Gut microbiota and intervertebral disc degeneration: a bidirectional two-sample Mendelian randomization study

Low back pain (LBP), which affects a staggering 70–85% of individuals at some point in their lives, presents a significant global public health challenge, resulting in a considerable financial burden on healthcare and social systems [1–3]. In general, LBP refers to discomfort, tension or inflexibility that is felt in the region of the body situated beneath the ribcage and above the inferior gluteal folds, often accompanied by leg pain (sciatica) and other neurological issues affecting the lower extremities [1, 4]. While various factors can play a role in the development of LBP, intervertebral disc degeneration (IVDD) stands out as one of the primary causes [4, 5]. IVDD serves as the pathological foundation for various spinal degenerative disorders and is a prevalent orthopedic condition that contributes to a reduced quality of life [6]. The intervertebral disc (IVD) consists of the nucleus pulposus (NP), annulus fibrosus, and the cartilage endplate, which are primarily composed of collagen and proteoglycan, imparting crucial properties to the disc. The NP is a critical component of the IVD, primarily made up of NP cells and the extracellular matrix (ECM). Intervertebral disc degeneration (IVDD) is a prevalent degenerative condition that is distinguished by the gradual reduction of proteoglycans and water content within NP [7]. As the disease progresses, the discs between the vertebrae may break down, rendering them more susceptible to herniation, which can cause compression of the spinal nerves and nerve roots. The irritation of nerves in the lower back as a result of IVDD is known as lumbar radiculopathy. If this occurs in the nerve roots of L4-S2, it commonly results in a distinct type of pain known as sciatica [8, 9].

The gut microbiota (GM) refers to the distinct microbial populations that inhabit the intestinal tract and coexist in a mutually beneficial relationship with the host organism, including bacteria, protozoa, fungi, archaea, and viruses [10]. It has the potential to influence multiple physiological processes, including metabolism, inflammation, and immune responses [11–14]. The identification of gut microbiota taxonomic characteristics and their potential role is mainly based on the utilization of 16S rRNA and metagenomic sequencing methods, which are commonly employed techniques [15]. In a recent study by Rajasekaran et al. [16], a total of 24 lumbar intervertebral discs (IVDs) were analyzed, revealing that the microbial makeup present in healthy IVDs contrasted with that of degenerated and herniated IVDs. Changes in the composition of the microbiome and the way hosts respond to microbiota, which can cause abnormal bone growth and resorption [17, 18], gave rise to the idea of the gut-bone marrow axis [19, 20] and the gut-bone axis [18]. Subsequent to the study conducted by Rajasekaran et al., a comparable gut-disc axis concept has emerged that could have significant implications in intervertebral disc degeneration and low back pain [16, 21]. As a result, the regulation of gut microbiota could potentially impact the diversity and quantity of microbiota within the intervertebral disc, ultimately helping to regulate intervertebral disc degeneration.

However, additional investigation is required to further explore the distinct role of various gut microbiota taxa in the development of intervertebral disc degeneration. Akin to randomized controlled trials (RCT), the Mendelian randomization (MR) study is a recent research approach that investigates the causal relationship between exposure and outcome [22]. Mendelian randomization is a genetic epidemiology technique that uses single nucleotide polymorphisms (SNPs) that are known to affect modifiable exposures as instrumental variables (IVs) to deduce the causal effect of an exposure on an outcome. This approach is advantageous because it can eliminate confounding bias and can help to distinguish between the causal pathways of phenotypically grouped risk variables that are difficult to randomize or that are prone to measurement error [23]. In this study, we utilized GWAS summary statistics of GM and IVDD to perform MR analysis, with the aim of identifying GM taxa that may have a significant impact. This approach can help to confirm existing evidence and offer fresh perspectives on the management and prevention of intervertebral disc degeneration.

To the best of our knowledge, this is the first Mendelian randomization study investigating the causal relationship between gut microbiota (GM) and intervertebral disc degeneration (IVDD). In this study, we utilized GM data derived from a GWAS meta-analysis conducted by the MiBioGen consortium and IVDD data from the R8 release of the FinnGen consortium. The causal effects of gut microbiota taxa (from phylum to genus level) on intervertebral disc degeneration were investigated, and nine gut microbiota taxa were identified to have a causal association with IVDD. Additionally, we performed reverse analysis to demonstrate the unidirectionality of the causal relationships and found no evidence of IVDD affecting the gut microbiota.

There is a growing interest in understanding the pathogenic impact of the microbiome in numerous human diseases. Dysbiosis of the gut microbiota can potentially impair the normal functioning of the gut microbial community in maintaining host health. It may also lead to the selective enrichment of certain microbial members, including pathogenic bacteria, resulting in a dysregulated production of microbial-derived products or metabolites that can be harmful to the host. This dysregulation can contribute to the development of various diseases in local, systemic, or distant organs [44]. Similarly, it has been established that alterations in the composition of the gastrointestinal, skin, and oral microbiota are associated with various musculoskeletal diseases, such as rheumatoid arthritis [45–47], osteoarthritis [48, 49], ankylosing spondylitis [47], spondyloarthritis [50]. As one of the most common complaints in orthopedic clinics, low back pain is a prevalent public health issue worldwide, causing severe lifelong disability and imposing significant economic burdens on both patients and society [51]. Although the etiology of LBP is diverse [52–54], IVDD is widely acknowledged as a prominent factor among the various causes [4, 55], which is responsible for approximately 40% of symptomatic LBP [56]. In the past, the intervertebral discs in healthy individuals were believed to be sterile. However, Rajasekaran et al. [16] conducted a comprehensive metagenomic analysis of lumbar intervertebral discs, revealing the presence of a human intervertebral disc microbiome, and documenting the existence of “dysbiosis”. Furthermore, it was discovered that the gut and intervertebral disc microbiomes share a total of 58 bacterial species. In addition, the intervertebral disc (IVD) is a complex fibrocartilaginous joint and is often referred to as the largest avascular structure in the human body. Blood vessels in the IVD are only present in the longitudinal ligaments and the outer layers of the annulus fibrosis. However, the formation of new blood vessels can occur when the following conditions arise: intervertebral disc herniation into the extradural space, physical injury and fractures, or local inflammation on the intervertebral disc and vertebral endplates [57]. In the mouse model of IVDD, the abundance of Muribaculaceae and Lactobacillus increased, while the abundance of Clostridia_UCG-014 decreased. Furthermore, fecal microbiota transplantation further increased the abundance of Lactobacillus and reduced the abundance of Clostridia_UCG-014 [58]. Su et al. [59] conducted a Mendelian randomization study on the potential causal effects of specific gut microbiota and gut microbiota metabolites on low back pain (LBP). Similar to our findings, they observed that the abundance of the genus Marvinbryantia is a potential risk factor for LBP, while the abundance of the family Rikenellaceae and family Ruminococcaceae is a potential protective factor for LBP. These studies provide evidence for the association between gut microbiota and intervertebral disc degeneration (IVDD).

The pathogenic mechanisms underlying the role of gut microbiota in IVDD have been widely discussed in previous studies, involving various aspects such as inflammatory response, gut barrier function, and nutrient metabolism. These mechanisms intersect and collectively contribute to the overall impact of gut microbiota in IVDD. The Escherichia Shigella is a group of bacteria capable of producing lipopolysaccharide (LPS). LPS is a glycolipid component found in the outer membrane of gram-negative bacteria [60]. LPS can activate the TLR4/MyD88/NF-κB signaling pathway, leading to the release of pro-inflammatory mediators such as IL-6, IL-1β, and TNF-α. This cascade of events triggers a series of inflammatory processes, ultimately resulting in chronic low-grade inflammation [61]. A significant decrease in the abundance of a short chain fatty acids-producer, Marvinbryantia spp, was observed in the low muscle mass elders [62]. Sarcopenia is postulated to be an influential factor in chronic low back pain [63, 64]. The GM taxon may potentially impact intervertebral disc degeneration and trigger lower back pain by regulating nutrient absorption in the intestinal epithelium, thereby affecting muscle mass. Ruminococcus, originally classified as a member of the core phylum Firmicutes in the human gut microbiota, accounting for approximately 30% of the total gut microbial population [65], has been recently reclassified as Blautia [66]. Blautia is a genus of anaerobic bacteria that exhibits probiotic characteristics. It has the potential to reduce the production of local inflammatory factors and alleviate inflammatory responses in the gut. In normal intervertebral disc (IVD) samples, Firmicutes are abundantly present, and their abundance is associated with intestinal barrier function and antimicrobial protection [16]. Both Firmicutes and Bacteroidetes are capable of fermenting dietary fiber to produce short-chain fatty acids (SCFAs), which then act on G-protein-coupled receptors, such as GPR43, GPR41, and GPR109a,to regulate immune responses [67]. This leads to an increase in regulatory T cells (Tregs) and dendritic cell precursors, improvement in epithelial barrier function, and an upregulation of anti-inflammatory cytokines such as IL-10 [68]. The ratio of Firmicutes and Bacteroidetes has been implicated that some diseases like T2DM, hyperlipidemia [69], obesity [70], and non-alcoholic fatty liver disease (NAFLD) [71] are associated with elevated Firmicutes to Bacteroidetes ratio (F/B) in the gut microbiota. Our research, along with previous studies, demonstrates a strong association between Firmicutes, Bacteroidetes and intervertebral disc degeneration (IVDD). Thus, the F/B ratio could potentially be utilized as an evaluative indicator for assessing the association with IVDD. In mouse models of lumbar disc herniation, the gut was found to have a higher abundance of Ruminococcaceae [72], which can alleviate chronic inflammation through the production of butyrate [73]. However, it contradicts our study results to some extent, which suggest that this bacterium acts as a protective factor against IVDD. The reason for this discrepancy may be attributed to the complex interactions that exist among gut microbiota. Further prospective randomized controlled trials may be necessary to validate it. The Gordonibacter, as a probiotic, has been shown to alleviate inflammatory reactions and prevent the occurrence of various diseases [74]. It is known to produce anti-inflammatory metabolites, including urolithins. Urolithin A, in particular, has been found to improve mitochondrial health, reduce cartilage degeneration, and alleviate pain in osteoarthritis [75]. Studies have indicated that Urolithin A affects intervertebral discs through mechanisms that include inhibiting the breakdown metabolism of nucleus pulposus cells by TNFα in the body [76].Additionally, it increases mitochondrial autophagy and reduces cell apoptosis in the nucleus pulposus cells [77]. These findings suggest that Gordonibacter may have potential characteristics in regulating intervertebral disc health through the modulation of anti-inflammatory metabolites. Supplementation of vitamin D has been shown to impact the gut microbiota [78]. Importantly, vitamin D is closely associated with the gut barrier and can improve barrier function by inducing the expression of E-cadherin and enhancing epithelial cell connections within the intestine [79]. Deficiency in vitamin D can lead to disruption of these connections, resulting in intestinal permeability and facilitating the passage of bacterial components and metabolites. In patients with vitamin D deficiency and osteoarthritis, there is an increase in the abundance of Gordonibacter [80]. This particular microbial group may alter intestinal permeability, thereby creating conditions for the gut microbiota to influence the microecology of intervertebral discs. Yang et al. discovered that Oscillospira sp. constitutes a significant proportion of the gut microbiota. This suggests that Oscillospira sp. may play a crucial role in maintaining microbial balance and human health [81]. Importantly, its abundance has been found to be negatively correlated with a range of inflammatory diseases [82].

In the subsequent analysis after removing the instrumental variables associated with confounding factors, the analysis results for Eubacterium coprostanoligenes group, Rikenellaceae, and Lachnoclostridium no longer showed statistical significance.Limited research has been conducted regarding the association between Eubacterium coprostanoligenes group,Rikenellaceae, Lachnoclostridium, and IVDD. Therefore, whether there is a causal relationship between these three microbial taxa and IVDD remains to be debated.In conclusion,our research findings suggest that gut microbiota a dual role, both inducing and protective, in the occurrence and progression of IVDD.The specific mechanisms underlying these effects and the intricate interplay between gut microbial communities require further elucidation.

In previous studies, the etiology of intervertebral disc degeneration (IVDD) has mainly focused on various pathological factors, such as aging, inflammation, oxidative stress, mitochondrial dysfunction, and abnormal mechanical load [83–86]. However, there is limited research on the role of gut microbiota in IVDD. To the best of our knowledge, this study represents the first investigation of this topic using a two-sample bidirectional Mendelian randomization (MR) approach. MR utilizes genetic variations as instrumental variables to analyze the causal relationship between exposure and disease outcomes. By leveraging the random allocation of genetic variations, which occurs before the onset of disease and is independent of environmental factors, MR overcomes the inherent limitations of traditional observational studies, such as confounding factors and reverse causality. In this study, we employed available exposure and outcome data from genome-wide association studies (GWAS) to obtain precise analytical results. This approach provides an efficient means of harnessing reliable genetic information without the need for additional experimental costs. Furthermore, we meticulously examined the instrumental variables associated with the positive microbial results using the Phenoscanner V2 website to mitigate the influence of confounding factors and applied false discovery rate (FDR) correction to control the occurrence of false positives, thereby enhancing the reliability and reproducibility of the analysis results. Our research results provide the initial evidence of a causal association between the phylum Bacteroidetes and IVDD, thereby further confirming the concept of the gut-disc axis.

Our study provides insights into the relationship between specific GM taxa and intervertebral disc degeneration from a genetic perspective. By exploring potential genetic factors and the role of GM, our research contributes significantly to understanding the complex interplay between host genetics, GM, and intervertebral disc degeneration. Based on our findings, clinicians can innovate in the diagnosis and treatment of intervertebral disc degeneration in future research. Firstly, our study identifies potential therapeutic targets: by pinpointing genes and microbial taxa associated with intervertebral disc degeneration, we provide potential targets for drug development and interventions. This could lead to new treatment approaches and preventive measures aimed at modulating the genetic regulation of the microbiome or specific GM taxa to alleviate intervertebral disc degeneration. Secondly, personalized medicine approaches: the genetic insights from our research may facilitate the application of personalized medicine in managing intervertebral disc degeneration. Further investigations can explore the significance of individual-specific microbial and genetic characteristics in assessing the risk of developing intervertebral disc degeneration, allowing tailored treatment strategies. Thirdly, interdisciplinary research: our findings may inspire further collaboration between microbiologists, geneticists, and orthopedic experts. This interdisciplinary approach can provide new insights into the role of intervertebral disc degeneration and different GM taxa, bridging knowledge gaps between fields. Lastly, influencing lifestyle and environmental factors: the associations between specific microbial taxa and intervertebral disc degeneration can stimulate research on the impact of lifestyle factors (such as diet, exercise, and stress) on the GM and their potential effects on intervertebral disc health. Ultimately, evidence-based recommendations can be provided for patients with intervertebral disc degeneration. In conclusion, the genetic perspective highlighted in our study on the GM's role in intervertebral disc degeneration can guide new diagnostic and treatment strategies, support personalized medicine approaches, and foster interdisciplinary research collaboration. The results also underscore the importance of considering environmental and lifestyle factors that may influence the GM in the management of intervertebral disc degeneration.

Nevertheless, the study has several limitations. First, due to the use of a stringent genome-wide significance threshold (P < 5 × 10⁻⁸), the number of SNPs available for analysis was limited. Only SNPs meeting the suggestive significance threshold (P < 1 × 10⁻⁵) were included, which may reduce the reliability and accuracy of the results. Furthermore, the sample size and strain-level information of gut microbiota GWAS data are still in the early stages. This resulted in the exclusion of certain microbial species from the study, and the limited number of instrumental variables used may lead to a decrease in statistical power and increase the potential impact of weak instrumental variables on reverse Mendelian randomization, making it difficult to fully exclude reverse causality. It is essential to acknowledge that the study's data were exclusively obtained from European populations, without considering factors like gender and ethnicity, limiting the generalizability of the findings to other populations. Additionally, it is important to recognize that the study focused solely on bacteria, disregarding the considerable diversity of eukaryotic viruses and prokaryotic phages present in the human microbiota [87]. Further investigations are warranted to explore their potential involvement in IVDD. Finally, the association between human microbiota and the host in both healthy and disease states is a complex interplay rather than a simple one-way “causal relationship” [44]. Therefore, future studies should consider the intricate coordination and crosstalk between the host and gut microbiota to gain a better understanding of the relationship between gut microbiota and disease.

Using publicly available GWAS data, we conducted a bidirectional two-sample Mendelian randomization analysis on the causal association between 211 gut microbiota taxa and intervertebral disc degeneration (IVDD). Our analysis resulted in the identification of eight nominal causal associations and one strong correlation, further providing a theoretical foundation for the concept of the gut-disc axis. This study was based on a GWAS meta-analysis dataset generated from 16S rRNA sequencing, thus highlighting the need for analyses based on more advanced large-scale studies using metagenomics sequencing. Nevertheless, our research also offers valuable biomarkers for understanding the progression, diagnosis, and potential therapeutic approaches for IVDD.

Not applicable.

ZMG and JM contributed to conception and design of the study. ZMG, JW and GDC organized the database. ZMG and JW performed the statistical analysis. ZG wrote the first draft of the manuscript. JCL, JL and ZPZ wrote sections of the manuscript. All authors contributed to manuscript revision, read, and approved the submitted version. All author(s) read and approved the final manuscript. Jun Miao conceived and designed the study.

The study is supported by the National Natural Science Foundation of China (81472140), the Natural Science Foundation of Tianjin City (S20ZDD484) and Health and Technology Project of Tianjin Municipal Commission, Grant No. zc20219.

Please contact the corresponding author for data requests.