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Exogenous BMP-4 amplifies asymmetric ureteric branching in the developing mouse kidney in vitro
Jason E. Cain, Thibauld Nion, Dominique Jeulin, John F. Bertram Kidney International Volume 67, Issue 2, Pages (February 2005) DOI: /j x Copyright © 2005 International Society of Nephrology Terms and Conditions
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Figure 1 Effects of recombinant human bone morphogenetic protein 4 (rhBMP-4) on mouse metanephroi in culture. Phase-contrast photomicrographs of whole kidneys at the commencement of culture in either control media (A) or media supplemented with 260 ng/mL rhBMP-4 (B). The same metanephroi after 48 hours culture in control media (C) or media supplemented with 260 ng/mL rhBMP-4 (D). The kidney cultured with exogenous BMP-4 is smaller and contains fewer branches. Calbindin-D28K immunostaining shows typical metanephric ureteric branching and patterning (E). In the metanephros cultured with rhBMP-4, ureteric branching is severely reduced, predominantly in the posterior region (F) (same orientation, bar = 500 μm). Abbreviations are: a, anterior; p, posterior. Kidney International , DOI: ( /j x) Copyright © 2005 International Society of Nephrology Terms and Conditions
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Figure 2 Effects of recombinant human bone morephogenetic protein (rhBMP-4) on renal symmetry in vitro. Whole metanephroi from embryos of different weights (A to C), from smallest to largest weight embryo, cultured for 48 hours in 260 ng/mL rhBMP-4. Calbindin-D28K immunostaining reveals a marked asymmetry involving a severe reduction of ureteric branching morphogenesis in the posterior region (same orientation, bar = 250 μm). Abbreviations are: a, anterior; p, posterior. Kidney International , DOI: ( /j x) Copyright © 2005 International Society of Nephrology Terms and Conditions
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Figure 3 Three-dimensional image analysis of 48-hour cultured kidneys in either control media (CON), bovine serum albumin (BSA) control, or 260 ng/mL recombinant human bone morphogenetic protein 4 (rhBMP-4)(N = 10/group). (A) Total ureteric length in metanephroi cultured in rhBMP-4 is approximately 40% less than in those cultured in control or BSA control media. (B) Similarly, the number of ureteric branches in metanephroi cultured in rhBMP-4 is approximately 45% less than in the control and BSA control media cultures. *P < compared to both other groups. Kidney International , DOI: ( /j x) Copyright © 2005 International Society of Nephrology Terms and Conditions
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Figure 4 Three-dimensional image analysis of kidneys cultured for 48 hours in either control media (CON), bovine serum albumin (BSA) control, or 260 ng/mL recombinant human bone morphogenetic protein 4 (rhBMP-4)(N = 10/group). (A) Distributions of branch lengths are similar between the groups with most branches 50 to 99 μm in length. (B) The total number of branches less than 200 μm long is markedly reduced in metanephroi cultured in the presence of BMP-4. (C) Contribution of specific branch lengths to total length of the ureteric tree is also similar between the three groups. These results suggest that the reduction in branching morphogenesis is the result of a slowing of the growth rate of the ureteric tree rather than the adoption of an alternative branching pattern. Kidney International , DOI: ( /j x) Copyright © 2005 International Society of Nephrology Terms and Conditions
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Figure 5 Three-dimensional image analysis of metanephroi cultured for 48 hours in the presence of 260 ng/mL recombinant human bone morphogenetic protein 4 (rhBMP-4). (A) Total ureteric length in the posterior region of metanephroi cultured in rhBMP-4 was approximately 60% less than in the anterior region. (B) Similarly, the posterior region of metanephroi cultured with rhBMP-4 contained approximately 65% fewer branches than did the anterior region. *P < Kidney International , DOI: ( /j x) Copyright © 2005 International Society of Nephrology Terms and Conditions
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Figure 6 Three-dimensional image analysis of metanephroi cultured for 48 hours in control media. (A) Total ureteric length in the posterior region of metanephroi cultured in control media was approximately 40% less than in the anterior region. *P < (B) Similarly, the posterior region of metanephroi cultured in control media contained approximately 30% fewer branches than did the anterior region. *P < 0.05. Kidney International , DOI: ( /j x) Copyright © 2005 International Society of Nephrology Terms and Conditions
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Figure 7 Analysis of in vivo ureteric branching morphogenesis of E11.5 Hoxb7/GFP embryos. (A) In vivo kidneys at embryonic day (E) E11.5 showing a single branching event consisting of an anterior and shorter posterior branch. (B) The posterior branch is approximately 50% shorter than the anterior branch (bar = 100 μm). Abbreviations are: a, anterior; p, posterior. *P < (N = 6). Kidney International , DOI: ( /j x) Copyright © 2005 International Society of Nephrology Terms and Conditions
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