core fiber was 10% to 50%, 50% had 3 cases, 2 patients showed the hypertrophy and cleavage muscle fibers, and 3 patients showed the type I fiber atrophy; 5 patients showed type I fiber superiority. Of the 2 patients, selective type I fiber dominance and atrophy were seen; 1 patients showed type II fiber dominance; 5 patients showed necrosis with phagocytic fibers; 4 patients showed regenerative muscle fibers; 2 cases showed a small number of nuclear bags; 4 cases showed infiltration of inflammatory cells; 1 patients showed infiltration of inflammatory cells around blood vessels; 3 patients were visible. The intimal hyperplasia and 1 mild hyperplasia were seen in 1 cases. In 5 patients, the side vacuoles were seen in 5 patients and 1 cases of intramuscular myelinated fibers decreased. In the SDH staining, the enzyme activity was reduced in 3 patients. In the ORO staining, the content of lipid droplets in the muscle fibers increased significantly in 7 cases, and the large round or oval lipid droplets and muscle fibers under the musculi membrane were observed. In immunohistochemical staining, 1 of the 7 cases of MHC-1 showed weak positive expression, 1 cases of peripheral inflammatory cells CD3 positive expression, 5 cases of necrotic and phagocytic CD68 positive expression in.7 patients with PNPLA2 gene detection. 6 cases were PNPLA2 homozygous mutation, respectively IVS6+1GT shear. Mutation, c.467del C base deletion mutation (2 cases), c.475_478dupCTCC, P. (Gln160Profs*19) code mutation, c.6dupT, P. (Pro3fs) shift mutation, c.434GA, P. (Ser145Asn) missense mutation; 1 cases were compound heterozygous mutation of IVS6+2TC shear mutation and missense mutation. S) the change of code mutation was not reported. In the 7 patients of this group, 6 patients had at least 1 serious mutations without normal functional protein expression. In this study, 7 patients with NLSDM took prednisone, vitamin B, vitamine B1, vitamin B2, vitamin B3, vitamin B6, vitamin B1. 2, vitamin E, coenzyme Q10, L-carnitine, thin ganoderma and so on, myasthenia and atrophy of myasthenia and amyotrophic symptoms have not been alleviated and improved in.7 patients. As of the last follow-up (March 2017), the symptoms of myasthenia and atrophy of myasthenia and myasthenia in 7 patients were slowly progressing and asymmetrical in the left and right side. 2 cases appeared bilateral at follow-up. Hearing loss, tinnitus in 2 cases, and no obvious heart involvement, diabetes and hyperTG in 7 cases. Conclusion: 1. of the cases were all onset and occult. The main clinical manifestations were progressive myasthenia and atrophy of extremities and extremities. More than half of the extremities were involved. Myasthenia and atrophy of myasthenia and myasthenia were slow. Slow progress.2., in addition to skeletal muscle involvement, other system involvement, not obvious.3.Jordans body can provide a basis for rapid clinical screening of NLSDM, combined with a large number of lipid droplets in two types of muscle fibers in muscle biopsy, and the pathological diagnosis is that more than half of the cases in the NLSDM. group are accompanied by border vacuoles, necrosis, phagocytosis, and regenerative muscle fibers. PNPLA2 gene mutations were present in all cases of inflammatory cell infiltration in.4., including 6 cases of homozygous mutation and 1 complex heterozygous mutations. 6 patients carried at least 1 severe mutations without normal functional protein expression,.C.434GA missense mutation and c.6dupT code shift mutation at home and abroad.

【学位授予单位】：山东大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R596;R746

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