RESUMO
OBJECTIVES: Diabetes is closely linked to hearing loss, yet the exact mechanisms remain unclear. Cochlear stria vascularis and pericytes (PCs) are crucial for hearing. This study investigates whether high glucose induces apoptosis in the cochlear stria vascularis and pericytes via elevated ROS levels due to oxidative stress, impacting hearing loss. METHODS: We established a type II diabetes model in C57BL/6J mice and used auditory brainstem response (ABR), Evans blue staining, HE staining, immunohistochemistry, and immunofluorescence to observe changes in hearing, blood-labyrinth barrier (BLB) permeability, stria vascularis morphology, and apoptosis protein expression. Primary cultured stria vascularis pericytes were subjected to high glucose, and apoptosis levels were assessed using flow cytometry, Annexin V-FITC, Hoechst 33342 staining, Western blot, Mitosox, and JC-1 probes. RESULTS: Diabetic mice showed decreased hearing thresholds, reduced stria vascularis density, increased oxidative stress, cell apoptosis, and decreased antioxidant levels. High glucose exposure increased apoptosis and ROS content in pericytes, while mitochondrial membrane potential decreased, with AIF and cytochrome C (CytC) released from mitochondria to the cytoplasm. Adding oxidative scavengers reduced AIF and CytC release, decreasing pericyte apoptosis. DISCUSSION: Hyperglycemia may induce mitochondrial apoptosis of cochlear stria vascularis pericytes through oxidative stress.
Assuntos
Fator de Indução de Apoptose , Apoptose , Citocromos c , Hiperglicemia , Camundongos Endogâmicos C57BL , Mitocôndrias , Estresse Oxidativo , Pericitos , Proteínas Proto-Oncogênicas c-bcl-2 , Espécies Reativas de Oxigênio , Estria Vascular , Animais , Pericitos/metabolismo , Pericitos/efeitos dos fármacos , Pericitos/patologia , Estria Vascular/metabolismo , Estria Vascular/patologia , Camundongos , Espécies Reativas de Oxigênio/metabolismo , Mitocôndrias/metabolismo , Citocromos c/metabolismo , Fator de Indução de Apoptose/metabolismo , Hiperglicemia/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Masculino , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patologia , Cóclea/metabolismo , Cóclea/patologiaRESUMO
Cisplatin (CDDP) is broadly employed to treat different cancers, whereas there are no drugs approved by the Food and Drug Administration (FDA) for preventing its side effects, including ototoxicity. Quercetin (QU) is a widely available natural flavonoid compound with anti-tumor and antioxidant properties. The research was designed to explore the protective effects of QU on CDDP-induced ototoxicity and its underlying mechanisms in male C57BL/6 J mice and primary cultured pericytes (PCs). Hearing changes, morphological changes of stria vascularis, blood labyrinth barrier (BLB) permeability and expression of apoptotic proteins were observed in vivo by using the auditory brainstem response (ABR) test, HE staining, Evans blue staining, immunohistochemistry, western blotting, etc. Oxidative stress levels, mitochondrial function and endothelial barrier changes were observed in vitro by using DCFH-DA probe detection, flow cytometry, JC-1 probe, immunofluorescence and the establishment in vitro BLB models, etc. QU pretreatment activates the PI3K/AKT signaling pathway, inhibits CDDP-induced oxidative stress, protects mitochondrial function, and reduces mitochondrial apoptosis in PCs. However, PI3K/AKT specific inhibitor (LY294002) partially reverses the protective effects of QU. In addition, in vitro BLB models were established by coculturing PCs and endothelial cells (ECs), which suggests that QU both reduces the CDDP-induced apoptosis in PCs and improves the endothelial barrier permeability. On the whole, the research findings suggest that QU can be used as a novel treatment to reduce CDDP-induced ototoxicity.
Assuntos
Cisplatino , Ototoxicidade , Camundongos , Animais , Masculino , Cisplatino/farmacologia , Cisplatino/metabolismo , Pericitos/metabolismo , Quercetina/farmacologia , Quercetina/química , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Células Endoteliais/metabolismo , Ototoxicidade/metabolismo , Camundongos Endogâmicos C57BL , Estresse Oxidativo , ApoptoseRESUMO
BACKGROUND: Gliomas, especially high-grade gliomas, are highly malignant with a poor prognosis. Although existing treatments have improved the survival rate of patients with glioma, the recurrence and mortality rates are still not ideal. The molecular mechanisms involved in the occurrence and development of glioma are still poorly understood. We previously reported that thrombospondin-2 (TSP2) expression was increased in tumor specimens from rat models, promoting excitatory synapse formation. However, little is known about the effect of TSP2 on the biological characteristics of glioma. METHODS: Glioma and cerebral cortex tissues were collected from 33 patients, and the expression of TSP2 in them was analyzed. Next, the proliferation and migration of TSP2 on glioma cells were analyzed in vitro. At last, a glioma transplantation model was constructed to explore the growth of TSP2 on glioma in vivo. RESULTS: The expression of TSP2 in surgical glioma specimens was increased compared to that in the normal cortex. Interestingly, the TSP2 protein level was higher in high-grade glioma (HGG, World Health Organization (WHO) grades 3-4) than in low-grade glioma (LGG, WHO grades 1-2) tissues. Exogenous addition of the TSP2 protein at an appropriate concentration promoted the migration of glioma cells but did not significantly affect their proliferation. Surprisingly, overexpression of TSP2 promoted both the migration and proliferation of cultured glioma cells. Moreover, in vivo experimental data implied that overexpression of TSP2 in C6 cells promoted the malignant growth of gliomas, while knockout of TSP2 slowed glioma growth. CONCLUSIONS: TSP2 promotes the migration and proliferation of glioma cells, which may provide new ideas for blocking glioma progression.
RESUMO
Objective: To investigate the role of Cav1.2 and its possible mechanism in the apoptosis of cochlear spiral ganglion neurons(SGNs) induced by cisplatin (CDDP) in C57BL/6J mice. Methods: Animal experiment: 8-week-old male C57BL/6J mice were randomly divided into the following two groups (10 mice/group) : normal saline group (Control group) and Cisplatin group (Cisplatin group). The Control group received daily intraperitoneal injections of normal saline, Cisplatin group was injected with cisplatin intraperitoneally at a dose of 3 mg/kg at the first 4 days of each cycle, and normal saline was injected daily at the last 10 days,repeat for 3 cycles. After administration, auditory threshold was detected by auditory brainstem response (ABR). Blood samples were collected from inner canthus of mice, and cochlea was cut off from neck. SOD and MDA kits were used to detect SOD activity and MDA content in serum and cochlea tissues. The expressions of apoptosis proteins in cochlear tissues were detected by Western blot. Morphological changes of spiral ganglion in mouse cochlea were observed by hematoxylin-eosin (HE) staining. TUNEL staining was used to observe the apoptosis of SGNs in cochlea of mice. The distribution and expression of Cav1.2 in SGNs of cochlea were observed by immunofluorescence. Cell experiment: Primary cultured SGNs were randomly divided into: control group (Control), solvent group (DMSO), Cav1.2 blocker group (N), cisplatin group, cisplatin and Cav1.2 blocker co-incubation group (Cisplatin+N). 5 µmol/L cisplatin was selected to treat SGNs based on the results of CCK8. Western blot was used to detect the protein expressions of Cav1.2.and apoptotic proteins. Hoechst33342 staining was used to observe the apoptosis of each group. Flow cytometry was used to detect the apoptosis rate of each group. Mitochondrial superoxide indicator (MitoSOXTM-Red) was used to detect the ROS release of mitochondria. Results: Animal experiments: Compared to the Control group, the hearing threshold was increased in Cisplatin group (P<0.01), the content of MDA in serum and cochlea tissues, apoptosis protein Cleaved caspase-3, Bax protein level, TUNEL positive rate, Cav1.2 protein expression level were increased significantly (P<0.05, P<0.01); the activity of SOD in serum and cochlear tissue, anti-apoptotic protein bcl-2 protein level and SGCs density in cochlear tissue were decreased significantly (P<0.05, P<0.01). Cell tests: Compared with the Control group, the expression of Cav1.2, apoptosis rate, Cleaved caspase-3, Bax protein level, intracellular calcium ion concentration, and ROS release were increased significantly only in Cisplatin group (P<0.05, P<0.01). The levels of bcl-2 protein and mitochondrial membrane potential were decreased significantly (P<0.01). Cav1.2 blockers could partially reverse the above changes (P<0.05). Conclusion: Cisplatin may increase intracellular Ca2+ concentration through up-regulation of Cav1.2, and then damage mitochondria, causing oxidative stress injury of SGNs and inducing neuronal apoptosis.
Assuntos
Cisplatino , Gânglio Espiral da Cóclea , Masculino , Camundongos , Animais , Gânglio Espiral da Cóclea/metabolismo , Cisplatino/farmacologia , Cisplatino/metabolismo , Proteína X Associada a bcl-2/metabolismo , Caspase 3/metabolismo , Camundongos Endogâmicos C57BL , Solução Salina , Espécies Reativas de Oxigênio/metabolismo , Cóclea/metabolismo , Apoptose , Neurônios , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteínas Reguladoras de Apoptose/metabolismo , Superóxido Dismutase/metabolismoRESUMO
It is widely accepted that the stria vascularis (SV) in cochlea plays a critical role in the generation of endocochlear potential (EP) and the secretion of the endolymph. 17ß-estradiol (E2) is the most potent and abundant endogenous estrogen during the premenopausal period, thus, considered as the reference estrogen. This study aimd to investigate the protective effect of E2 by promoting the expression of vascular endothelial growth factor (VEGF) and thus promoting the vascular regeneration of the SV in elderly mice. After being treated with E2 either in vivo or in vitro, the hearing threshold changes of C57BL/6J elder mice continuously reduced, endothelial cell morphology improved, the number of endothelial cells (ECs) tubular nodes increased significantly, the ability of tubular formation enhanced significantly and the expression of VEGF increased. In vitro, cell model in conjunction with in vivo ovariectomized model was established to demonstrate for the first time that E2 promotes angiogenesis by promoting the secretion of VEGF through the phosphatidylinositol 3-kinase (PI3K)/AKT pathway (PI3K/AKT). In conclusion, E2 demonstrated potent angiogenesis properties with significant protection against Age-Related Hearing Loss (ARHL), which provides a new idea for the improvement of ARHL.
Assuntos
Indutores da Angiogênese/farmacologia , Estradiol/farmacologia , Perda Auditiva/prevenção & controle , Neovascularização Fisiológica/efeitos dos fármacos , Estria Vascular/efeitos dos fármacos , Envelhecimento/fisiologia , Indutores da Angiogênese/uso terapêutico , Animais , Modelos Animais de Doenças , Células Endoteliais/metabolismo , Estradiol/uso terapêutico , Feminino , Perda Auditiva/fisiopatologia , Humanos , Camundongos , Técnicas de Cultura de Órgãos , Regeneração/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Estria Vascular/fisiologia , Fator A de Crescimento do Endotélio Vascular/agonistas , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
Seizures are common in patients with glioma, especially low-grade glioma (LGG). However, the epileptogenic mechanisms are poorly understood. Recent evidence has indicated that abnormal excitatory synaptogenesis plays an important role in epileptogenesis. The thrombospondin (TSP) family is a key regulator of synaptogenesis. Thus, this study aimed to elucidate the role of TSP2 in epileptogenesis in glioma-related epilepsy. The expression of TSP2 was increased in tumor tissue specimens from LGG patients, and this increase may have contributed to an increase in the density of spines and excitatory synapses in the peritumoral area. A glioma cell-implanted rat model was established by stereotactic implantation of wild-type TSP2-expressing, TSP2-overexpressing or TSP2-knockout C6 cells into the neocortex. Similarly, an increase in the density of excitatory synapses was also observed in the peritumoral area of the implanted tumor. In addition, epileptiform discharges occurred in the peritumoral cortex and were positively correlated with the TSP2 level in glioma tissues. Moreover, α2δ1/Rac1 signaling was enhanced in the peritumoral region, and treatment with the α2δ1 antagonist gabapentin inhibited epileptiform discharges in the peritumoral cortex. In conclusion, glioma-derived TSP2 promotes excitatory synapse formation, probably via the α2δ1/Rac1 signaling pathway, resulting in hyperexcitability in the peritumoral cortical networks, which may provide new insight into the epileptogenic mechanisms underlying glioma-related epilepsy.