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BACKGROUND: A recent sham-controlled clinical study has shown that low-intensity pulsed ultrasound twice per week can safely and effectively treat patients with mild-to-moderate erectile dysfunction (ED). However, large-scale clinical trials are needed to verify its efficacy and safety and determine a reasonable treatment interval. AIM: To study whether low-intensity pulsed ultrasound therapy thrice per week is non-inferior to twice per week in patients with mild-to-moderate ED. METHODS: A randomized, open-label, parallel-group, non-inferiority clinical trial was conducted in 7 hospitals in China. A total of 323 patients with mild-to-moderate ED were randomized (1:1) into thrice per week (3/W) and twice per week (2/W) groups. Low-intensity pulsed ultrasound was applied on each side of the penis for 16 sessions. OUTCOMES: The primary outcome was response rate using the minimal clinically important difference in the International Index of Erectile Function (IIEF-EF) score at week 12. Secondary outcomes included Erection Hardness Score (EHS), Sexual Encounter Profile, Global Assessment Question, and Self Esteem and Relationship Questionnaire. RESULTS: Response rates in 3/W and 2/W groups were 62.0% and 62.5%, respectively. Treatment effect in the 3/W group was noninferior to that of the 2/W group, with rate difference lower bound of -0.01% [95% confidence interval -0.11 to 0.10%] within the acceptable margin (-14.0%). No significant difference was found among secondary outcomes. IIEF-EF score showed a significant increase from baseline in the 3/W group (16.8 to 20.7) and 2/W group (17.8 to 21.7), and the percentage of patients with EHS ≥3 increased in the 3/W (54.9% to 84.0%) and 2/W (59.5% to 83.5%) groups. There was no significant difference in response rate between the 2 groups after controlling for strata factors and homogeneous tests. No treatment-related adverse events were reported. CLINICAL IMPLICATIONS: Low-intensity pulsed ultrasound therapy displays similar efficacy and safety for mild-to-moderate ED when administered thrice or twice per week for 16 sessions. This study provides two options to suit patients' needs. STRENGTHS & LIMITATIONS: This is a large-sample, randomized, controlled, noninferiority trial study. Short-term follow-up and mostly younger patients are the main limitations. CONCLUSION: Low-intensity pulsed ultrasound therapy thrice and twice per week showed equivalent therapeutic effects and safety for mild-to-moderate ED in a young and generally healthy population. This therapy warrants further investigation of its potential value in rehabilitation of ED. Chen, H., Li Z., Li X., et al. The Efficacy and Safety of Thrice vs Twice per Week Low-Intensity Pulsed Ultrasound Therapy for Erectile Dysfunction: A Randomized Clinical Trial. J Sex Med 2022;19:1536-1545.
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Disfunção Erétil , Método Duplo-Cego , Humanos , Masculino , Ereção Peniana , Pênis , Resultado do Tratamento , Ondas UltrassônicasRESUMO
OBJECTIVE: To determine the outcomes and mechanisms of microenergy acoustic pulse (MAP) therapy in an irreversible rat model of female stress urinary incontinence. MATERIALS AND METHODS: Twenty-four female Sprague-Dawley rats were randomly assigned into four groups: sham control (sham), vaginal balloon dilation and ovariectomy (VBDO), VBDO + ß-aminopropionitrile (BAPN), and VBDO + ß-aminopropionitrile treated with MAP (MAP). MAP therapy was administered twice per week for 4 weeks. After a 1-week washout period, all 24 rats were evaluated with functional and histological studies. The urethral vascular plexus was examined by immunofluorescence staining with antibodies against collagen IV and von Willebrand factor (vWF). The urethral smooth muscle stem/progenitor cells (uSMPCs) were isolated and functionally studied in vivo and in vitro. RESULTS: Functional study with leak point pressure (LPP) measurement showed that the MAP group had significantly higher LPPs compared to VBDO and BAPN groups. MAP ameliorated the decline in urethral wall thickness and increased the amount of extracellular matrix within the urethral wall, especially in the urethral and vaginal elastic fibers. MAP also improved the disruption of the urethral vascular plexus in the treated animals. In addition, MAP enhanced the regeneration of urethral and vaginal smooth muscle, and uSMPCs could be induced by MAP to differentiate into smooth muscle and neuron-like cells in vitro. CONCLUSION: MAP appears to restore urethral wall integrity by increasing muscle content in the urethra and the vagina and by improving the urethral vascular plexus and the extracellular matrix.
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Incontinência Urinária por Estresse , Acústica , Aminopropionitrilo , Animais , Modelos Animais de Doenças , Feminino , Ratos , Ratos Sprague-Dawley , UretraRESUMO
BACKGROUND: Neurogenic erectile dysfunction (ED) is often refractory to treatment because of insufficient functional nerve recovery after injury or insult. Noninvasive mechano-biological intervention, such as microenergy acoustic pulse (MAP), low-intensity pulsed ultrasound, and low-intensity extracorporeal shockwave treatment, is an optimal approach to stimulate nerve regeneration. AIM: To establish a new model in vitro to simulate nerve injury in neurogenic ED and to explore the mechanisms of MAP in vitro. METHODS: Sprague-Dawley rats were used to isolate Schwann cells (SCs), major pelvic ganglion (MPG), and cavernous nerve with MPG (CN/MPG). SCs were then treated with MAP (0.033 mJ/mm2, 1 Hz, 100 pulses), and SC exosomes were isolated. The MPG and CN/MPG were treated with MAP (0.033 mJ/mm2, 1 Hz) at different dosages (25, 50, 100, 200, or 300 pulses) or exosomes derived from MAP-treated SCs in vitro. OUTCOMES: Neurite growth from the MPG fragments and CN was photographed and measured. Expression of neurotropic factors (brain-derived neurotrophic factor, nerve growth factor, and neurotrophin-3) was checked. RESULTS: Neurite outgrowth from MPG and CN/MPG was enhanced by MAP in a dosage response manner, peaking at 100 pulses. MAP promoted SC proliferation, neurotropic factor (brain-derived neurotrophic factor, nerve growth factor, and neurotrophin-3) expression, and exosome secretion. SC-derived exosomes significantly enhanced neurite outgrowth from MPG in vitro. CLINICAL IMPLICATIONS: MAP may have utility in the treatment of neurogenic ED by SC-derived exosomes. STRENGTH & LIMITATIONS: We confirmed that MAP enhances penile nerve regeneration through exsomes. Limitations of this study include that our study did not explore the exact mechanisms of how MAP increases SC exosome secretion nor whether MAP modulates the content of exosomes. CONCLUSION: This study revealed that neurite outgrowth from MPG was enhanced by MAP and by SC-derived exosomes which were isolated after MAP treatment. Our findings indicate that one mechanism by which MAP induces nerve regeneration is by stimulation of SCs to secrete exosomes. Peng D, Reed-Maldonado AB, Zhou F, et al. Exosome Released From Schwann Cells May Be Involved in Microenergy Acoustic Pulse-Associated Cavernous Nerve Regeneration. J Sex Med 2020;17:1618-1628.
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Exossomos , Acústica , Animais , Humanos , Masculino , Regeneração Nervosa , Ratos , Ratos Sprague-Dawley , Células de SchwannRESUMO
INTRODUCTION: The human placenta provides a bountiful and noncontroversial source of stem cells which have the potential for regeneration of injured tissue. These cells may restore erectile function after neurovascular tissue injury such as that seen in radical pelvic surgeries and pelvic trauma. AIM: To determine the effect of human placenta-derived stem cells on erectile function recovery and histological changes at various time points in a cavernous nerve injury rat model and to study the fate of injected stem cells throughout the regenerative process. METHODS: Human placental stem cells (PSCs) were dual labeled with monomeric Katushka far red fluorescent protein (mKATE)-renLUC using a lentivirus vector. A pelvic neurovascular injury-induced erectile dysfunction model was established in male, athymic rats by crushing the cavernous nerves and ligating the internal pudendal neurovascular bundles, bilaterally. At the time of defect creation, nonlabeled PSCs were injected into the corpus cavernosum at a concentration of 2.5 × 106 cells/0.2 mL. The phosphate-buffered saline-treated group served as the negative control group, and age-matched rats (age-matched controls) were used as the control group. Erectile function, histomorphological analyses, and Western blot were assessed at 1, 6, and 12 weeks after model creation. The distribution of implanted, dual-labeled PSCs was monitored using an in vivo imaging system (IVIS). Implanted cells were further tracked by detection of mKATE fluorescence in histological sections. MAIN OUTCOME MEASURE: The main outcome measure includes intracavernous pressure/mean arterial pressure ratio, neural, endothelial, smooth muscle cell regeneration, mKATE fluorescence, and IVIS imaging. RESULTS: The ratio of intracavernous pressure to mean arterial pressure significantly increased in PSC-injected rats compared with phosphate-buffered saline controls (P < 0.05) at the 6- and 12-week time points, reaching 72% and 68% of the age-matched control group, respectively. Immunofluorescence staining and Western blot analysis showed significant increases in markers of neurons (84.3%), endothelial cells (70.2%), and smooth muscle cells (70.3%) by 6 weeks in treatment groups compared with negative controls. These results were maintained through 12 weeks. IVIS analysis showed luminescence of implanted PSCs in the injected corpora immediately after injection and migration of cells to the sites of injury, including the incision site and periprostatic vasculature by day 1. mKATE fluorescence data revealed the presence of PSCs in the penile corpora and major pelvic ganglion at 1 and 3 days postoperatively. At 7 days, immunofluorescence of penile PSCs had disappeared and was diminished in the major pelvic ganglion. CLINICAL IMPLICATIONS: Placenta-derived stem cells may represent a future "off-the-shelf" treatment to mitigate against development of erectile dysfunction after radical prostatectomy or other forms of pelvic injury. STRENGTH & LIMITATIONS: Single dose injection of PSCs after injury resulted in maximal functional recovery and tissue regeneration at 6 weeks, and the results were maintained through 12 weeks. Strategies to optimize adult stem cell therapy might achieve more effective outcomes for human clinical trials. CONCLUSION: Human PSC therapy effectively restores the erectile tissue and function in this animal model. Thus, PSC therapy may provide an attractive modality to lessen the incidence of erectile dysfunction after pelvic neurovascular injury. Further improvement in tissue regeneration and functional recovery may be possible using multiple injections or systemic introduction of stem cells. Gu X, Thakker PU, Matz EL, et al. Dynamic Changes in Erectile Function and Histological Architecture After Intracorporal Injection of Human Placental Stem Cells in a Pelvic Neurovascular Injury Rat Model. J Sex Med 2020;17:400-411.
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Disfunção Erétil/fisiopatologia , Placenta/citologia , Transplante de Células-Tronco/métodos , Traumatismos do Sistema Nervoso/complicações , Animais , Modelos Animais de Doenças , Células Endoteliais/metabolismo , Feminino , Humanos , Plexo Hipogástrico/metabolismo , Masculino , Pelve/patologia , Ereção Peniana/fisiologia , Gravidez , Prostatectomia/efeitos adversos , Ratos , Ratos Nus , Recuperação de Função FisiológicaRESUMO
Human stem cell therapy for type 2 diabetes/obesity (T2D/O) complications is performedwith stem cell autografts, exposed to the noxious T2D/O milieu, often with suboptimal results.We showed in the Obese Zucker (OZ) rat model of T2D/O that when their muscle-derived stemcells (MDSC) were from long-term T2D/O male rats, their repair ecacy for erectile dysfunctionwas impaired and were imprinted with abnormal gene- and miR-global transcriptional signatures(GTS). The damage was reproduced in vitro by short-term exposure of normal MDSC to dyslipidemicserum, causing altered miR-GTS, fat infiltration, apoptosis, impaired scratch healing, and myostatinoverexpression. Similar in vitro alterations occurred with their normal counterparts (ZF4-SC) fromthe T2D/O rat model for female stress urinary incontinence, and with ZL4-SC from non-T2D/O leanfemale rats. In the current work we studied the in vitro eects of cholesterol and Na palmitate aslipid factors on ZF4-SC and ZL4-SC. A damage partially resembling the one caused by the femaledyslipidemic serum was found, but diering between both lipid factors, so that each one appears tocontribute specifically to the stem cell damaging eects of dyslipidemic serum in vitro and T2D/Oin vivo, irrespective of gender. These results also confirm the miR-GTS biomarker value forMDSC damage.
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Colesterol/metabolismo , Diabetes Mellitus Tipo 2/patologia , Obesidade/patologia , Ácido Palmítico/metabolismo , Células-Tronco/patologia , Incontinência Urinária por Estresse/patologia , Animais , Apoptose , Células Cultivadas , Diabetes Mellitus Tipo 2/metabolismo , Modelos Animais de Doenças , Feminino , Obesidade/metabolismo , Ratos , Ratos Zucker , Células-Tronco/metabolismo , Incontinência Urinária por Estresse/metabolismoRESUMO
OBJECTIVE: To determine the long-term therapeutic effect amongst three human cell types on erectile function recovery in a rat model of dual neurovascular-injury erectile dysfunction (NVED). MATERIALS AND METHODS: A dual NVED model was established in athymic rats by crushing the bilateral cavernous nerves and ligating the bilateral internal pudendal neurovascular bundles. At the time of defect creation, three different types of human cell populations (2.5 × 106 cells/0.2 mL: umbilical vein endothelial cells, adipose-derived stem cells, and amniotic fluid-derived stem cells) were injected intracavernously into the penile tissue. Saline injection (0.2 mL) served as a control group. Erectile function and histomorphological analyses of penile tissues were assessed 12 weeks after defect creation and cell or saline injection. RESULTS: The ratio of intracavernous pressure to mean arterial pressure (functional indicator) was significantly higher in the cell therapy groups compared to the saline-injected control group (P < 0.05). Immunofluorescence staining showed more cells expressing biomarkers of endothelial, smooth muscle, and nerve cells within the penile tissue in the cell therapy groups when compared to the control group. CONCLUSIONS: Cell therapy enhanced erectile function and ameliorated the histological changes 12 weeks after pelvic neurovascular injury in vivo, indicating that cell therapy may improve the long-term outcomes in neurogenic, myogenic and vascular tissue regeneration in the treatment of NVED.
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Terapia Baseada em Transplante de Células e Tecidos , Células Endoteliais , Disfunção Erétil/terapia , Células-Tronco , Animais , Modelos Animais de Doenças , Disfunção Erétil/etiologia , Masculino , Traumatismos dos Nervos Periféricos , Ratos , Ratos Nus , Lesões do Sistema VascularRESUMO
INTRODUCTION: Modulating tissue-resident stem and progenitor cells with a non-invasive, mechanobiological intervention is an optimal approach for tissue regeneration. Stem cell antigen-1 (Sca-1) has been identified as a stem cell marker within many organs but never within the penis. AIM: To localize and isolate penile stem/progenitor cells (PSPCs) and to evaluate cellular differentiation after exposure to induction medium and microenergy acoustic pulse (MAP) therapy. METHODS: Six male Sprague-Dawley rats were used to isolate PSPCs. Isolation was followed by stem cell characterization and differentiation assays. The PSPCs were then treated with MAP (0.033 mJ/mm2, 1 Hz) at various dosages (25, 50, 100, and 200 pulses) and for different durations (1, 2, 4, 6, or 8 hours) in vitro. MAIN OUTCOME MEASURE: The PSPCs (Sca-1-positive cells) were isolated using the magnetic-activated cell sorting system. PSPC cellular differentiation was assessed after induction with induction medium and with MAP in vitro. Wnt/ß-catenin signaling was also assayed. RESULTS: The PSPCs were successfully localized within the penile subtunic and perisinusoidal spaces, and they were successfully isolated using magnetic-activated cell sorting. The stemness of the cells was confirmed by stem cell marker characterization and by multiple differentiation into smooth muscle cells, endothelial cells, adipocytes, and neurons. MAP-induced PSPCs differentiated into smooth muscle cells by activating the Wnt/ß-catenin signaling pathway in a time- and dosage-dependent manner. CLINICAL IMPLICATIONS: By modulating resident PSPCs, MAP may have utility in the treatment of erectile dysfunction (ED). STRENGTHS & LIMITATIONS: This study provides solid evidence in support of microenergy therapies, including both MAP and low-intensity extracorporeal shock wave therapy, for the treatment of ED. Additional studies are needed and should include additional stem cells markers. Furthermore, studies exploring the underling mechanisms for PSPC activation and differentiation are required. CONCLUSION: PSPCs were successfully identified, localized, and isolated. Additionally, MAP provoked PSPCs to differentiate into smooth muscle cells via the Wnt/ß-catenin signaling pathway. As such, MAP provides a novel method for activating endogenous tissue-resident stem/progenitor cells and might facilitate stem cell regenerative therapy targeting ED. Peng D, Yuan H, Liu T, et al. Smooth Muscle Differentiation of Penile Stem/Progenitor Cells Induced by Microenergy Acoustic Pulses In Vitro. J Sex Med 2019; 16:1874-1884.
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Acústica , Tratamento por Ondas de Choque Extracorpóreas/métodos , Miócitos de Músculo Liso/metabolismo , Células-Tronco/metabolismo , Animais , Células Endoteliais/metabolismo , Disfunção Erétil/terapia , Masculino , Músculo Liso , Ereção Peniana/fisiologia , Pênis , Ratos , Ratos Sprague-DawleyRESUMO
BACKGROUND: Erectile dysfunction (ED) caused by pelvic neurovascular injury (PNVI) is often refractory to treatment. In many cases, erectogenic therapy is administered in a delayed fashion. AIM: To evaluate penile hemodynamic effects and histologic changes associated with delayed low-intensity extracorporeal shock wave therapy (Li-ESWT) after PNVI ED in a rat model. We visualized images using immunofluorescence and 3-dimensional imaging of solvent-cleared organs (3DISCO), a novel imaging technique. METHODS: A total of 32 Sprague-Dawley male rats aged 12 weeks were divided equally into 4 groups: sham surgery as normal controls (NC), PNVI controls (PC), PNVI with very-low-energy Li-ESWT (PVL), and PNVI with low-energy Li-ESWT (PL). Bilateral cavernous nerve crush and internal pudendal bundle ligation were performed in the 3 PNVI groups. Li-ESWT was administered twice a week for 4 weeks in the PL and PVL groups starting at 4 weeks after PNVI. OUTCOMES: Intracavernous pressure (ICP) studies (normalized to mean arterial pressure [MAP]) were conducted in all subject animals. After testing, tissue was harvested for immunofluorescence staining and 3DISCO analysis. RESULTS: Mean ICP/MAP was lower in PC animals compared with NC animals (0.37 ± 0.03 vs 0.91 ± 0.03, respectively; P = .001). The ICP/MAP ratio was significantly higher in PVL and PL animals (0.66 ± 0.07 and 0.82 ± 0.05, respectively) compared with PC animals (P = .002 and .001, respectively). Detailed microstructures and trajectories of nerves and vessels were identified with immunofluorescence and 3DISCO. The PC group had lower density of nerves, axons, neuronal nitric oxide synthase-positive nerves, and Schwann cells in the dorsal penis. Animals in the PL group had significantly higher expression of all of these markers compared with PC animals. CLINICAL IMPLICATIONS: Li-EWST may have utility in the management of severe ED related to PNVI from severe pelvic injury or radical pelvic surgeries, even when administered in a delayed fashion. STRENGTH & LIMITATIONS: This study of a severe ED phenotype involved treatment administered in a delayed fashion, which is more consistent with how therapy likely would be delivered in a real-world clinical context. Moreover, because the treatment commenced at 4 weeks after injury, when nerve and tissue atrophy have already occurred, the results imply that Li-ESWT can be used for regenerative therapy. Additional studies on dose optimization and treatment interval are needed to inform the design of human clinical trials. CONCLUSION: Li-ESWT ameliorates the negative functional and histologic effects of severe pelvic neurovascular injury in a rat model system. 3DISCO provides high-resolution images of neuroanatomy and neural regeneration. Wang HS, Ruan Y, Banie L, et al. Delayed Low-Intensity Extracorporeal Shock Wave Therapy Ameliorates Impaired Penile Hemodynamics in Rats Subjected to Pelvic Neurovascular Injury. J Sex Med 2019;16:17-26.
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Disfunção Erétil/terapia , Tratamento por Ondas de Choque Extracorpóreas/métodos , Ereção Peniana/fisiologia , Pênis/irrigação sanguínea , Animais , Modelos Animais de Doenças , Disfunção Erétil/etiologia , Hemodinâmica , Masculino , Regeneração Nervosa , Óxido Nítrico Sintase Tipo I/metabolismo , Ratos , Ratos Sprague-Dawley , Células de Schwann/metabolismo , Traumatismos do Sistema Nervoso/complicaçõesRESUMO
AIM: To investigate the possibility and mechanism of microenergy acoustic pulses (MAP) for activating tissue resident stem/progenitor cells within pelvic and urethral muscle and possible mechanism. METHODS: The female Zucker Lean and Zucker Fatty rats were randomly divided into four groups: ZL control, ZLMAP, ZF control, and ZFMAP. MAP was applied at 0.033 mJ/mm2 , 3 Hz for 500 pulses, and the urethra and pelvic floor muscles of each rat was then harvested for cell isolation and flow cytometry assay. Freshly isolated cells were analyzed by flow cytometry for Pax-7, Int-7α, H3P, and EdU expression. Meanwhile, pelvic floor muscle-derived stem cells (MDSCs) were harvested through magnetic-activated cell sorting, MAP was then applied to MDSCs to assess the mechanism of stem cell activation. RESULTS: Obesity reduced EdU-label-retaining cells and satellite cells in both pelvic floor muscle and urethra, while MAP activated those cells and enhanced cell proliferation, which promoted regeneration of striated muscle cells of the pelvic floor and urethral sphincter. Activation of focal adhesion kinase (FAK)/AMP-activated protein kinase (AMPK) /Wnt/ß-catenin signaling pathways by MAP is the potential mechanism. CONCLUSIONS: MAP treatment activated tissue resident stem cells within pelvic floor and urethral muscle in situ via activating FAK-AMPK and Wnt/ß-catenin signaling pathway.
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Músculo Esquelético/fisiologia , Obesidade/fisiopatologia , Diafragma da Pelve/fisiopatologia , Células Satélites de Músculo Esquelético/fisiologia , Uretra/fisiopatologia , Incontinência Urinária por Estresse/fisiopatologia , Estimulação Acústica , Acústica , Animais , Antígenos CD/metabolismo , Proliferação de Células , Desoxiuridina , Modelos Animais de Doenças , Feminino , Citometria de Fluxo , Proteína-Tirosina Quinases de Adesão Focal/metabolismo , Cadeias alfa de Integrinas/metabolismo , Contração Muscular/fisiologia , Músculo Esquelético/citologia , Músculo Estriado/citologia , Músculo Estriado/fisiologia , Mioblastos/fisiologia , Obesidade/complicações , Fatores de Transcrição Box Pareados , Ratos , Ratos Zucker , Regeneração , Células-Tronco , Uretra/citologia , Incontinência Urinária por Estresse/etiologia , Via de Sinalização WntRESUMO
AIM: Obesity is a strong independent risk factor for urinary incontinence. Effective therapeutic approaches for obesity-associated stress urinary incontinence (OA-SUI) are lacking as the mechanisms remain unclear. The aim of our study is to explore the impacts of microenergy acoustic pulse (MAP) therapy on urethral and pelvic floor muscle structure and function in female lean and fatty rats. METHODS: A total 24 Zucker fatty (ZF) and 24 Zucker lean (ZL) female 24-week-old rats were grouped into four groups: ZL control, ZLMAP, ZF control, and ZFMAP. For MAP treatment, 500 pulses were delivered at an energy level of 0.033 mJ/mm 2 and a frequency of 3 Hz and were applied twice a week for 4 weeks. After a 1-week washout, all rats underwent conscious cystometry and leak-point pressure (LPP) measurements followed by ex vivo organ-bath assay and histological study. RESULTS: ZF rats had lower LPP as compared to ZL rats, and MAP treatment significantly improved LPP in ZF rats (P < .05). Impaired muscle contractile activity (MCA) in organ-bath study was noted in ZF rats. MAP treatment significantly increased MCA in ZF rats (P < .05) and also increased the thickness of the striated muscle layer and the number of neuromuscular junctions (NMJs). In situ, MAP activated muscle satellite cells significantly (P < .05). CONCLUSIONS: Obesity impairs the function of both the urethral sphincter and the pelvic floor and leads to atrophy and distortion of the striated muscle in obese female rats. These issues contribute to OA-SUI. MAP improves continence by stimulating muscle regeneration and nerve innervation as well as by activating satellite cells.
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Estimulação Acústica , Músculo Esquelético/fisiopatologia , Obesidade/fisiopatologia , Diafragma da Pelve/fisiopatologia , Bexiga Urinária/fisiopatologia , Incontinência Urinária por Estresse/fisiopatologia , Acústica , Animais , Modelos Animais de Doenças , Feminino , Contração Muscular/fisiologia , Músculo Estriado/fisiopatologia , Obesidade/complicações , Ratos , Ratos Zucker , Uretra/fisiopatologia , Incontinência Urinária por Estresse/etiologiaRESUMO
Female stress urinary incontinence (FSUI) is prevalent in women with type 2 diabetes/obesity (T2D/O), and treatment is not optimal. Autograph stem cell therapy surprisingly has poor efficacy. In the male rat model of T2D/O, it was demonstrated that epigenetic changes, triggered by long-term exposure to the dyslipidemic milieu, led to abnormal global transcriptional signatures (GTS) of genes and microRNAs (miR), and impaired the repair capacity of muscle-derived stem cells (MDSC). This was mimicked in vitro by treatment of MDSC with dyslipidemic serum or lipid factors. The current study aimed to predict whether these changes also occur in stem cells from female 12 weeks old T2D/O rats, a model of FSUI. MDSCs from T2D/O (ZF4-SC) and normal female rats (ZL4-SC) were treated in vitro with either dyslipidemic serum (ZFS) from late T2D/O 24 weeks old female Zucker fatty (ZF) rats, or normal serum (ZLS) from 24 weeks old female Zucker lean (ZL) rats, for 4 days and subjected to assays for fat deposition, apoptosis, scratch closing, myostatin, interleukin-6, and miR-GTS. The dyslipidemic ZFS affected both female stem cells more severely than in the male MDSC, with some gender-specific differences in miR-GTS. The changes in miR-GTS and myostatin/interleukin-6 balance may predict in vivo noxious effects of the T2D/O milieu that might impair autograft stem cell (SC) therapy for FSUI, but this requires future studies.
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Diabetes Mellitus Tipo 2/patologia , Dislipidemias/patologia , Células-Tronco/patologia , Incontinência Urinária/patologia , Animais , Apoptose , Células Cultivadas , Modelos Animais de Doenças , Dislipidemias/sangue , Feminino , Masculino , Ratos , Ratos Zucker , Transplante de Células-TroncoRESUMO
OBJECTIVES: To investigate the feasibility of the Zucker fatty (ZF) rat as a model for research in to obesity-associated erectile dysfunction (OAED) and to determine the effect of low-intensity extracorporeal shockwave therapy (Li-ESWT) on penile tissue and function in these rats. MATERIALS AND METHODS: Eight new-born male Zucker lean (ZL group) rats (ZUC-Leprfa 186) and 16 new-born male ZF rats (ZUC-Leprfa 185) were injected with 5-ethynyl-2'-deoxyuridine (EdU) at birth to identify and monitor endogenous stem cells. Insulin tolerance testing was performed at 10 weeks of age. Beginning at 12 weeks of age, eight ZF rats were kept as controls, and the remaining eight ZF rats were treated with Li-ESWT (0.02 mJ/mm2 , 3 Hz, 500 pulses; ZF + SW group) twice a week for 4 weeks. Following a 1-week washout period, erectile function was evaluated by measuring intracavernosal pressure (ICP) and mean arterial pressure (MAP). Penile tissues were then harvested for histological study to assess smooth muscle/collagen content and endothelium content in the corpora cavernosum. LipidTOX™ staining was used to evaluate lipid accumulation. EdU, as a marker of cell activation, and phosphorylated histone 3 (H3P), as a marker of cell mitosis, were also assessed. RESULTS: The ICP/MAP indicated that erectile function was severely impaired in the ZF group as compared with the ZL group. In the ZF + SW group, erectile function was significantly improved (P < 0.05). Muscle atrophy was seen in the ZF group, while Li-ESWT increased the muscle content in ZF + SW group. Moreover, the penile endothelium was damaged in the ZF group, and Li-ESWT enhanced the regeneration of endothelial cells (P < 0.01) in the ZF + SW group. Lipid accumulation was seen in the penile tissue of ZF rats. Li-ESWT significantly reduced both the amount and the distribution pattern of LipidTOX, suggesting decreased overall lipid infiltration. Furthermore, Li-ESWT increased EdU-positive cells and markedly enhanced the phosphorylation level of H3P at Ser-10 in the ZF + SW group. Most H3P-positive cells were located within smooth muscle cells, with some located in the endothelium suggesting that these tissues are the reservoirs of penile stem/progenitor cells. CONCLUSION: ZF rats can serve as an animal model in which to study OAED. This study reveals that obesity impairs erectile function by causing smooth muscle atrophy, endothelial dysfunction, and lipid accumulation in the corpus cavernosum. Li-ESWT restored penile haemodynamic parameters in the ZF rats by restoring smooth muscle and endothelium content and reducing lipid accumulation. The underlying mechanism of Li-ESWT appears to be activation of stem/progenitor cells, which prompts cellular proliferation and accelerates penile tissue regeneration. Our findings are of interest, not just as a validation of this emerging treatment for erectile dysfunction, but also as a novel and potentially significant method to modulate endogenous stem/progenitor cells in other disease processes.
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Disfunção Erétil/terapia , Tratamento por Ondas de Choque Extracorpóreas/métodos , Obesidade/complicações , Animais , Glicemia/metabolismo , Proliferação de Células/fisiologia , Colágeno/fisiologia , Modelos Animais de Doenças , Endotélio Vascular/fisiologia , Disfunção Erétil/sangue , Disfunção Erétil/etiologia , Hipoglicemiantes/farmacologia , Insulina/farmacologia , Metabolismo dos Lipídeos/fisiologia , Masculino , Músculo Liso Vascular/fisiologia , Pênis/fisiologia , Ratos Zucker , Células-Tronco/fisiologiaRESUMO
OBJECTIVES: To evaluate the therapeutic effect of once-weekly low-intensity extracorporeal shock wave therapy (Li-ESWT) on underactive bladder (UAB) in the streptozotocin (STZ)-induced diabetic rat model. MATERIALS AND METHODS: In all, 36 female Sprague-Dawley rats were divided into three groups: normal control (NC), diabetes mellitus control (DMC), and DM with Li-ESWT (DM Li-ESWT). The two DM groups received an intraperitoneal 60 mg/kg STZ injection to induce DM. The Li-ESWT was applied toward the pelvis of the rats starting 4 weeks after STZ administration and lasting for 4 weeks. The Li-ESWT was given once weekly, with an energy flux density of 0.02 mJ/mm2 at 3 Hz for 400 pulses. All rats underwent conscious cystometry, leak-point pressure (LPP) assessment, ex vivo organ-bath study, histology, immunofluorescence, and Western Blot analysis. RESULTS: Conscious cystometry revealed voiding dysfunction in the DMC group, whereas the DM Li-ESWT group showed significantly improved voiding function, reflected in a reduced post-void residual urine volume and increased LPP compared to the DMC group. Ex vivo organ-bath studies showed that Li-ESWT enhanced muscle contractile activity of the bladder and urethra during electrical-field stimulation and drug stimulation. Histologically, Li-ESWT significantly restored bladder morphology, reflected by a reduction in the intravesical lumen area and increased muscle proportion of the bladder wall. Western Blot analysis showed higher smooth muscle actin expression in the bladder wall in the DM Li-ESWT group compared to the DMC group. Immunofluorescence showed decreased nerve-ending distribution, and destroyed and shortened nerve fibres in the DMC group, and recovery of neuronal integrity and innervation in the DM Li-ESWT group. CONCLUSIONS: In conclusion, Li-ESWT ameliorated UAB and urinary incontinence in the diabetic UAB rat model. The improvement appears to be the result of restoration of bladder and urethral structure and function by Li-ESWT. Li-ESWT is non-invasive and may become a better alternative therapy for UAB. Further investigations are warranted.
Assuntos
Diabetes Mellitus Experimental/complicações , Tratamento por Ondas de Choque Extracorpóreas/métodos , Bexiga Inativa/terapia , Bexiga Urinária/fisiopatologia , Animais , Western Blotting , Feminino , Imunofluorescência , Ratos , Ratos Sprague-Dawley , Estreptozocina/farmacologia , Bexiga Inativa/etiologiaRESUMO
AIM: Stress urinary incontinence (SUI) is a significant health problem for women. Treatments employing muscle derived stem cells (MDSCs) may be a promising approach to this prevalent, bothersome condition, but these treatments are invasive and require collection of cells from one site for injection into another. It is also unknown whether or not these cells establish themselves and function as muscle cells in the target tissues. Alternatively, low-intensity extracorporeal shock wave therapy (Li-ESWT) is non-invasive and has shown positive outcomes in the treatment of multiple musculoskeletal disorders, but the biological effects responsible for clinical success are not yet well understood. The aim of this study is to explore the possibility of employing Li-ESWT for activation of MDSCs in situ and to further elucidate the underlying biological effects and mechanisms of action in urethral muscle. METHODS: Urethral muscle derived stem cells (uMDSCs) were harvest from Zucker Lean (ZUC-LEAN) (ZUC-Leprfa 186) rats and characterized with flow cytometry. Li-ESWT (0.02 mJ/mm2 , 3 Hz, 200 pulses) and GSK2656157, an inhibitor of PERK pathway, were applied to L6 rat myoblast cells. To assess for myotube formation, we used immunofluorescence staining and western blot analysis in uMDSCs and L6 cells. RESULTS: The results indicate that uMDSCs could form myotubes. Myotube formation was significantly increased by the Li-ESWT as was the expression of muscle heavy chain (MHC) and myogenic factor 5 (Myf5) in L6 cells in vitro. Li-ESWT activated protein kinase RNA-like ER kinase (PERK) pathway by increasing the phosphorylation levels of PERK and eukaryotic initiation factor 2a (eIF2α) and by increasing activating transcription factor 4 (ATF4). In addition, GSK2656157, an inhibitor of PERK, effectively inhibited the myotube formation in L6 rat myoblast cells. Furthermore, GSK2656157 also attenuated myotube formation induced by Li-ESWT. CONCLUSION: In conclusion, this experiment reveals that rat uMDSCs can be isolated successfully and can form myotubes in vitro. PERK/ATF4 pathway was involved in myotube formation, and L6 rat myoblast cells were activated by Li-ESWT to form myotubes. These findings suggest that PERK/ATF4 pathway is activated by Li-ESWT. This study elucidates one of the biochemical pathways responsible for the clinical improvements seen after Li-ESWT. It is possible that this information will help to establish Li-ESWT as an acceptable treatment modality and may help to further refine the use of Li-ESWT in the clinical practice of medicine.
Assuntos
Fator 4 Ativador da Transcrição/metabolismo , Tratamento por Ondas de Choque Extracorpóreas , Desenvolvimento Muscular/fisiologia , Mioblastos/metabolismo , eIF-2 Quinase/metabolismo , Adenina/análogos & derivados , Adenina/farmacologia , Animais , Fator de Iniciação 2 em Eucariotos , Indóis/farmacologia , Desenvolvimento Muscular/efeitos dos fármacos , Mioblastos/efeitos dos fármacos , Fosforilação/efeitos dos fármacos , Ratos , Ratos Zucker , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Células-TroncoRESUMO
OBJECTIVE: To compare the effect of complete transection (tSCI) and contusion spinal cord injury (cSCI) on bladder function and bladder wall structure in rats. MATERIALS AND METHODS: A total of 30 female Sprague-Dawley rats were randomly divided into three equal groups: an uninjured control, a cSCI and a tSCI group. The cSCI group underwent spinal cord contusion, while the tSCI group underwent complete spinal cord transection. At 6 weeks post-injury, 24-h metabolic cage measurement and conscious cystometry were performed. RESULTS: Conscious cystometry analysis showed that the cSCI and tSCI groups had significantly larger bladder capacities than the control group. The cSCI group had significantly more non-voiding detrusor contractions than the tSCI group. Both injury groups had more non-voiding contractions compared with the control group. The mean threshold pressure was significantly higher in the tSCI group than in the control and cSCI groups. The number of voids in the tSCI group was lower compared with the control group. Metabolic cage analysis showed that the tSCI group had larger maximum voiding volume as compared with the control and cSCI groups. Vesicular acetylcholine transporter/smooth muscle immunoreactivity was higher in the control than in the cSCI or tSCI rats. The area of calcitonin gene-related peptide staining was smaller in the tSCI group than in the control or cSCI groups. CONCLUSIONS: Spinal cord transection and contusion produce different bladder phenotypes in rat models of SCI. Functional data suggest that the tSCI group has an obstructive high-pressure voiding pattern, while the cSCI group has more uninhibited detrusor contractions.
Assuntos
Traumatismos da Medula Espinal/complicações , Doenças da Bexiga Urinária/etiologia , Bexiga Urinária/patologia , Bexiga Urinária/fisiopatologia , Animais , Feminino , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Doenças da Bexiga Urinária/patologia , Doenças da Bexiga Urinária/fisiopatologiaRESUMO
OBJECTIVE: To study and compare the function and structure of the urethral sphincter in female Zucker lean (ZL) and Zucker fatty (ZF) rats and to assess the viability of ZF fats as a model for female obesity-associated stress urinary incontinence (SUI). MATERIALS AND METHODS: Two study arms were created: a ZL arm including 16-week-old female ZL rats (ZUC-Leprfa 186; n = 12) and a ZF arm including 16-week-old female ZF rats (ZUC-Leprfa 185; n = 12). I.p. insulin tolerance testing was carried out before functional study. Metabolic cages, conscious cystometry and leak point pressure (LPP) assessments were conducted. Urethral tissues were harvested for immunofluorescence staining to check intramyocellular lipid (IMCL) and sphincter muscle (smooth muscle and striated muscle) composition. RESULTS: The ZF rats had insulin resistance, a greater voiding frequency and lower LPP compared with ZL rats (P < 0.05), with more IMCL deposition localized in the urethral striated muscle fibres of the ZF rats (P < 0.05). The thickness of the striated muscle layer and the ratio of striated muscle to smooth muscle were lower in ZF than in ZL rats. CONCLUSION: Obesity impairs urethral sphincter function via IMCL deposition and leads to atrophy and distortion of urethral striated muscle. The ZF rats could be a consistent and reliable animal model in which to study obesity-associated SUI.
Assuntos
Obesidade/complicações , Uretra/fisiopatologia , Incontinência Urinária por Estresse/etiologia , Animais , Animais Geneticamente Modificados , Modelos Animais de Doenças , Feminino , Ratos , Ratos ZuckerRESUMO
BACKGROUND: We previously reported that progenitor cells, or stem cells, exist within penile tissue. We hypothesized that acoustic wave stimulation by low-intensity extracorporeal shockwave therapy (Li-ESWT) would activate local stem or progenitor cells within the penis, producing regenerative effects. AIMS: To study the feasibility of in situ penile progenitor cell activation by Li-ESWT. METHODS: We performed a cohort analysis of young and middle-age male Sprague-Dawley rats treated with 5-ethynyl-2'-deoxyuridine (EdU) pulse followed by Li-ESWT. In addition, Li-ESWT was applied to cultured Schwann cells and endothelial cells to study the molecular mechanism involved in cell proliferation. Thirty minutes before Li-ESWT, each rat received an intraperitoneal injection of EdU. Li-ESWT was applied to the penis at very low (0.02 mJ/mm2 at 3 Hz for 300 pulses) or low (0.057 mJ/mm2 at 3 Hz for 500 pulses) energy levels. The endothelial and Schwann cells were treated with very low energy (0.02 mJ/mm2 at 3 Hz for 300 pulses) in vitro. OUTCOMES: At 48 hours or 1 week after Li-ESWT, penile tissues were harvested for histologic study to assess EdU+ and Ki-67+ cells, and cell proliferation, Ki-67 expression, Erk1/2 phosphorylation, translocation, and angiogenesis were examined in cultured Schwann and endothelial cells after Li-ESWT. RESULTS: Li-ESWT significantly increased EdU+ cells within penile erectile tissues (P < .01) at 48 hours and 1 week. There were more cells activated in young animals than in middle-age animals, and the effect depended on dosage. Most activated cells were localized within subtunical spaces. In vitro studies indicated that Li-ESWT stimulated cell proliferation through increased phosphorylation of Erk1/2. CLINICAL TRANSLATION: The present results provide a possible explanation for the clinical benefits seen with Li-ESWT. STRENGTHS AND LIMITATIONS: The main limitation of the present project was the short period of study and the animal model used. Li-ESWT could be less effective in improving erectile function in old animals because of the decreased number and quality of penile stem or progenitor cells associated with aging. CONCLUSION: Li-ESWT activation of local penile progenitor cells might be one of the mechanisms that contribute to the beneficial effects of shockwave treatment for erectile dysfunction, which represents a non-invasive alternative to exogenous stem cell therapy. Lin G, Reed-Maldonado AB, Wang B, et al. In Situ Activation of Penile Progenitor Cells With Low-Intensity Extracorporeal Shockwave Therapy. J Sex Med 2017;14:493-501.
Assuntos
Desoxiuridina/análogos & derivados , Disfunção Erétil/terapia , Ondas de Choque de Alta Energia/uso terapêutico , Animais , Desoxiuridina/uso terapêutico , Modelos Animais de Doenças , Células Endoteliais/metabolismo , Disfunção Erétil/metabolismo , Humanos , Masculino , Ratos , Ratos Sprague-Dawley , Células de Schwann/metabolismo , Células-TroncoRESUMO
AIM: Obesity has been an independent risk factor for female stress urinary incontinence (SUI), the mechanism of this association remains unknown. The aim of this study is to validate the hypothesis that urethral dysfunction is a possible contributor to SUI in obese women. METHODS: Ten Zucker Fatty (ZF) (ZUC-Leprfa 185) and 10 Zucker Lean (ZL) (ZUC-Leprfa 186) female rats at 12-week-old were used in this experiment. The urethral sphincter rings were harvested from the bladder neck through to the most proximal 2/3 regions. In the organ bath study, single pulses of electrical field stimulation (EFS) were applied. For the fatiguing stimulation, repeated multi-pulse EFS with 70 mA were applied at frequency of 5 Hz for 5 min. Caffeine-containing Krebs' solution was administrated to contract the urethra until the contraction began to reach a plateau for 10 min. We performed immunofluorescence staining of the urethra after the experiment was finished. RESULTS: Compared to ZL controls, ZF rats had significantly impaired muscle contractile activity (MCA) (P < 0.05). Also, ZF rats presented early fatiguing of MCA and had a significantly greater percentage of MCA decline from baseline in the fatiguing test (37.7% vs 25.6%, P < 0.05). The plateau of maximal MCA induced by caffeine in ZF rats was significantly lower than ZL controls (0.22 vs 0.36, P < 0.05). CONCLUSIONS: This novel study showed that obese female rats had significantly impaired contractile properties of striated urethral sphincter, suggesting urethral dysfunction could be an important contributor to SUI in obesity.
Assuntos
Contração Muscular/fisiologia , Músculo Esquelético/fisiopatologia , Uretra/fisiopatologia , Bexiga Urinária/fisiopatologia , Incontinência Urinária por Estresse/fisiopatologia , Animais , Estimulação Elétrica , Feminino , Obesidade/fisiopatologia , Ratos , Ratos ZuckerRESUMO
Low-intensity extracorporeal shock wave therapy (Li-ESWT) is used in the treatment of erectile dysfunction, but its mechanisms are not well understood. Previously, we found that Li-ESWT increased the expression of brain-derived neurotrophic factor (BDNF). Here we assessed the underlying signaling pathways in Schwann cells in vitro and in penis tissue in vivo after nerve injury. The result indicated that BDNF were significantly increased by the Li-ESWT after nerve injury, as well as the expression of BDNF in Schwann cells (SCs, RT4-D6P2T) in vitro. Li-ESWT activated the protein kinase RNA-like endoplasmic reticulum (ER) kinase (PERK) pathway by increasing the phosphorylation levels of PERK and eukaryotic initiation factor 2a (eIF2α), and enhanced activating transcription factor 4 (ATF4) in an energy-dependent manner. In addition, GSK2656157-an inhibitor of PERK-effectively inhibited the effect of Li-ESWT on the phosphorylation of PERK, eIF2α, and the expression of ATF4. Furthermore, silencing ATF4 dramatically attenuated the effect of Li-ESWT on the expression of BDNF, but had no effect on hypoxia-inducible factor (HIF)1α or glial cell-derived neurotrophic factor (GDNF) in Schwann cells. In conclusion, our findings shed new light on the underlying mechanisms by which Li-ESWT may stimulate the expression of BDNF through activation of PERK/ATF4 signaling pathway. This information may help to refine the use of Li-ESWT to further improve its clinical efficacy.
Assuntos
Fator 4 Ativador da Transcrição/metabolismo , Fator Neurotrófico Derivado do Encéfalo/genética , Transdução de Sinais , Ondas Ultrassônicas , eIF-2 Quinase/metabolismo , Fator 4 Ativador da Transcrição/genética , Adenina/análogos & derivados , Adenina/farmacologia , Animais , Modelos Animais de Doenças , Inativação Gênica , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Indóis/farmacologia , Masculino , Pênis/metabolismo , Traumatismos dos Nervos Periféricos , Fosforilação/efeitos dos fármacos , Ratos , Células de Schwann/metabolismo , Células de Schwann/efeitos da radiação , Transdução de Sinais/efeitos dos fármacosRESUMO
INTRODUCTION: Erectile dysfunction (ED) caused by pelvic injuries is a common complication of civil and battlefield trauma with multiple neurovascular factors involved, and no effective therapeutic approach is available. AIMS: To test the effect and mechanisms of low-energy shock wave (LESW) therapy in a rat ED model induced by pelvic neurovascular injuries. METHODS: Thirty-two male Sprague-Dawley rats injected with 5-ethynyl-2'-deoxyuridine (EdU) at newborn were divided into 4 groups: sham surgery (Sham), pelvic neurovascular injury by bilateral cavernous nerve injury and internal pudendal bundle injury (PVNI), PVNI treated with LESW at low energy (Low), and PVNI treated with LESW at high energy (High). After LESW treatment, rats underwent erectile function measurement and the tissues were harvested for histologic and molecular study. To examine the effect of LESW on Schwann cells, in vitro studies were conducted. MAIN OUTCOME MEASUREMENTS: The intracavernous pressure (ICP) measurement, histological examination, and Western blot (WB) were conducted. Cell cycle, Schwann cell activation-related markers were examined in in vitro experiments. RESULTS: LESW treatment improves erectile function in a rat model of pelvic neurovascular injury by leading to angiogenesis, tissue restoration, and nerve generation with more endogenous EdU(+) progenitor cells recruited to the damaged area and activation of Schwann cells. LESW facilitates more complete re-innervation of penile tissue with regeneration of neuronal nitric oxide synthase (nNOS)-positive nerves from the MPG to the penis. In vitro experiments demonstrated that LESW has a direct effect on Schwann cell proliferation. Schwann cell activation-related markers including p-Erk1/2 and p75 were upregulated after LESW treatment. CONCLUSION: LESW-induced endogenous progenitor cell recruitment and Schwann cell activation coincides with angiogenesis, tissue, and nerve generation in a rat model of pelvic neurovascular injuries.