| The Background and Objective:Neutral lipid storage disease with myopathy（NLSDM）is a rare and recessive genetic disease which is characterized by neutral lipid deposits in different tissues and organs.NLSDM is caused by mutations in the gene encoding adipose triglyceride lipase（ATGL）,which is also known as PNPLA2.ATGL catalyzes the first step of triglyceride hydrolysis,whose mutation leads to a large amount of neutral fat deposition in different tissues and cells（such as skeletal muscle cells and granulocytes in blood and skin）.Skeletal muscle is the main target,while myocardium and multisystem could be involved sometimes.The skeletal muscle symptoms slowly progresses.Both upper and lower limbs,proximal and distal muscles can.be affected.Neurological examinations shows severe asymmetric muscle weakness and atrophy.Some patients complain of fatigue and muscle spasm.Unfortunately,to date,there is few effective drugs to impro-ve the muscle symptoms.In this research,the clinical,pathological and genetic features of 7 NLSDM cases which were diagnosed at Qilu Hospital during 2006.1～2016.11 will be retrospectively reviewed.The patients will be followed-up and course will be recorded and analysed.The main objective of this study is to summarize and discuss the clinical,pathological and genetic features of NLSDM patients and provide a reference for the diagnosis and treatment of NLSDM in the future.Materials and methods:All the 7 cases were diagnosed with NLSDM by the Research Institute of Neuromascular andNeurodegenerative Disease of Shandong University during 2006.1-2016.11.All the 7 patients underwent serologic test,electromyography（EMG）,muscle biopsy and PNPLA2 geen screening.The H-E/MGT/NADH-TR/SDH/COX/PAS/ORO/ATPase staining and immunohistochemical staining were performed on transverse muscle sections.The medical records of the patients were obtained from the record room of Qilu Hospital.The course of these patients and their outcomes were achieved through 1～10.5 years follow-up by telephone.We summarize and discuss the clinical features,electrophysiological characteristics,laboratory findings,courses and outcomes of the patients.Outcomes:Of the 7 patients,2 were female and 5 male.The onset age of the patients was between 22～45,with an average age of 36.7 years old.Patients aged of 28～49 years old when diagnosed,and the average age was 40.6 years old.The interval between onset and diagnosis was 1～7 years,of which 2 were about 1 years,3 were more than 6 years,the longest was 7 years,and the average was 4 years.The symptom of all the7 patients showed adult and inconspicuous onset.It progressed slowly,which was characterized by unilateral or bilateral asymmetric limb weakness and muscle atrophy,both proximal and distal limbs affected.The major clinical manifestations of the 7 patients at initial visit were fatigue,upper limb weakness presenting as difficult to lift handle and handle objects,lower limb weakness presenting as unable to walk,abnormal walking posture,easy to fall,difficult to squat down,stand up and walk upstairs,etc.Of the 7 patients,5 cases（5/7）exhibited bilateral limb weakness,while 2 cases（2/7）showed unilateral limb weakness.The upper and lower limbs were both involved in 4 cases（4/7）.2 patients complained only the upper limb fatigue（2/7）.1 patient showed with only the lower limb involvement（1/7）.Distal muscles were worse than proximal muscles in 4 cases（4/7）,milder in 3 cases（3/7）.In all 7 cases,tendon reflexes were weak or disappeared（7/7）.Interestingly,multisystematic symptoms were not observed in all these patients.None of them suffered cardiac disease,diabetes mellitus,hearing loss or hypertriglyceridemia.Jordans bodies were observed in white blood cells in 7 patients（7/7）.The serum creatine kinase levels was 651-2419U/L,the average value was 1527.9 U/L.They all did electromyography examinaiton,the results were as follow:2 cases showed normal EMG;3 patients showed myogenic EMG damage;1 patients showed neurogenic EMG damage,and tonic potential visible in upper and lower linbs;1 patient showed myogenic EMG damage（mainly in the upper limbs）,and neurogenic EMG damage（mainly in the lower limbs）.Muscle biopsy was performed in 7 patients.The results of frozen section staining showed myogenic pathological damage changes.In the H-E straining,the size of the muscle fibers of all the 7 patients were obviously different,and the shape of the small fibers were round,polygon,long strip and angle.Many small and large vacuoles were found in both two muscle fibers in all patients.5 patients showed slight to moderate increase of the centrally placed nuclei（10%～50%）,of which 3 patients were 50%.2 patients showed hypertrophic and splitting muscle fibers.3 patients showed Type 1 fiber atrophy;5 patients showed type 1 fiber advantage;2 patient showed type 1 fiber predominance and atrophy;1 patient showed type 2 fiber predominance.5 patients showed muscle fiber necrosis.4 patients showed muscle regeneration.2 patients showed pyknotic nuclear clump.4 patients showed infiltration of inflammatory cells.1 patient showed perivascular inflammatory cells infiltration.3 patients showed significant endomysium hyperplasia,1 patient showed mild hyperplasia.In the MGT staining,5 patients showed rimmed vacuoles.1 patient showeddecreased intramuscular nerve myelinated fibers.In the SDH staining,3 patients showed general decrease of enzyme activity.In the ORO staining,all 7 patients showed numerous lipid droplets in both types of fibers but more predominant in type 1 fibers.The lipid droplets under the membrane were large round or oval,while the centrel muscle fibers were small.With regard to immunohistochemical staining,MHC-I was weakly expressed in the sarcolemma of non-necrotic muscle fibers in 1 patient.Moreover,CD3 positive T cells infiltrated in perivascular area in 1 patient and CD68 positive macrophage cells were observed in or around necrotic fibers in 5 patients.7 patients all underwent PNPLA2 gene screening.The mutations of 6 patients were homozygous,which were IVS6 + 1G>T cut mutation,c.467del C base deletion mutation,c.475478dupCTCC,p.（Gln160Profs*19）frameshift mutation,c.6dupT,p.（Pro3fs）frameshift mutation,c.434G>A,p.（Serl45Asn）missense mutation.The mutation of 1 patient was heterozygous,which contained IVS6 + 2T>C splicing mutation and c.749A>C（Q250P）missense mutation.Among them.c.434G>A,P.（Ser145Asn）missense mutation and c.6dupT,P.（Pro3fs）frameshift mutation have not been reported.We analyzed them with bioinformatics software,and speculated that they were both pathogenic.The 7 patients were treated with prednisone,vitamin B complex,vitamin B2,vitamin B3,vitamin B6,vitamin B12,vitamin E,coenzyme Q10,L-carnitine,Bozhi Glycopeptide Injection,but we did not find any improvement in muscle weakness and muscle atrophy.We did a recent follow-up in 2017.3,and we found that all the 7 patients weren’t taking any medicine.The initial follow-up was short to 1 year,up to 10.5 years,with an average of 5 years,of which 4 was longer than 5 years.During the follow-up study.the manifestations of all the patients progressed slowly and asymmetrically.2 patients developed bilateral deafness and 2 patients had tinnitus,while cardiomyopathy,diatete mellitus or hypertriglyceridemia were not found.Conclusions:1.All the cases were adult,with insidious onset and slow disease progression.Asymmetric muscle weakness and muscle atrophy were observed.Both distal and proximal muscle groups were involved.Distal involvement was seen in more than half of these cases.Muscle weakness and muscle atrophy slowly progressed.2.Extramuscular systems were not obviously affected in all 7 cases.3.Jordans bodies were very useful for clinical rapid screening.The deposition of a large nrmber of lipid droplets were meaningful for diagnosis.Rimmed vacuoles,necrotic muscle fibers,regenerated muscle fibers and infiltration of inflammatory cells were seen in more than half of the cases.4.All the cases had mutations of the PNPLA2 gene,including 6 homozygous mutations and 1 compound heterozygous mutation.There were at least 6 cases having 1 severe mutation which leading to abnormal function of the protein.c.434G>A missense mutation and c.6dupT,P.（Pro3fs）frameshift mutation hadn’t been reported.5.Now specific therapy of NLSDM was deficient. |
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