Electrotherapy for urethral modulation: Are extracellular matrix molecules and growth factors potential targets?

AIMS: To evaluate the expression of genes and proteins involved in the urethral components: vessels, nerves, and extracellular matrix, in female rats after trauma by vaginal distension (VD) and after electrical stimulation therapy (electrotherapy). METHODS: We analyzed the urethras of three groups of 18 female rats 30 days posttrauma by VD: control (no interventions); trauma (animals that had VD); and electrotherapy group (those that had VD and were treated with electrical stimulation). We compared the expression of vascular endothelial growth factor (VEGF), nerve growth factor (NGF), collagen types I and III (COL1a1 and COL3a1), and lysyl-oxidase like 1 (LOXL1) among the groups. Real-time reverse transcription-polymerase chain reaction, Western blot, and immunohistochemistry were used for molecule quantification. We used the Kruskal-Wallis test and analysis of variance for statistical analyses with p < 0.05 for significance. RESULTS: The COL1a1 gene expression was higher in the electrotherapy group than the trauma group (p = 0.036). COL3a1, VEGF, NGF, LOXL1 messenger RNA (mRNA) expression did not differ among the groups (p ≥ 0.05). COL1a1, COL3a1, VEGF, NGF, LOXL1 protein levels did not significantly differ among the groups (p ≥ 0.05) in Western blot analysis or immunohistochemistry assays. CONCLUSIONS: Electrotherapy caused a long-term increase in the COL1a1 mRNA level but did not change COL1a1 protein expression or VEGF, NGF, COL3a1, and LOXL1 genes and proteins in the urethras of rats after trauma by VD.

The molecular effects of electrical stimulation on the muscle components of the urethra of female rats after trauma by vaginal distention.

AIMS: To evaluate the expression of genes and proteins related to the urethral muscles of female rats after trauma by vaginal distention (VD) and after electrical stimulation therapy (EST). METHODS: We compared the urethras of four groups of 20 animals each: control without trauma (C), 7 (recent-trauma) and 30 days (late-trauma) post-VD, and VD-treated with EST. We evaluated the expression of myogenic regulatory factors MYOD1 and myogenin (MYOG); skeletal muscle myosin heavy chain 1, 2, and 3 (MYH1, MYH2, and MYH3); smooth muscle MYH11; and myosin light chain 9 (MYL9). We used real-time quantitative polymerase chain reaction, Western blot analysis, and immunohistochemistry. RESULTS: MYOD1 and MYOG genes were overexpressed in the recent-trauma group compared with the other groups (P < .05). MYH1 and MYH3 genes were upregulated in the recent-trauma group compared with the control and EST groups (P < .05). The MYH2 gene was overexpressed in the late-trauma group (P < .05), while the MYH2 protein was significantly increased in the EST group compared with control, recent-trauma and late-trauma groups by 5-, 3-, and 2.7-fold change, respectively (P < .05). MYL9 and MYH11 messenger RNA were overexpressed in both trauma groups compared with control and EST groups (P < .05). MYH11 protein was not different among the study groups (P > .05). CONCLUSIONS: EST enhances the recovery of the damaged urethral tissue of rats mainly by acting on the striated-muscle components. The MYH2 pathway underlies the positive effects of EST in the external urethral sphincter.

Molecular and immunohistochemical analysis of the urethra of female rats after induced trauma and intravenous therapy with muscle derived stem cells.

AIMS: To identify the urethral migration of muscle derived stem cells (MDSCs) after intravenous (IV) injection in rats that underwent vaginal distension (VD) and to analyze the effects of MDSC in the urethra of rats after trauma in regards to: (1) mRNA expression of collagens, Vegf, Ngf, Ki67, Myh11, and Myh2; (2) expression of smooth and striated muscle proteins. METHODS: MDSCs expressing green fluorescent protein (GFP) were injected into the tail vein of rats 3 days after VD. The location of GFP cells was verified at 2 h and at 7 days following IV injection. Urethras of three groups were analyzed: Control, Trauma 7D, and MDSC 7D. Real-time RT-qPCR and immunohistochemistry were performed. RESULTS: MDSCs were identified only after 2 h of the procedure in the urethra. Myh11 gene was overexpressed in the Trauma group in relation to Control. Ki67 gene expression was increased in the MDSC group relative to Trauma and Control. Col1a1 and Col3a1 genes expression were increased in the MDSC group relative to Control. Ngf mRNA level was decreased in the MDSC group in relation to Trauma. Protein expression of Mhy11, Myh2, and Desmin were increased in the MDSC group in relation to Trauma and decreased in the Trauma in relation to Control. CONCLUSION: MDSCs migrated early to the traumatized urethra, but did not integrate into the tissue. MDSC alters the expression of genes related to cell proliferation, neural growth factor and extracellular matrix and the expression of smooth and striated muscle proteins in the traumatized rat urethra.