RESUMO
Evidence suggests that damage to the ribbon synapses (RS) may be the main cause of auditory dysfunction in noise-induced hearing loss (NIHL). Oxidative stress is implicated in the pathophysiology of synaptic damage. However, the relationship between oxidative stress and RS damage in NIHL remains unclear. To investigate the hypothesis that noise-induced oxidative stress is a key factor in synaptic damage within the inner ear, we conducted a study using mice subjected to single or repeated noise exposure (NE). We assessed auditory function using auditory brainstem response (ABR) test and examined cochlear morphology by immunofluorescence staining. The results showed that mice that experienced a single NE exhibited a threshold shift and recovered within two weeks. The ABR wave I latencies were prolonged, and the amplitudes decreased, suggesting RS dysfunction. These changes were also demonstrated by the loss of RS as evidenced by immunofluorescence staining. However, we observed threshold shifts that did not return to baseline levels following secondary NE. Additionally, ABR wave I latencies and amplitudes exhibited notable changes. Immunofluorescence staining indicated not only severe damage to RS but also loss of outer hair cells. We also noted decreased T-AOC, ATP, and mitochondrial membrane potential levels, alongside increased hydrogen peroxide concentrations post-NE. Furthermore, the expression levels of 4-HNE and 8-OHdG in the cochlea were notably elevated. Collectively, our findings suggest that the production of reactive oxygen species leads to oxidative damage in the cochlea. This mitochondrial dysfunction consequently contributes to the loss of RS, precipitating an early onset of NIHL.
RESUMO
Adeno-associated virus (AAV)-mediated gene transfer is an efficient method of gene over-expression in the vestibular end organs. However, AAV has limited usefulness for delivering a large gene, or multiple genes, due to its small packaging capacity (< 5 kb). Co-transduction of dual-AAV vectors can be used to increase the packaging capacity for gene delivery to various organs and tissues. However, its usefulness has not been well validated in the vestibular sensory epithelium. In the present study, we characterized the co-transduction of dual-AAV vectors in mouse utricles following inoculation of two AAV-serotype inner ear (AAV-ie) vectors via canalostomy. Firstly, co-transduction efficiencies were compared between dual-AAV-ie vectors using two different promoters: cytomegalovirus (CMV) and CMV early enhancer/chicken ß-actin (CAG). In the group of dual AAV-ie-CAG vectors, the co-transduction rates for striolar hair cells (HCs), extrastriolar HCs, striolar supporting cells (SCs), and extrastriolar SCs were 23.14 ± 2.25%, 27.05 ± 2.10%, 57.65 ± 7.21%, and 60.33 ± 5.69%, respectively. The co-transduction rates in the group of dual AAV-ie-CMV vectors were comparable to those in the dual AAV-ie-CAG group. Next, we examined the co-transduction of dual-AAV-ie-CAG vectors in the utricles of neonatal mice and damaged adult mice. In the neonatal mice, co-transduction rates were 52.88 ± 3.11% and 44.93 ± 2.06% in the striolar and extrastriolar HCs, respectively, which were significantly higher than those in adult mice. In the Pou4f3+/DTR mice, following diphtheria toxin administration, which eliminated most HCs and spared the SCs, the co-transduction rate of SCs was not significantly different to that of normal utricles. Transgene expression persisted for up to 3 months in the adult mice. Furthermore, sequential administration of two AAV-ie-CAG vectors at an interval of 1 week resulted in a higher co-transduction rate in HCs than concurrent delivery. The auditory brainstem responses and swim tests did not reveal any disruption of auditory or vestibular function after co-transduction with dual-AAV-ie vectors. In conclusion, dual-AAV-ie vectors allow efficient co-transduction in the vestibular sensory epithelium and facilitate the delivery of large or multiple genes for vestibular gene therapy.
RESUMO
Wumei Pills originates from Treatise on Cold Damage. A total of 128 records on it were screened out, involving 102 ancient books, 110 modern clinical studies, and 48 diseases. According to the records, the prescription origin, prescription composition, prescription explanation, main indications, dosage, medicinal processing, preparation, and usage, contraindications, and mo-dern clinical applications were analyzed. The result shows that Wumei Pills is composed of Mume Fructus, Asari Radix et Rhizoma, Zingiberis Rhizoma, Coptidis Rhizoma, Angelicae Sinensis Radix, Aconiti Lateralis Radix Praeparata, Zanthoxyli Pericarpium, Cinnamomi Ramulus, Ginseng Radix et Rhizoma, and Phellodendri Chinensis Cortex. The main indications expand over time, and it can be applied to diarrhea, dysentery, retching, chest pain, cough, Qi ascending from lower abdomen, and reversal cold of hands and feet with the syndromes of cold and heat in complexity and hyperactivity of liver Yang and spleen deficiency. According to modern clinical records, it is mainly used for the treatment of diseases in the digestive system, nervous system, endocrine system, metabolic system, etc., such as ulcerative colitis, diarrhea, insomnia, and type 2 diabetes mellitus. The dosage of Wumei Pills has gradually reduced from the Han Dynasty to the Qing Dynasty, but the proportions of the medicinals has remained basically unchanged. In this prescription, Aconiti Lateralis Radix Praeparata and Zanthoxyli Pericarpium need to be processed, while the rest medicinals are used in raw form. As for the medicinal selection, Zanthoxyli Pericarpium is examinable. Asari Radix et Rhizoma is derived from Aristolochiaceae, which is toxic to the liver and kidney, so the dosage should be kept in a safe range. In summary, Wumei Pills has great clinical value. The textual research on Wumei Pills helps clarify the development of Wumei Pills, which provides evidence in-depth research and development and rational clinical application of Wumei Pills.