Urolithiasis is a common urinary disease with an increasing incidence worldwide, of which the occurrence varies among geographical regions and ethnicities. Kidney stone formation is reported to contribute to obesity, type 2 diabetes and hypertension (1–3). A previous study demonstrated a series of events leading to kidney stone formation, including oxalate/crystal-induced oxidative stress, tubular cell injury, inflammation and fibrosis (4). A positive family history predisposes an individual to nephrolithiasis, which suggests that genetic factors may have a critical role in the development of kidney stone formation (5).

Osteopontin (OPN), also known as or secreted phosphoprotein 1, is a secreted phosphoprotein expressed in various human body fluids, notably plasma, bile and urine (6). The OPN protein is synthesized within the kidney and is subsequently secreted into the urine by epithelial cells, including the papillary epithelium, distal convoluted tubule and loop of Henle (7). However, several studies have showed a controversial function of OPN, among which OPN could have a role in repressing the nucleation and aggregation of calcium oxalate crystals in vitro (8); however, OPN knockdown in the kidneys of hyperoxaluric rats could lead to reduction in renal calcium oxalate crystal deposition (9). OPN, a constituent of stone matrix, is also known as a specific monocyte chemoattractant to the renal interstitium (10). The expression of OPN mRNA is unregulated in stone-forming rats (11). In addition, OPN could promote the adherence of calcium oxalate crystals to Madin-Darby canine kidney cells (11). Furthermore, OPN could increase the formation and aggregation of calcium oxalate crystals in several experimental systems (12). Taken together, inhibitory and stimulatory roles in calcium stone formation have been proposed for OPN.

Single-nucleotide polymorphisms (SNPs) have been proposed as a worthwhile tool for identifying key genes associated with complex disease, such as those involved in urolithiasis (13). Polymorphisms in the OPN gene have been reported to exhibit functional implications and have been evaluated in several conditions (14), including the associations with autoimmune lymphoproliferative syndrome (15), type 1 diabetes (16), multiple sclerosis (17), systemic lupus erythematosus (18), rheumatoid arthritis (19,20) and Crohn's disease (21). However, the associations between OPN gene polymorphisms and urolithiasis are not widely reported (22,23), and have not been investigated in Chinese populations.

Considering OPN is linked to formation of calcium oxalate crystal deposition in the kidneys, we hypothesized OPN as a candidate gene linked to the genetic predisposition to develop nephrolithiasis. Additionally, the genetic variants in the promoter affecting the transcriptional level has attracted increasing attention; therefore in the present study, three SNPs (rs28357094 −170 T>G, rs11439060 −260 del/ins and rs11730582 −546 T>C) located on promoter of OPN were genotyped in a total of 480 individuals, including 230 nephrolithiasis patients and 250 matched healthy controls, and the associations between these SNPs and nephrolithiasis risk in different genetic models in the Chinese population were assessed.

Genetic variations are involved in the development and progression of nephrolithiasis (22). In this candidate gene study, polymorphisms in the OPN gene were associated with the presence of nephrolithiasis. A total of three polymorphisms were genotyped. This case-control study identified an association between a 1-base pair ins/del polymorphism rs11439060 (−/G) within the OPN promoter and nephrolithiasis susceptibility, and found that individuals with the rs11439060 variant genotypes (ins/del + ins/ins) had a significantly increased nephrolithiasis risk compared with the del/del genotypes.

OPN, an acidic phosphorylated glycoprotein, has been demonstrated to cause multiple biological processes such as the regulation of normal and abnormal calcification (12,24). An animal experiment revealed that overexpressed OPN could cause diabetic glomerulosclerosis (25). However, with regards to the urinary tract, OPN, mainly synthesized within the kidney and subsequently secreted into the urine, serves as a normal element for urine (7,26). Previous studies performed in cell lines have approved the inhibitory activities of OPN on aggregation, calcium crystal nucleation and adhesion to renal epithelial cells (8,27). Additional in vivo studies discovered that mice with OPN-knockdown were more susceptible to calcium urolithiasis compared to wild-type mice (28). Furthermore, emerging evidence has revealed that the products of the OPN gene, including mRNA and protein expression levels could be elevated and assembled in renal epithelial cells results of calcium oxalate crystals exposure (29,30), and the increased products of the OPN gene were considered to prevent urolithiasis formation (8,27). In addition, the imbalanced effect among the urinary-promoting/-inhibiting factors has been widely considered as more important in urinary stone formation compared to the disturbance of any other single substance (31).

Considering OPN has a crucial role in stone formation, polymorphism in the gene controlling the synthesis of OPN may significantly affect the risk of urolithiasis. The studies for the association between OPN genetic variants and the calcium urolithiasis risk are limited. There is an association of two common SNPs of OPN [non-synonymous amino acid located at positions +9402 (Arg/His) and +9171 (Asn/Ser) in exon 7] with calcium urolithiasis risk, and the SNP at position +9402 is significantly associated with the increased risk of calcium urolithiasis (32). Furthermore, a novel SNP at −156 (delG/G) of the OPN gene has been considered as a candidate genetic marker for calcium urolithiasis risk (33). Recently, Gao et al (34) have performed sequencing analysis on the entire OPN gene for the stone cases and normal controls of Japanese ancestry, and identified that two novel SNPs (−145T/G and −144G/T) resided at the OPN promoter region were also significantly associated with the risk of calcium urolithiasis. Additionally, Gögebakan et al (35) identified the significant association between the T-593A and C6982T polymorphism and nephrolithiasis risk in the Turkish population; however, the small sample size limited its extensive use as biomarkers. Even though the present study did not detect the expression of OPN, the findings show the notable association between SNP rs11439060 and the increased nephrolithiasis risk. It is biologically plausible that the allele of rs11439060 could affect the expression level of OPN, and contribute to the development of nephrolithiasis. Furthermore, the association between smoking and the nephrolithiasis risk was also identified, suggesting that there may be a risk effect of smoking on nephrolithiasis development and progression. In the subsequent stratified analysis, smokers with rs11439060 ins/del + ins/ins have a higher risk of nephrolithiasis, indicating the interaction effect of SNP rs11439060 and smoking on nephrolithiasis.

Considered the complex process for kidney stone formation, it is impossible that any single SNP or gene would exert a clear influence on nephrolithiasis risk. There was a limitation in the present study; it is unclear that aberrant expression of OPN is the cause or the consequence of crystal deposition. Certain studies have found the association between expression of OPN and development of nephrolithiasis in vivo, but the present results are not substantial in themselves to clarify the causal link between rs11439060 and nephrolithiasis risk. Although a strong correlation between rs11439060 and nephrolithiasis risk was observed, how genetic variability at this locus influences OPN function remains to be fully elucidated at functional levels. Additional large well-designed studies in multiple populations of different ethnicities and further functional studies are warranted.

In conclusion, the present study has provided the initial evidence that individuals carrying the SNP rs11439060 ins/del + ins/ins have a significantly increased nephrolithiasis risk compared with the del/del genotypes, indicating that the ins G allele was a deleterious effect on the nephrolithiasis. The results suggest that the SNP rs11439060 may serve as a potential marker to predict the nephrolithiasis risk in Chinese populations.