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. 2021 Oct;32(10):2579–2594. doi: 10.1681/ASN.2020030263

Figure 7.

Figure 7.

Similar interference in myoglobin uptake in iMegKO and cilastatin-treated mice. To characterize the role of megalin in myoglobin endocytosis, iMegKO mice and controls were injected with FITC-myoglobin (0.5 mg). Then 15 minutes later, mice were killed and kidneys prepared for histologic examination. (A) Controls demonstrated abundant tubular epithelial FITC signal, primarily organized in punctae within the apical brush border and the adjacent cytoplasm, whereas (B), iMegKO mice demonstrated attenuated FITC-myoglobin signal, and far fewer punctae. (C) Mean fluorescence of all puncta (n=371,156) quantified by strain demonstrates that puncta from iMegKO mice exhibit reduced fluorescence compared with those from controls. Horizontal lines depict the mean. (D–F) To characterize the effect of cilastatin on myoglobin endocytosis, wild-type mice were injected with FITC-myoglobin (0.5 mg). Then 15 minutes later, mice were killed and kidneys prepared for histologic examination. Vehicle-treated mice exhibited abundant FITC-myoglobin punctae, whereas (D) cilastatin-treated mice exhibited fewer punctae and overall reduced FITC-fluorescence. (H) Mean fluorescence of all puncta (n=2,298,499) compared by drug treatment demonstrates that puncta from cilastatin-treated mice exhibit reduced fluorescence compared with those from vehicle-treated mice. Horizontal lines depict the mean. (G–I) To determine whether cilastatin interfered with overall renal myoglobin uptake, wild-type mice received vehicle or cilastatin injection 1 hour before glycerol intramuscular injection. (G, H) Then 2 hours after glycerol injection, myoglobin-directed immunofluorescence demonstrates abundant myoglobin-positive punctae at the brush border and within the cytoplasm of proximal tubules of vehicle injected mice, this signal appeared attenuated in tubules from cilastatin-injected mice. (I) Unbiased stereology demonstrates reduced total intrarenal myoglobin content in cilastatin-treated mice. Scale bars are 20 µm. Statistical analysis presented is derived from the t test.