Introduction

According to a 2019 annual report, over 178 million fractures occur universally, with more than 450 million pending patients with acute and chronic complications related to fractures every year.1 One of the more calcitrant complications is fracture nonunion (FNU), which occurs in about 15% of all fractures.2

It is estimated that about nine million FNU occurs yearly the world over and causes tremendous morbidity and clinical consequences.3 Many causes have been implicated in FNU, including host factors, diseases, site of fractures, mechanical, infection, and surgical failures.4–9 FNU provokes prolonged morbidity, decreased productivity, disability, impaired quality of life, and soaring healthcare costs. In Great Britain, the direct hospital cost of treating each FNU costs between £7000 and £79,000.10,11

It was accepted long before that individual genetic variations influence all human diseases, but it was difficult to identify specific genes and single nucleotide polymorphisms (SNP). In the past 20 years since the inception of the Human Genome Project and technological advances and analytical approaches, it can be pinpointed in the genetic variations that cause the specific disease process.12–17

In the last decade, few of the genes and SNPs that cause FNU have been described, which otherwise would have united. This systematic review aims to compile all the genes and SNPs associated with FNU to provide information to orthopedic surgeons, so they can confirm the genetic cause of FNU and take appropriate action.

Methods

Our review protocol was designed according to the Preferred Reporting Items of Systematic Reviews and Meta-Analysis for Protocols (PRISMA-P). The authors followed the current best practices for a systematic review and the protocol was registered in PROSPERO18 (ID395302).

Information Sources

An extensive literature search was designed and performed by a central medical librarian of the Imam Abdul Rahman Bin Faisal University, Dammam, for the concepts of nonunion of traumatic fractures and genes and SNPs. Pertinent publications were identified by searching the following databases with preset keywords: We searched between 2000 and July 2022 in PubMed, EMBASE, the Cochrane Central Register of Controlled Trials and Cochrane Database of Systematic Reviews, Genome Wide Association Studies (GWASs) Catalog, Embase (via Elsevier, 2 to present), MEDLINE, Web of Science Core Collection (via Thomson Reuters), including the Science Citation Index. The keywords for the search were nonunion of fractures, genetic influence, SNPs, and GWASs. Searches were limited to English language literature. The data was analyzed as soon as consensus was reached to finally include the full-text articles.

Inclusion and Exclusion Criteria

The inclusion criteria were all genetic studies reporting specific variants (ie, GWASs, SNPs, whole genome sequencing, whole exome sequencing) published in English literature with case cohorts of at least 100. The exclusion criteria were review articles and correspondence.

Data Extraction and Synthesis

The authors created the data in an Excel spreadsheet. They collected the pertinent clinical and demographic data, type of fractures studied, year of publication, size of cohorts, controls, genes and SNPs, nucleotide level, candidate genes, GWASs, targeted sequencing, whole genome and exome sequencing. The data was finally put to analysis when the authors agreed with the data and methodology used for the review.

Study Outcomes

The primary study outcome was the presence of a nonunion of the long bones. Analyses will include odds ratios and relative risk assessments, and subgroup analysis (ie, males vs females, age groups) was applied. The retrieved data was analyzed using SPSS Inc Version 26.

Results

A total of 79 studies on the nonunion of fractures were reviewed, with a focus on the genetic influence. After the inclusion and exclusion criteria were implemented, ten studies with 4402 patients’ data were analyzed. Nine studies were case controlled and 1GWAS. Figure 1 shows the PRISMA flow chart depicting the eliminated and the final ten in-depth studies examined. Table 1 gives the studies included in the analysis with the year of publication, SNPs identified, and the number of study subjects.

Table 1 List of All Published Data Analyzed

Figure 1 PRISMA flow chart of the review.

Notes: PRISMA figure adapted from Moher D, Shamseer L, Clarke M, et al. Preferred Reporting Items for Systematic Review and Meta-Analysis Protocols (PRISMA-P) 2015 statement. Syst Rev. 2015;4(1):1. Creative Commons.28

Three ethnic population groups were studied: Asians, Caucasians, and South Americans.

Table 2 gives the genes and SNPs strongly prone to developing FNUs. The table indicates that patients carrying these genes and SNPs are prone to get FNUs. Genes ANXA3 SNPs GSE95849, GSE93213, and GSE93215; BMP4 gene polymorphism rs17563, CYR61 polymorphism rs3753793; FGFR1 gene polymorphism rs1331 and IL1β gene polymorphism rs2853550; NOS2 gene polymorphisms rs2297514; rs2297514 and rs2248814; NOG gene polymorphisms rs1372857 and rs2053423, TLR2 rs5743708; and TACR1 gene with SNPs are located in Calcyon (CALY).

Table 2 Influencing Genes, SNPs and Ethnicities in the Review

Discussion

Our analysis covered ten publications and 4402 patients. It showed that specific genes and SNPs contribute to a higher risk of FNU. Three studies reported similar genes and SNPs.

Three genes located on chromosome 4: ANXA3 GSE95849, TACR1 rs229812 and TLR2 rs5743708; on chromosome 17, NOG rs2053423; and 2248814 and NOS2 rs2297514.

These SNPs have shown that in fracture or nonunion, not one but many genes and SNPs are involved. This should help surgeons dealing with nonunion that prove resistant to commonly used treatments consider genetic analysis to rule out the presence of genes and SNPs that influence nonunion.

Several factors have been found to contribute to the nonunion of fractures. These are due to the patient’s general health and secondly due to the type of fracture, locations, and biological and mechanical environment at the site.29–32 Recently, Mills et al (2016)4 reported that in their series of patients with proven nonunion, 75% of patients had multiple causes that could cause a nonunion. These results suggest that, in addition to genetic causes, there are other contributing factors to nonunion of fractures. There is no definitive treatment for nonunion fractures except to investigate and find the real cause or causes of contributing factors and to treat them. In many instances, despite all the efforts in implementing the available treatments, fractures do not heal even after treating the host factors. In some other patients without any known risk factors, fractures do not unite and develop into full-fledged nonunions.33

This review covers work published on how genes and their mutations and SNPs could cause or increase the risk of developing nonunion. There is ample evidence from the reported studies in the literature that reveals in FNU that genetics play a significant role. As per our review, different genes and SNPs influence different known factors that cause FNU. Szczesny et al (2011)20 were the first to highlight that subjects with a mutant TLR 4 gene are unable or have difficulty recognizing the infecting pathogens and eradicating them from the fracture site, causing nonunion. Dimitriou et al (2011)22 after a case–control study, showed that genes and SNPSs could cause nonunion by interfering with the BMP cascade, which is essential in the molecular and cellular regulation of fracture union. Other studies have shown how genes and SNPs can interfere with the various steps of normal fracture healing. The only GWAS in this review reported is that McCoy et al (2018)34 found that the TACR1 gene SNP rs2298122 is strongly associated with DNU, and the mode of the effect is related to pain sensitivity, which may interfere with fracture healing. In a recent review, Yan et al (2020)35 concluded that there are a number of genes and SNPs that cause nonunion of fractures and better understanding of this concept will allow to intervene early to heal fractures. Panteli et al (2022)36 further emphasized that there is ample evidence that patients with nonunion of fractures have genetic vulnerability and additional focused research is needed to identify biomarkers which can be utilized in the prediction of nonunion of fractures which also help in the development of innovative gene therapies.

The study has limitations due to the small number of studies and the simultaneous number of subjects in the analysis. Secondly, only three ethnic populations have been studied. Conversely, this review sheds light on the target genes with the study population of over 4000 patients screened. In conclusion, our review has implications in that it accentuates the importance of further studies with robust numbers, and the results of such studies will give definitive conclusions for the best clinical practice. To achieve this perspective, GWAS, with a large sample size, will shed light on which aspects of factors cause FNU to encourage caution in avoiding such complications.

Funding

No funding to report.

Disclosure

The authors report no conflicts of interest in this work.

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