The Role of mRNA Metabolism Regulator Human Antigen R (HuR) in Age-Related Hearing Loss in Aged Mice

Geriatrics Bioengineering Basic Medical Sciences Cell Biology Biochemistry Genetics Life Sciences Medicine
mRNA metabolism HuR age-related hearing loss hair cells auditory function AAV gene therapy cochlea

mRNA Metabolism Regulator Human Antigen R (HuR) Regulates Age-Related Hearing Loss in Aged Mice — A Systematic Study of Molecular Pathological Mechanisms and Therapeutic Intervention

1. Academic Background and Scientific Questions

Age-related hearing loss (ARHL), also known as presbycusis, is one of the most common sensory disorders in the global elderly population. Statistics show that more than 25% of people over 60 years old suffer from hearing impairment, with the incidence of hearing loss rising rapidly with age, making it the third most common health issue after heart disease and arthritis. Moreover, ARHL not only affects individuals’ auditory perception, but is also closely associated with serious health problems such as cognitive decline, social isolation, depression, and dementia. Therefore, the prevention and treatment of ARHL have significant social and medical value.

On the pathological level, ARHL mainly manifests as hair cell (HC) loss, spiral nerve degeneration, and stria vascularis dysfunction. Hair cells are highly susceptible to age-related damage due to their high energy consumption and sensitivity to mechanical stimulation. However, lost hair cells in adult mammals cannot regenerate, and existing clinical interventions are limited—protecting hair cells from damage becomes the key to ARHL prevention and treatment.

In recent years, abnormal RNA metabolism has emerged as a new genetic regulatory mechanism closely related to a variety of aging-associated diseases. RNA-binding proteins (RBPs), as core regulators of RNA metabolism, can modulate RNA splicing, stability, and translation. Among the RBP family, HuR (Human antigen R, also known as ELAVL1) has been confirmed to play a key role in mRNA fate in various tissues, but its role and mechanisms in the development of ARHL remain unclear.

Addressing this issue, this study systematically reveals the expression dynamics, cellular localization, action mechanisms, and intervention potential of HuR in the auditory system of aged mice, based on high-throughput single-cell transcriptomics, gene knockout/knock-in, animal models, and molecular biology approaches.

2. Paper Source and Author Introduction

This paper, titled “mrna metabolism regulator human antigen r (hur) regulates age-related hearing loss in aged mice,” was published in May 2025 in the top international academic journal Nature Aging (nature aging, volume 5, may 2025, pages 848–867). The first authors include Siwei Guo, Jieying Cao, Guodong Hong, et al., with Jiangan Gao, Renjie Chai, and Xiaolong Fu as corresponding authors; the team comes from well-known domestic universities and medical research institutions including Shandong University, Soochow University, Southeast University, and Shandong First Medical University.

3. Research Process and Detailed Methods

1. Overall Study Design

This study systematically tracked HuR expression dynamics in cochlear cells during aging using multi-aged C57BL/6J and SAMP8 mice and a variety of gene knockout and knock-in models, and elucidated its molecular mechanisms, cellular consequences, and intervention potentials via behavioral, molecular, histological, functional, and transcriptomic analyses.

The main research workflow includes:

  • Single-cell transcriptome analysis revealing HuR expression dynamics and localization changes;
  • Knockdown/overexpression models to validate HuR’s effects on hair cell survival and hearing;
  • Exploration of molecular mechanisms, downstream targets, and regulatory networks of HuR;
  • Evaluation of gene therapy strategies of HuR and its major targets in ARHL intervention.

2. Single-Cell Transcriptomics and Protein Detection—Dynamic Expression and Localization

Using C57BL/6J mice, cochleae were collected at 1, 2, 5, 12, and 15 months of age; single-cell RNA sequencing (scRNA-seq) was used to map the transcriptome at cellular resolution. Dimensionality reduction (tSNE) and clustering uncovered a significant increase in HuR mRNA expression with age, with an upward trend in all ARHL-related cochlear tissues (hair cells, supporting cells, spiral ganglion cells, etc.). Further, paraffin-section immunofluorescence (IF) staining resolved the cellular localization of HuR protein, showing that in young mice, HuR is expressed inside the hair cell nuclei, whereas in older mice, it is found both in the nucleus and cytoplasm, suggesting age-dependent nuclear–cytoplasmic shuttling; other cochlear cell types had no significant localization shifts.

In addition, the team performed cross-species validation in non-human primate cochlear tissue, observing conserved hair cell–specific HuR nuclear–cytoplasmic shuttling.

3. Establishment of ARHL Animal Model and Pharmacological Intervention

SAMP8 accelerated aging mice were chosen as an ARHL model, manifesting hearing loss as early as 2 months and nearly complete deafness by 5 months. Immunoblotting (Western Blot) and real-time quantitative PCR showed significantly higher aging markers (p16, p21, p53) in 5-month-old cochleae, with SA-β-gal staining indicating comprehensive cochlear cell senescence.

Upon observing the age-related cytoplasmic translocation of HuR in hair cells, the team used the HuR translocation inhibitor SRI-42127 (i.p., 10 days) to block this process, resulting in more rapid and severe hearing loss—suggesting that increased HuR nuclear–cytoplasmic shuttling helps resist ARHL progression.

4. AAV-Mediated HuR Gene Overexpression and Safety Evaluation

An AAV-ie (efficient inner ear cell transduction vector) carrying HuR and HA-tag was constructed and injected through the round window into neonatal SAMP8 mouse cochleae. Immunostaining showed that AAV-ie-HuR efficiently transduced inner ear hair cells. By P60, a subset of treated mice underwent additional SRI-42127 blockade; results at P90 showed that the AAV-ie-HuR–only group had significantly lower hearing thresholds and less hair cell loss, whereas this effect was absent in mice also receiving SRI-42127, indicating a tight correlation between HuR nuclear–cytoplasmic translocation and its protective role.

5. Hair Cell–Specific and Cochlear-Specific HuR Knockout Models

Using the Cre-LoxP system, Atoh1-cre and Pax2-cre enabled creation of hair cell–specific and cochlear-specific HuR knockout mice (Atoh1-HuR−/− and Pax2-HuR−/−). Immunohistochemistry confirmed HuR deletion; behavioral testing (ABR and DPOAE) found no differences at P30 (normal hearing development) but high-frequency hearing loss at P60, near-complete deafness at P90. DPOAE supported outer hair cell dysfunction. Supporting cell–specific knockout (SOX2-CreER induction) did not cause overt hearing loss, confirming HuR’s action is hair cell–specific.

6. Gene Compensation Experiment (AAV Rescue of HuR Expression)

Using AAV-ANC80L65, a highly efficient hair cell transduction vector, AAV-ANC80L65-HuR was injected through the round window into neonatal knockout mice; at P90, both ABR and DPOAE were markedly improved (especially in mid-high frequencies), and hair cell loss was greatly reduced—demonstrating that exogenous HuR can partially restore lost function.

7. Mechanistic Studies—Impact of HuR Inactivation on Hair Cell Structure and Function

FM1-43 staining (for mechanotransduction/MET channel function) showed much weaker uptake in hair cells of knockout mice, indicating compromised MET channels. Scanning and transmission electron microscopy confirmed that HuR deletion caused the outer hair cell stereocilia to become disorganized, fused, and internalized, progressing with age (most severe in the basal turn); inner hair cells were less affected. The cuticular plate structure and key ion channel proteins (e.g., KCNQ4, BK, Prestin) were unaffected, further implicating HuR loss specifically in hair bundle pathology.

At the protein level, TRIOBP (rootlet protein) expression was unaffected, but levels of and localization for supporting proteins TPRN, RDX, Baiap2l2, and EPS8 were reduced and disordered. Furthermore, hair cells of knockout mice showed pronounced aging markers (SA-β-gal, p21, γH2A.X) upregulation, with mitochondrial degeneration and increased lysosomes, indicating accelerated aging in the absence of HuR.

8. Exploration of HuR Targets and Molecular Mechanism Validation

Using RNA immunoprecipitation sequencing (RIP-seq) on Pax2-Cre background cochlear cells, 1,122 HuR binding targets were identified, enriched in gene expression and RNA metabolic pathways. Focusing on GNAI3 (a signaling protein family member), which plays a pivotal role in stereocilia maintenance and hair cell survival, its mRNA level sharply dropped with HuR loss. Subsequent RIP-RT-qPCR confirmed HuR binding to GNAI3 mRNA, and overexpression of HuR in vitro significantly extended GNAI3 mRNA half-life, demonstrating enhanced stability. Further, RNA pull-down of uridine-rich elements in the GNAI3 3’UTR verified direct binding by HuR.

9. GNAI3 Gene Therapy for ARHL Intervention

AAV-ANC80L65 encapsulating GNAI3 was administered to knockout mice early on; at P90, these mice exhibited obvious improvement in hearing and hair cell survival. RNA-seq and qPCR showed that HuR loss suppresses the expression of many inner ear development and stereocilia functional genes (e.g., MYO3A, MYO15, TMC1, TOMT), and supplementing GNAI3 upregulates their expression, partially restoring hair cell function and structure.

4. Major Findings and Their Significance

This study provides a comprehensive depiction and confirmation of the core role of HuR and its regulatory molecular network in the prevention and control of age-related hearing loss:

  1. Reveals age-dependent upregulation of HuR and nuclear–cytoplasmic shuttling in hair cells as a feature of adaptive stress responses.
  2. Loss of HuR results in hair cell (stereocilia/hair bundle) disorganization, functional impairment, upregulation of aging markers, and accelerated apoptosis—key pathogenic factors for ARHL.
  3. HuR maintains outer hair cell stereocilia homeostasis by binding and stabilizing GNAI3 mRNA; GNAI3 in turn supports a group of crucial stereocilia proteins necessary for mechanotransduction.
  4. AAV-mediated overexpression of HuR or GNAI3 can greatly alleviate or even reverse the ARHL phenotype in rapidly aging mice, providing a new direction for genetic therapies for deafness.
  5. The research utilizes highly flexible genetic engineering interventions, multi-omic single-cell integration, and cross-species and multi-model verifications—demonstrating multi-dimensional innovative experimental design.

5. Conclusions, Scientific and Applied Value

This study is the first to reveal the “molecular guardian” role of the RNA-binding protein HuR in aging progression, providing a brand-new perspective on ARHL pathogenesis. Not only is HuR upregulated and relocalized spatially, it also specifically targets GNAI3 mRNA to regulate hair bundle homeostasis, preventing premature hair cell aging and loss of function due to instability.

On the clinical application level, AAV-mediated HuR/GNAI3 gene enhancement therapy opens a new door for hearing loss prevention and treatment, offering a precise interventional approach for elderly hearing loss, which currently lacks effective therapies. In the future, the development of more specific, safer gene vectors and more effective regulatory elements for expression control holds promise for the precision medicine of hearing impairment in the elderly.

6. Research Highlights and Innovations

  • Originality: This is the first systematic analysis of ARHL pathological mechanisms from the perspective of RNA-binding proteins, and proposes molecular-level intervention strategies.
  • Technical Innovation: Combines single-cell sequencing, efficient cochlear-targeting AAV vectors, RIP-seq, and multi-level transcriptome integration.
  • Prominent Clinical Significance: Establishes a closed-loop from “molecule–cell–behavior–therapy,” directly addressing clinical needs.
  • Rich Animal Models: Covers multiple strains, ages, tissues, and includes cross-species validation.

7. Other Points of Interest

The paper notes that HuR is not limited to regulating GNAI3; more cochlear targets of HuR await discovery. Although a highly efficient AAV expression system was used, further optimization of specific promoters and dosage control is needed to improve gene therapy efficiency and safety. In addition, the research focused mainly on early–mid stages of aging; future deep studies in 18–24-month super-aged cochlear models will better mirror late-stage human pathology.

8. Summary

This study, collaboratively completed by top domestic teams such as Shandong University and published in Nature Aging, is acclaimed for its scientific rigor, well-designed methodology, innovative workflow, and reliable results. It contributes a “Chinese solution” integrating theory, experimentation, and clinical translation for conquering age-related hearing loss. It is of great reference and pioneering significance for future molecular interventions in deafness and for preventing and controlling more age-associated degenerative diseases.