Association of apolipoprotein A5 genetic polymorphisms with steroid-induced osteonecrosis of femoral head in a Chinese Han population
© Cui et al.; licensee BioMed Central. 2014
Received: 16 October 2014
Accepted: 8 December 2014
Published: 17 December 2014
Previous studies suggested that apolipoprotein A5 (ApoA5) genetic polymorphisms (SNPs) may result in lipid metabolism disorders. Therefore, genetic polymorphisms in ApoA5 may be associated with the occurrence of osteonecrosis of femoral head (ONFH).
We designed a case–control study including 223 patients of osteonecrosis and 201 age- and sex-matched control subjects to analyze the association between ApoA5 polymorphisms and susceptibility of steroid-induced ONFH. We utilized polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method to genotype two SNPs (rs662799 and rs3135506) in ApoA5 gene.
We found both rs662799 and rs3135506 were associated with the risk of ONFH in codominant, dominant, and recessive model, respectively. Haplotype analyses suggested that T-C haplotype was associated with decreased risk of ONFH, whereas the haplotype C-C was significantly associated with an increased risk of ONFH.
Our study suggested that ApoA5 genetic polymorphisms were associated with susceptibility to ONFH in Chinese population. However, our results need further investigation with large sample size and various populations.
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Osteonecrosis of femoral head (ONFH) is a kind of orthopedic refractory disease that cellular death happens within femoral head owing to damage of blood supply to the anterior-superior-lateral part of the femoral head . In China, there are approximately 7 million people with ONFH currently. And, new cases reached to 100–200 thousand each year . ONFH is a complex disorder may result from various risk factors such as trauma, alcoholism, coagulation defects, and abnormal lipid metabolism. Recent study suggested that abnormal lipid metabolism is main pathogenesis of osteonecrosis . Hyperlipidemia affects the microcirculation of the femoral head to result in femoral necrosis from multiple links, such as affecting blood coagulation solvent systems, influencing bone fat embolism, and affecting the formation of bone micro-thrombosis ,. ApoA5 is an important apolipoprotein which involved in plasma lipid metabolism. The most significant impact on plasma triglycerides (TG) levels seems to be associated with ApoA5 gene (ID 116519, OMIM accession number 606368) variants ,. ApoA5 is located on TG-rich and high density lipoprotein (HDL) particles, enhances the activity of lipoprotein lipase ,, and recombinant apoA5 binds to the LDL receptor family members . Previous studies suggested that minor alleles of two SNPs (rs662799 and rs3135506) in ApoA5 gene were associated with elevated plasma TG levels, regardless of ethnicity and sex –.
In 2007, Hirata et al.  found ApoB gene polymorphism was associated with osteonecrosis. Subsequently, Wang et al.  found -75G > A polymorphism in ApoAI gene was associated with osteonecrosis in Chinese population. Recently, Yin et al. also found SNPs in ApoAI gene were associated with ONFH in a Chinese Han population . However, the relation between the polymorphisms of ApoA5 gene and ONFH has not been studied.
In the present study, we designed a case–control study to reveal the relation between ApoA5 genetic polymorphisms and ONFH in a Chinese population.
Subjects and methods
The present study has been performed with the approval of the ethics committee of the 5th Affiliated Hospital of Xinjiang Medical University and was in compliance with the Helsinki Declaration. The informed consents of the study were collected from all the candidate subjects.
Characteristics of the participants
SNPs selection and genotyping
Two SNPs (rs662799 and rs3135506), which were reported to be associated with plasma lipid level, in the ApoA5 gene were selected in this study. Blood samples were collected using a standard venipuncture technique and EDTA-containing tubes. DNA was extracted from peripheral vein blood leukocytes using a whole blood genome extraction kit (Beijing Boiteke Corporation, Beijing, China). SNPs rs662799 and rs3135506 were genotyped using PCR-RFLP as described in details elsewhere –. Briefly, polymerase chain reaction (PCR) was performed in a volume of 25 ml containing 200 ng of genomic DNA. The amounts of Mg2+, dNTP, and DNA polymerase (Bangalore Genei, India) used in each reaction were 1.5 mM, 200 mM, and 1 U, respectively. The thermal cycles started with 94°C for 4 min and were followed by 35 cycles of 94°C for 30 s, 55°C for 30 s, and 72°C for 30 s. A total volume of 20 ul containing 20 U endonuclease was added directly to the PCR product and digested at 37°C overnight. After electrophoresis, the digested products were visualized on a 3% polyacrylamide gel with ethidium bromide staining.
Data were analyzed using SPSS 17.0 software package (Chicago, IL, USA).The genotype and allele frequencies were calculated by direct counting method. The differences of genotype and allele distributions between case and control groups were compared using χ2 test, OR value and its 95% CI was calculated according to χ2 test. Normality was assessed by plotting the residuals. Statistical significance was set at p <0.05.
Characteristics of participants
The characteristics of 223 patients and 201 control subjects were shown in Table 1. After the statistical analysis, there were no significant differences in the distribution of age, sex, body mass index (BMI), GLU, and HDL-C between the two groups. However, there were significant differences in TG, TC and LDL-C between these two groups.
The genotype distributions were in Hardy-Weinberg equilibrium in both case group and control group (both P > 0.05, data not shown).
Genotype and allele frequency distributions
Genotypic distribution of ApoA5 polymorphisms
OR (95% CI)
TC + CC
TT + TC
CG + GG
CC + CG
Haplotype analysis of rs662799 and rs3135506
Distribution of haplotypes
OR (95% CI)
0.605 [0.430 ~ 0.852]
1.455 [0.796 ~ 4.905]
0.936 [0.811 ~ 1.982]
2.223 [1.144 ~ 3.361]
ApoA5 genetic polymorphisms and lipids levels
Comparison of lipids levels between each genotype
5.0 2.7 (n = 146)
5.2 2.5 (n = 77)
5.2 2.6 (n = 159)
5.0 2.2 (n = 65)
4.4 2.1 (n = 192)
4.9 2.4 (n = 9)
4.6 2.3 (n = 176)
4.8 2.1 (n = 25)
In the present study, we found that in patients with ONFH, C allele of rs662799 and G allele of rs3135506 in apoA5 gene were significantly higher than that in the control group, the T-C haplotype frequency was significantly lower than that in the control group (P <0.0001) and C-G haplotype was common in the control group. This is the first study to clarify the relation between ApoA5 polymorphism and ONFH.
The abnormal lipid metabolism and intravascular coagulation composed the main pathogenesis of osteonecrosis . Hyperlipidemia affected the microcirculation of the femoral head resulting in femoral necrosis from multiple links, such as affecting blood coagulation solvent systems, influencing bone fat embolism, and affecting the formation of bone micro-thrombosis ,. Many evidences showed that ApoA5 gene is associated with serum lipid levels . Recent findings indicate that ApoA5 could also influence cholesterol homeostasis and probably could play a role in hypertriglyceridemia . In our study, we found polymorphisms of ApoA5 gene are associated with TG levels. The carriers with mutant alleles have higher levels of TG in the ONFH patients. Therefore, we consider the mechanism of the association of ApoA5 polymorphisms with ONFH may resulting from lipids level changes caused by ApoA5 genetic polymorphism.
Although we found a positive association between ApoA5 polymorphisms and ONFH, the present study was limited by the relatively small sample size. This may have led to weak statistical significance and wide CIs when estimating odds ratios. In addition, we did not perform functional study of these two SNPs, which may be another limitation of our study.
In conclusion, this study showed that ApoA5 polymorphism may be associated with ONFH in Han Chinese population.
- Zhang H, Xiao F, Liu Y, Zhao D, Shan Y, Jiang Y: A higher frequency of peripheral blood activated B cells in patients with non-traumatic osteonecrosis of the femoral head. Int Immunopharmacol. 2014, 20 (1): 95-100. 10.1016/j.intimp.2014.02.016.PubMedView ArticleGoogle Scholar
- Li ZR: Scientific diagnosis and treatment of femoral head necrosis. Zhongguo Xiufu Chongjian Waike Zazhi. 2005, 19 (9): 685-686. (In Chinese)Google Scholar
- Gangji V, De Maertelaer V, Hauzeur JP: Autologous bone marrow cell implantation in the treatment of non-traumatic osteonecrosis of the femoral head: Five year follow-up of a prospective controlled study. Bone. 2011, 49 (5): 1005-1009. 10.1016/j.bone.2011.07.032.PubMedView ArticleGoogle Scholar
- Graham LS, Tintut Y, Parhami F, Kitchen CM, Ivanov Y, Tetradis S, Effros RB: Bone density and hyperlipidemia: the T-lymphocyte connection. J Bone Miner Res. 2010, 25 (11): 2460-2469. 10.1002/jbmr.148.PubMedPubMed CentralView ArticleGoogle Scholar
- Xiao Y, Cui J, Li YX, Shi YH, Wang B, Le GW, Wang ZP: Dyslipidemic high-fat diet affects adversely bone metabolism in mice associated with impaired antioxidant capacity. Nutrition. 2011, 27 (2): 214-220. 10.1016/j.nut.2009.11.012.PubMedView ArticleGoogle Scholar
- Chen TZ, Xie SL, Jin R, Huang ZM: A novel lipoprotein lipase gene missense mutation in Chinese patients with severe hypertriglyceridemia and pancreatitis. Lipids Health Dis. 2014, 13: 52-10.1186/1476-511X-13-52.PubMedPubMed CentralView ArticleGoogle Scholar
- Martín-Campos JM, Julve J, Roig R, Martínez S, Errico TL, Martínez-Couselo S, Escolà-Gil JC, Méndez-González J, Blanco-Vaca F: Molecular analysis of chylomicronemia in a clinical laboratory setting: diagnosis of 13 cases of lipoprotein lipase deficiency. Clin Chim Acta. 2014, 429: 61-68. 10.1016/j.cca.2013.11.025.PubMedView ArticleGoogle Scholar
- Long S, Chen Z, Han Y, Christopher DM, Zhang C, Yang Y, Tian Y: Relationship between the distribution of plasma HDL subclasses and the polymorphisms of APOA5 in hypertriglyceridemia. Clin Biochem. 2013, 46 (9): 733-739. 10.1016/j.clinbiochem.2013.03.003.PubMedView ArticleGoogle Scholar
- Zhang X, Qi Q, Bray GA, Hu FB, Sacks FM, Qi L: APOA5 genotype modulates 2-y changes in lipid profile in response to weight-loss diet intervention: the Pounds Lost Trial. Am J Clin Nutr. 2012, 96 (4): 917-922. 10.3945/ajcn.112.040907.PubMedPubMed CentralView ArticleGoogle Scholar
- Di Taranto MD, Staiano A, D’Agostino MN, D’Angelo A, Bloise E, Morgante A, Marotta G, Gentile M, Rubba P, Fortunato G: Association of USF1 and APOA5 polymorphisms with Familial Combined Hyperlipidemia in an Italian population.Mol Cell Probes 2014, doi: 10.1016/j.mcp.2014.10.002. [Epub ahead of print].,Google Scholar
- De Castro-Orós I, Cenarro A, Tejedor MT, Baila-Rueda L, Mateo-Gallego R, Lamiquiz-Moneo I, Pocoví M, Civeira F.: Common Genetic Variants Contribute to Primary Hypertriglyceridemia without Differences between Familial Combined Hyperlipidemia and Isolated Hypertriglyceridemia.Circ Cardiovasc Genet. 2014 Aug 30. [Epub ahead of print] PMID: 25176936 [PubMed - as supplied by publisher],Google Scholar
- Ariza MJ, Sánchez-Chaparro MA, Barón FJ, Hornos AM, Calvo-Bonacho E, Rioja J, Valdivielso P, Gelpi JA, González-Santos P: Additive effects of LPL, APOA5 and APOE variant combinations on triglyceride levels and hypertriglyceridemia: results of the ICARIA genetic sub-study. BMC Med Genet. 2010, 11: 66-10.1186/1471-2350-11-66.PubMedPubMed CentralView ArticleGoogle Scholar
- Kraja AT, Province MA, Straka RJ, Ordovas JM, Borecki IB, Arnett DK: Fenofibrate and metabolic syndrome. Endocr Metab Immune Disord Drug Targets. 2010, 10 (2): 138-148. 10.2174/187153010791213047.PubMedView ArticleGoogle Scholar
- Hirata T, Fujioka M, Takahashi KA, Arai Y, Asano T, Ishida M, Kuribayashi M, Akioka K, Okamoto M, Yoshimura N, Satomi Y, Nishino H, Fukushima W, Hirota Y, Nakajima S, Kato S, Kubo T: ApoB C7623T polymorphism predicts risk for steroid-induced osteonecrosis of the femoral head after renal transplantation. J Orthop Sci. 2007, 12: 199-206. 10.1007/s00776-007-1110-9.PubMedView ArticleGoogle Scholar
- Wang XY, Niu XH, Chen WH, Lin N, Song JN, Chen B, Jin H: Effects of apolipoprotein A1 and B gene polymorphism on avascular necrosis of the femoral head in Chinese population. Zhongguo Gu Shang. 2008, 21 (2): 99-102.PubMedGoogle Scholar
- Yin JM, Zhao L, Zhao SC, Guo YJ, Liu ZT, Liu ZT: Relationship between the Apolipoprotein AI, B gene polymorphism and the risk of non-traumatic osteonecrosis. Lipids Health Dis. 2014, 13 (1): 149-10.1186/1476-511X-13-149.PubMedPubMed CentralView ArticleGoogle Scholar
- Mont MA, Hungerford DS: Non-traumatic avascular necrosis of the femoral head. J Bone Joint Surg Am. 1995, 77 (3): 459-474.PubMedGoogle Scholar
- Oinuma K, Harada Y, Nawata Y, Takabayashi K, Abe I, Kamikawa K, Moriya H: Osteonecrosis in patients with systemic lupus erythematosus develops very early after starting high dose corticosteroid treatment. Ann Rheum Dis. 2001, 60 (12): 1145-1148. 10.1136/ard.60.12.1145.PubMedPubMed CentralView ArticleGoogle Scholar
- Koo KH, Kim R, Kim YS, Ahn IO, Cho SH, Song HR, Park YS, Kim H, Wang GJ: Risk period for developing osteonecrosis of the femoral head in patients on steroid treatment. Clin Rheumatol. 2002, 21 (4): 299-303. 10.1007/s100670200078.PubMedView ArticleGoogle Scholar
- Mont MA, Jones LC, Hungerford DS: Nontraumatic osteonecrosis of the femoral head: ten years later. J Bone Join t Surg(Am). 2006, 88 (5): 1117-1132. 10.2106/JBJS.E.01041.View ArticleGoogle Scholar
- Ouatou S, Ajjemami M, Charoute H, Sefri H, Ghalim N, Rhaissi H, Benrahma H, Barakat A, Rouba H: Association of APOA5 rs662799 and rs3135506 polymorphisms with arterial hypertension in Moroccan patients. Lipids Health Dis. 2014, 13: 60-10.1186/1476-511X-13-60.PubMedPubMed CentralView ArticleGoogle Scholar
- Hubacek JA, Peasey A, Kubinova R, Pikhart H, Bobak M: The association between APOA5 haplotypes and plasma lipids is not modified by energy or fat intake: the Czech HAPIEE study. Nutr Metab Cardiovasc Dis. 2014, 24 (3): 243-247. 10.1016/j.numecd.2013.08.008.PubMedPubMed CentralView ArticleGoogle Scholar
- Horvatovich K, Bokor S, Baráth A, Maász A, Kisfali P, Járomi L, Polgár N, Tóth D, Répásy J, Endreffy E, Molnár D, Melegh B: Haplotype analysis of the apolipoprotein A5 gene in obese pediatric patients. Int J Pediatr Obes. 2011, 6 (2–2): e318-e325. 10.3109/17477166.2010.490268.PubMedView ArticleGoogle Scholar
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