Single-Cell Analysis Reveals Differences Between Aged and Young Mouse Prostates

Academic Background

The prostate is a vital component of the male reproductive system. As men age, the prostate undergoes hyperplasia, which can lead to urethral compression, resulting in voiding dysfunction and associated symptoms, as well as an increased risk of prostate cancer. Despite the importance of prostate aging, its molecular mechanisms remain incompletely understood. Single-cell RNA sequencing (scRNA-seq) has emerged as a powerful tool for uncovering cellular heterogeneity and gene expression changes during aging. Although numerous single-cell studies on the prostate have been conducted, they have not been systematically applied to characterize prostate aging in mice. Due to the difficulty in obtaining disease-free human prostate tissue, mice, which share genetic and physiological similarities with humans, serve as an alternative model for studying aging mechanisms.

Source of the Paper

This study was conducted by Yang Li, Yuhong Ding, and their team from Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, and was published in 2024 in the journal Biomarker Research. The research comprehensively mapped the cellular landscape of the mouse prostate using scRNA-seq and explored changes in specific cell types during aging.

Research Process

1. Animal and Tissue Isolation

The study used 2-month-old (young) and 24-month-old (aged) male C57BL/6J mice. Prostate tissues were isolated and divided into the anterior prostate (AP) and ventral/dorsal/lateral prostate (VLP).

2. Cell Dissociation

Prostate tissues were minced and transferred to a centrifuge tube containing tissue storage buffer. After enzymatic digestion and filtration, cells were resuspended in PBS.

3. Single-Cell RNA Sequencing Library Preparation and Sequencing

Cells were loaded into a microfluidic chip to generate droplets containing cell barcodes. cDNA amplification and library construction were performed, followed by sequencing on the Illumina NovaSeq 6000 system.

4. Data Processing and Quality Control

Raw data were quantified using Mobivision v3.0, and quality control was performed using Seurat software to remove low-quality cells. Batch effects were eliminated using Harmony.

5. Cell Type Identification and Differential Expression Analysis

Unsupervised clustering was performed using the UMAP algorithm to identify nine major cell types. Differential expression analysis was conducted to identify age-related gene expression changes.

6. Functional Enrichment Analysis and Cell-Cell Communication Analysis

Gene Ontology (GO) enrichment analysis was performed on differentially expressed genes, and cell-cell communication was analyzed using CellChat to reveal interactions between cells.

7. Western Blot and Immunofluorescence Staining

Western blot and immunofluorescence staining were used to validate the expression changes of certain genes.

Key Findings

1. Construction of Single-Cell Transcriptome Profiles

The study analyzed 68,214 cells and identified nine major cell types, including luminal cells, basal cells, mesenchymal cells, endothelial cells, lymphatic endothelial cells, myeloid cells, T cells, B cells, and plasma cells. The proportion of mesenchymal and immune cells significantly increased in aged mice.

2. Aging-Induced Molecular Changes

The study found significant changes in genes and pathways related to cellular senescence, oxidative stress, and regeneration in prostate cells from aged mice. Basal cells may undergo epithelial-mesenchymal transition (EMT) to become mesenchymal cells, particularly in aged mice.

3. Transcriptional Classification of Epithelial Cells

Epithelial cells were classified into five luminal cell subsets, two basal cell subsets, and one proliferative cell subset. The proportion of basal cell subsets increased in aged mice, while the proportion of proliferative cells decreased.

4. Transcriptional Classification of Mesenchymal Cells

Mesenchymal cells were classified into six fibroblast subsets, two smooth muscle cell subsets, and one pericyte subset. The proportion of certain fibroblast subsets significantly increased in aged mice.

5. Basal Cells Transform into Mesenchymal Cells via EMT

The study found that basal cells may transform into mesenchymal cells through EMT, particularly in aged mice. This process may play an important role in prostate aging and disease development.

6. Changes in Immune Cells

The proportion of myeloid cells, T/B cells, and plasma cells increased in aged mice, with macrophages and T cells exhibiting a stronger inflammatory response.

Conclusion

This study compared the transcriptomes of aged and young mouse prostates at single-cell resolution, revealing cellular and molecular changes. The proportion of mesenchymal and immune cells significantly increased in aged mice, and basal cells may transform into mesenchymal cells through EMT. Immune cells exhibited a stronger inflammatory response. These findings provide new insights into the molecular mechanisms of prostate aging and lay the foundation for developing therapeutic strategies for age-related prostate diseases.

Research Highlights

1. Single-Cell Resolution: The study revealed cellular and molecular changes in the mouse prostate during aging at single-cell resolution for the first time.
2. EMT Mechanism: Basal cells may transform into mesenchymal cells through EMT, a mechanism that plays an important role in prostate aging and disease development.
3. Role of Immune Cells: The proportion of immune cells increased in aged mice, with macrophages and T cells exhibiting a stronger inflammatory response, highlighting the role of the immune system in prostate aging.

Research Value

This study not only enhances our understanding of the molecular mechanisms of prostate aging but also provides a valuable foundation for identifying potential biomarkers and developing therapeutic strategies for age-related prostate diseases. By revealing the heterogeneity of different cell types in prostate aging, the study offers new directions for future functional research and therapeutic development.

Additional Valuable Information

The study also identified genes that were generally upregulated or downregulated in aged mouse prostates, such as Ly6a and SPINK1, which may play important roles in prostate aging and disease development. Additionally, the study highlighted the unique role of mesenchymal cells in prostate aging, particularly certain fibroblast subsets closely associated with immune responses.