Background: In the past decade, it has been established that dynamic communication exists among nerves, macrophages, and podocytes (PODO) within the kidney, which plays a crucial role in renal homeostasis and the response to acute and chronic renal injury. However, the exact molecular mechanisms of the interplay among these three elements in the progression of diabetic nephropathy (DN) remain to be elucidated. The aim of this study is to explore the intercellular communication via signaling molecules among neurons, macrophages, and PODO, and to determine their mechanism of action in renal inflammation in DN.

Methods: Multiple DN-related single-cell transcriptome sequencing datasets were merged. Using the Seurat pipeline, cells were standardized, underwent principal component analysis (PCA), dimensionality reduction, clustering, and annotation. Cells transmitting neural signals, macrophages, and PODO were identified using biomarkers from CellMarker 2.0 and SingleR method. The interactions of ligand-receptor pairs between these cells were analyzed using Celltalker and CellphoneDB. Additionally, differentially expressed genes (DEGs) in macrophages, and PODO in DN were obtained and annotated for Gene Ontology and Kyoto Encyclopedia of Genes and Genomes functions.

Results: A total of 2394 cells from kidneys exhibiting transcriptome characteristics highly similar to neurons were identified. These neuron-like cells were clustered into 11 cell clusters. Among these, cluster 7 (GABBR1^hiCHRM3^hi) showed a significant decrease in relative proportion in DN tissue (p < 0.05). A total of 202 macrophages were identified, clustered into 4 cell clusters. Notably, cluster C3 (RYR1⁺ESRRG⁺BICC1⁺) was significantly reduced in DN. Functional analysis indicated that RYR1⁺ESRRG⁺BICC1⁺ macrophages were associated with negative regulation of inflammatory responses. Cell communication analysis further revealed weakened interactions between GABBR1^hiCHRM3^hi neuron-like cells and RYR1⁺ESRRG⁺BICC1⁺ macrophages involving Amyloid β precursor protein (APP)-CD74. Additionally, interactions of PODO with RYR1⁺ESRRG⁺BICC1⁺ macrophages involving SLIT1-ROBO2, EFNA5-EPHA4, and PDGFB-PDGFRB were weakened, as was the interaction of GABBR1^hiCHRM3^hi neuron-like cells involving VEGFA-FLT1.

Conclusion: In DN, there is signal crosstalk between GABBR1^hiCHRM3^hi cells, anti-inflammatory phenotype macrophages, and PODO. The communication between these cells is primarily mediated through signaling pathways such as APP-CD74, PDGFB-PDGFRB, and VEGFA-FLT1.