Velvet antler is the only renewable bone tissue of mammalian animals, which consists of a variety of growth factors, amino acids and polypeptides. But the mechanism of high-speed proliferation without carcinogenesis is still mystifying. The previous study of this work found that the velvet antler peptides (VAP) could not only inhibit the proliferation and migration of osteosarcoma cell lines MG-63 and U2OS, but also induced U2OS apoptosis and inhibited MG-63 epithelial-mesenchymal transition (EMT) through TGF-β and Notch pathways. These results lead us to conclude that VAP has the potential ability to mediate osteosarcoma cells by regulating related signaling pathways and growth factors. Therefore, finding a new appropriate inhibitor for OS is a valuable research direction, which will give patients a better chance to receive proper therapy. From an applied perspective, this review summarized the effects of velvet antler, genes, growth factors and research progress of relative pathways and genes of osteosarcoma, which are poised to help link regenerative molecular biology and regenerative medicine in osteosarcoma pathogenesis.

Abusive head trauma (AHT) is the leading cause of death from trauma in infants and young children. An AHT animal model was developed on 12-day-old mice subjected to 90° head extension-flexion sagittal shaking repeated 30, 60, 80, and 100 times. The mortality and time until return of consciousness were dependent on the number of repeats and severity of the injury. Under 60 times of repeated head shakings, the pups demonstrated apnea and/or bradycardia immediately after injury. Acute oxygen desaturation was observed by pulse oximetry during respiratory and cardiac suppression. The cerebral blood perfusion was assessed by laser speckle contrast analysis (LASCA) using the PeriCam PSI system. There was a severe reduction in cerebral blood perfusion immediately after the trauma that did not significantly improve within 24 hours. The injured mice began to experience reversible sensorimotor function at 9 days post-injury (dpi) which completely recovered at 28 dpi. However, cognitive deficits and anxiety-like behavior remained. Subdural/subarachnoid hemorrhage, damage to the brain-blood barrier, and parenchymal edema were found in all pups subjected to 60 insults. Pro-inflammatory response and reactive gliosis were up-regulated 3 dpi. Degenerated neurons were found in the cerebral cortex and olfactory tubercles at 30 dpi. This mouse model of repetitive brain injury by rotational head acceleration-deceleration partially mimics the major pathophysiological and behavioral events that occur in children with AHT. The resultant hypoxia/ischemia suggests a potential mechanism underlying the secondary rotational acceleration-deceleration induced brain injury in developing mice.

Objective To analyze plasma vitamin E and CoQ10 levels in patients with autosomal recessive cerebellar ataxia for finding the evidence of the related pathogenesis research and therapeutic strategies.Methods The plasma vitamin E and CoQ10 levels were detected by high performance liquid chromatography (HPLC) with diode array detector in 123 probands of autosomal recessive cerebellar ataxia pedigrees.Quantitation was performed using vitamin E and CoQ10 external standard and two 5-point calibration curve;clinical manifestations were analyzed simuhaneously.Results Vitamin E and CoQ10 levels of healthy subjects in the plasma were (8.77 ± 2.28) μg/ml and (1.31 ± 0.38) μg/ml,respectively;the plasma vitamin E and CoQ10 levels of patients were (5.61 ± 2.04) μg/ml and (0.79 ± 0.26) μg/ml,respectively,which were significantly lower than those in healthy controls (t =11.87,13.15;all P＜ 0.01).Clinical manifestations were characterized by cerebellar symptoms,and gait instability was usually the first recognized abnormality.Most of early onset occurred before the age of 25 years (111/123);dysarthria and abnormal eye movement were observed,with cerebellar atrophy on MRI;concomitant symptoms were also present.Conclusions HPLC analysis shows that the plasma vitamin E and CoQ10 levels of patients with autosomal recessive cerebellar ataxia are generally lower than those in the healthy controls.Several patients with significant reductions in these two levels have genetic defects.The combination of clinical phenotypes,biochemical indexes and genetic analyses will be helpful for the establishment of diagnosis and specific treatment.

Our preliminary proteomics analysis suggested that expression of microtubule-associated protein tau is elevated in the spinal cord after injury. Therefore, the first aim of the present study was to examine tau expression in the injured spinal cord. The second aim was to determine whether tau can regulate neural stem cell migration, a critical factor in the successful treatment of spinal cord injury. We established rat models of spinal cord injury and injected them with mouse hippocampal neural stem cells through the tail vein. We used immunohistochemistry to show that the expression of tau protein and the number of migrated neural stem cells were markedly increased in the injured spinal cord. Furthermore, using a Transwell assay, we showed that neural stem cell migration was not affected by an elevated tau concentration in the outer chamber, but it was decreased by changes in intracellular tau phosphorylation state. These results demonstrate that neural stem cells have targeted migration capability at the site of injury, and that although tau is not a chemokine for targeted migration of neural stem cells, intracellular tau phosphorylation/dephosphorylation can inhibit cell migration.

Background: The molecular mechanism for osteolysis around prostheses remains unclear. Artificial joint loosening is typified by osteolytic resorption. Interleukin-1 affects bone resorption via the MAPK signaling pathway. The aim of this study was to observe the effects of siRNA-silenced expression of interleukin-1 receptor-associated kinase-4 (IRAK-4) gene on the mitogen-activated protein kinase (MAPK) signaling transduction pathway in human osteoblast-like cells MG63, and to provide experimental evidence for the prevention and treatment of osteolysis around prostheses after artificial joint replacement. Methods: IRAK-4-siRNA was transfected into MG63 cells by using Lipofectamine 2000 as the vector. The cells were divided into a blank group, a control group and a silencing group, which were not transfected, transfected with scrambled siRNA and with specific IRAK-4-siRNA respectively. The protein expressions of extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (c-JNK) and p38MAPK in target cells were detected by Western blot. Results: Compared with the control group, the IRAK-4 mRNA and protein expression levels of the silencing group were significantly lower (P< 0.05). After IRAK-4 expression was down-regulated, the expressions of c-JNK1/2 p46, ERK1/2 and p-p38 MAPK were also down-regulated by 62%, 64% and 68% respectively (P<0.05). Conclusion: Silencing IRAK-4 gene expression with siRNA inhibited the expressions of ERK, c-JNK and p38 MAPK in human osteoblast-like cells MG63.

Osteosarcoma (OS) is the most common primary malignant tumor of bone and the third most common cancer in childhood and adolescence. However, controversy concerning the ideal combination of chemotherapy agents ensued throughout the last quarter of the 20th century because of conflicting and often nonrandomized data. Collaborative efforts to increase understanding of the biology of osteosarcoma and the use of preclinical models to test novel protein targets will be critical to identify the path toward improving outcomes for patients. We attempted to identify potential protein markers or therapy targets of osteosarcoma and give a glance at tumorigenesis of osteosarcoma. A sensitive and accurate method was employed in comparative proteomic analysis between benign tumor and osteosarcoma. Tumor tissues obtained by open biopsy before induction chemotherapy were investigated With 2D DIGE and MALDI-TOF/TOF MS, 22 differentially expressed proteins were identified after database searching, including 8 up-regulated and 14 down-regulated proteins. We also validated the expression levels of interesting proteins(have higher Ratios(tumor/normal)) by Western blotting assay. Annotating by bioinformatic tools, we found structural and signal transduction associated proteins were in large percentage among altered level proteins. In particular, some low abundant proteins involving translation and transcription, such as EEF2(Elongation Factor 2), LUM Lumican 23 kDa Protein) and GTF2A2(Transcription Initiation Factor Iia Gamma Chain.), were firstly reported by our study comparing to previous observations. Our findings suggest that these differential proteins may be potential biomarkers for diagnosis or molecules for understanding of osteosarcoma tumorigenesis, coming with biologic, preclinical, and clinical trial efforts being described to improve outcomes for patients.

Objective To investigate the characteristics of mitofusin2(MFN2) mutation and the clinical variability of Charcot-Marie-Tooth disease type 2(CMT2) patients. Methods MFN2 mutation analysis were performed in 8 autosomal dominant CMT2 families and 24 sporadic CMT2 cases by PCR and direct sequencing. The clinical features of the positive cases were precisely analyzed. Results Sequencing of the MFN2 gene revealed 4 different missense mutations，and detection rate was 4 of 32 (12.5%) in patients with CMT2. c.281G＞A (R94Q) and c.2240 T＞C（M747T）were detected in 2 autosomal dominant pedigrees, and intrafamilial clinical phenotype variability was present；c.1090 C＞T （R364W）and c.2198 T＞C （L733P）were detected in 2 sporadic cases，and the latter was a novel mutation which had not been reported. In addition to weakness and atrophy in the distal limbs，the former sporadic case was optic atrophy，and the latter case was hyperkeratosis of the skin. Conclusions Divergent MFN2 mutations are frequently found among axonal varieties of CMT2 in the Chinese population，and analysis of phenotypes in the CMT2A2 patients indicate the high clinical variability of this disease. Key words: Charcot-Marie-Tooth disease; Mitochondrial proteins; Mutation; Phenotype

Objective To investigate the clinical features and genetic mutations of ataxia with isolated vitamin E deficiency (AVED). Methods The TTPA gene was analyzed by polymerase chain reaction combined with sanger sequencing in the probands of 160 autosomal recessive cerebellar ataxia families, which had been excluded from GAA triplet repeat expansions of Frataxin gene. Furthermore, serum vitamin E was detected by high performance liquid chromatography. The clinical features of the positive cases were precisely analyzed. Results Sequencing of the TTPA gene revealed 2 patients (12.5‰) carried mutations, one with homozygous nonsense mutation (c.400C>T, p.R134X) in exon 3, and the other with homozygous insertional mutations (c.83_84insGCGGCGCC) in exon 1. Serum vitamin E were 1.95 mg/L and 0.19 mg/L separately (normal (10.8±3.3) mg/L). In addition to the typical characteristics of AVED, the cases presented with scoliosis. Conclusions We reported two cases with homozygous mutations of TTPA gene in Chinese autosomal recessive ataxia families, and c.83_84insGCGGCGCC in exon 1 is a novel mutation. Severe decrease of serum vitamin E was detected. The two cases presented with scoliosis，indicating the clinical variability of the disease. Key words: Ataxia; Vitamin E deficiency; Mutation; Scoliosis

Objective To investigate the inheritance principle of the expanded GAG repeat allele and the clinical features of spinocerebellar ataxias 3 (SCA3) in a consanguinity family with first cousin marriage.Methods The CAG repeats of SCA3 gene were amplified by means of polymerase chain reaction.Fragment analysis with laser-induced fluorescence in capillary electrophoresis were performed for the positive samples detected by agarose gel electrophoresis.Furthermore,the clinical features were analyzed carefully.Results Fragment analysis revealed that the proband carried 2 alleles with 56 and 72 CAG repeats separately.The proband' s father carried 28 and 66,and the expanded CAG repeat allele inherited from his grandfather.The proband' s mother carried 33 and 56,and the expanded CAG repeat allele inherited from his grandmother.The proband' s son carried 27 and 85 and presented with dystonia besides ataxia.Conclusions The proband' s parents have the common ancestors.Their alleles with expanded CAG repeats probably come from the same allele of their ancestor.The GAG repeat is more unstable in the paternal inheritance than in the maternal inheritance.The 71-year-old asymptomatic family member carry the allele with 56 CAG repeats,which indicates the 56 CAG repeats may be not associated with the disease.The patients within this family have variable clinical features,especially the juvenile-onset case presents with apparent dystonia.

Spinal cord ischemia/reperfusion injury is a stress injury to the spinal cord. Our previous studies using differential proteomics identified 21 differentially expressed proteins (n > 2) in rabbits with spinal cord ischemia/reperfusion injury. Of these proteins, stress-related proteins included protein disulfide isomerase A3, stress-induced-phosphoprotein 1 and heat shock cognate protein 70. In this study, we established New Zealand rabbit models of spinal cord ischemia/reperfusion injury by abdominal aorta occlusion. Results demonstrated that hind limb function initially improved after spinal cord ischemia/reperfusion injury, but then deteriorated. The pathological morphology of the spinal cord became aggravated, but lessened 24 hours after reperfusion. However, the numbers of motor neurons and interneurons in the spinal cord gradually decreased. The expression of protein disulfide isomerase A3, stress-induced-phosphoprotein 1 and heat shock cognate protein 70 was induced by ischemia/reperfusion injury. The expression of these proteins increased within 12 hours after reperfusion, and then decreased, reached a minimum at 24 hours, but subsequently increased again to similar levels seen at 6-12 hours, showing a characterization of induction-inhibition-induction. These three proteins were expressed only in cytoplasm but not in the nuclei. Moreover, the expression was higher in interneurons than in motor neurons, and the survival rate of interneurons was greater than that of motor neurons. It is assumed that the expression of stress-related proteins exhibited a protective effect on neurons.

Objective To investigate whether Chinese multiple system atrophy(MSA) patients have premutation of fragile X mental retardation 1 gene(FMR1).Methods FMR1 CGG repeats were analyzed in 157 MSA patients by polymerase chain reaction,agarose gel electrophoresis and capillary electrophoresis.The patients were collected from Movement Disorder & Neurogenetics Research Center of China-Japan friendship hospital.There were 83 male cases and 74 female cases,including 51 MSA-C patients,12 MSAP patients and 94 MSA-P + C patients.Results No FMR1 CGG repeat premutation was detected in 157 MSA patients.The repeats ranged from 11-49,most common allele was 22.A MSA-C case carried 35/49 alleles did not have middle cerebellar peduncles(MCP) sign which was necessary for the diagnosis of fragile X associated tremor ataxia syndrome (FXTAS).Conclusion The FMR1 premutation in Chinese MSA patients might be very rare.

OBJECTIVE To investigate genetics and clinical characteristics of dentatorubral-pallidoluysian atrophy (DRPLA) in Chinese kindreds. METHODS Fragment analysis with laser-induced fluorescence in capillary electrophoresis was performed for the cytosine-adenine-guanine (CAG) repeats of DRPLA gene in 708 probands of autosomal dominant ataxia pedigrees and 119 sporadic ataxia cases. RESULTS Expanded CAG repeats of DRPLA gene were detected in probands of three ataxia pedigrees, with the numbers of repeats being 16/58, 16/58 and 14/54, respectively. In addition to ataxia, patients with adult-onset disease also exhibited spasm and neck torsion. CONCLUSION Only three cases of DRPLA have been identified among 827 cases, which suggested that DRPLA is a relatively rare subtype of SCA in Chinese population. Clinical variation among the patients suggested DRPLA has a wide spectrum of phenotype.

Background Spinocerebellar ataxia type 2 (SCA2) is an autosomal dominant genetic disease characterized by cerebellar ataxia, ophthalmoplegia, slow saccade, hyporeflexia, action tremor, cognitive decline and peripheral neuropathy. The brain MRI shows obvious atrophy of cerebellum and brainstem, indicating typical change of olivopontocerebellar atrophy. SCA2 is caused by an expanded cytosine-adenine-guanine (CAG) trinucleotide repeat in the encoding region of ATXN2. The normal CAG repeats range from 13 to 31, and ataxic phenotype occurs when the repeats are more than 34. This study focused on the clinical and imaging features of 5 SCA2 families confirmed by genetic testing. The correlation between phenotype and genotype was analyzed. Methods The pathological CAG triplet repeat expansions of SCA1-3, 6, 7, 17 and dentatorubral-pallidoluysian atrophy (DRPLA) genes were analyzed in the probands of 708 autosomal dominant SCA families and 119 sporadic SCA cases. The CAG repeat of ATXN2 gene was amplified by polymerase chain reaction (PCR) and agarose gel electrophoresis. Fragment analysis based on CEQ8000 sequencer were applied to analyze expanded alleles. Results Expanded CAG repeats of ATXN2 gene were detected in 45 probands of SCA2 families. Most of the patients manifested with the typical clinical features of SCA2 such as cerebellar ataxia, ophthalmoplegia, slow saccade and hyporeflexia. Some of them also associated with action tremor. The brain MRI showed obvious atrophy of cerebellum and brainstem. The correlation between clinical features and CAG repeat of ATXN2 gene wasstudied carefully in 5 families. Conclusion Genetic analysis provides the basis for the diagnosis of SCA2. Clinical and neuroimaging features are very helpful in the diagnosis and differential diagnosis of this disease. For the identification of cases carrying intermediate allele, it is important to combine clinical, imaging features with dynamic mutation analysis in the affected cases within the family.

Objective To summarize electromyographic features of external anal sphincter(EAS) in patients with motor neuron disease(MND).Methods EAS-EMG was performed in 20 MND patients with different subtypes,as well as 22 negative controls(cases with neither symptoms nor signs of bowel and bladder dysfunction).Electromyographic characteristics of the two groups were recorded and analyzed statistically.Results Significantly prolonged mean duration [(12.21±2.26) ms vs.(14.55±2.27)ms,P=0.000] and increased area of motor unit potentials(MUPs) [(512.89±363.55)(ms·μV)vs.(718.47±236.57)(ms·μV),P=0.05] were shown in the MND group,when comparing with the negative control group(P≤0.05).No significant differences were found in the amplitude,percentage of polyphasic wave and turns of MUPs between the two groups(P0.05).Conclusions Asymptomatic or subclinical neurogenic lesion may occur in EAS of MND patients.Mean duration and area of MUPs are the most sensitive parameters exhibiting MUPs characters.

目的探讨周围髓鞘蛋白22（PMP22）基因重复突变阳性的夏科-马里-图斯病（CMT）lA亚型患者临床和神经电生理改变特点。方法总结21例PMP22基因重复突变阳性的CMTlA患者的临床特点,并分析其神经电生理特征。结果 21例患者中,10例临床特征符合四肢远端萎缩无力的典型CMTl型表现,另外11例呈不典型性,如仅有头晕、合并听力障碍、上肢姿势性震颤、反复发作性肢体无力、伴有小脑性共济失调及癫疒间等。10例患者肌电图出现纤颤电位和（或）正锐波,15例患者运动单位电位时限延长。神经传导存在广泛异常,所有患者被检的运动或感觉神经传导速度存在不同程度的减慢或消失。结论 PMP22重复突变阳性的CMTlA患者具有较高的临床异质性,其电生理特点为肌电图呈神经源性损害,感觉神经病变重于运动神经,下肢受累程度重于上肢,神经电生理检查对CMT1A的诊断很重要。

Objective To investigate the clinical features and genetic mutations of spinocerebellar ataxia type 17 (SCA17).Methods The pathological CAG triplet repeat expansions of the SCA3,SCA1,SCA2,SCA6,SCA7,SCA8,SCA12,SCA17 and dentatorubral pallidoluysian atrophy genes were analyzed in 708 probands of autosomal dominant familial SCA and 1 19 sporadic SCA cases.The CAG repeats of TATA-binding protein (TBP) gene were amplified by means of polymerase chain reaction and agarose gel electrophoresis.For the samples with two alleles,fragment analysis based on CEQ8000 sequencer was applied to analyze the CAG repeat numbers.Furthermore,the correlation between clinical features and CAG repeat in the TBP gene was studied carefully.Results The expanded CAG repeats in the TBP gene was detected in 5 cases with 37/50,36/45,38/52,38/53,36/54 separately.And the main clinical manifestations were ataxia and memory impairment.Conclusion These findings indicate that SCA17 might be a rare subtype of SCA in the Chinese population and the clinical features of SCA17 cover a wider spectrum than previously reviewed.

脑干梗死是指椎基底动脉及其分支血管狭窄或闭塞引起中脑、桥脑或延髓缺血、坏死、软化,出现相应的神经系统症状,临床上较为常见,占脑梗死的20％～30％[1].经典脑干梗死常有眩晕、复视、构音障碍、吞咽困难、进食水呛咳、交叉性瘫痪、四肢瘫痪、交叉性感觉障碍等临床表现[2-3].随着影像技术特别是磁共振技术的发展,临床上可以观察到一些脑干梗死无交叉定位体征,而有中枢性面瘫、偏身运动或感觉障碍等类似对侧大脑半球病变体征,单纯依据经典理论和临床表现可能对病灶定位错误,对病情判断失当.本文对此类脑梗死的临床特点进行了初步回顾性分析,以期对减少临床误诊有所帮助.

Objective To investigate the change of the level of platelet 5-hydroxytryptamine(HT) and its correlation with depression and cognitive dysfunction in patients with Parkinson disease(PD). Methods The levels of platelet 5-HT was measured in 54 PD patients and 54 healthy aged and elderly people(normal control group) by high performance liquid chromatography.PD patients were measured by Hamilton depression rating scale(HAMD) and Mini-mental state examination(MMSE) scale.The correlation analyed between the level of platelet 5-HT and HAMD,MMSE. Results The level of platelet 5-HT in PD group was significantly lower than that in normal control group(P<0.05),and the level of 5-HT in PD patients with depression or cognitive dysfunction was significantly lower than that in PD patients without depression or cognitive dysfunction(all P<0.05).The level of platelet 5-HT was negative correlation with HAMD score(r=-0.3310,P<0.05),and positive correlation with MMSE score(r=0.5415,P<0.05).The MMSE score in PD patients with depression was significantly lower than that in PD patients without depression(P<0.05). Conclusion The level of platelet 5-HT in PD patient is decreased significantly,which even lower in PD patients with depression and cognitive dysfunction.

<正>多巴反应性肌张力不全(DRD)是一种较少见的锥体外系疾病,现报告2例如下。1病例1.1例1女,17岁,高中学生。因"间断走路困难6年余"于2009年6月14日住院。患者大约自10岁起逐步出现双下

Objective To establish eukaryotic expression system of full ⁃ length gene in normal [CAG repeats (Q19)] and abnormal [CAG repeats (Q82 and Q66)] atrophin ⁃ 1 [the pathogenic gene of dentatorubral⁃pallidoluysian atrophy (DRPLA)]. To compare the expression system of stable and transient transfected cell line by observing the cellular appearance and atrophin⁃1 fusion protein expression and its distribution for further research on the establishment of a fine cell model. Methods Based on the 2 established stable transfected green⁃fluoresent protein (GFP)⁃atrophin⁃1⁃Q19/Flp⁃In TREx293 and GFP⁃ atrophin ⁃ 1 ⁃ Q82/Flp ⁃ In TREx293 cell lines, Tet ⁃ on system was used to induce the expression, which contained normal and abnormal atrophin ⁃ 1 full ⁃ length gene and GFP fusion proteins, respectively. Meanwhile, GFP ⁃ atrophin ⁃ 1 ⁃ Q19 and GFP ⁃ atrophin ⁃ 1 ⁃ Q66 recombinant plasmids were transiently transfected into 293T cells and SH⁃SY5Y cells respectively with LipofectamineTM 2000. The expression and localization of normal and abnormal atrophin ⁃ 1 fusion proteins in the cells were detected by light microscope with HE staining and fluorescence microscope; the ultrastructure of the cells were observed by electron microscope; the expression of GFP ⁃ atrophin ⁃ 1 fusion proteins were detected by Western blotting. Results In stable transfected system, the green fluorescence signals were detected in the nucleus, and abnormal proteins were not uniformly distributed while some were aggregated. Electron microscope showed shrinkaged nuclear membrane and abnormal intranuclear structure in abnormal cells. In transient transfection of 293T, Q66 protein aggregated in the nucleus which was detected as acidophilic bodies by HE staining⁃light microscope and atrophin⁃1 fusion protein as punctuate pattern by fluorescence microscope. Under the observation of electron microscope, great quantity of electron ⁃ dense substance accumulation, distinct shrinkage nuclear membrane and nucleus disappearance were seen in abnormal cells. The expression of GFP ⁃ atrophin ⁃ 1 fusion proteins were detected by Western blotting. The location of protein expression and Western blotting results in transient transfected SH⁃SY5Y cells system were essentially similar to those in transfected 293T cells. Conclusion In eukaryotic cell expression system, atrophin⁃1 with amplified polyglutamine chain may induce obvious aberrant aggregation in nucleus, and alteration of cellular appearance. The establishment of atrophin⁃1 expression eukaryotic cell model will provide the foundation for the research of pathogenesis of DRPLA, and therotic basis for further investigation of clincial intervention. DOI：10.3969/j.issn.1672-6731.2011.01.016