[Evaluation of the lower urinary tract function using voiding spot assay in mice].

The objective of present study was to develop a simple and reliable voiding spot assay (VSA) system to evaluate the lower urinary tract function of mice, and to establish it as a standardized protocol. Ultraviolet (UV) light was used to screen out the filter paper without autofluorescence and with optimal urine diffusion properties. Next, the appropriate wavelength of UV was determined based on the quality of the photographic image of urine spots on the filter paper. To confirm that the urine stain area on the filter paper was correlated with the amount of urine, a volume-area standard curve was constructed. The utility of this VSA system was validated using female wild-type C57BL/6J mice aged 12-13 weeks, and the data generated under identical procedural settings were compared among laboratories. Furthermore, this VSA system was employed to analyze the changes in voiding patterns in mice with urinary tract infections or transportation stress. No. 4 filter paper with a thickness of 0.7 mm was identified as the most suitable material for VSA, exhibiting no autofluorescence and facilitating optimal urine diffusion. The filter paper retained its integrity during the assay, and there was a linear correlation between urine volume and stained area under 365 nm UV light. Utilizing this VSA system, we determined that female wild-type C57BL/6J mice produced approximately 695.8 µL total urine and 5.5 primary voiding spots (PVS) with an average size of 126.4 µL/spot within 4-h period. Over 84% of PVS volumes ranged from 20 to 200 µL. Notably, PVS volumes of mice were similar across different laboratories. Mice with urinary tract infections or transportation stress exhibited significant changes in VSA parameters, including increased voiding frequency, PVS number, and decreased PVS volume. Therefore, this VSA system can be used to evaluate the urinary function of normal mice, as well as those with urinary tract infection or transportation stress.

Suppressive stroma-immune prognostic signature impedes immunotherapy in ovarian cancer and can be reversed by PDGFRB inhibitors.

BACKGROUND: As the most lethal gynecologic cancer, ovarian cancer (OV) holds the potential of being immunotherapy-responsive. However, only modest therapeutic effects have been achieved by immunotherapies such as immune checkpoint blockade. This study aims to propose a generalized stroma-immune prognostic signature (SIPS) to identify OV patients who may benefit from immunotherapy. METHODS: The 2097 OV patients included in the study were significant with high-grade serous ovarian cancer in the III/IV stage. The 470 immune-related signatures were collected and analyzed by the Cox regression and Lasso algorithm to generalize a credible SIPS. Correlations between the SIPS signature and tumor microenvironment were further analyzed. The critical immunosuppressive role of stroma indicated by the SIPS was further validated by targeting the major suppressive stroma component (CAFs, Cancer-associated fibroblasts) in vitro and in vivo. With four machine-learning methods predicting tumor immune subtypes, the stroma-immune signature was upgraded to a 23-gene signature. RESULTS: The SIPS effectively discriminated the high-risk individuals in the training and validating cohorts, where the high SIPS succeeded in predicting worse survival in several immunotherapy cohorts. The SIPS signature was positively correlated with stroma components, especially CAFs and immunosuppressive cells in the tumor microenvironment, indicating the critical suppressive stroma-immune network. The combination of CAFs' marker PDGFRB inhibitors and frontline PARP inhibitors substantially inhibited tumor growth and promoted the survival of OV-bearing mice. The stroma-immune signature was upgraded to a 23-gene signature to improve clinical utility. Several drug types that suppress stroma-immune signatures, such as EGFR inhibitors, could be candidates for potential immunotherapeutic combinations in ovarian cancer. CONCLUSIONS: The stroma-immune signature could efficiently predict the immunotherapeutic sensitivity of OV patients. Immunotherapy and auxiliary drugs targeting stroma could enhance immunotherapeutic efficacy in ovarian cancer.

Minimizing the variables of voiding spot assay for comparison between laboratories.

The voiding spot assay (VSA) is increasingly being adopted as a standard method for assessing mouse urinary function. However, VSA outcomes are highly sensitive to housing environment and procedural parameters. Many variables exist among laboratories, including analytical software, type of daily housing cage, transportation, and the time of the day. Some of these variables, such as the time of VSA and analytical software, have been shown to result in inconsistency and incomparability of data. In this study, we evaluated whether the results of VSA can be compared across laboratories by minimizing these variables. We found that analytical tools between Fiji and MATLAB are in good agreement in the quantification of VSA parameters, especially primary voiding spot (PVS) parameters. Unexpectedly, we found that mice housed in different daily home cages did not alter voiding patterns in a standard VSA cage. Nonetheless, we still recommend acclimation when performing VSA in unfamiliar cages. Notably, mice are highly sensitive to transportation and the time in the morning versus afternoon, which can induce significant changes in voiding patterns. Therefore, a standardized period among laboratories and allowing 2-3 days of rest for mice acclimation after transportation are necessary for VSA. Finally, we performed VSA using identical procedural parameters in two laboratories from two geographical locations to compare the results of VSA and concluded that it is possible to generate limited comparable VSA data, such as PVS volume.

[Structural changes and functional evaluation of the thymus in aging mouse induced by D-galactose].

Objective To explore the structural changes and functional changes of the thymus in aging mouse induced by D-galactose, and to explore a suitable method for establishing an aging mouse model of the thymus. Methods Thymus aging mouse models were established, female C57BL/6 mice were randomly divided into control group, [500 mg/(kg.d)] D-galactose treatment group, and [1000 mg/(kg.d)] D-galactose treatment group, with 8 mice in each group. The mice in the D-galactose treatment group were injected with 500 mg/kg and 1000 mg/kg of D-galactose subcutaneously on the back of the neck every day, while the mice in the control group were injected with the same amount of normal saline every day. After 56 days of continuous administration, the mice were sacrificed to take the thymus to observe the gross thymus morphology and calculate the thymus index. Then the thymus structure were observed by HE staining, and CD4/CD8 positive thymocytes were detected by flow cytometry to evaluate the immune function of the thymus. Later, thymus aging mouse models with different treatment time were established. Female C57BL/6 mice were randomly divided into control group and [1000 mg/(kg.d )] D-galactose treatment group, with 24 mice in each group. The mice were sacrificed after 6 weeks, 9 weeks, and 12 weeks treatment. The structure of thymus was observed by HE staining. The contents of thymosin ß4, thymosin α1, and thymopoietin in plasma were determined by ELISA. Results D-galactose treatment can induce mouse thymus senescence, atrophy of thymus, decrease of thymus index, disorder of thymus structure and impaired immune function. In the [1000 mg/(kg.d)] D-galactose treatment group, the atrophy of the thymic medulla of mice was more obvious, with the disappeared cortical and medullary boundary, decreased CD4+CD8+ thymocytes and increased CD4+CD8- thymocytes. The thymus aging mouse models with different treatment time showed the atrophied thymus, decreased thymus index, constricted thymus medulla, blurred boundary of cortex and medulla, decreased plasma thymosin α1 and impaired thymic secretion function. Thymus senescence was most obvious 12 weeks after D-galactose treatment. Conclusion D-galactose can induce the atrophy of the thymus, the thymus index decreases, consticted thymus medulla and blurred boundary of the cortex and medulla,and result in impaired thymic immune and secretory functions.A subcutaneous injection of 1000 mg/kg D-galactose on the back of the neck every day for 12 consecutive weeks is a suitable method to establish a thymus aging model.

A Soft Robotic Intervention for Gait Enhancement in Older Adults.

Falls continue to be a major safety and health concern for older adults. Researchers reported that increased gait variability was associated with increased fall risks. In the present study, we proposed a novel wearable soft robotic intervention and examined its effects on improving gait variability in older adults. The robotic system used customized pneumatic artificial muscles (PAMs) to provide assistive torque for ankle dorsiflexion during walking. Twelve older adults with low fall risks and twelve with medium-high fall risks participated in an experiment. The participants were asked to walk on a treadmill under no soft robotic intervention, inactive soft robotic intervention, and active soft robotic intervention, and their gait variability during treadmill walking was measured. The results showed that the proposed soft robotic intervention could reduce step length variability for elderly people with medium-high fall risks. These findings provide supporting evidence that the proposed soft robotic intervention could potentially serve as an effective solution to fall prevention for older adults.

Dual Role of Reactive Oxygen Species and their Application in Cancer Therapy.

Reactive oxygen species (ROS) play a dual role in the initiation, development, suppression, and treatment of cancer. Excess ROS can induce nuclear DNA, leading to cancer initiation. Not only that, but ROS also inhibit T cells and natural killer cells and promote the recruitment and M2 polarization of macrophages; consequently, cancer cells escape immune surveillance and immune defense. Furthermore, ROS promote tumor invasion and metastasis by triggering epithelial-mesenchymal transition in tumor cells. Interestingly, massive accumulation of ROS inhibits tumor growth in two ways: (1) by blocking cancer cell proliferation by suppressing the proliferation signaling pathway, cell cycle, and the biosynthesis of nucleotides and ATP and (2) by inducing cancer cell death via activating endoplasmic reticulum stress-, mitochondrial-, and P53- apoptotic pathways and the ferroptosis pathway. Unfortunately, cancer cells can adapt to ROS via a self-adaption system. This review highlighted the bidirectional regulation of ROS in cancer. The study further discussed the application of massively accumulated ROS in cancer treatment. Of note, the dual role of ROS in cancer and the self-adaptive ability of cancer cells should be taken into consideration for cancer prevention.

NMDAR in bladder smooth muscle is not a pharmacotherapy target for overactive bladder in mice.

Overactive bladder (OAB) is a common condition that affects a significant patient population. The N-methyl-D-aspartate receptor (NMDAR) has a role in developing bladder overactivity, pharmacological inhibition of which inhibits bladder overactivity. The common pathogenesis of OAB involves bladder smooth muscle (BSM) overactivity. In this study, a smooth muscle-specific NMDAR knockout (SMNRKO) mouse model was generated. The bladders from SMNRKO mice displayed normal size and weight with an intact bladder wall and well-arranged BSM bundles. Besides, SMNRKO mice had normal voiding patterns and urodynamics and BSM contractility, indicating that NMDAR in BSM was not essential for normal physiological bladder morphology and function. Unexpectedly, cyclophosphamide (CYP)-treated SMNRKO and wild-type (WT) mice had similar pathological changes in the bladder. Furthermore, SMNRKO mice displayed similar altered voiding patterns and urodynamic abnormalities and impaired BSM contractility compared with WT mice after CYP treatment. MK801 partially reversed the pathological bladder morphology and improved bladder dysfunction induced by CYP, but did not cause apparent differences between WT mice and SMNRKO mice, suggesting that NMDAR in BSM was not involved in pathological bladder morphology and function. Moreover, the direct instillation of NMDAR agonists or antagonists into the CYP-induced OAB did not affect bladder urodynamic function, indicating that NMDAR in BSM was not the pharmacotherapy target of MK801 for CYP-induced cystitis. The findings indicated that NMDAR in BSM was not essential for normal physiological or pathological bladder morphology and function, and MK801 improving pathological bladder function was not mediated by an action on NMDAR in BSM.

Molecular pathways underlying tissue injuries in the bladder with ketamine cystitis.

Ketamine cystitis (KC) is a chronic bladder inflammation leading to urinary urgency, frequency, and pain. The pathogenesis of KC is complicated and involves multiple tissue injuries in the bladder. Recent studies indicated that urothelium disruption, lamina propria fibrosis and inflammation, microvascular injury, neuropathological alterations, and bladder smooth muscle (BSM) abnormalities all contribute to the pathogenesis of KC. Ketamine has been shown to induce these tissue injuries by regulating different signaling pathways. Ketamine can stimulate antiproliferative factor, adenosine triphosphate, and oxidative stress to disrupt urothelium. Lamina propria fibrosis and inflammation are associated with the activation of cyclooxygenase-2, nitric oxide synthase, immunoglobulin E, and transforming growth factor ß1. Ketamine contributes to microvascular injury via the N-methyl-D aspartic receptor (NMDAR), and multiple inflammatory and angiogenic factors such as tumor necrosis factor α and vascular endothelial growth factor. For BSM abnormalities, ketamine can depress the protein kinase B, extracellular signal-regulated kinase, Cav1.2, and muscarinic receptor signaling. Elevated purinergic signaling also plays a role in BSM abnormalities. In addition, ketamine affects neuropathological alterations in the bladder by regulating NMDAR- and brain-derived neurotrophic factor-dependent signaling. Inflammatory cells also contribute to neuropathological changes via the secretion of chemical mediators. Clarifying the role and function of these signaling underlying tissue injuries in the bladder with KC can contribute to a better understanding of the pathophysiology of this disease and to the design of effective treatments for KC.

Comparison of open and intracorporeal modified ureterosigmoidostomy (Mainz II) after laparoscopic radical cystectomy with bladder cancer.

OBJECTIVE: To compare perioperative and oncologic outcomes of open modified ureterosigmoidostomy urinary diversion (OMUUD) and intracorporeal modified ureterosigmoidostomy urinary diversion (IMUUD) following laparoscopic radical cystectomy (LRC). PATIENTS AND METHODS: We retrospectively reviewed our single institutional collected database patients undergoing LRC from October 2011 to October 2019. The perioperative characteristics were compared between OMUUD and IMUUD, and overall survival (OS) and progression-free survival (PFS) were evaluated by the Kaplan-Meier method. RESULTS: Overall, 84 patients were included. OMUUD and IMUUD were performed in 63 (75%) and 21 (25%) patients, respectively. IMUUD patients demonstrated shorter postoperative length of stay (16.24 ± 3.91 days vs. 18.98 ± 7.41 days, P = 0.033), similar operation time (498.57 ± 121.44 vs. 462.24 ± 99.71, P = 0.175), similar estimated blood loss [400 (200-475) ml vs. 400 (200-700) ml, P = 0.095], and similar overall complication rate within 30 days (19.05% vs. 25.40%, P = 0.848) and 90 days (23.81% vs. 17.46%, P = 0.748). Complete urinary control rate was 87.3% (55/63) in the OMUUD group. In IMUUD, the complete urinary control rate was 90.5% (19/21). There was no significant difference in OS (χ2 = 0.015, P = 0.901) and PFS (χ2 = 0.107, P = 0.743) between the two groups. CONCLUSION: IMUUD postoperative recovery is faster; other perioperative outcomes and oncology results are not significantly different with OMUUD. It is indicated that IMUUD can be utilized safely and effectively in the urinary diversion after LRC.

Fabrication, antibacterial activity and cytocompatibility of quaternary ammonium chitooligosaccharide functionalized polyurethane membrane via polydopamine adhesive layer.

In this study, to enhance the antibacterial activity and cytocompatibility of the electrospinning polyurethane (PU) fibrous membrane, quaternary ammonium chitooligosaccharide (G-COS) was immobilized on the fibrous membrane surface via an intermediate layer of polydopamine (PDOPA) to obtain the G-COS functionalized PU (G-C-D-PU), as a control, chitooligosaccharide (COS) functionalized PU fibrous membrane (C-D-PU) was prepared, too. Surface composition, morphology, hydrophilicity and surface energy of the original and modified PU fibrous membranes were characterized, which revealed that the surface roughness and hydrophilicity of the PU fibrous membrane were obviously increased by modified with COS and G-COS, respectively. Antibacterial experiment against E. coli and S. aureus indicated that antibacterial activity of the G-C-D-PU fibrous membrane was markedly superior to that of pure PU and C-D-PU fibrous membranes. In vitro cells culture experiments revealed that the adhesion and proliferation of NIH-3T3 cells on the PU fibrous membrane were improved by successively immobilized with PDOPA and COS as well as G-COS with the concentration of 2 g/L and 6 g/L. Moreover, the G-C-D-PU fibrous membranes with relative high G-COS content were more beneficial to the enhancement of antibacterial activity, but on the contrary, those with relative low G-COS content were more in favor of cell attachment and proliferation.

Thermoelectric properties of n-type ZrNiSn prepared by rapid non-equilibrium laser processing.

The traditional manufacturing of thermoelectric (TE) modules is a complex process that requires a long processing time and is high cost. In this work, we introduce a novel one-step 3D printing technique for TE module manufacturing, which integrates the Self-propagation High-temperature Synthesis (SHS) with the Selective Laser Melting (SLM) method. As a demonstration of this technique, bulk ZrNiSn samples were successfully fabricated on a Ti substrate. The effect of SLM processing parameters, such as the laser power and the scanning speed, on the quality of the forming ZrNiSn layers was systematically studied and analyzed, and the optimal processing window for the SLM process was determined. Transport property measurements indicate that the SLM-processed ZrNiSn possesses the maximum thermoelectric figure of merit ZT of 0.39 at 873 K. The interface of the ZrNiSn with the Ti substrate shows good adherence and low contact resistivity. The work demonstrates the viability of the SHS-SLM method for rapid fabrication of TE materials, legs and even modules.

Antibacterial activity and cytocompatibility of chitooligosaccharide-modified polyurethane membrane via polydopamine adhesive layer.

The aim of this study was to provide a convenient surface modification method for polyurethane (PU) membrane and evaluate its influence on hydrophilicity, antibacterial activity and cell functions, which are the most important factors for wound dressings. For this purpose, chitooligosaccharide (COS) was modified onto the surface of PU membrane based on the self-polymerization of dopamine (DOPA). Surface composition, morphology, hydrophilicity and surface energy of the original and modified PU membranes were characterized. Surface roughness and hydrophilicity of the PU membrane were obviously increased by modified with polydopamine (PDOPA) and COS. Antibacterial experiment against Escherichia coli and Staphylococcus aureus indicated that antibacterial activity of PU membrane increased only slightly by modified with PDOPA, but increased significantly by further modified with COS. Cells culture results revealed that COS-functionalized PU membrane is more beneficial to the adhesion and proliferation of NIH-3T3 cells compared to the original and PDOPA-modified PU membranes.