Single nucleus sequencing of human spinal dorsal and ventral horn

Study Purpose: The purpose was to provide a valuable resource of the cellular and molecular diversity in the human spinal cord.

Data Collection: We generated a high-resolution single-nucleus RNA sequencing (snRNA-seq) dataset from the lumbar spinal cords of 19 human donors. Each spinal cord was dissected into dorsal horn and ventral horn regions prior to sequencing, resulting in 38 anatomically distinct samples. Data processing and downstream analysis were performed using the Seurat package, enabling robust integration, clustering, and cell type annotation across samples. We identified eight major cell types consistently present across the dataset: neurons, lymphocytes, microglia, fibroblasts, oligodendrocytes, endothelial cells, ependymal cells, and astrocytes. Notably, eight of the 19 donors met our lab’s criteria for classification as pain donors.

Primary Conclusion: Comparative analysis revealed a unique subpopulation of lymphocytes localized exclusively to the dorsal horn of these pain-associated samples, suggesting a potential role in pain-related neuroimmune interactions. This dataset provides a valuable resource for exploring cellular and molecular diversity in the human spinal cord, particularly in the context of pain. We identified eight major cell types consistently present across the dataset: neurons, lymphocytes, microglia, fibroblasts, oligodendrocytes, endothelial cells, ependymal cells, and astrocytes.

Curator's Notes

Experimental Design: Neuronal nuclei were isolated from lumbar spinal cord tissue of 19 human donors for single-nucleus RNA sequencing (snRNA-seq). Each spinal cord was dissected into dorsal horn and ventral horn regions, resulting in 38 anatomically distinct samples. Tissue was microdissected into small pieces and homogenized using a tissue grinder with lysis buffer, followed by filtration and sucrose cushion centrifugation to purify intact nuclei. Nuclei were then fixed with chromium-compatible buffer for 18 hours at 4°C. Fixed nuclei were dual-stained with anti-NeuN-Alexa-647 antibody and 7-AAD for fluorescence-activated cell sorting (FACS). Double-positive neuronal nuclei were sorted from non-neuronal populations at low pressure, with compensation controls ensuring clear population separation. Data processing and downstream analysis were performed using the Seurat package, enabling robust integration, clustering, and cell type annotation across samples.

Completeness: This dataset is complete.

Subjects & Samples: Female (n=10), male (n=9) adult human patients (ages 20-63 years) were used in this study.

Primary vs. derivative data: Primary data is organized hierarchically by subject folders containing spinal cord sample folders subdivided into dorsal horn (DH) and ventral horn (VH) regions. Each anatomical region contains 10x Genomics Cell Ranger pipeline outputs with sequencing data mapped to the human genome (GRCh38). For eight samples with multiple sequencing runs, additional run folders (run004 and run008) separate duplicate sequencing attempts. Raw FASTQ files are not included. There is no derivative data folder.