Oxalobacter formigenes–Derived Bioactive Factors Stimulate Oxalate Transport by Intestinal Epithelial Cells

Abstract

Hyperoxaluria is a major risk factor for kidney stones and has no specific therapy, although colonization is associated with reduced stone risk. interacts with colonic epithelium and induces colonic oxalate secretion, thereby reducing urinary oxalate excretion, an unknown secretagogue. The difficulties in sustaining colonization underscore the need to identify the derived factors inducing colonic oxalate secretion. We therefore evaluated the effects of culture conditioned medium (CM) on apical C-oxalate uptake by human intestinal Caco-2-BBE cells. Compared with control medium, CM significantly stimulated oxalate uptake (>2.4-fold), whereas CM from did not. Treating the CM with heat or pepsin completely abolished this bioactivity, and selective ultrafiltration of the CM revealed that the -derived factors have molecular masses of 10-30 kDa. Treatment with the protein kinase A inhibitor H89 or the anion exchange inhibitor 4,4'-diisothiocyano-2,2'-stilbenedisulfonic acid completely blocked the CM-induced oxalate transport. Knockdown of the oxalate transporter SLC26A6 also significantly restricted the induction of oxalate transport by CM. In a mouse model of primary hyperoxaluria type 1, rectal administration of CM significantly reduced (>32.5%) urinary oxalate excretion and stimulated (>42%) distal colonic oxalate secretion. We conclude that -derived bioactive factors stimulate oxalate transport in intestinal cells through mechanisms including PKA activation. The reduction in urinary oxalate excretion in hyperoxaluric mice treated with CM reflects the retention of biologic activity and the therapeutic potential of these factors.

Microbial contributions to oxalate metabolism in health and disease

“…We present the first comprehensive study of human oxalate-degrading microbes, with multi-omics evidence, and quantitatively characterized their contributions. Our finding that multiple human gut microbes encode ODPs is consistent with multiple prior studies [22,43,64,80,81] [31,88,89] for direct bacterial access.…”

“…Multiple gut microbes can degrade oxalate [22], including Oxalobacter formigenes, an oxalate autotroph [23][24][25][26][27]. When given to animals, O. formigenes has reduced host oxalate burden [28][29][30][31][32][33]. [44][45][46].…”

The role of intestinal oxalate transport in hyperoxaluria and the formation of kidney stones in animals and man

“…A confounding issue is the oxalate-degrading activity of Oxalobacter enzymes and degradation of the 14 C-oxalate tracer used to measure intestinal fluxes together with the highly oxygenated buffers used in Ussing chamber experiments [7]. A number of intriguing recent reports have shown that exposure to Oxalobacter conditioned media (CM) at a 1:50 dilution for 6–24 h stimulates oxalate influx by Caco-2 cells and a > 4-fold increase in DRA mRNA expression, with no changes to DTDST or PAT1 [170, 171]. We repeated this experiment measuring unidirectional oxalate fluxes across Caco-2 monolayers in Ussing chambers using a 1:50 dilution (and also a 1:25 dilution; see Supplementary Table 1) of CM from the cultured human strain of Oxalobacter , HC-1.…”

“…6). In addition to these contrasting results, it is difficult to reconcile why Oxalobacter CM targets DRA [170, 171], and not PAT1, since the latter is the major oxalate transporter in Caco-2 cells [47], and given the respective contributions of PAT1 and DRA to oxalate secretion and absorption in mice [16–18]. It is also notable that Hassan et al reported no effect of CM from Lactobacillus acidophilus [170, 171], whereas several studies have previously shown that Lactobacillus significantly upregulates the functional expression of DRA in both Caco-2 and mouse models [172–174].…”

The role of the microbiome in kidney stone formation

“…While the responsible molecule has not yet been identified, culture media exposed to Oxf , administered rectally to knockout mice with primary hyperoxaluria type 1, reduced urinary oxalate excretion (>32.5%) and stimulated distal colonic oxalate secretion (>42%) in Ussing chamber-mounted bowel cross-sections [29]. The mechanism of action included activation of the SLC26A6 transporter by protein kinase A without any increased expression of the protein.…”