Safeguarding intestinal stem cells during aging through balanced signaling

A recent study led by Associate Professor Takuya Yamamoto and Researcher May Nakajima-Koyama has revealed that maintaining a delicate balance between interferon-gamma (IFN-γ) and extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinase (MAPK) signaling is essential for preserving the intestinal stem cell population during aging. By comparing young and aged mouse intestinal tissues, the researchers uncovered critical insights into the interplay between these signaling pathways in supporting stem cell maintenance over time.

The intestinal epithelium exhibits the highest cell turnover rate in the adult body, requiring constant and precise regulation of intestinal stem cells (ISCs) to sustain the production of new absorptive enterocytes and secretory cells, including Paneth, goblet, tuft, and enteroendocrine cells. Driven by growth factors and cytokines derived from the stem cell niche and surrounding immune cells, multiple signaling pathways tightly regulate ISC self-renewal and differentiation. Despite aging-related changes in the intestinal environment, ISCs continuously generate new cells to maintain this high turnover throughout life, suggesting the existence of protective mechanisms that shield ISCs from microenvironmental fluctuations. However, the underlying mechanisms remain largely unknown.

To understand how aging influences ISCs, the research team examined intestinal tissues from young (2–4 months old) and aged (22–27 months old) mice, which have an average lifespan of approximately two years. Using mice with an Lgr5 reporter, a marker gene for ISCs, the researchers found that the ISC pool is maintained even with aging. Histological analyses further revealed that while the proliferative capacity of ISCs remains unchanged, the proliferative capacity of progenitor cells declines with age. Additionally, they observed an enhanced differentiation of ISCs into enteroendocrine cells, which produce gut hormones and play a key role in regulating whole-body metabolism.

To gain deeper insights into aging-related changes, the researchers analyzed the gene expression profiles of over 10,000 individual cells from young and aged intestines to examine their gene expression patterns at the single-cell level. Their analysis identified cell-type-specific, age-related alterations in gene expression, with notable changes in enterocyte maturation. Additionally, they observed an age-dependent upregulation of metabolic genes, suggesting that aged enterocytes may undergo enhanced maturation and metabolic adaptations compared to their younger counterparts. These findings imply that differentiated cells may be more susceptible to the effects of aging than ISCs.

To uncover the regulatory mechanisms underlying age-dependent cellular and transcriptomic changes in the intestinal epithelium, the researchers first identified aging markers to pinpoint upstream factors by using their expression as indicators. By performing an integrative analysis with public datasets, they identified MHC class II genes, Ceacam10, and Ly6e as aging markers for ISCs upregulated prominently during aging. Although the expression of these genes was elevated in aged ISCs, they did not correlate at the single-cell level, thus suggesting the involvement of multiple independent regulatory mechanisms.

Next, the researchers conducted an in silico analysis to predict upstream regulatory factors and identified several candidate signaling pathways that may contribute to the induction of aging markers. To validate the involvement of these pathways, they treated intestinal organoids—mini-intestines in culture—with bioactive molecules known to influence the candidate pathways. Their findings revealed that IFN-γ signaling activation and ERK/MAPK signaling inactivation contribute to age-related changes in the intestinal epithelium. Using mouse models, the researchers further confirmed that these signaling alterations occur in the small intestine with aging. However, the combined effects of these signaling changes on intestinal homeostasis remain to be fully elucidated.

To examine how changes in these pathways affect ISCs, the researchers treated intestinal organoids with IFN-γ and iMEK (an inhibitor of MEK/ERK MAPK signaling). Notably, iMEK induced a quiescent state in ISCs, which was reversed by IFN-γ treatment. Moreover, they demonstrated that combined iMEK and IFN-γ treatment mitigates organoid damage caused by either molecule alone, suggesting that these two signaling pathways function in a compensatory manner to support organoid growth and survival. Mechanistically, transcriptome analysis revealed that IFN-γ reversed most iMEK-induced global transcriptomic changes, partly by regulating Wnt/β-catenin signaling and the transcription factor Myc. Conversely, iMEK or IFN-γ treatment independently triggered age-related changes in enteroendocrine cells and enterocytes. These findings suggest that while the balance between IFN-γ and ERK/MAPK signaling is crucial for ISC maintenance, it may simultaneously drive age-related changes and dysfunctions in differentiated cells.

This study revealed that the interplay between ERK/MAPK and IFN-γ signaling plays a key role in intestinal epithelial aging. The findings suggest that the mammalian intestinal epithelium has evolved a signaling mechanism that prioritizes the preservation of ISCs, potentially at the expense of differentiated cells, which may contribute to age-related metabolic changes in the body. Restoring ERK/MAPK and IFN-γ signaling activities simultaneously to a more youthful state could represent a promising anti-aging therapeutic strategy with minimum impacts on ISCs.

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