Sympathetic Reinnervation is Associated With Antihypertensive Treatment Burden and Interstitial Fibrosis After Kidney Transplantation

Figures

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  Figure 1. Representative case of a 25-year-old male recipient with IgA nephropathy who received an ABOincompatible kidney transplant from his mother, categorized into the sympathetic nerve regeneration group (Group 1). At 1-hour post-transplantation, numerous TH-positive sympathetic nerve bundles were observed along arteries and around Bowman’s capsules (A–C). At 3 months, no TH-positive fibers were detected in perivascular or periglomerular areas, despite the presence of large arteries, indicating complete denervation (D–F). By 1 year, TH-positive fibers had reappeared around Bowman’s capsules, confirming sympathetic reinnervation, which persisted at 3 years (G–I). Scale bars = 100 μm.

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  Figure 2. Tyrosine hydroxylase (TH) immunostaining results from serial renal allograft biopsies (0 h, 1 h, 3 months, 1 year, 3 years, 5 years, and 10 years) in 50 kidney transplant recipients. Each horizontal line represents 1 patient, and red “+” and blue “–” symbols indicate TH-positive and TH-negative findings, respectively. Sympathetic nerve regeneration was histologically confirmed in 23 recipients (Group 1: regeneration group), absent in 6 (Group 2: non-regeneration group), and indeterminate in 21 due to persistent TH positivity from early post-transplant biopsies (Group 3: histologically non-evaluable group). These categories were used for subsequent morphometric, clinical, and transcriptomic analyses.

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  Figure 3. Quantitative analysis of interstitial fibrosis (upper panel) and sympathetic nerve regeneration rates (lower panel) in serial renal allograft biopsies from Group 1: sympathetic nerve regeneration group (n = 23), Group 2: non-regeneration group (n = 6), and Group 3: histologically non-evaluable group (n = 21). Fibrosis area was calculated as the proportion of Sirius Red–positive cortical area (%). In the regeneration group, fibrosis increased significantly from 7.78% ± 2.84% at 1 year to 10.56% ± 3.12% at 3 years, whereas the non-regeneration group showed minimal change (6.21% ± 2.07% vs. 6.22% ± 1.98%). The non-evaluable group demonstrated intermediate fibrosis (6.82%± 2.31 at 1 year vs. 9.18± 2.85% at 3 years). The difference in fibrotic progression between the regeneration and non-regeneration groups was statistically significant (P = 0.037). Bar graphs below show the percentage of cases with TH-positive nerve fibers (“regeneration rate”) at each time point. Data are presented as mean ± SD. P values were calculated using the Kruskal–Wallis test with Holm-adjusted pairwise comparisons.

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  Figure 4. The mean number of antihypertensive drugs was evaluated at baseline, 3 months, 1 year, and 3 years after kidney transplantation in the Group 1: sympathetic nerve regeneration group (n = 23), Group 2: non-regeneration group (n = 6), and Group 3: histologically non-evaluable group (n = 21). All groups showed a transient decrease at 3 months, followed by a gradual increase thereafter. At 1 year, the Group 1 tended to require more antihypertensive agents than the Group 2 (∗P = 0.066). By 3 years, this difference became significant, with the Group 1 using significantly more agents than both the Group 2 (∗∗P = 0.012) and Group 3 (††P = 0.042). Data are expressed as mean ± SD. Statistical analysis was performed using the Kruskal–Wallis test followed by Holm-adjusted pairwise comparisons.

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  Figure 5. mRNA expression profiling was performed using the NanoString nCounter Human Fibrosis V2 Panel on representative 3-year biopsy specimens to evaluate fibrosis-associated transcripts. The volcano plot (left) displays differentially expressed genes between the Group 1: sympathetic nerve regeneration group (n = 4) and the Group 2: non-regeneration group (n = 4). Genes with |log 2 fold change| > 1 and P < 0.05 are highlighted (green, upregulated; purple, downregulated). Among 760 fibrosis-related genes, POSTN (periostin) and SERPINE1 (PAI-1) were significantly upregulated in the Group 1. The box plots (right) show normalized expression levels (log 2 scale) of these 2 genes, each dot representing an individual sample. Statistical comparison was performed using unpaired t-test.

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  Figure 6. mRNA expression profiling was performed on paired biopsy samples obtained from the same recipients in the Group 1: sympathetic nerve regeneration group at 3 months and 3 years after kidney transplantation. The volcano plot (left) shows differentially expressed genes between the 2 time points, with |log 2 fold change| > 1 and P < 0.05 considered significant (green, upregulated at 3 years; purple, downregulated at 3 years). Among fibrosis-related genes, FABP4 (fatty acid–binding protein 4) was significantly upregulated at 3 years, whereas CTGF (connective tissue growth factor) showed a modest increase. The box plot (right) demonstrates normalized expression levels of FABP4, confirming significant within-individual upregulation at 3 years compared to 3 months. Each dot represents a paired sample from the same individual. Statistical analysis was conducted using paired t-tests.