A neonatal rat model of progressive left ventricular pressure overload induced by abdominal aortic banding microsurgery

Figures

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  Neonatal ABS.

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  Figure 1. ABS procedure. A, Local anatomy at the site of the ABS. B, Schematic diagram of ABS. C, Key steps of ABS. C1, Expose the abdominal aorta. C2 and C3, A 12-0 suture needle was passed through the AA. The needle was held with a needle holder on the opposite side as it was passed through and then gently pulled until the suture thread was completely under the AA. C4 and C5, The padding needle was placed on the AA, and the AA and padding needle were ligated with 12-0 thread. Then, the padding needle was gently removed using a micro holder to create a lumen with a diameter equal to that of the needle. C6, The ends of the sutures were cut. ABS, Abdominal aortic banding surgery; AA, abdominal aorta.

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  Figure 2. Verification of LVPO. A, At postnatal day (P)7, a banding suture around the AA was visible under a microscope. B, Representative abdominal ultrasonography at the banding site at P21. C, Quantification of peak velocity at the banding site. The lower and upper borders of the box represent the lower and upper quartiles (25th percentile and 75th percentile). The middle horizontal line represents the median. The lower and upper whiskers represent the minimum and maximum values of nonoutliers. N = 6 rats. D, Systolic BP increased over time from postoperative day 21 to 35 because of the ABS. E, Diastolic BP increased over time from postoperative day 21 to 35 because of the ABS. F, Mean BP increased over time from postoperative day 21 to 35 because of the ABS. LVPO, Left ventricular pressure overload; AA, abdominal aorta; ABS, abdominal aortic banding surgery. ∗P < .05; ∗∗P < .01.

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  Figure 3. A progressive LVPO indicated by a gradually increasing number of DEGs from P3 to P7. A, Volcano plot of DEGs at P3. B, Volcano plot of DEGs at P7. C, Quantification of DEGs at P3 and P7. LVPO, Left ventricular pressure overload; DEG, differentially expressed gene; ABS, abdominal aortic banding surgery.

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  Figure 4. High reproducibility of ABS at the transcriptome level. A, Cluster analysis of DEGs at P3. B, Cluster analysis of DEGs at P7. ABS, Abdominal aortic banding surgery; DEG, differentially expressed gene.

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  Figure 5. Gene ontology (GO) analysis of upregulated DEGs. A, Cell proliferation-associated GO terms at P3. B, Cell proliferation–associated GO terms at P7. C, Heatmap of genes associated with cell proliferation at P3. D, Heatmap of genes associated with cell proliferation at P7. DEG, Differentially expressed gene.

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  Figure 6. More CMs were active in the cell cycle at P7 than at P3. A, Representative Ki67-positive CMs at P3 and P7. Ki67 (green); cTnT (red); DAPI (blue). B, Quantification of Ki67-positive CMs. C, Representative Aurora B-positive CMs at P3 and P7. D, Quantification of middle body Aurora B-positive CMs. E, Quantification of nuclear Aurora B-positive CMs. The lower and upper borders of the box represent the lower and upper quartiles (25th percentile and 75th percentile). The middle horizontal line represents the median. The lower and upper whiskers represent the minimum and maximum values of nonoutliers. N = 6 rats. CM, Cardiomyocyte; ABS, abdominal aortic banding surgery.

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  Figure E1. The surgical tools and materials for neonatal ABS. A, Surgical Items. B, Self-made tools required for neonatal ABS include padding needle, rubble band, and ice bed for operation. ABS, Abdominal aortic banding surgery.

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  Figure E2. ABS process. A, Longitudinal incision of approximately 1.5 to 2 cm was made at the lower midline of the abdomen. B, The small intestine was pulled to expose the AA. C, The suture needle was passed under the AA. D, The padding needle was placed and ligated. E, The small intestine was placed back in the abdomen. F, The abdomen was closed layer-by-layer. AA, Abdominal aorta; IVC, inferior vena cava.

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  Figure E3. Key biological terms enriched by ABS at different time points. A, Enriched GO terms at P3. B, Enriched GO terms at P7. ABS, Abdominal aortic banding surgery.

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  Figure E4. ABS induces an increase of LV thickness on postoperative day 35. A, Representative hematoxylin–eosin staining. B, Quantification of LV thickness. The lower and upper borders of the box represent the lower and upper quartiles (25th percentile and 75th percentile). The middle horizontal line represents the median. The lower and upper whiskers represent the minimum and maximum values of nonoutliers. N = 6 rats. ABS, Abdominal aortic banding surgery.