Effects of RSC96 Schwann Cell-Derived Exosomes on Proliferation, Senescence, and Apoptosis of Dorsal Root Ganglion Cells In Vitro.

BACKGROUND Stress urinary incontinence is a common condition in women and can be associated with peripheral nerve injury. Exosomes. derived from Schwann cells, can enhance the regeneration of axons of the peripheral nervous system. This study aimed to investigate the effects of RSC96 Schwann cell-derived exosomes in a novel in vitro model of dorsal root ganglion (DRG) cell injury induced by cyclic mechanical strain (CMS). MATERIAL AND METHODS RSC96 Schwann cells and DRG cells were cultured in vitro. CMS in DRG cells involved mechanical stretch trauma with 5333 µ strain. ExoQuick-TC polymer was used to precipitate exosomes from RSC96 Schwann cell culture medium and identified by nanoparticle tracking analysis, electron microscopy, and Western blot for detection of CD9 and tumor susceptibility gene 101 (Tsg101) protein. Cultured DRG cells were treated with RSC96 Schwann cell-derived exosomes, followed by measurement of cell viability, proliferation, senescence, and apoptosis using the cell counting kit-8 (CCK-8) assay, senescence-associated beta-galactosidase (SA-ß-gal) staining, and Hoechst 33258 (blue) fluorescence nucleic acid staining using flow cytometry. RESULTS Mechanical stretch with 5333 µ strain for 8 hours at 1 Hz decreased the activity of cultured DRG cells. RSC96 Schwann cell-derived exosomes promoted cell proliferation and significantly inhibited apoptosis and senescence of DRG cells following injury induced by CMS. CONCLUSIONS An in vitro model of DRG cell injury induced by CMS, showed that RSC96 Schwann cell-derived exosomes had a protective effect. The effects of Schwann cell-derived exosomes on peripheral nerve injury, including in stress urinary incontinence, require future in vivo studies.

Effects of electrical stimulation on cell activity, cell cycle, cell apoptosis and ß­catenin pathway in the injured dorsal root ganglion cell.

The present study aimed to investigate the effects of electrical stimulation (ES) on cell activity, cell cycle and apoptosis in injured rat dorsal root ganglion (DRG) cells induced by cyclic mechanical stretching (CMS). The present study also investigated whether the Wnt/ß­catenin pathway is involved in this process. Injury and ES models were established in DRG cells. Then, cell activity was detected using a Cell Counting Kit­8 and 5­ethynyl­2'­deoxyuridine­594 cell proliferation assay kit. Cell cycle distribution was detected using a cell cycle detection kit. Apoptosis was detected using an Annexin V­FITC apoptosis detection kit, and Wnt/ß­catenin pathway­associated proteins were detected using western blotting. The present study demonstrated that CMS decreased DRG cell activity, increased the number of cells in the S phase, promoted cell apoptosis and inhibited the Wnt/ß­catenin pathway. In addition, ES significantly increased the proliferation activity of DRG cells, increased the number of cells in the G2 phase, decreased the apoptotic rate and activated the Wnt/ß­catenin pathway, ultimately reversing the injury caused by CMS. Following inhibition of the Wnt/ß­catenin signaling pathway using XAV939, the effects of ES were weakened. In conclusion, the present study demonstrated that ES may reverse CMS­induced injury in DRG cells, and that the Wnt signaling pathway may be involved in this process.

Electrical stimulation enhances neuronal cell activity mediated by Schwann cell derived exosomes.

Electrical stimulation (ES) therapy has good effects in patients with nervous system injury-related diseases. ES promotes nerve cell regeneration and stimulates Schwann cells to express neurotrophic factors. The incidence of stress urinary incontinence (SUI) among elderly people is increasing. Some studies suggest that damage to the pudendal nerve is closely related to the pathogenesis of SUI. It has also been found that pelvic ES can reduce SUI symptoms in a rat model of SUI caused by pudendal nerve injury. Clinically, pelvic floor electrical stimulation is effective in patients with mild to moderate SUI. These studies indicate that ES may ameliorate damage to the pudendal nerve and thus achieve the goal of SUI treatment, although the mechanism of action of this treatment remains unclear. Therefore, the purpose of the present study was to clarify the relationships among ES, neural cells and Schwann cells at the cellular level. We applied ES to nerve cells at 100 mV/mm or 200 mV/mm for 0, 0.5, 1, or 2 h to investigate changes in nerve cell activity. We then co-cultured the nerve cells with Schwann cells to explore the influence of single-culture and co-culture conditions on the nerve cells. Compared to non-ES, ES of the nerve cells increased their activity. Compared to those in single culture, co-cultured nerve cells exhibited an additional increase in activity. We also found that Schwann cell derived exosomes could promote the activity of nerve cells, with glutamate and calcium ions playing a potential role in this process. These results suggest that the mutual regulation of neural cells and Schwann cells plays an important role in the process by which ES ameliorates neurological function, which may provide a basis for subsequent studies.

Protective Role of Nuclear Factor Erythroid-2-Related Factor 2 against Mechanical Trauma-Induced Apoptosis in a Vaginal Distension-Induced Stress Urinary Incontinence Mouse Model.

Apoptosis and oxidative damage are involved in the pathogenesis and progression of stress urinary incontinence (SUI). Our previous results indicate that cell apoptosis and oxidative damage increase in a mouse model of mechanical injury-induced SUI and in fibroblasts treated with excessive mechanical strain. Nuclear factor erythroid-2-related factor 2 (Nrf2) is a well-characterized global antioxidant gene inducer that can reduce oxidative damage and apoptosis. Therefore, we predicted that Nrf2 may have a protective role in mechanical trauma-induced SUI. To test this hypothesis, a mouse model of vaginal distension- (VD-) induced SUI was established. Leak point pressure (LPP); levels of apoptosis, apoptosis-related proteins, and peroxidation products; and the activities of antioxidative proteins in the anterior vaginal wall were measured in wild-type (Nfe2l2+/+) C57BL/6 mice and Nrf2-knockout mice (Nfe2l2-/-). The results showed that Nrf2 knockout aggravated VD-induced reduction in LPP, increase in cell apoptosis and peroxidation product levels, decrease in antioxidative protein activities, and alterations in apoptosis-related protein levels in the vaginal walls of mice. To further confirm the role of Nrf2 in mechanical trauma-induced apoptosis and SUI, VD was performed on mice overexpressing Nrf2 via in vivo transfection of LV-Nfe2l2. The results showed that Nrf2 overexpression significantly alleviated VD-induced abnormalities in the anterior vaginal wall. Taken together, our data suggested that Nrf2 is a potential protective factor in mechanical trauma-induced apoptosis in a mouse model of SUI. Antioxidative therapy may be a promising treatment for mechanical trauma-related SUI.

Effect of puerarin on collagen metabolism of fibroblasts in pelvic tissue of women with pelvic organ prolapse.

The aim of the present study was to investigate the protective effect of puerarin on pelvic organ prolapse (POP) and the underlying mechanisms that regulate the metabolism of human parametrial ligament fibroblasts (HPLFs). HPLFs obtained from the pelvic tissue of patients with (n=10) or without (n=8) POP during hysterectomy were isolated by enzymatic digestion and subsequently identified by immunocytochemistry in a previous study of the authors. Following this, cultured HPLFs were treated with 0.01, 0.10 or 1.00 mmol/l puerarin, followed by detection of proliferation rate by Cell Counting kit­8 assay. Following incubation with puerarin for 48 h, mRNA and protein expression levels of tissue inhibitor of metalloproteinase­1 (TIMP­1), matrix metalloproteinase (MMP)­2 and ­9, and collagen (COL)I and III in HPLFs were quantified by reverse transcription­quantitative polymerase chain reaction, and western blot and gelatin zymography analyses, respectively. MMP­2 and ­9 expression levels were increased, whereas expression levels of TIMP­1, and COL I and III were decreased, in patients with POP compared with healthy controls. Following puerarin treatment, the expression levels of TIMP­1, and COL I and III were enhanced, whereas MMP­2 and ­9 were inhibited. In conclusion, the present study demonstrated evidence increased degradation of the extracellular matrix in pelvic tissues of patients with POP compared with controls, and the protective effect of puerarin against POP via its anti­degradation effect on collagen. These results provide evidence for puerarin as a novel approach for the treatment of POP.

Effectiveness of Traditional Chinese Medicine (TCM) treatments on the cognitive functioning of elderly persons with mild cognitive impairment associated with white matter lesions.

BACKGROUND: Cerebral white matter lesion (WML) is a pathological change of the white matter which is considered an early sign of brain impairment in elderly individuals, so it is reasonable to administer early dementia prevention programs to individuals with WML.Traditional Chinese Medicine (TCM) has developed several approaches to prevent or delay the onset of dementia that have, as yet, not been formally tested. AIM: Evaluate the effects of a 6-month TCM intervention for elderly persons with mild cognitive impairment and WML. METHODS: Eighty individuals 65 years of age or older with radiological evidence of WML and mild cognitive impairment based on the Montreal Cognitive Assessment (MoCA) were classified into the four main TCM constitutional types (qi deficiency, yang deficiency, phlegm dampness, or blood stasis) and randomly assigned to a treatment group or a treatment-as-usual control group. The treatment group participated in training focused on diet, lifestyle, exercises, and emotional regulation adjustment; they also received six monthly courses of moxibustion (heating acupoints by burning the moxa of dried mugwort), each of which involved 10 daily 15-minute sessions focused on three targeted acupoints (one of which was specific to the constitutional type). Changes in the MoCA and in the score of each of the four constitutional types were the main outcomes assessed. RESULTS: Two participants dropped out of each group over the 6 months, leaving 38 in each group. Based on repeated measures analysis of variance, the total MoCA score, four of the six MoCA subscales scores (visual space and executive function, naming, attention and calculation, and delayed memory), and all four of the TCM constitution type scores showed significantly greater improvement over the 6 months in the treatment group than in the control group. CONCLUSION: This study shows that TCM interventions can improve both the cognitive functioning and the severity of symptoms considered in the TCM assessment of constitutional types among elderly individuals with mild cognitive impairment and WML. Long-term follow-up studies that use blinded evaluation of the outcome are needed to determine whether or not constitution-specific TCM treatments can prevent the onset of dementia.