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Background <p>​ The S100 protein family constitutes a group of calcium-signaling proteins involved in numerous vital physiological and pathological functions. The enteric nervous system, often termed the second brain, regulates the maintenance of gastrointestinal motility, secretion, and immune functions. S100A4 is one member of this extensive family. To date, no reports exist concerning the distribution, cellular localization, or functional role of S100A4 within the gastrointestinal tract.</p> Methods <p>​ The double or triple labeling immunofluorescence technique was employed to investigate the localization and cytochemical characteristics of S100A4-positive neurons in the enteric nervous system. A semi-quantitative analysis was conducted on the changes in the number of S100A4 neurons in the gastrointestinal system of young and aged rats. Colchicine was intraperitoneally injected into aged rats to optimize the visualization of S100A4-positive neurons in their gastrointestinal system. The expression of S100A4 in the gastrointestinal tissues of young, aged, and colchicine-treated aged rats was studied using Western blotting and quantitative real-time reverse transcription PCR techniques.</p> Results <p>​ S100A4 was widely localized in neurons of the enteric nervous system (ENS). Specifically, all S100A4-immunoreactive cells in the myenteric and submucosal ganglia showed co-localization with HuD and calretinin. Partial co-localization was observed with choline acetyltransferase (CHAT) and calbindin, while only a minimal fraction of S100A4-positive cells co-existed with NF200-positive neurons. Notably, aging resulted in a significant reduction in S100A4-positive cell numbers within the ENS. However, the staining intensity of S100A4 markedly increased in nerve bundles and terminals. In contrast, the population of HuD-positive cells remained stable between young and aged rats. Following intraperitoneal injection of colchicine, aged rats exhibited a significant increase in S100A4-positive neurons in the ENS 24&#xa0;h post-treatment, reaching levels comparable to those in young rats. Quantitative real-time reverse transcription polymerase chain reaction(qRT-qPCR) and Western blot analysis revealed no significant changes in S100A4 mRNA and corresponding protein levels in the muscularis propria and submucosa of the colon, small intestine, and stomach between young and aged rats.</p> Conclusion <p>​ S100A4 is widely and specifically localized within neurons of the rat gastrointestinal nervous system, with all S100A4 neurons expressing calretinin. In aged enteric neurons, S100A4 is transported from the cell body to neuronal processes and nerve endings, indicating a dynamic redistribution rather than mere depletion during aging.</p>

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S100A4 expression in the enteric nerve system of young and aged rats

  • 期刊论文

  • 发表日期：

    2026年4月21日

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  • Fan Xiaoyi,
  • Han Yiman,
  • Xia Tianhao,
  • Huang Kesheng,
  • Xiang Zhenghua,
  • Yuan Hongbin

摘要

Background

​ The S100 protein family constitutes a group of calcium-signaling proteins involved in numerous vital physiological and pathological functions. The enteric nervous system, often termed the second brain, regulates the maintenance of gastrointestinal motility, secretion, and immune functions. S100A4 is one member of this extensive family. To date, no reports exist concerning the distribution, cellular localization, or functional role of S100A4 within the gastrointestinal tract.

Methods

​ The double or triple labeling immunofluorescence technique was employed to investigate the localization and cytochemical characteristics of S100A4-positive neurons in the enteric nervous system. A semi-quantitative analysis was conducted on the changes in the number of S100A4 neurons in the gastrointestinal system of young and aged rats. Colchicine was intraperitoneally injected into aged rats to optimize the visualization of S100A4-positive neurons in their gastrointestinal system. The expression of S100A4 in the gastrointestinal tissues of young, aged, and colchicine-treated aged rats was studied using Western blotting and quantitative real-time reverse transcription PCR techniques.

Results

​ S100A4 was widely localized in neurons of the enteric nervous system (ENS). Specifically, all S100A4-immunoreactive cells in the myenteric and submucosal ganglia showed co-localization with HuD and calretinin. Partial co-localization was observed with choline acetyltransferase (CHAT) and calbindin, while only a minimal fraction of S100A4-positive cells co-existed with NF200-positive neurons. Notably, aging resulted in a significant reduction in S100A4-positive cell numbers within the ENS. However, the staining intensity of S100A4 markedly increased in nerve bundles and terminals. In contrast, the population of HuD-positive cells remained stable between young and aged rats. Following intraperitoneal injection of colchicine, aged rats exhibited a significant increase in S100A4-positive neurons in the ENS 24 h post-treatment, reaching levels comparable to those in young rats. Quantitative real-time reverse transcription polymerase chain reaction(qRT-qPCR) and Western blot analysis revealed no significant changes in S100A4 mRNA and corresponding protein levels in the muscularis propria and submucosa of the colon, small intestine, and stomach between young and aged rats.

Conclusion

​ S100A4 is widely and specifically localized within neurons of the rat gastrointestinal nervous system, with all S100A4 neurons expressing calretinin. In aged enteric neurons, S100A4 is transported from the cell body to neuronal processes and nerve endings, indicating a dynamic redistribution rather than mere depletion during aging.