Protein restriction in pregnancy is associated with increased apoptosis of mesenchymal cells at the start of rat metanephrogenesis

Abstract

Background: In rats, offspring born to mothers supplied low protein diets during pregnancy have fewer glomeruli than normal. We hypothesized that such nephron deficits are associated with altered cell turnover in the metanephros, the embryonic precursor of the adult kidney.

Methods: Wistar rats were supplied with one of three isocaloric diets from day 0 of pregnancy: control (18% protein) or low protein (9% or 6%) diets. All had a normal chow after birth. Groups were compared by multilevel statistical modeling.

Results: At two weeks postnatally, when nephrogenesis has finished, controls had 16.8 x 103 +/- 0.7 x 10(3) (mean +/- SEM) glomeruli/kidney, whereas offspring exposed to 9% diet had 5.1 x 10(3) +/- 1.2 x 10(3) fewer and those exposed to 6% diet had 6.9 x 10(3) +/- 1.7 x 10(3) fewer glomeruli/kidney (P < 0.001, both diets). At embryonic day 13 (E13), when the metanephros has just formed, control metanephroi contained 2.35 x 10(4) +/- 0.15 x 10(4) cells, with no significant differences in low protein groups. At E15, when mesenchyme begins forming primitive nephrons but glomeruli are still absent, controls had 2.00 x 10(6) +/- 0.13 x 10(6) cells. E15 embryos exposed to 9% protein had 1.09 x 10(6) +/- 0.36 x 10(6) fewer cells/metanephros than controls, while those exposed to 6% diet had 1.45 x 10(6) +/- 0.37 x 10(6) fewer (P < 0.01, both diets). Apoptotic cells were detected by molecular (in-situ end-labeling) and morphological (propidium iodide staining) techniques. In all diets, apoptosis was noted in condensing mesenchyme (nephron precursors) and loose mesenchyme (interstitial precursors). Control E13 metanephroi had 63 +/- 7 apoptotic cells/mm2, whereas those exposed to 9% diet had an increase of 77 +/- 26 cells/mm2 (P < 0.01) and those exposed to 6% diet had an increase of 55 +/- 26 cells/mm2 (P < 0.05). By E15, apoptosis was similar in all groups but metanephric mitosis was significantly increased in the 6% protein diet group. No change was found in the level of apoptosis in E13 mesonephroi.

Conclusions: Maternal low protein diets reduce final numbers of glomeruli in association with enhanced deletion of mesenchymal cells at the start of kidney development. Whether aberrant nephrogenesis is a direct effect from deletion of nephron precursors, or an indirect effect from loss of supportive interstitial precursors, requires further investigation.