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SMPD1 variants in Chinese Han patients with sporadic Parkinson's disease
Parkinsonism and Related Disorders xxx (2016) 1e3
Contents lists available at ScienceDirect
Parkinsonism and Related Disorders journal homepage: www.elsevier.com/locate/parkreldis
Short communication
SMPD1 variants in Chinese Han patients with sporadic Parkinson's disease Cheng-yuan Mao a, b, 1, Jing Yang a, b, 1, Hui Wang a, b, 1, Shu-yu Zhang a, b, Zhi-hua Yang a, b, Hai-yang Luo a, b, Fang Li a, b, Mengmeng Shi a, b, Yu-tao Liu a, Zheng-ping Zhuang c, Pan Du d, Yao-he Wang d, Chang-he Shi a, b, **, Yu-ming Xu a, * a
Department of Neurology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou University, Zhengzhou, 450000, Henan, China Institute of Clinical Medicine, The First Affiliated Hospital of Zhengzhou University, Zhengzhou University, Zhengzhou, 450000, Henan, China Surgical Neurology Branch, National Institute of Neurological Disorders and Stroke, Bethesda, 20892, MD, USA d International Joint Research Laboratory for Cell and Gene Therapy of Henan Province, The First Affiliated Hospital of Zhengzhou University, Zhengzhou University, 1 Jian-she East Road, Zhengzhou, 450000, Henan, China b c
a r t i c l e i n f o
a b s t r a c t
Article history: Received 1 July 2016 Received in revised form 4 October 2016 Accepted 18 October 2016
Introduction: A founder mutation, p.L302P, in sphingomyelin phosphodiesterase 1, acid lysosomal (SMPD1), causing Niemann-Pick disease, a recessive lysosomal storage disorder, was reported to be associated with increased risk of Parkinson's disease (PD) in Ashkenazi Jewish population. Several other studies about the association between SMPD1 variants and PD were performed afterward in other populations. However, the results on the role of SMPD1 mutations for PD have been conflicting. This study aimed to investigate the role of mutations in SMPD1 in Chinese PD patients. Methods: We sequenced all the exons of this gene in 512 Chinese Han cases with sporadic Parkinson's disease and 495 matched healthy control subjects. Results: We identified Leu-Ala (Val) repeat variants and six known single nucleotide variants (p.A36V, p.D212D, p.P332R, p.G508R, p.P533L, p.T544T) in SMPD1 in both patients and normal controls. Casecontrol analysis showed the association between Leu-Ala (Val) repeat variants in SMPD1and Chinese Han patients with PD (c2 ¼ 8.771, p ¼ 0.012), and the allele with less than seven LeuAla (Val) repeats may increase the risk of PD (p ¼ 0.010). Conclusion: We identified association between Leu-Ala (Val) repeat variants in SMPD1 and Chinese Han patients with sporadic Parkinson's disease. Our results provide further support for the role of lysosomal pathways in PD development. © 2016 Elsevier Ltd. All rights reserved.
Keywords: Parkinson's disease SMPD1 Variant Chinese population
1. Introduction Parkinson's disease (PD) is the second most common neurodegenerative condition after Alzheimer's disease, with an estimated worldwide prevalence of 50e200 cases per 100,000 inhabitants [1].
* Corresponding author. Department of Neurology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou University. 1 Jian-she east road, Zhengzhou, 450000, Henan, China. ** Corresponding author. Department of Neurology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou University. 1 Jian-she east road, Zhengzhou, 450000, Henan, China. E-mail addresses: [email protected] (C.-h. Shi), [email protected] (Y.-m. Xu). 1 These authors contributed equally to this article.
The phenotype of bradykinesia, resting tremor, rigidity and postural instability is mainly caused by the massive death of dopaminergic neurons in the substantia nigra pars compacta. Surviving neurons show the Lewy bodies (LBs), typical cytoplasmic inclusions containing ubiquitin, alpha-synuclein (a-syn) and other proteins [1]. Although familial forms only account for approximately 10% of PD cases, exploring the genetic factors underlying PD is essential to provide clues about the molecular mechanisms involved, with the ultimate goal to develop treatments aimed at disease prevention and cure. Previous studies have shown that lysosomal dysfunction may play an important role in the pathogenesis of PD. Genetic causes of PD including a-synuclein, LRRK2, Parkin, PINK1, DJ1, and ATP13A2 encode proteins that reside in the lysosome or function in lysosomal-related pathways, such as autophagy and mitophagy [2].
http://dx.doi.org/10.1016/j.parkreldis.2016.10.014 1353-8020/© 2016 Elsevier Ltd. All rights reserved.
Please cite this article in press as: C.-y. Mao, et al., SMPD1 variants in Chinese Han patients with sporadic Parkinson's disease, Parkinsonism and Related Disorders (2016), http://dx.doi.org/10.1016/j.parkreldis.2016.10.014
2
C.-y. Mao et al. / Parkinsonism and Related Disorders xxx (2016) 1e3
And carriers of mutations in GBA causing Gaucher's disease, the most common lysosomal storage disorder, are at significantly increased risk for developing PD, with an earlier age of onset compared with PD patients who do not carry these mutations. Recently, a founder mutation, p.L302P, in sphingomyelin phosphodiesterase 1, acid lysosomal (SMPD1), causing Niemann-Pick disease, another recessive lysosomal storage disorder, was reported to be associated with increased risk of PD in Ashkenazi Jewish population [2]. Several other studies about the association between SMPD1variants and PD were performed afterward in other populations [3e7]. However, the results on the role of SMPD1 mutations for PD have been conflicting. In this study, to investigate the role of mutations in SMPD1 in PD, we sequenced all the exons of this gene in 512 Chinese Han cases with sporadic Parkinson's disease and 495 matched healthy control subjects.
2. Materials and methods 512 unrelated Chinese Han patients with sporadic Parkinson's disease and 495 gender-, age-, and ethnicity matched healthy controls were enrolled in this study. This study was approved. by the Ethics Committee of First Affiliated Hospital of Zhengzhou University, and all subjects provided written informed consent in genetic research. The cases were defined using the United Kingdom Parkinson's Disease Society Brain Bank criteria. Genomic DNA was extracted from peripheral blood collected from the patients and controls according to the manufacturer's protocol. The coding and flanking intronic regions of the SMPD1 gene were amplified (Table S1). Sanger sequencing was performed to identify SMPD1 variants. Allele frequencies in case and control subjects were compared using Pearson's c2 or c2 with Yates correction test, and we used the Bonferroni correction to adjust for multiple testing. The Hardy-Weinberg equilibrium was performed as to ascertain the normal heterogeneity of the population.
3. Results The mean age of the 512 PD patients was 64.6 ± 9.6 years (male/ female: 327/185) and the mean age at onset of PD symptoms was 58.8 ± 11.3 years. And the mean age of the 495 unrelated healthy controls was 65.3 ± 9.3 years (male/female: 313/182). From the sequencing of all the subjects, we identified Leu-Ala (Val) repeat variants and six known single nucleotide variants (p.A36V, p.D212D, p.P332R, p.G508R, p.P533L, p.T544T) in both PD and control cohorts. Distributions of the genotypes in PD and control groups were both in Hardy-Weinberg equilibrium. Four of these variants were missense and have previously been reported (p.A36V, p.P332R, p.G508R, p.P533L) [7e9]. All the four variants were predicted by the Sorting Intolerant From Tolerant program (SIFT), and the results were tolerated ((p.A36V, p.G508R) and deleterious (p.P332R, p.P533L). The other two variants (p.D212D, p.T544T) were synonymous. There were no significant differences in genotypic distribution between the PD cohort and control cohort for the 6 variants after Bonferroni correction (all p > 0.008, Table 2), indicating that they were not pathogenic variants. We further found the association between Leu-Ala (Val) repeat variants (Fig. S1) in SMPD1 and Chinese Han patients with PD (p ¼ 0.012, c2 ¼ 8.771, Table 1, Table S2). The allele with less than seven LeuAla (Val) repeats may increase the risk of PD (p ¼ 0.01). No significant clinical differences were observed when PD patients with less than 7 Leu-Ala repeat and non-mutation patients were compared (Table S3).
Table 1 Allelic distribution of the SMPD1 gene Leu-Ala (Val) repeat variants in 512 patients and 495 controls. Leu-Ala (Val) repeats
Patients Controls
N<7
N¼7
N>7
221 169
796 807
7 14
c2
P value
8.771
0.012
4. Discussion In this study, we identified Leu-Ala (Val) repeat variants and six known single nucleotide variants (p.A36V, p.D212D, p.P332R, p.G508R, p.P533L, p.T544T) in SMPD1 in both 512 unrelated Chinese Han patients with sporadic Parkinson's disease and 495 normal controls. Case-control analysis showed the association between Leu-Ala (Val) repeat variants in SMPD1and Chinese Han patients with PD, and the allele with less than seven LeuAla (Val) repeats may increase the risk of PD. Recently, a founder mutation, p.L302P, in SMPD1 was reported to be associated with increased risk of PD in Ashkenazi Jewish population [2]. But the association between the SMPD1 p.L302P mutation and PD has not been validated in additional population, suggesting possible ethnic-specific effect of the variant [3e7]. Especially, Li et al. did not find an association between the p.L302P mutation and Chinese PD patients in Mainland China [6]. Foo et al. identified a rare variant p.R591C which indicated an increased risk of PD in Chinese population from Singapore [4]. Our study did not find p.R591C, possibly due to limited sample size. In this study, we identified six known single nucleotide variants. Even though two of these variants (p.P332R, p.P533L) were predicted by the Sorting Intolerant From Tolerant program (SIFT) to be deleterious, they did not increase the risk of PD. Additionally, we found the association between Leu-Ala (Val) repeat variants in SMPD1and Chinese Han patients with PD, which is consistent with Sheng Deng et al.’s result [7]. However, it remains unclear whether more than or less than seven LeuAla (Val) repeats may increase the risk of PD. Further validation in larger samples will be needed. The SMPD1 gene, located on chromosome 11p15.1-p15.4, is 4.6 kb in length and composed of six exons encoding 631 amino acid proteins. The SMPD1 gene encodes sphingomyelin phosphodiesterase 1 (acid sphingomyelinase, ASM) which is a lysosomal enzyme that cleaves the phosphocholine head group of sphingomyelin to generate ceramide [10]. The deficient activity of ASM may result in types A or B Niemann-Pick disease (NPD). The levels of ceramide changes have been considered to play an important role in neurodegeneration diseases. The ceramide metabolism pathway may be involved in PD pathogenesis. Furthermore, the Leu-Ala (Val) repeat variant has been reported in Niemann-Pick disease in several previous studies [11,12]. The repeated sequence located within the putative ASM signal peptide, which contained a unique hydrophobic core comprised of leucine/alanine or leucine/valine repeats. Rhein et al. provided evidence that the polymorphic ASM signal peptide regulates ASM secretion [12]. Leu-Ala repeat variants may affect formation of the secreted form (S-ASM) which is finally released into the extracellular space, influencing the lysosomal form (L-ASM) which is targeted to the endo-lysosomal compartment. That may disturb the lysosomal pathways in PD development. However, further studies are needed to validate this hypothesis. In conclusion, we identified association between Leu-Ala (Val) repeat variants in SMPD1 and Chinese Han patients with sporadic Parkinson's disease. Our results provide further support for the role of lysosomal pathways in PD development. Further genetic analysis
Please cite this article in press as: C.-y. Mao, et al., SMPD1 variants in Chinese Han patients with sporadic Parkinson's disease, Parkinsonism and Related Disorders (2016), http://dx.doi.org/10.1016/j.parkreldis.2016.10.014
C.-y. Mao et al. / Parkinsonism and Related Disorders xxx (2016) 1e3
3
Table 2 Genotypes and alternative minor allele frequencies of identified SMPD1 variants. Variant
p.A36V p.D212D p.P332R p.G508R p.P533L p.T544T
Sequencing
Alternative minor allele frequency
ExAC allele frequence
Sporadic PD vs controls
Case(n ¼ 512)
Controls (n ¼ 495)
Sporadic PD
Controls
(East Asian)
OR(95%CI)
P
142 5 5 153 7 3
153 2 2 99 10 5
0.247 0.005 0.005 0.167 0.007 0.003
0.225 0.002 0.002 0.131 0.010 0.005
0.783 0.005 0.004 0.139 0.007 0.002
1.097(0.937e1.284) 2.417(0.470e12.429) 2.417(0.470e12.429) 1.272(1.030e1.570) 0.677(0.259e1.771) 0.580(0.139e2.421)
0.249 0.476 0.476 0.025 0.423 0.688
PD, Parkinson's disease; OR, odds ratio; CI, confidence interval.
and function studies of SMPD1 variants are needed to understand the role of SMPD1 in the pathogenesis of PD.
dx.doi.org/10.1016/j.parkreldis.2016.10.014.
Potential conflict of interest References No conflict of interest. Author contributions Study conception, design and, organization (Chengyuan Mao, Jing Yang, Changhe Shi and Yuming Xu); acquisition of data (Chengyuan Mao, Jing Yang, Hui Wang, Shuyu Zhang, Haiyang Luo, Fang Li, Mengmeng Shi, Yutao Liu, Changhe Shi and Yuming Xu); analysis and interpretation of data (Chengyuan Mao, Jing Yang, Hui Wang, Zhi-hua Yang, Pan Du, Yaohe Wang and Yuming Xu); drafting of the manuscript (Chengyuan Mao, Jing Yang, Hui Wang, Changhe Shi and Yuming Xu); obtained funding (Jing Yang, Yutao Liu, Changhe Shi and Yuming Xu); administrative, technical, and material support (Changhe Shi, Pan Du, Yaohe Wang and Zhengping Zhuang) and study supervision (Changhe Shi, Zhengping Zhuang and Yuming Xu). Acknowledgements This work was supported by grant 81530037 and 81471158from the National Natural Science Foundation of China (to Dr. Yu-ming Xu), grant U1404311 from the National Natural Science Foundation of China (to Dr. Chang-he Shi), grant 81600946 from the National Natural Science Foundation of China (to Dr. Jing Yang), grant 201503038 from the Medical Science and Technique Foundation of Henan Province (to Dr. Yu-tao Liu), grant 16A320052 from the Key Scientific Research Projects of Universities in Henan Province (to Dr. Yu-tao Liu). We would like to thank the patients and normal controls for their participation in this study. Appendix A. Supplementary data Supplementary data related to this article can be found at http://
[1] P. De Rosa, E.S. Marini, V. Gelmetti, E.M. Valente, Candidate genes for Parkinson disease: lessons from pathogenesis, Clin. Chim. Acta Int. J. Clin. Chem. 449 (2015) 68e76. [2] Z. Gan-Or, L.J. Ozelius, A. Bar-Shira, R. Saunders-Pullman, A. Mirelman, R. Kornreich, M. Gana-Weisz, D. Raymond, L. Rozenkrantz, A. Deik, T. Gurevich, S.J. Gross, N. Schreiber-Agus, N. Giladi, S.B. Bressman, A. OrrUrtreger, The p.L302P mutation in the lysosomal enzyme gene SMPD1 is a risk factor for Parkinson disease, Neurology 80 (17) (2013) 1606e1610. [3] R.M. Wu, C.H. Lin, H.I. Lin, The p.L302P mutation in the lysosomal enzyme gene SMPD1 is a risk factor for Parkinson disease, Neurology 82 (3) (2014) 283. [4] J.N. Foo, H. Liany, J.X. Bei, X.Q. Yu, J. Liu, W.L. Au, K.M. Prakash, L.C. Tan, E.K. Tan, Rare lysosomal enzyme gene SMPD1 variant (p.R591C) associates with Parkinson's disease, Neurobiol. Aging 34 (12) (2013) 2890 e13-5. [5] E. Dagan, I. Schlesinger, M. Ayoub, A. Mory, M. Nassar, A. Kurolap, J. PeretzAharon, R. Gershoni-Baruch, The contribution of Niemann-Pick SMPD1 mutations to Parkinson disease in Ashkenazi Jews, Park. Relat. Disord. 21 (9) (2015) 1067e1071. [6] K. Li, B.S. Tang, N.N. Yang, J.F. Kang, Z.H. Liu, R.Q. Liu, et al., Association study between SMPD1 p.L302P and sporadic Parkinson's disease in ethnic Chinese population, Int. J. Clin. Exp. Med. 8 (2015) 13869e13873. [7] S. Deng, X. Deng, Z. Song, X. Xiu, Y. Guo, J. Xiao, H. Deng, Systematic genetic analysis of the SMPD1 gene in Chinese patients with Parkinson's disease, Mol. Neurobiol. 53 (7) (Sep 2016) 5025e5029. [8] M.D. Manshadi, B. Kamalidehghan, F. Keshavarzi, O. Aryani, S. Dadgar, A. Arastehkani, et al., Four novel p.N385K, p.V36A, c.1033-1034insT and c.1417-1418delCT mutations in the sphingomyelin Phosphodiesterase 1 (SMPD1) gene in patients with types A and B Niemann-Pick disease (NPD), Int. J. Mol. Sci. 16 (2015) 6668e6676. [9] G. Murgia, D. Firinu, R. Meleddu, M.M. Lorrai, P.E. Manconi, S.R. Del Giacco, Systemic lupus erythematosus occurring in a patient with Niemann-Pick type B disease, Lupus 24 (2015) 1332e1334. [10] E.H. Schuchman, O. Levran, L.V. Pereira, R.J. Desnick, Structural organization and complete nucleotide sequence of the gene encoding human acid sphingomyelinase (SMPD1), Genomics 12 (2) (1992) 197e205. [11] Q. Wan, E.H. Schuchman, A novel polymorphism in the human acid sphingomyelinase gene due to size variation of the signal peptide region, Biochim. Biophys. Acta 1270 (1995) 207e210. [12] C. Rhein, M. Reichel, C. Muhle, A. Rotter, S.G. Schwab, J. Kornhuber, Secretion of acid Sphingomyelinase is affected by its polymorphic signal peptide, Cell. Physiol. Biochem. Int. J. Exp. Cell. Physiol. Biochem. Pharmacol. 34 (2014) 1385e1401.
Please cite this article in press as: C.-y. Mao, et al., SMPD1 variants in Chinese Han patients with sporadic Parkinson's disease, Parkinsonism and Related Disorders (2016), http://dx.doi.org/10.1016/j.parkreldis.2016.10.014
Contents lists available at ScienceDirect
Parkinsonism and Related Disorders journal homepage: www.elsevier.com/locate/parkreldis
Short communication
SMPD1 variants in Chinese Han patients with sporadic Parkinson's disease Cheng-yuan Mao a, b, 1, Jing Yang a, b, 1, Hui Wang a, b, 1, Shu-yu Zhang a, b, Zhi-hua Yang a, b, Hai-yang Luo a, b, Fang Li a, b, Mengmeng Shi a, b, Yu-tao Liu a, Zheng-ping Zhuang c, Pan Du d, Yao-he Wang d, Chang-he Shi a, b, **, Yu-ming Xu a, * a
Department of Neurology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou University, Zhengzhou, 450000, Henan, China Institute of Clinical Medicine, The First Affiliated Hospital of Zhengzhou University, Zhengzhou University, Zhengzhou, 450000, Henan, China Surgical Neurology Branch, National Institute of Neurological Disorders and Stroke, Bethesda, 20892, MD, USA d International Joint Research Laboratory for Cell and Gene Therapy of Henan Province, The First Affiliated Hospital of Zhengzhou University, Zhengzhou University, 1 Jian-she East Road, Zhengzhou, 450000, Henan, China b c
a r t i c l e i n f o
a b s t r a c t
Article history: Received 1 July 2016 Received in revised form 4 October 2016 Accepted 18 October 2016
Introduction: A founder mutation, p.L302P, in sphingomyelin phosphodiesterase 1, acid lysosomal (SMPD1), causing Niemann-Pick disease, a recessive lysosomal storage disorder, was reported to be associated with increased risk of Parkinson's disease (PD) in Ashkenazi Jewish population. Several other studies about the association between SMPD1 variants and PD were performed afterward in other populations. However, the results on the role of SMPD1 mutations for PD have been conflicting. This study aimed to investigate the role of mutations in SMPD1 in Chinese PD patients. Methods: We sequenced all the exons of this gene in 512 Chinese Han cases with sporadic Parkinson's disease and 495 matched healthy control subjects. Results: We identified Leu-Ala (Val) repeat variants and six known single nucleotide variants (p.A36V, p.D212D, p.P332R, p.G508R, p.P533L, p.T544T) in SMPD1 in both patients and normal controls. Casecontrol analysis showed the association between Leu-Ala (Val) repeat variants in SMPD1and Chinese Han patients with PD (c2 ¼ 8.771, p ¼ 0.012), and the allele with less than seven LeuAla (Val) repeats may increase the risk of PD (p ¼ 0.010). Conclusion: We identified association between Leu-Ala (Val) repeat variants in SMPD1 and Chinese Han patients with sporadic Parkinson's disease. Our results provide further support for the role of lysosomal pathways in PD development. © 2016 Elsevier Ltd. All rights reserved.
Keywords: Parkinson's disease SMPD1 Variant Chinese population
1. Introduction Parkinson's disease (PD) is the second most common neurodegenerative condition after Alzheimer's disease, with an estimated worldwide prevalence of 50e200 cases per 100,000 inhabitants [1].
* Corresponding author. Department of Neurology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou University. 1 Jian-she east road, Zhengzhou, 450000, Henan, China. ** Corresponding author. Department of Neurology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou University. 1 Jian-she east road, Zhengzhou, 450000, Henan, China. E-mail addresses: [email protected] (C.-h. Shi), [email protected] (Y.-m. Xu). 1 These authors contributed equally to this article.
The phenotype of bradykinesia, resting tremor, rigidity and postural instability is mainly caused by the massive death of dopaminergic neurons in the substantia nigra pars compacta. Surviving neurons show the Lewy bodies (LBs), typical cytoplasmic inclusions containing ubiquitin, alpha-synuclein (a-syn) and other proteins [1]. Although familial forms only account for approximately 10% of PD cases, exploring the genetic factors underlying PD is essential to provide clues about the molecular mechanisms involved, with the ultimate goal to develop treatments aimed at disease prevention and cure. Previous studies have shown that lysosomal dysfunction may play an important role in the pathogenesis of PD. Genetic causes of PD including a-synuclein, LRRK2, Parkin, PINK1, DJ1, and ATP13A2 encode proteins that reside in the lysosome or function in lysosomal-related pathways, such as autophagy and mitophagy [2].
http://dx.doi.org/10.1016/j.parkreldis.2016.10.014 1353-8020/© 2016 Elsevier Ltd. All rights reserved.
Please cite this article in press as: C.-y. Mao, et al., SMPD1 variants in Chinese Han patients with sporadic Parkinson's disease, Parkinsonism and Related Disorders (2016), http://dx.doi.org/10.1016/j.parkreldis.2016.10.014
2
C.-y. Mao et al. / Parkinsonism and Related Disorders xxx (2016) 1e3
And carriers of mutations in GBA causing Gaucher's disease, the most common lysosomal storage disorder, are at significantly increased risk for developing PD, with an earlier age of onset compared with PD patients who do not carry these mutations. Recently, a founder mutation, p.L302P, in sphingomyelin phosphodiesterase 1, acid lysosomal (SMPD1), causing Niemann-Pick disease, another recessive lysosomal storage disorder, was reported to be associated with increased risk of PD in Ashkenazi Jewish population [2]. Several other studies about the association between SMPD1variants and PD were performed afterward in other populations [3e7]. However, the results on the role of SMPD1 mutations for PD have been conflicting. In this study, to investigate the role of mutations in SMPD1 in PD, we sequenced all the exons of this gene in 512 Chinese Han cases with sporadic Parkinson's disease and 495 matched healthy control subjects.
2. Materials and methods 512 unrelated Chinese Han patients with sporadic Parkinson's disease and 495 gender-, age-, and ethnicity matched healthy controls were enrolled in this study. This study was approved. by the Ethics Committee of First Affiliated Hospital of Zhengzhou University, and all subjects provided written informed consent in genetic research. The cases were defined using the United Kingdom Parkinson's Disease Society Brain Bank criteria. Genomic DNA was extracted from peripheral blood collected from the patients and controls according to the manufacturer's protocol. The coding and flanking intronic regions of the SMPD1 gene were amplified (Table S1). Sanger sequencing was performed to identify SMPD1 variants. Allele frequencies in case and control subjects were compared using Pearson's c2 or c2 with Yates correction test, and we used the Bonferroni correction to adjust for multiple testing. The Hardy-Weinberg equilibrium was performed as to ascertain the normal heterogeneity of the population.
3. Results The mean age of the 512 PD patients was 64.6 ± 9.6 years (male/ female: 327/185) and the mean age at onset of PD symptoms was 58.8 ± 11.3 years. And the mean age of the 495 unrelated healthy controls was 65.3 ± 9.3 years (male/female: 313/182). From the sequencing of all the subjects, we identified Leu-Ala (Val) repeat variants and six known single nucleotide variants (p.A36V, p.D212D, p.P332R, p.G508R, p.P533L, p.T544T) in both PD and control cohorts. Distributions of the genotypes in PD and control groups were both in Hardy-Weinberg equilibrium. Four of these variants were missense and have previously been reported (p.A36V, p.P332R, p.G508R, p.P533L) [7e9]. All the four variants were predicted by the Sorting Intolerant From Tolerant program (SIFT), and the results were tolerated ((p.A36V, p.G508R) and deleterious (p.P332R, p.P533L). The other two variants (p.D212D, p.T544T) were synonymous. There were no significant differences in genotypic distribution between the PD cohort and control cohort for the 6 variants after Bonferroni correction (all p > 0.008, Table 2), indicating that they were not pathogenic variants. We further found the association between Leu-Ala (Val) repeat variants (Fig. S1) in SMPD1 and Chinese Han patients with PD (p ¼ 0.012, c2 ¼ 8.771, Table 1, Table S2). The allele with less than seven LeuAla (Val) repeats may increase the risk of PD (p ¼ 0.01). No significant clinical differences were observed when PD patients with less than 7 Leu-Ala repeat and non-mutation patients were compared (Table S3).
Table 1 Allelic distribution of the SMPD1 gene Leu-Ala (Val) repeat variants in 512 patients and 495 controls. Leu-Ala (Val) repeats
Patients Controls
N<7
N¼7
N>7
221 169
796 807
7 14
c2
P value
8.771
0.012
4. Discussion In this study, we identified Leu-Ala (Val) repeat variants and six known single nucleotide variants (p.A36V, p.D212D, p.P332R, p.G508R, p.P533L, p.T544T) in SMPD1 in both 512 unrelated Chinese Han patients with sporadic Parkinson's disease and 495 normal controls. Case-control analysis showed the association between Leu-Ala (Val) repeat variants in SMPD1and Chinese Han patients with PD, and the allele with less than seven LeuAla (Val) repeats may increase the risk of PD. Recently, a founder mutation, p.L302P, in SMPD1 was reported to be associated with increased risk of PD in Ashkenazi Jewish population [2]. But the association between the SMPD1 p.L302P mutation and PD has not been validated in additional population, suggesting possible ethnic-specific effect of the variant [3e7]. Especially, Li et al. did not find an association between the p.L302P mutation and Chinese PD patients in Mainland China [6]. Foo et al. identified a rare variant p.R591C which indicated an increased risk of PD in Chinese population from Singapore [4]. Our study did not find p.R591C, possibly due to limited sample size. In this study, we identified six known single nucleotide variants. Even though two of these variants (p.P332R, p.P533L) were predicted by the Sorting Intolerant From Tolerant program (SIFT) to be deleterious, they did not increase the risk of PD. Additionally, we found the association between Leu-Ala (Val) repeat variants in SMPD1and Chinese Han patients with PD, which is consistent with Sheng Deng et al.’s result [7]. However, it remains unclear whether more than or less than seven LeuAla (Val) repeats may increase the risk of PD. Further validation in larger samples will be needed. The SMPD1 gene, located on chromosome 11p15.1-p15.4, is 4.6 kb in length and composed of six exons encoding 631 amino acid proteins. The SMPD1 gene encodes sphingomyelin phosphodiesterase 1 (acid sphingomyelinase, ASM) which is a lysosomal enzyme that cleaves the phosphocholine head group of sphingomyelin to generate ceramide [10]. The deficient activity of ASM may result in types A or B Niemann-Pick disease (NPD). The levels of ceramide changes have been considered to play an important role in neurodegeneration diseases. The ceramide metabolism pathway may be involved in PD pathogenesis. Furthermore, the Leu-Ala (Val) repeat variant has been reported in Niemann-Pick disease in several previous studies [11,12]. The repeated sequence located within the putative ASM signal peptide, which contained a unique hydrophobic core comprised of leucine/alanine or leucine/valine repeats. Rhein et al. provided evidence that the polymorphic ASM signal peptide regulates ASM secretion [12]. Leu-Ala repeat variants may affect formation of the secreted form (S-ASM) which is finally released into the extracellular space, influencing the lysosomal form (L-ASM) which is targeted to the endo-lysosomal compartment. That may disturb the lysosomal pathways in PD development. However, further studies are needed to validate this hypothesis. In conclusion, we identified association between Leu-Ala (Val) repeat variants in SMPD1 and Chinese Han patients with sporadic Parkinson's disease. Our results provide further support for the role of lysosomal pathways in PD development. Further genetic analysis
Please cite this article in press as: C.-y. Mao, et al., SMPD1 variants in Chinese Han patients with sporadic Parkinson's disease, Parkinsonism and Related Disorders (2016), http://dx.doi.org/10.1016/j.parkreldis.2016.10.014
C.-y. Mao et al. / Parkinsonism and Related Disorders xxx (2016) 1e3
3
Table 2 Genotypes and alternative minor allele frequencies of identified SMPD1 variants. Variant
p.A36V p.D212D p.P332R p.G508R p.P533L p.T544T
Sequencing
Alternative minor allele frequency
ExAC allele frequence
Sporadic PD vs controls
Case(n ¼ 512)
Controls (n ¼ 495)
Sporadic PD
Controls
(East Asian)
OR(95%CI)
P
142 5 5 153 7 3
153 2 2 99 10 5
0.247 0.005 0.005 0.167 0.007 0.003
0.225 0.002 0.002 0.131 0.010 0.005
0.783 0.005 0.004 0.139 0.007 0.002
1.097(0.937e1.284) 2.417(0.470e12.429) 2.417(0.470e12.429) 1.272(1.030e1.570) 0.677(0.259e1.771) 0.580(0.139e2.421)
0.249 0.476 0.476 0.025 0.423 0.688
PD, Parkinson's disease; OR, odds ratio; CI, confidence interval.
and function studies of SMPD1 variants are needed to understand the role of SMPD1 in the pathogenesis of PD.
dx.doi.org/10.1016/j.parkreldis.2016.10.014.
Potential conflict of interest References No conflict of interest. Author contributions Study conception, design and, organization (Chengyuan Mao, Jing Yang, Changhe Shi and Yuming Xu); acquisition of data (Chengyuan Mao, Jing Yang, Hui Wang, Shuyu Zhang, Haiyang Luo, Fang Li, Mengmeng Shi, Yutao Liu, Changhe Shi and Yuming Xu); analysis and interpretation of data (Chengyuan Mao, Jing Yang, Hui Wang, Zhi-hua Yang, Pan Du, Yaohe Wang and Yuming Xu); drafting of the manuscript (Chengyuan Mao, Jing Yang, Hui Wang, Changhe Shi and Yuming Xu); obtained funding (Jing Yang, Yutao Liu, Changhe Shi and Yuming Xu); administrative, technical, and material support (Changhe Shi, Pan Du, Yaohe Wang and Zhengping Zhuang) and study supervision (Changhe Shi, Zhengping Zhuang and Yuming Xu). Acknowledgements This work was supported by grant 81530037 and 81471158from the National Natural Science Foundation of China (to Dr. Yu-ming Xu), grant U1404311 from the National Natural Science Foundation of China (to Dr. Chang-he Shi), grant 81600946 from the National Natural Science Foundation of China (to Dr. Jing Yang), grant 201503038 from the Medical Science and Technique Foundation of Henan Province (to Dr. Yu-tao Liu), grant 16A320052 from the Key Scientific Research Projects of Universities in Henan Province (to Dr. Yu-tao Liu). We would like to thank the patients and normal controls for their participation in this study. Appendix A. Supplementary data Supplementary data related to this article can be found at http://
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Please cite this article in press as: C.-y. Mao, et al., SMPD1 variants in Chinese Han patients with sporadic Parkinson's disease, Parkinsonism and Related Disorders (2016), http://dx.doi.org/10.1016/j.parkreldis.2016.10.014