脊髓性肌萎縮癥患兒的臨床特征分析

劉維亮，李 芳，何志旭，艾 戎，麻宏偉(貴陽醫學院附屬醫院兒科，貴陽 550004;中國醫科大學附屬盛京醫院發育兒科;貴陽醫學院附屬醫院眼科;通訊作者，E-mail:mahongwei960@6.com)

Spinal muscular atrophy(SMA)is a common autosomal recessive disease affecting 1/6 000 to 1/10 000 births.The carrier frequency is 1/35［1，2］.The clinical characteristics of disease are basically caused by the progressive loss of alpha motor neurons in spinal cord anterior horn cells，which leads to lower motor neuron degeneration and weakness and wasting of voluntary muscle.SMA is clinically subdivided into three types based on age of onset and clinical severity.In 1995，the SMN1 was identified as the SMA disease-determining gene［3］.In SMA，96%of patients showed homozygous deletion of SMN1 exon 7 and/or exon 8［3］.The finding of homozygous deletions of SMN1 exons 7 and/or 8 of patients with consistent clinical characteristics is generally diagnosed as SMA［4］.SMA patients have many typical and atypical characteristics，therefore，this study analyzed the clinical data of patients who had been diagnosed by SMN gene deletion，and expected to more sufficiently understand the disease.

1 Materials and methods

1.1 Subjects

From 1996 to 2011，we selected 66 Chinese SMA patients(34 boys and 32 girls，including 30 of typeⅠ，24 of typeⅡ，12 of type Ⅲ，their onset ages were from 4 days to 6.2 years)who fulfilled the diagnostic criteria of the homozygous deletion of SMN1 exons 7 and/or 8 by PCR-restriction fragment length polymorphism(RFLP).We analyzed the clinical characteristics and examinations including creatases，electromyography(EMG)，and muscle biopsy.The study was performed according to the Declaration of Helsinki and approved by the Ethics Committees of Affiliated Shengjing Hospital of China Medical University and Affiliated Hospital of Guiyang Medical College.Informed consent was obtained from the patients’parents.

1.2 Diagnostic criteria

①All of the SMA patients fulilled the genetic diagnostic criteria by genetic analysis showing homozygous deletion of the SMN1 gene［3，4］;②All of the SMA cases fulilled the clinical diagnostic criteria defined by the International SMA Workshop［5］.

1.3 Statistical analysis

Data were expressed as mean±SD.Variance analysis，Chi-square test and Fisher’s exact test were used for the data analysis，and the statistical significance was set as P≤0.05.

2 Results

2.1 Laboratory results

2.1.1 Creatases Values of each type creatases and positive rates of above normal values were shown in Table 1，Table 2，respectively.The difference of each type creatase values between groups was not significant by analysis of variance(P ＞0.05).The values of creatases including creatine kinase(CK)，MB isoenzyme of creatine kinase(CKMB)，aspartate aminotransferase(AST)，hydroxybutyrate dehydrogenase(HBD)，lactate dehydrogenase(LDH)in each type were normal or slightly higher than the normal.Abnormal rate of CKMB and HBD was higher than those of CK，AST and LDH in every type by Chi-square test(P ＜0.001).

2.1.2 EMG The results showed neurogenic damage in 58 of 66 patients(87.9%，26 of typeⅠ，21 of typeⅡ，11 of typeⅢ)by EMG.Of 58 patients with neurogenic damage，3(2 of typeⅠand 1 of type Ⅲ)were accompanied by features of peripheral(motor and/or sensory)neuropathy.EMG results showed that the evoked potential of motor nerve conduction of double peroneal nerve and double tibial nerve was not elicited in 1 of typeⅠpatient;the evoked potentials of sensory nerve conduction of left sural nerve and motor nerve conduction of double peroneal nerve and right tibial nerve were not elicited in another case of typeⅠ;andthe stepped down motor nerve conduction velocity of right peroneal nerve and motor nerve conduction latency of right femoral nerve were extended in 1 of typeⅢ.Another 2 patients had peripheral neuropathy but no neurogenic damage，and EMG results of the 2 patients showed that the evoked potential of sensory nerve conduction of double sural nerve was not elicited in 1 of type Ⅰ patient，and that myogenic damage with stepped down sensory nerve conduction velocity was in 1 of typeⅢ.

Table 1Values of creatases in spinal muscular atrophy patients with different types(±s，U/L)

Table 1Values of creatases in spinal muscular atrophy patients with different types(±s，U/L)

P ＞0.05 for the comparison between typeⅠ，Ⅱ，Ⅲ by Variance analysis;the normal values are showed as follows:CK 29-200 U/L，CKMB 0-24 U/L，AST 5-34 U/L，HBD 72-182 U/L，LDH 125-243 U/L

Type CK CKMB AST HBD LDHⅠ163.0 ±138.7 41.5 ±22.6 62.4 ±102.2 252.6 ±89.77.7 ±72.5 1 325.6 ±176.5Ⅱ135.8 ±77.5 41.5 ±30.8 32.7 ±7.9 247.8 ±98.4 272.2 ±113.7Ⅲ188.4 ±87.1 35.1 ±11.4 30.5 ±5.9 276.1 ±136.0 2

Table 2 Positive rates of abnormal(higher tham normal)creatases in spinal muscular atrophy patients with different types cases(%)

2.1.3 Muscle biopsy Muscle biopsy was performed on 11 patients，of whom 10 showed neurogenic damage.Another one was not ascertained damage condition.The difference of neurogenic damage positive rate in the result of comparing muscle biopsy with EMG was not significant(Fisher’s exact test P=1.00 ＞0.05).

2.2 Clinical characteristics

2.2.1 Typical clinical characteristics Typical clinical characteristics originated in classic SMA as a purely motor disorder.Type I patients had the inability to sit and abolition of tendon reflex before the age of 6 months.Patients of SMA type Ⅱ usually presented onset in 6-8 months，they were able to sit but not able to walk unaidedly，had usual limb weakness(the lower limbs were more severer than the upper ones)，and had weakness or abolition of tendon reflex.Onset of SMA type Ⅲ was usually after 18 months，and there were some characteristics including the ability to walk but amyasthenia，weakness or abolition of tendon reflex.Intelligence of patients with different types was normal.Creatases were normal or mildly higher.EMG or muscle biopsy finding was usual neurogenic damage in patients of each type.

2.2.2 Atypical clinical characteristics We found some atypical clinical characteristics as concomitant diseases of SMA patients by analyzing the clinical data.Five patients(7.6%，3 of typeⅠ，2 of type Ⅲ)accompanied with periphery(motor and/or sensory)neuropathy manifestation，and thedifferencewasnot significant between typeⅠ and Ⅲ［(Fisher’s exact test P=0.613 ＞ 0.05)］.Of the 5 patients，2(1 of typeⅠ，1 of type Ⅲ)had motor neuropathy，1(1 of type Ⅲ)had sensory neuropathy，and 2(2 of typeⅠ)had mixed neuropathy by EMG.Four patients(6.1%)accompanied with acleistocardia(3 of typeⅠand 1of type Ⅲ);4 patients(6.1%)had skeletal deformity，1 of typeⅠhad polydactylism(hexadactyly of right foot)，1 of type Ⅱ had pes valgus，1 of type Ⅲ had pes varus and dislocation of the hip，and 1 of type Ⅲ had knee contracture.Two patients(3.0%)had central nervous system(CNS)concomitant diseases，including subarachnoid hemorrhage and brain edema in 1 of typeⅠand delayed myelinization in cerebrum in 1 of typeⅢ.One(1.5%)of type Ⅲ had lingual muscle hypertrophy.

3 Discussion

In the study，we analyzed clinical data of SMA patients who fulfilled genetic diagnosis to summarize full clinical symptoms，estimate routine examinations，and further guide the understanding and research about the disease.Preliminary diagnosis of SMA may be per-formed by typical characteristics.The elevated CK，CKMB，AST，HBD，LDH in some patients might reflect minor damage of skeletal muscle and myocardial，and abnormal creatase values might be associated with progression of SMA.Both of EMG and muscle biopsy can be easier to detect neurogenic damage in patients.

Reports of concomitant diseases of SMA were very rare.We found some concomitant diseases mainly in typeⅠ andⅢ SMA.Peripheral neuropathy manifestation in typeⅠ andⅢ was the most one of them，and 7.6%of SMA patients was found with peripheral neuropathy manifestation.Although classic SMA is believed to be a purely motor disorder，affecting neurons of the spinal anterior horn and nuclei of the lower cranial nerves.Other organ malformations or peripheral nerve involvement have been regarded as exclusion criteria for SMA［6］.But in patients with severe forms of SMA，additional sensory abnormalities have been reported.In some patients of the confirmed diagnosis by the presence of a homozygous deletion of the SMN1 gene，electrophysiological test showed inexcitability of motor and sensory nerves，the presence of a severe sensorymotor axonal neuropathy and myographic signs of denervation［7，8］.Nerve biopsies and postmortem examination showed loss of myelinated fibers and axonal damage in sensory and mixed nerves［7］.Particularly，71.4%of type Ⅰ SMA showed abnormal sensory conduction，and there was significant sensory nerve pathology in severely affected patients with typeⅠSMA by biopsy，whereas there had been no found sensory nerve alterations clinically or morphologically in patients with milder type Ⅱ or Ⅲ SMA in the past［6］.And animal experiments indicated that pathological alterations in severe forms of SMA were not restricted to motor neurons，but the defects in the sensory neurons were milder than those in the motor neurons［9］.

Here we found the motor and/or sensory neuropathy by EMG not only in typeⅠ but also in typeⅢ.Motor and/or sensory neuropathy might be part of the clinical spectrum of minority of SMA.So molecular genetic test of SMA including those SMA patients with additional periphery neuropathy should be enforced as a further definitive diagnosis particularly.To the congenital cardiopathy，the existed findings showed that typeⅠSMA patient with congenital heart defects was a feature of severe infantile spinal muscular atrophy，and SMN protein was relevant to normal cardiogenesis［10］.SMA accompanied with congenital cardiopathy was found in wider typeⅠ and Ⅲ SMA in the study，which may be a new mechanism to explain the phenomenon.And we should continue to expand the investigation because heart ultrasonography was not performed in every patient.Mechanism is not clear about other concomitant diseases including skeletal deformities，CNS and lingual muscle changes.

Molecular genetic basis of SMA is deletion or subtle mutation of SMN gene，and may also be a wide range of deletion of genes including SMN gene.The SMA region on 5q13 shows a complex genomic structure，containing many repeated sequences，pseudogenes，and retrotransposition elements.The region is instable，and there may be homologous recombination，deletion，duplication，gene substitution，et al［11］.Ubiquitously expressed SMN protein is involved in the biogenesis of uridine-rich small nuclear ribonucleoproteins(UsnRNPs)and facilitates cytoplasmic assembly of UsnRNPs into the spliceosome，a large snRNA-protein complex that catalyzes pre-mRNA splicing［12，13］.About molecular pathogenesis of concomitant diseases including peripheral neuropathy，congenital cardiopathy，skeletal deformities，CNS and lingual muscle changes，we conclude that obvious SMN deficiency in typeⅠmight lead to decrease of functional SMN protein expression，abnormal SMN protein might affect downstream related gene regulation，and abnormal activity of chromosomal region including SMN gene in typeⅢmight cause relative gene abnormality or susceptible changes.

In summary，typical characteristics may be used as a preliminary diagnostic criteria for SMA.Atypical characteristics including peripheral neuropathy，central nervous system diseases，and development malformations(skeleton，heart，lingual muscle，et al)in patients with typeⅠandⅢSMA should be paid high attention.We think concomitant diseases of SMA are not coincidental，because each type SMA might have associa-tion with different concomitant diseases，the causes of concomitant diseases might be abnormal unique genetic function of SMN protein that may be associated with development of the multiple systems leading to changes of additional or unknown contiguous genes to SMN，abnormal activity of chromosomal region of including SMN gene urging abnormality of other genes.The results of this study may be useful in research of future precise molecular pathogenesis of SMA.

［1］ Feldk?tter M，Schwarzer V，Wirth R，et al.Quantitative analyses of SMN1 and SMN2 based on real-time lightCycler PCR:fast and highly reliable carrier testing and prediction of severity of spinal muscular atrophy［J］.Am J Hum Genet，2002，70:358-368.

［2］ Cusin V，Clermont O，Gérard B，et al.Prevalence of SMN1 deletion and duplication in carrier and normal populations:implication for genetic counseling［J］.J Med Genet，2003，40:e39.

［3］ Lefebvre S，Bürglen L，Reboullet S，et al.Identification and characterization of a spinal muscular atrophy-determining gene［J］.Cell，1995，80:155-165.

［4］ Prior TW.Spinal muscular atrophy diagnostics［J］.J Child Neurol，2007，22(8):952-956.

［5］ Zerrres K，Davies KE.59th ENMC International Workshop:Spinal Muscular Atrophies:recent progress and revised diagnostic criteria 17-19 April，1998，Soestduinen，The Netherlands［M］.Neuromusc Disord，1999，9:272-278.

［6］ Rudnik-Sch?neborn S，Goebel HH，Schlote W，et al.Classical infantile spinal muscular atrophy with SMN deficiency causes sensory neuronopathy［J］.Neurology，2003，60(6):983-987.

［7］ Korinthenberg R，Sauer M，Ketelsen UP，et al.Congenital axonal neuropathy caused by deletions in the spinal muscular atrophy region［J］.Ann Neurol，1997，42(3):364-368.

［8］ Fernández-Torre JL，Teja JL，Castellanos A，et al.Spinal muscular atrophy type I mimicking critical illness neuropathy in a paediatric intensive care neonate:electrophysiological features［J］.Brain Dev，2008，30(9):599-602.

［9］ Jablonka S，Karle K，Sandner B，et al.Distinct and overlapping alterations in motor and sensory neurons in a mouse model of spinal muscular atrophy［J］.Hum Mol Genet，2006，15(3):511-518.

［10］ Rudnik-Sch?neborn S，Heller R，Berg C，et al.Congenital heart disease is a feature of severe infantile spinal muscular atrophy［J］.J Med Genet，2008，45(10):635-638.

［11］ Chen Q，Baird SD，Mahadevan M，et al.Sequence of a 131kb region of 5q13.1 containing the spinal muscular atrophy candidate genes SMN and NAIP［J］.Genomics，1998，48:121-127.

［12］ Frugier T，Nicole S，Cifuentes-Diaz C，et al.The molecular bases of spinal muscular atrophy［J］.Curr Opin Genet Dev，2002，12:294-298.

［13］ Paushkin S，Gubitz AK，Massenet S，et al.The SMN complex，an assemblyosome of ribonucleoproteins［J］.Curr Opin Cell Biol，2002，14:305-312.