Patients with urolithiasis in our study exhibited several known risk factors for stone formation, such as increased urinary excretion of calcium and oxalate coupled with reduced urinary citrate excretion. Notably, the urinary supersaturation index was significantly elevated—approximately 3.8-fold higher in patients with stones compared to healthy individuals, underscoring it as a critical risk factor for urolithiasis. Echoing previous research, our investigation also identified gut dysbiosis in patients with urolithiasis, characterized mainly by an increased abundance of Bacteroidota. This contrasts with earlier reports that did not find significant changes in populations of Escherichia, Shigella, or Prevotella 11,12. The consistent rise in Bacteroidota abundance emerges as a hallmark of gut dysbiosis associated with urolithiasis.
Further, our study extended these findings to an animal model, where rats receiving fecal microbiota transplants from urolithiasis patients also showed a higher prevalence of Muribaculaceae of the phylum Bacteriodota compared to those receiving transplants from healthy controls. This parallel increase in Bacteroidota suggests that the fecal microbiota transplantation was effective in replicating the gut dysbiosis observed in human urolithiasis within the rat model. Our results lend support to the idea that gut dysbiosis could play a contributory role in the pathogenesis of urolithiasis, and they affirm the potential of FMT as a tool for modeling disease-associated microbiota alterations in experimental settings.
In our study, urolithiasis-FMT rats subsequently developed gut dysbiosis, exhibited specific disorders that are recognized as risk factors for calcium oxalate urolithiasis. These included elevated urinary oxalate levels, decreased urinary magnesium, an increase in urine pH, and a urinary supersaturation index that was approximately threefold higher compared to controls. These findings underscore the substantial influence of gut dysbiosis on the pathogenesis of urolithiasis, particularly implicating alterations in oxalate metabolism.
Oxalate in the body originates from two primary sources: dietary intake and endogenous synthesis. For the endogenous pathway, oxalate is a byproduct of the metabolism of various biomolecules, including ascorbic acid, L-hydroxy-lysine, and glyoxalate. In our rat model, we maintained a consistent diet across the board, which should standardize the production of endogenous oxalate. However, the absorption of exogenous, or dietary, oxalate may differ significantly due to gut dysbiosis.
It is well-established that gut dysbiosis can lead to increased intestinal epithelial permeability, a process often linked to the downregulation of tight junction proteins, pro-inflammatory responses, endotoxin production, and bacterial translocation 13. The proliferation of gram-negative bacteria in the gut, particularly Bacteroides spp., which constitutes the largest gram-negative phylum in the gut microbiota, correlates with elevated synthesis of endotoxins, especially lipopolysaccharides (LPS), thereby exacerbating gut permeability 14. Despite some Bacteroides species producing less endotoxin or even inhibiting toxin production from E. coli, the whole Bacteroidota phylum generates more endotoxin relative to other gram-positive bacteria, such as Firmicutes 15,16. In this context, we hypothesize that gut dysbiosis observed in urolithiasis could lead to compromised gut barrier integrity, enhancing the paracellular absorption of both LPS and oxalate. Our findings confirm this hypothesis, demonstrating that rats receiving FMT from urolithiasis patients not only exhibited gut dysbiosis but also showed reduced expression of the intestinal tight junction protein ZO-1 and increased expression of intrarenal NF-κB, indicative of potential gut leakage and consequent renal inflammation.
The chloride/oxalate transporter SLC26A6 is highly expressed on the renal epithelium, apical regions of the salivary glands and the villi of the small bowel, spanning from the duodenum through the jejunum and ileum, yet it is markedly reduced in the large intestine 17. The process of oxalate transport in the intestine is bidirectional, involving both net absorption—primarily through transcellular (via SLC26A1 and SLC26A3 transporters) and paracellular routes—and secretion, which is regulated by the SLC26A6 and, to a lesser extent, SLC26A2 transporters 18. The role of intestinal SLC26A6 is pivotal in facilitating the clearance of serum oxalate, thereby minimizing the oxalate load on the body 19. Evidence from previous research indicates that wild-type mice with intact intestinal SLC26A6 expression exhibit resistance to urolithiasis 20, whereas SLC26A6 knockout (KO) mice are prone to hyperoxalemia, hyperoxaluria, and the formation of kidney stones 21. The expression of SLC26A6 is modulated by various factors, including inflammation, microRNAs, and metabolites. A study by Liu Y. in 2021 highlighted that short-chain fatty acids such as acetate and propionate enhance the expression of intestinal SLC26A6 and reduced intestinal oxalate absorption 22. Recent research has shown that FMT rats with increased Muribaculaceae, Lactobacillus, and Bifidobacterium exhibited significantly lower urinary oxalate excretion and reduced calcium oxalate crystal deposition 23.
Our findings align with these observations, showing an upregulation of intestinal SLC26A6 expression. This upregulation may be attributed to either a high influx of oxalate or the influence of an increased abundance of Bacteroidota, particularly Muribaculaceae, a prominent propionate-producing bacterial family 24,25. The augmentation of the intestinal oxalate-secreting transporter SLC26A6 could represent an adaptive mechanism aimed at mitigating elevated serum oxalate levels resulting from enhanced intestinal absorption and gut leakage, thereby reducing the overall oxalate burden to the body.
Conclusively, our study reveals a critical link between gut microbiota dysbiosis and urolithiasis, emphasizing the importance of intestinal oxalate management and clearance mechanisms. Notably, an increased abundance of Bacteroidota in patients with calcium oxalate stone disease is considered a risk factor for urolithiasis. This is because gut dysbiosis correlates with intestinal leakage, which stimulates oxalate absorption and hyperoxaluria. Additionally, it leads to increased urinary supersaturation and renal inflammation (Fig. 5). Furthermore, the upregulation of the SLC26A6 transporter in the context of urolithiasis underscores potential compensatory responses to heightened oxalate levels, possibly influenced by Muribaculaceae prevalence. Addressing gut dysbiosis emerges as a strategic consideration for mitigating urolithiasis in high-risk individuals.