Nuclear Hormone Receptor Expression in Mouse Kidney and Renal Cell Lines

Nuclear hormone receptors (NHRs) are transcription factors that regulate carbohydrate and lipid metabolism, immune responses, and inflammation. Although several NHRs, including peroxisome proliferator-activated receptor-γ (PPARγ) and PPARα, demonstrate a renoprotective effect in the context of diabetic nephropathy (DN), the expression and role of other NHRs in the kidney are still unrecognized. To investigate potential roles of NHRs in the biology of the kidney, we used quantitative real-time polymerase chain reaction to profile the expression of all 49 members of the mouse NHR superfamily in mouse kidney tissue (C57BL/6 and db/m), and cell lines of mesangial (MES13), podocyte (MPC), proximal tubular epithelial (mProx24) and collecting duct (mIMCD3) origins in both normal and high-glucose conditions. In C57BL/6 mouse kidney cells, hepatocyte nuclear factor 4α, chicken ovalbumin upstream promoter transcription factor II (COUP-TFII) and COUP-TFIII were highly expressed. During hyperglycemia, the expression of the NHR 4A subgroup including neuron-derived clone 77 (Nur77), nuclear receptor-related factor 1, and neuron-derived orphan receptor 1 significantly increased in diabetic C57BL/6 and db/db mice. In renal cell lines, PPARδ was highly expressed in mesangial and proximal tubular epithelial cells, while COUP-TFs were highly expressed in podocytes, proximal tubular epithelial cells, and collecting duct cells. High-glucose conditions increased the expression of Nur77 in mesangial and collecting duct cells, and liver x receptor α in podocytes. These data demonstrate NHR expression in mouse kidney cells and cultured renal cell lines and suggest potential therapeutic targets in the kidney for the treatment of DN

Comparative expression levels of NHRs in renal cell lines under high-glucose conditions.

<p>Mesangial (MES13), podocyte (MPC), proximal tubular epithelial (mProx24), and collecting duct (mIMCD3) cell lines. Each panel displays NHR expression under high-glucose conditions compared with low-glucose conditions. Values depict the means ± SEM of three independent samples. HG, high-glucose. LG, low-glucose.</p

Orphan receptors expressed in renal cell lines.

<p>(A) Refer to the legend for <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0085594#pone-0085594-g003" target="_blank">Figure 3A</a> for details. (B) Representative photomicrographs of immunofluorescent staining. Chicken ovalbumin upstream promoter transcription factor II (COUP-TFII) was expressed in both MES13 and mProx24 cell lines.</p

Comparative expression levels of 49 nuclear hormone receptors (NHRs) in mouse kidney tissue and renal cell lines.

<p>The relative mRNA levels are depicted for mouse kidney (C57BL/6 (A) and db/m (B)) and mesangial (MES13) (C), podocyte (MPC) (D), proximal tubular epithelial (mProx24) (E) and collecting duct (mIMCD3) (F) cell lines. All values are expressed relative to 18S and arithmetically adjusted to depict the highest-expressed NHR for each tissue/cell line as a unit of 100. Values represent the means ± SEM of three independent samples of each tissue or cell line. Setting arbitrary cutoffs at Ct<31 (present) or Ct>31 (absent), as shown by broken lines in the C57BL/6 mouse kidney panel, reveals that 25 NHRs were expressed and six NHRs were not detected in C57BL/6 mouse kidney.</p

Adopted orphan receptors expressed in renal cell lines.

<p>(A) Refer to the legend for <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0085594#pone-0085594-g003" target="_blank">Figure 3A</a> for details. (B) Representative photomicrographs of immunofluorescent staining. Peroxisome proliferator-activated receptor-δ (PPARδ) was expressed in both MES13 and mProx24 cell lines.</p

Representative photomicrographs of double immunofluorescent staining in diabetic C57BL/6 mice.

<p>NOR1 expression was localized in mesangial, proximal tubular epithelial, and collecting duct cells, but not in podocytes in the kidneys of diabetic C57BL/6 mice.</p

Endocrine receptors expressed in renal cell lines.

<p>(A) The relative mRNA levels are depicted for mesangial (MES13), podocyte (MPC), proximal tubular epithelial (mProx24), and collecting duct (mIMCD3) cell lines. All values are expressed relative to 18S and arithmetically adjusted to depict the lowest-expressing sample as a unit of 1. Values represent the means ± SEM of three independent samples of each cell lines, and the results are representative of two independent studies. (B) Representative photomicrographs of immunofluorescent staining. Vitamin D receptor (VDR) was predominantly expressed in mProx24 cells, and to a lesser extent in MES13 cells.</p

Influence of dapagliflozin on physiologic and metabolic parameters in <i>db/db</i> and <i>db/m</i> mice at 20 weeks.

<p><i>db/m</i>, nondiabetic control mice; <i>db/db</i>, untreated diabetic mice; <i>db/db</i>+0.1 dapa, dapagliflozin (0.1 mg/kg)-treated diabetic mice; <i>db/db</i>+1.0 dapa, dapagliflozin (1.0 mg/kg)-treated diabetic mice; HbA1c, hemoglobin A1c. Data are presented as mean ± SEM;</p>a<p><i>P</i><0.05 vs. <i>db/m</i>,</p>b<p><i>P</i><0.05 vs. <i>db/db</i>,</p>c<p><i>P</i><0.05 vs. <i>db/db</i>+0.1 dapa.</p