Acute kidney injury (AKI) is a critical syndrome characterized by rapid decline in renal function. During the previous ten years, single-cell RNA sequencing (scRNA-seq) technology has evolved from a groundbreaking method to a fundamental tool in life science research. By integrating data from cellular spatial positioning with molecular omics, single-cell spatial omics offers unparalleled insight into how heterogeneous cells assemble and interact, illuminating complex networks governing cellular behavior in tissues and organs and uncovering dynamic processes of cell-cell communication. Single-cell sequencing and spatial transcriptomics have not only facilitated investigations of fundamental biological questions such as developmental differentiation but also enabled exploration of clinical issues such as cardiovascular diseases and tumor microenvironments, fundamentally transforming biomedical research paradigms. This review outlines the progression of sequencing methods, with emphasis on bulk RNAseq, scRNA-seq, and spatially resolved transcriptomics. Additionally, we review relevant published studies to discuss the ongoing cutting-edge applications of single-cell transcriptomic profiling coupled with spatial omics in AKI research, focusing on cellular heterogeneity, personalized treatment, and other related research directions. Finally, we discuss current constraints of scRNA-seq coupled with spatial omics in AKI research. From a multi-dimensional analysis perspective, we studied scRNA-seq and spatial transcriptomics, highlighting their critical role in advancing AKI understanding.