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. 2011 Mar;300(3):G461-9.
doi: 10.1152/ajpgi.00434.2010. Epub 2010 Dec 16.

Enteric Oxalate Elimination Is Induced and Oxalate Is Normalized in a Mouse Model of Primary Hyperoxaluria Following Intestinal Colonization With Oxalobacter

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Enteric Oxalate Elimination Is Induced and Oxalate Is Normalized in a Mouse Model of Primary Hyperoxaluria Following Intestinal Colonization With Oxalobacter

Marguerite Hatch et al. Am J Physiol Gastrointest Liver Physiol. .
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Abstract

Oxalobacter colonization of rat intestine was previously shown to promote enteric oxalate secretion and elimination, leading to significant reductions in urinary oxalate excretion (Hatch et al. Kidney Int 69: 691-698, 2006). The main goal of the present study, using a mouse model of primary hyperoxaluria type 1 (PH1), was to test the hypothesis that colonization of the mouse gut by Oxalobacter formigenes could enhance enteric oxalate secretion and effectively reduce the hyperoxaluria associated with this genetic disease. Wild-type (WT) mice and mice deficient in liver alanine-glyoxylate aminotransferase (Agxt) exhibiting hyperoxalemia and hyperoxaluria were used in these studies. We compared the unidirectional and net fluxes of oxalate across isolated, short-circuited large intestine of artificially colonized and noncolonized mice. In addition, plasma and urinary oxalate was determined. Our results demonstrate that the cecum and distal colon contribute significantly to enteric oxalate excretion in Oxalobacter-colonized Agxt and WT mice. In colonized Agxt mice, urinary oxalate excretion was reduced 50% (to within the normal range observed for WT mice). Moreover, plasma oxalate concentrations in Agxt mice were also normalized (reduced 50%). Colonization of WT mice was also associated with marked (up to 95%) reductions in urinary oxalate excretion. We conclude that segment-specific effects of Oxalobacter on intestinal oxalate transport in the PH1 mouse model are associated with a normalization of plasma oxalate and urinary oxalate excretion in otherwise hyperoxalemic and hyperoxaluric animals.

Figures

Fig. 1.
Fig. 1.
Comparison of 24-h urinary oxalate excretion in wild-type (WT, n = 18 pools) mice and mice deficient in liver alanine-glyoxylate aminotransferase (Agxt, n = 11 pools) fed standard mouse chow. Hyperoxaluria in Agxt mice (*P < 0.05) is significantly increased when the mice (n = 5 pools) are fed 1.5% oxalate-0.5% calcium diet (§P < 0.05). In colonized Agxt mice that are also fed the oxalate-supplemented diet, urinary oxalate excretion (n = 6 pools) is significantly reduced compared with noncolonized Agxt mice fed the same diet (†P < 0.05) and not statistically different from WT mice fed the standard diet. OxWR, wild rat strain of Oxalobacter formigenes.
Fig. 2.
Fig. 2.
Unidirectional [mucosal-to-serosal (Jms) and serosal-to-mucosal (Jsm)] and net transepithelial fluxes of oxalate across isolated, short-circuited segments of cecum, proximal colon, and distal colon from noncolonized (n = 11 tissue pairs) and artificially colonized (n = 9 tissue pairs) Agxt mice fed a diet containing 1.5% oxalate-0.5% calcium. *Significant difference between the two groups. Transepithelial conductance was not affected by colonization of any segment (20.2 ± 1.2, 19.2 ± 0.9, and 14.9 ± 0.7 mS/cm2 across cecum, proximal colon, and distal colon, respectively). Compared with noncolonized segments, short-circuit current (Isc) was significantly higher in colonized Agxt cecum (from 0.9 ± 0.1 to 1.6 ± 0.3 μeq/cm2) and distal colon (from 3.2 ± 0.4 to 5.8 ± 0.7 μeq/cm2) but remained unchanged in proximal colon (from 2.2 ± 0.3 to 2.9 ± 0.4 μeq/cm2).
Fig. 3.
Fig. 3.
Normal 24-h urinary oxalate excretion in WT mice (n = 18 pools) fed standard mouse chow and WT mice (n = 8 pools) fed oxalate-supplemented diet (1.5% oxalate-0.5% calcium). Asterisk above the middle bar denotes a significant difference (P < 0.05) between these two groups. In colonized WT mice that are also fed the oxalate-supplemented diet (n = 10 pools), urinary oxalate is significantly reduced compared with the noncolonized group (§P < 0.05) and compared with WT mice fed the standard diet (*P < 0.05).
Fig. 4.
Fig. 4.
Unidirectional and net transepithelial fluxes of oxalate across isolated, short-circuited segments of cecum, proximal colon, and distal colon from noncolonized (n = 7 tissue pairs) and artificially colonized (n = 9 tissue pairs) WT mice fed 1.5% oxalate-0.5% calcium diet. *Significant difference between the two groups. Transepithelial conductance was not affected by colonization of any segment (19.9 ± 2.7, 24.1 ± 2.7, and 14.5 ± 1.5 mS/cm2, across cecum, proximal colon, and distal colon, respectively). Compared with the noncolonized segments, short-circuit current was not affected by colonization (3.1 ± 0.3, 4.6 ± 1.1, and 1.9 ± 0.3 μeq/cm2 in cecum, proximal colon, and distal colon, respectively).
Fig. 5.
Fig. 5.
Immunoblot analyses of slc26a6 in protein extracts of large intestine from colonized (Agxt + OxWR and WT + OxWR) and noncolonized (Agxt and WT) mice. Membranes were reprobed for GAPDH as an internal loading control. Results were quantified by densitometry from 4–12 tissues in each group. *Significant difference between noncolonized Agxt and noncolonized WT mice. §Significant difference between colonized and noncolonized Agxt mice. PAT-1, protein associated with topoisomerase II.
Fig. 6.
Fig. 6.
Temporal characterization of loss of Oxalobacter colonization in Agxt and WT mice. In the first series, 18 artificially colonized WT mice (○) and 22 artificially colonized Agxt mice (▾) were fed 1.5% oxalate-0.5% calcium diet prior to gavage (G) with OxWR. On G + 12 days, this diet was replaced with standard chow containing 1% calcium and no oxalate supplementation. In the next series, artificially colonized Agxt mice (n = 8, ●) were fed 1.5% oxalate-0.5% calcium diet prior to gavage with OxWR. On G + 12 days, this diet was replaced with chow containing 0.5% calcium and no oxalate supplementation. The last series involved naturally colonized Agxt pups (n = 11) fed 1.5% oxalate-0.5% calcium diet before and after the day of weaning (W). On W + 12 days, this diet was replaced with standard chow containing 1% calcium and no oxalate supplementation.

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