Morphogenesis during mouse embryonic kidney explant culture.

BACKGROUND

Renal organogenesis is routinely studied using cultured murine embryonic kidneys, but the application of this model has not yet been subjected to rigorous standards.

METHODS

We measured ex vivo growth and morphogenesis of day 13 murine kidneys and evaluated the importance of culture conditions and biological variables.

RESULTS

Kidney size was measured in two dimensions as planar surface area and was shown to correlate highly with volume (R2 = 0.60, P < 0.005). The final surface area of kidneys was directly dependent on the initial starting size (R2 = 0.61, P < 0.05), suggesting that the final surface area is not a valid outcome measurement unless starting size is equal among treatments. Relative growth rate, defined as (final surface area - initial surface area)/initial surface area, was a good measure of growth and independent of size and anatomical position (P> 0.05). Significant differences in size and growth rates were observed among litters (P < 0.05), implying that kidneys from a given litter must be randomized to avoid confounding results. Planar surface area of each explant increased in proportion to ureteric bud branching (R2 = 0.6854, P < 0.05). In a comparison of a variety of base media and supplements, kidney explants were observed to grow best in Dulbecco's modified Eagle's medium (DMEM)/F12 with 5% fetal bovine serum and to sustain growth for up to 96 hours, despite decreased proliferation and increased apoptosis at this time point.

CONCLUSIONS

These results represent an important step in establishing standardized procedures for the use of cultured embryonic kidneys and will improve our ability to apply the model to better understand kidney morphogenesis.