Scholarlink

Purpose <p>CYP27B1 is the rate-limiting enzyme in the activation of 1,25-dihydroxyvitamin D<sub>3</sub> [1,25(OH)<sub>2</sub>D<sub>3</sub>], the biologically active form of vitamin D<sub>3</sub>. This study aimed to characterize the spatial expression of CYP27B1 in mouse and human kidneys and to investigate the contribution of renal CYP27B1 to systemic vitamin D metabolism using inducible conditional knockout models.</p> Methods <p>Immunofluorescent co-staining with segment-specific markers was used to localize CYP27B1 in kidney tissue. Mice with selective <i>Cyp27b1</i> knockdown in renal tubular cells were generated by crossing <i>Cyp27b1</i><sup><i>lox/lox</i></sup> mice, in which exon 8 was flanked with loxP sites, with inducible <i>Pax8-Cre</i> or <i>Cdh16-Cre</i> lines.</p> Results <p>In mice, CYP27B1 immunoreactivity was detected in proximal and distal tubules under basal conditions but increased specifically in proximal tubules under elevated enzyme activity. In healthy human kidneys, CYP27B1 staining was confined to proximal convoluted tubules. <i>Cyp27b1</i><sup><i>lox/lox</i></sup> mice showed no overt phenotype, however, loxP insertion significantly reduced full-length <i>Cyp27b1</i> mRNA expression in the kidney and various other tissues, which in some cases was accompanied by a PTH-independent increase in incomplete <i>Cyp27b1</i> transcripts, as detected by exon 2–3 spanning primers. In <i>Pax8-Cre+;Cyp27b1</i><sup><i>lox/lox</i></sup> mice, doxycycline-induced <i>Cre</i> expression further reduced full-length <i>Cyp27b1</i> transcripts selectively in the kidney. In contrast, tamoxifen-induced deletion in <i>Cdh16-Cre+;Cyp27b1</i><sup><i>lox/lox</i></sup> mice was less effective and showed more off-target effects. Circulating 1,25(OH)₂D₃ levels remained stable, and calcium and bone homeostasis were preserved or minimally affected.</p> Conclusion <p>Kidney-selective <i>Cyp27b1</i> knockdown by inducible <i>Cre</i>-mediated recombination is insufficient to silence 1,25(OH)<sub>2</sub>D<sub>3</sub> synthesis, suggesting potent compensatory mechanisms that buffer loss of renal <i>Cyp27b1</i> transcription.</p>

错误：搜索内容不能为空，请输入英文关键词
错误：关键词超出字数限制，请精简
高级检索

Inducible kidney-selective knockdown of Cyp27b1 does not compromise calcium and bone homeostasis

  • 期刊论文

  • 发表日期：

    2026年2月18日

查看全文
  • Lieve Verlinden,
  • Stefanie Doms,
  • Iris Janssens,
  • Kayleigh Rillaerts,
  • René St-Arnaud,
  • Pieter Evenepoel,
  • Karel David,
  • Brigitte Decallonne,
  • Annemieke Verstuyf

摘要

Purpose

CYP27B1 is the rate-limiting enzyme in the activation of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3], the biologically active form of vitamin D3. This study aimed to characterize the spatial expression of CYP27B1 in mouse and human kidneys and to investigate the contribution of renal CYP27B1 to systemic vitamin D metabolism using inducible conditional knockout models.

Methods

Immunofluorescent co-staining with segment-specific markers was used to localize CYP27B1 in kidney tissue. Mice with selective Cyp27b1 knockdown in renal tubular cells were generated by crossing Cyp27b1lox/lox mice, in which exon 8 was flanked with loxP sites, with inducible Pax8-Cre or Cdh16-Cre lines.

Results

In mice, CYP27B1 immunoreactivity was detected in proximal and distal tubules under basal conditions but increased specifically in proximal tubules under elevated enzyme activity. In healthy human kidneys, CYP27B1 staining was confined to proximal convoluted tubules. Cyp27b1lox/lox mice showed no overt phenotype, however, loxP insertion significantly reduced full-length Cyp27b1 mRNA expression in the kidney and various other tissues, which in some cases was accompanied by a PTH-independent increase in incomplete Cyp27b1 transcripts, as detected by exon 2–3 spanning primers. In Pax8-Cre+;Cyp27b1lox/lox mice, doxycycline-induced Cre expression further reduced full-length Cyp27b1 transcripts selectively in the kidney. In contrast, tamoxifen-induced deletion in Cdh16-Cre+;Cyp27b1lox/lox mice was less effective and showed more off-target effects. Circulating 1,25(OH)₂D₃ levels remained stable, and calcium and bone homeostasis were preserved or minimally affected.

Conclusion

Kidney-selective Cyp27b1 knockdown by inducible Cre-mediated recombination is insufficient to silence 1,25(OH)2D3 synthesis, suggesting potent compensatory mechanisms that buffer loss of renal Cyp27b1 transcription.