Stress urinary incontinence (SUI) is the involuntary leakage of urine during physical activity caused by bladder pressure exceeding urethral closure pressure. Slings are commonly used as a surgical treatment for SUI. However, the sling procedure is invasive and inserted into the body, resulting in mesh infection and chronic pain. The current study aims to evaluate the feasibility of laser treatment for SUI by using 980nm laser light with a balloon-integrated diffusing applicator as a minimally invasive and non-permanent procedure. A female porcine urethra was dilated using a balloon, and then the tissue was irradiated with the 980nm laser at 20W for 15s. The laser irradiated urethra were harvested 0, 14 and 28 days after the laser irradiation, respectively. Both Masson’s trichrome and Sirius red staining were applied to confirm structural changes and collagen distribution. There was no change in mucosal thickness between the laser irradiation group and the control group. In the laser irradiation group, the thickness of the muscle layer increased by approximately 35%, compared to the control group. Furthermore, the laser irradiation group showed a 30% increase in collagen distribution in the external urethral sphincter compared to the SUI model. The current study demonstrated the feasibility of the therapeutic effect of the 980nm laser in the porcine urethra. To confirm the safety and efficacy of the proposed photothermal treatment, we will conduct in vivo studies to assess the extent of nerve damage and molecular changes using PCR assays in chronic and disease-induced animal models.
Several therapeutic effects of photobiomodultion (PBM) on variable mucosal lesions of the upper aerodigestive tract has been reported. However, the biomodulatory effect of PBM are rarely reported on tracheostomy and tracheal fenestration, which has been increased with the spread of coronavirus disease (COVID-19). In this study, we developed a translaryngeal PBM therapy by using a basket-integrated diffusing applicator (BIDA) to ameliorate the wound healing of fenestrated tracheostoma after surgery. Tracheostomy was performed by using an electrocautery unit (Bovie) to develop an in vivo porcine model of fenestrated tracheostoma with impaired wounds. A 635 nm laser light (200 mW/cm2 for 3 min, 36 J/cm2) was applied to the wounds by using BIDA once daily for five days. BIDA was inserted in tracheostomy tube and emitted homogeneous circumferential light into the fenestrated wound without thermal damage. Control highly induced fibrotic expressions of alpha-smooth muscle actin and type-1 collagen with acute inflammation. In contrast, PBM reduced the expressions up to less than 82% of control. Histological scores presented that PBM significantly decreased acute inflammation and overgranulation to two-thirds of control with a small portion of abscess. In addition, a substantial difference in the lumen area was observed between the control and PBM due to the thickened wall. The current study demonstrated that the proposed PBM therapy could ameliorate the impaired wound healing of fenestrated tracheostoma as a result of modulated inflammation and fibrotic responses. Therefore, the translaryngeal PBM with BIDA can be an effective adjuvant therapy for managing the fenestrated wound after tracheostomy.
Cervical cancer is a fatal disease with over 50% of modality rate. Although chemo and external beam (EB) radiation are used for cancer treatment, radiation therapy has a limited effect and toxicity. Interstitial-photodynamic therapy (I-PDT) is a promising method for locally advanced cervical cancer (LACC). However, there are still challenges to reducing the totoxicity of photosensitizer and improving photoactivation. Thus, we propose a sodium copper chlorophyll (SCC), organic compound, as a photosensitizer. The purpose of this study is to investigate the feasibility of SCC as a photosensitizer, and develop an effective LACC treatment through SCC mediated I-PDT. The proposed therapy was evaluated on human cervical carcinoma cells and HeLa xenograft tumor models. After SCC injection (4 μg/ml and 0.1 g/kg), a 405 nm blue light (BL; 800 mW/cm2) was applied using a frontal (EB-PDT) and diffusing (I-PDT) fiber. SCC or BL was hardly cytotoxic (less than 10%) on cancer cells independently, whereas SCC mediated PDT rapidly and strongly reduced the viability. High intensity of ROS was measured within 20 min after treatment. In addition, SCC mediated PDT stimulated the expressions of cleaved caspase 3, 8, and 9 with downregulated pro-caspase. Moreover, the proposed I-PDT decreased tumor size by up to 37% less than EB-PDT with minimal thermal damage around tissue. The current study demonstrated the potential antitumor effect of SCC mediated I-PDT in human cervical carcinoma cell and xenograft tumor model as a result of biocompatible photosensitizer and improved photoactivation. Therefore, SCC mediated I-PDT can warrant an effective treatment of LACC.
Stress urinary incontinence (SUI) is the unintentional loss of urine. Currently, catheter and artificial urinary sphincters are applied for SUI treatment. However, these treatments are invasive and require continuous usage. While Er:YAG and CO2 lasers are used as a minimal or non-invasive method, their treatment effects are limited to the superficial vaginal tissue and can often cause severe burns, leading to tissue destruction. The purpose of this study to investigate the feasibility of non-ablative 980-nm laser treatment for SUI. Numerical simulations were performed to confirm thermal effects in urethral tissue by employing the Pennes bioheat transfer and partial differential equations. Rabbit urethral tissue was used for experimental validations. A non-compliant balloon was used to expand the urethral tissue, and the 980 nm laser light was irradiated on the tissue at 20 W for 15 s. After the laser irradiation, the treated samples were stained with hematoxylin and eosin (HE) to evaluate any physical changes in the overall urinary structure. Masson trichrome (MT) staining was performed to analyze the extent of thermal injury in the collagen of the urethral tissue. A uniform and symmetrical temperature distribution was observed around the balloon surface in the simulation. The numerical simulations and experiments indicated that mucosal and muscle layers in the urethral tissue reached the temperatures of 24 °C and 31 °C, respectively. Histology analysis presented the overall urinary structure with a total outer diameter of 10 mm from the mucosal layer to the muscle layer. The MT staining revealed that both control and treated groups had similar amounts of collagen components without thermal damage. The current study demonstrated that the non-ablative 980 nm laser could warrant an effective method of treating SUI with no or minimal thermal damage.
Tracheal stenosis or granuloma is one of the most frequent complications after tracheostomy as a result of inflammatory, traumatic and fibrotic responses. Recently, multidisciplinary treatment has been used for tracheal stenosis and granuloma such as bronchoscopy, balloon dilation and tracheal surgeries. However, current treatments have risk of morbidity and may worsen the situation with high recurrence rate. The purpose of this study is to develop a novel combined treatment of photobiomodulation (PBM) and phlorotannin (PT) to prevent stenosis and granuloma formation after tracheal injury. The therapeutic effect of the combined treatment was evaluated on transforming growth factor (TGF)-beta-stimulated human tracheal fibroblasts and the developed tracheostomy rodent models. A 405 nm wavelength light was applied for PBM in a continuous-wave mode after treatment with Ecklonia cava-derived PT. MTT assay and western blot analysis showed that 12 J/cm2 of PBM and 100 µg/ml of PT were hardly cytotoxic (less than 20%). The combined treatment significantly inhibited cell migration and suppressed the expressions of alpha-smooth muscle actin and type-1 collagen via the downregulation of SMAD 2/3 and MAPK signaling pathways. Moreover, the proposed combined treatment showed promoted healing of tracheal fenestration wounds by modulating inflammation and overexpressed fibrotic activities on the developed tracheostomy rodent models. Therefore, the combination of PBM and PT demonstrates therapeutic potential for preventing tracheal stenosis and granuloma after tracheostomy.
Tissue analysis is pivotal research to determine the pathological properties that occur after the wound healing process. Several staining techniques to understand the morphology of scar tissue are widely used, such as staining with HE (Hematoxylin and Eosin), picrosirius red, and Masson's Trichome. Tissue staining using hematoxylin and eosin has several limitations: labor-intensive, time-consuming, high memory, and cost. Besides that, used a whole slide image to analyze the scar lesion can be more challenging. Hence, we used deep learning to automatically classify and localize scar lesions in the whole slide image based on object instance segmentation. Deep learning trained the patterns from the data representation through a neural network and convolution equations. Deep learning recognized 384 images in less than a minute with 99.89% accuracy. Therefore, the proposed deep learning method can be time- and cost-effective to characterize the pathological feature of scar tissue for the objective histological analysis. In addition to confirming the scar's recognition in the qualitative analysis, the authors also performed a quantitative analysis to obtain information from the scar tissue, such as collagen density from color extraction and collagen directional variance. Segmentation analysis is also used to determine the morphological structure in scar tissue compared to normal tissue. The analysis results can determine various further therapeutic methods to reduce or even eliminate scars on urological tissues in future works.
Tissue analysis needs to determine the pathological properties that occur after wound healing process. Several staining techniques are widely used to understand the morphology of scar tissue, such as staining with hematoxylin and eosin (HE), picrosirius red, and Masson’s Trichome. In spite of the common staining technique, the tissue staining using HE has several limitations, such as labor-intensive, time-consuming, high memory and cost. Due to the limited of view, using the whole slide image is quite challenging to analyze the scar lesion. Hence, we developed a deep learning technique to simultaneously classify and characterize a scar lesion in the whole slide image, based on object instance segmentation. The deep learning trained the patterns from the data representation through neural network and convolution equations. The proposed technique recognized 384 images in less than a minute with 99.9% accuracy. Based on classification, quantitative analysis was performed to confirm the recognition of the scar based on the important features, such as collagen density and directional variance of collagen in scar area. After created the density map and directional variance map of collagen, the differences were almost 50% in normal and scar tissue. Therefore, the proposed deep learning method can be time- and cost-effective to characterize the pathological feature of scar tissue for the objective histological analysis. The analyses are expected to optimize various therapeutic methods to reduce or even eliminate scars on the skin.
Tracheal stomal stenosis is the most common complication that exists in patients after tracheostomy and prolonged intubation due to inflammatory and fibrotic responses. Various surgical techniques, such as Y-V and two circular incisions, have been attempted to minimize the stenosis. However, the current incision methods still have a risk of infection related to the enlarged stoma. The aim of this study is to develop a novel combined treatment of photobiomodulation (PBM) and phlorotannin (PT) to prevent the stenosis after tracheostomy. The therapeutic effect of the combined treatment was evaluated in tracheal fibroblast and the tracheostomy-developed rodent models. A 405 nm wavelength light was applied for PBM in a continuous-wave mode after treatment with Ecklonia cava-derived PT. MTT assay and western blot analysis showed that 12 J/cm2 of PBM and 100 μg/ml of PT were an insignificant cytotoxicity (less than 20%). The combined treatment significantly inhibited cell migration (51.7%) and suppressed the expressions of alpha-smooth muscle actin (38.3%) and type-1 collagen (54.2%) due to the downregulation of SMAD 2/3 and MAPK signalling pathways. In vivo tests confirmed that control group showed obstruction by tissue granulation in the incised tissue seven days after tracheostomy, whereas the combined group still maintained the stoma with minimal fibrosis. The current study demonstrated that PT-combined PBM could minimize the tracheal stomal stenosis in fibrotic conditions as a result of modulated fibrotic responses. Therefore, the combination of PBM and PT can warrant an effective prevention of the tracheal stomal stenosis post-tracheostomy.
Endoscopic biliary drainage including stent are a well-accepted clinical method for treating common bile duct (CBD) stenosis, but the treatment is only successful in the short term with stent exchange at 3 to 6 months. The current study aims to demonstrate a novel endoscopic retrograde cholangiopancreatography (ECRP)-guided laser ablation technique on CBD stenosis by using a balloon catheter-integrated diffusing applicator (BCDA) for uniform treatment of tubular tissue. A 532 nm laser system was employed with BCDA to thermally coagulate the CBD tissue. For performance assessments, photothermal and mechanical responses of the tissue were evaluated in both ex vivo and in vivo mini-pig models in terms of structural and histological variations after the laser treatment. The ex vivo results confirmed the irreversible thermal denaturation in liver tissue without carbonization. The in vivo mini-pig tests confirmed high durability of BCDA during the ERCP procedures and no evidence of CBD perforation. Histological analysis validated a thin circumferential coagulation in the CBD with no or minimal injury to the peripheral organs. The proposed BCDA-assisted treatment may be a feasible endoscopic ablation method for treating disease or disorder in a tubular tissue structure. Further in vivo studies will investigate CBD stenosis-developed animal models to evaluate the chronic response of the laser-treated CBD tissue as well as the recurrence of CBD stenosis.
Urethral stricture, also called urethral stenosis, is an abnormal narrowing of the urethra due to scarring on the urinary wall caused by inflammation or trauma. Although various surgical treatments have been developed such as urethrotomy and urethroplasty, the recurrence rate is almost 40 percent due to development of fibrosis in the urethra. The purpose of the current study was to evaluate the effect of phloroglucinol-assisted low-level laser therapy (LLLT) on inhibition of fibrosis in in vitro. LLLT is one of the promising methods for fibrosis treatment, and phloroglucinol has an anti-fibrotic effect. NIH/3T3 fibroblast cells were cultured in a medium containing transforming growth factor (TGF)-β1 to induce differentiation of the fibroblasts. Laser (635 nm at 375 mJ/cm2) and phloroglucinol (100 μg/ml) were treated separately or simultaneously after TGF-β1 was treated. MTT and BrdU assay presented the inhibitory effect of the combined treatment on cell proliferation. An inhibitory effect on migration speed was confirmed by wound healing assay. Confocal immunofluorescence images showed that the combined treatment inhibited the expression of α-smooth muscle action (sma) and type-1 collagen. Moreover, Western blot analysis demonstrated that α-sma, collagen-1, and TGF-β1 as the key factors of fibrosis were suppressed by the combined treatment. Therefore, phloroglucinol-assisted LLLT may be an effective treatment to prevent the recurrence of urethral stricture. In vivo tests will be conducted in canine models for pre-clinical evaluations.
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