Integrating spatial transcriptomics and single-cell RNA-sequencing reveals the alterations in epithelial cells during nodular formation in benign prostatic hyperplasia.

OBJECTIVE: Proliferative nodular formation represents a characteristic pathological feature of benign prostatic hyperplasia (BPH) and serves as the primary cause for prostate volume enlargement and consequent lower urinary tract symptoms (LUTS). Its specific mechanism is largely unknown, although several cellular processes have been reported to be involved in BPH initiation and development and highlighted the crucial role of epithelial cells in proliferative nodular formation. However, the technological limitations hinder the in vivo investigation of BPH patients. METHODS: The robust cell type decomposition (RCTD) method was employed to integrate spatial transcriptomics and single cell RNA sequencing profiles, enabling the elucidation of epithelial cell alterations during nodular formation. Immunofluorescent and immunohistochemical staining was performed for verification. RESULTS: The alterations of epithelial cells during the formation of nodules in BPH was observed, and a distinct subgroup of basal epithelial (BE) cells, referred to as BE5, was identified to play a crucial role in driving this progression through the hypoxia-induced epithelial-mesenchymal transition (EMT) signaling pathway. BE5 served as both the initiating cell during nodular formation and the transitional cell during the transformation from luminal epithelial (LE) to BE cells. A distinguishing characteristic of the BE5 cell subgroup in patients with BPH was its heightened hypoxia and upregulated expression of FOS. Histological verification results confirmed a significant association between c-Fos expression and key biological processes such as hypoxia and cell proliferation, as well as the close relationship between hypoxia and EMT in BPH tissues. Furthermore, a strong link between c-Fos expression and the progression of BPH was also been validated. Additionally, notable functional differences were observed in glandular and stromal nodules regarding BE5 cells, with BE5 in glandular nodules exhibiting enhanced capacities for EMT and cell proliferation characterized by club-like cell markers. CONCLUSIONS: This study elucidated the comprehensive landscape of epithelial cells during in vivo nodular formation in patients, thereby offering novel insights into the initiation and progression of BPH.

A modified sling mid-urethral suspension + subcutaneous tunnel-double point fixation technique for male stress urinary incontinence: a pilot study.

OBJECTIVES: The aim of this study was to demonstrate the feasibility of a modified sling mid-urethral suspension + subcutaneous tunnel-double point fixation technique for the treatment of male urinary incontinence and to preliminarily evaluate the short-term clinical efficacy of this technique. PATIENTS AND METHODS: The clinical data of patients treated with the modified sling mid-urethral suspension + subcutaneous tunnel-double point fixation technique using a Pelvimesh pelvic floor repair patch as a sling were collected. The primary evaluation criteria were surgery-related indicators and daily urinal pad usage before and after treatment, and the secondary evaluation criteria were the corresponding incontinence scores and the results of surgery-related questionnaires. RESULTS: After 1-12 months of follow-up, seven patients were clinically cured. Follow-up 1 month after surgery showed that one patient (14.3%) used one pad daily, and six patients (85.7%) did not need pads. The incontinence quality of life questionnaire (I-QOL) scores at 3 days and 1 month postoperatively were 89.4 ± 2.5 and 88.1 ± 6.7, respectively, which were significantly higher than the preoperative scores (31.5 ± 18.9) (P < 0.05). The scores of the International Continence Control Association Incontinence Questionnaire Short Form (ICI-Q-SF) at 3 days and 1 month postoperatively were 3.2 ± 0.9 and 4.2 ± 1.7, respectively, which were significantly lower than the preoperative scores of 19.4 ± 5.0 (P < 0.05). In addition, the results of the surgery-related questionnaires were positive. No serious complications occurred in any of the patients. CONCLUSION: The modified sling mid-urethral suspension + subcutaneous tunnel-double point fixation technique for the treatment of male urinary incontinence patients is safe, effective, minimally invasive, and has few complications. However, further validation in large sample, randomized, comparative, and longer-term follow-up studies is still needed.

ISGylation of NF-κBp65 by SCFFBXL19 E3 Ligase Diminishes Endothelial Inflammation.

BACKGROUND: NF-κB (nuclear factor kappa B) plays a pivotal role in endothelial cell (EC) inflammation. Protein ISGylation is regulated by E3 ISG15 (interferon-stimulated gene 15) ligases; however, ISGylation of NF-κBp65 and its role in EC functions have not been investigated. Here, we investigate whether p65 is ISGylated and the role of its ISGylation in endothelial functions. METHODS: In vitro ISGylation assay and EC inflammation were performed. EC-specific transgenic mice were utilized in a murine model of acute lung injury. RESULTS: We find that NF-κBp65 is ISGylated in resting ECs and that the posttranslational modification is reversible. TNFα (tumor necrosis factor alpha) and endotoxin stimulation of EC reduce p65 ISGylation, promoting its serine phosphorylation through reducing its association with a phosphatase WIP1 (wild-type p53-induced phosphatase 1). Mechanistically, an SCF (Skp1-Cul1-F-box) protein E3 ligase SCFFBXL19 is identified as a new ISG15 E3 ligase that targets and catalyzes ISGylation of p65. Depletion of FBXL19 (F-box and leucine-rich repeat protein 19) increases p65 phosphorylation and EC inflammation, suggesting a negative correlation between p65 ISGylation and phosphorylation. Moreover, EC-specific FBXL19 overexpressing humanized transgenic mice exhibit reduced lung inflammation and severity of experimental acute lung injury. CONCLUSIONS: Together, our data reveal a new posttranslational modification of p65 catalyzed by a previously unrecognized role of SCFFBXL19 as an ISG15 E3 ligase that modulates EC inflammation.

A low-cost compact blood enzyme analyzer based on optical sensing for point-of-care liver function testing.

Serum alanine aminotransferase (ALT) is the most sensitive indicator of liver function; therefore, in clinical practice, its detection has diagnostic significance. However, hepatotoxicity monitoring is constrained in resource-limited areas due to the high cost and expensive equipment. This article describes a low-cost, compact blood enzyme analyzer (BEA) for point-of-care (PoC) liver function testing that uses reflection photometry to quantitate the colorimetric enzyme assay results. Moreover, the analyzer incorporates rapid and constant 37 °C temperature control to ensure that human serum enzymes remain active. The BEA was used to evaluate ALT in 50 whole blood samples using PoC photochemical test strips. The test results showed a linear correlation coefficient of 0.9749 compared with a clinical-specific biochemical analyzer and coefficients of variation (CV)% less than 5%. It has a low detection limit of 3.62 U L-1 and a wide detection range of 4-480 U L-1. In addition, the BEA enables smartphone access to the Internet of Medical Things (IoMT) via Bluetooth to facilitate active chronic disease management or remote diagnosis in PoC settings. Therefore, the BEA is a promising system for health management using the IoMT.

Organ-on-a-chip: A new tool for in vitro research.

Organ-on-a-chip (OOC, organ chip) technology can closely simulate the human microenvironment, synthesize organ-like functional units on a fluidic chip substrate, and simulate the physiology of tissues and organs. It will become an increasingly important platform for in vitro drug development and screening. Most importantly, organ-on-a-chip technology, incorporating 3D cell cultures, overcomes the traditional drawbacks of 2D (flat) cell-culture technology in vitro and in vivo animal trials, neither of which generate completely reliable results when it comes to the actual human subject. It is expected that organ chips will allow huge reductions in the incidence of failure in late-stage human trials, thus slashing the cost of drug development and speeding up the introduction of drugs that are effective. There have been three key enabling technologies that have made organ chip technology possible: 3D bioprinting, fluidic chips, and 3D cell culture, of which the last has allowed cells to be cultivated under more physiologically realistic growth conditions than 2D culture. The fusion of these advanced technologies and the addition of new research methods and algorithms has enabled the construction of chip types with different structures and different uses, providing a wide range of controllable microenvironments, both for research at the cellular level and for more reliable analysis of the action of drugs on the human body. This paper summarizes some research progress of organ-on-a-chip in recent years, outlines the key technologies used and the achievements in drug screening, and makes some suggestions concerning the current challenges and future development of organ-on-a-chip technology.

3D printed microfluidic Coulter counter for blood cell analysis.

Coulter counters are ubiquitous in everyday life; however, with reduced orifice size shrinkage, there is an increased risk of clogging. Herein, a 3D microfluidic printing-based single-use kit is presented for analyzing biological samples and performing accurate Coulter cell count analysis (e.g., white blood cells in the blood). The gem hole eliminates the traditional design concept of integration inside the detection instrument, innovatively causing it to be independent from the analysis instrument. Further, integrating the hole in the analysis box enables the design of a separate detection module. The analysis box is disposable, convenient, and hygienic; avoids cross-infection; solves the problem of clogging of tiny holes from a new perspective; and no longer requires uneconomical and inconvenient methods, such as flushing, cauterization, and fluid focusing, through solid water flow. Further development of the newly designed 3D printing analysis box can enable its extensive use in POCT (point-of-care) detection scenarios. Moreover, through mass production, the issue of cost will be eliminated.

CTHRC1 facilitates bladder cancer cell proliferation and invasion through regulating the PI3K/Akt signaling pathway.

INTRODUCTION: Emerging evidence has illustrated that Collagen triple helix repeat containing 1 (CTHRC1) is crucial for tumorigenesis and development. However, the effects of CTHRC1 on bladder cancer progression remain largely unclear. Here, we aim to investigate the function and mechanism of CTHRC1 in behaviors of bladder cancer cells in vitro and in vivo. MATERIAL AND METHODS: Interference assays were applied to determine the biological functions of CTHRC1. The expression of CTHRC1 was examined by quantitative real time-PCR (qRT-PCR), Western blot and immunohistochemical (IHC) analysis. Effects of CTHRC1 on proliferation, migration and invasion were evaluated by CCK-8, colony formation, flow cytometry, EdU staining, wound healing, transwell and western blot assays. Bladder cancer cells transfected with sh-CTHRC1 were injected into nude mice to explore the effect of CTHRC1 on tumorigenesis in vivo. RESULTS: CTHRC1 expression was increased in bladder cancer tissues and cell lines compared with normal controls, and associated with advanced clinical stage and lymph node metastasis. Also, patients with high levels of CTHRC1 expression were found to have a poor prognosis. Knockdown of CTHRC1 alleviated bladder cancer cell proliferation, migration and invasion in vitro and impeded tumorigenesis in vivo. Moreover, mechanistic investigation indicated that CTHRC1 could regulate the PI3K/Akt signaling pathway. CONCLUSIONS: Our data demonstrated that CTHRC1 played an oncogenic role in bladder cancer by modulating the PI3K/Akt signaling pathway, which sheds novel light on diagnosis and treatment of bladder cancer.

Hsa_circ_0065217 promotes growth and metastasis of renal cancer through regulating the miR-214-3p-ALPK2 axis.

Circular RNA (circRNA) deregulation impacts on normal cell physiology leading to malignant phenotypic changes. Here, we determined the function of the circRNA, hsa_circ_0065217 in malignant renal cell carcinoma (RCC). Hsa_circ_0065217 was abundantly expressed in RCC tissue and cell lines, and its expression linked to advanced TNM stages, large tumor sizes, and lymph-node metastasis. Hsa_circ_0065217 silencing reduced in vitro RCC cell-line growth and aggressiveness. Mechanistically, hsa_circ_0065217 promoted alpha protein kinase 2 (ALPK2) expression via its competing endogenous RNA (ceRNA) activity toward miR-214-3p. Moreover, ALPK2 overexpression reversed hsa_circ_0065217 knockdown effects on RCC cell-line malignancy. Thus, hsa_circ_0065217/miR-214-3p/ALPK2 signaling putatively promotes RCC tumorigenesis and is a putative RCC treatment target.

Bilateral electrical pudendal nerve stimulation as additional therapy for lower urinary tract dysfunction when stage II sacral neuromodulator fails: a case report.

BACKGROUND: Sacral neuromodulation (SNM) has become an effective therapy for patients with lower urinary tract dysfunction (LUTD) who do not respond to conservative treatment. However, an effective treatment strategy for patients who fail SNM has not yet been identified. An option for LUTD is needed when the clinical response to the SNM diminishes. CASE PRESENTATION: A 51-year-old Chinese man presented to an outpatient clinic complaining of difficulty in urination for > 3 years. The patient also complained of urinary frequency and urgency, accompanied by perineal discomfort. He was diagnosed with LUTD based on his symptoms and previous examinations. The patient underwent sacral neuromodulation with a permanent implantable pulse generator (IPG) (provided free of charge by Chengnuo Medical Technology Co., Ltd.; General Stim, Hangzhou, China) in the left buttock, as he participated in the company's clinical trial to test the long-term effects of IPG. He reported loss of efficacy of the device 3 months after the implantation. We performed bilateral electrical pudendal nerve stimulation (EPNS) therapy for him. After 2 weeks of treatment, he began to report smooth voiding within 2 h after EPNS, and a moderate improvement in urinary frequency, urgency, and perineal discomfort. After 4 weeks of EPNS, the patient reported > 50% improvement in his urination, evaluated with the short form of the International Consultation on Incontinence Questionnaire for Male Lower Urinary Tract Symptoms. He reported smooth voiding, moderate improvements in urinary frequency and urgency, and the disappearance of the perineal discomfort. He also reported improved sleep and erections. The patient was discharged after 8 weeks of EPNS treatment. CONCLUSION: EPNS could be an option as an additional therapy for patients with LUTD who have failed SNM.

Clinical correlation between serum YKL-40 protein level and recurrence of non-muscle invasive bladder cancer.

BACKGROUND: To explore the clinical correlation between serum YKL-40 protein level and recurrence of non-muscle invasive bladder cancer (NMIBC). METHODS: Totally 76 NMIBC patients (including 34 patients with confirmed recurrence and 42 patients without recurrence during the 2-year post-operative follow-up) and 31 healthy volunteers were recruited in this study. Blood samples were collected early in the morning, and serum YKL-40 protein levels of all these patients were analysed by using enzyme-linked immunosorbent assay (ELISA). RESULTS: Serum YKL-40 protein levels were significantly higher in NMIBC patients than in healthy controls (P<0.001). Meanwhile, serum YKL-40 protein levels were found to be significantly higher in patients with recurrent NMIBC than those without tumor recurrence (P=0.001). CONCLUSIONS: Serum YKL-40 protein can be a reliable molecular marker for the clinical diagnosis of NMIBC recurrence. In particular, it can inform the post-operative management.

Metformin inhibits cell growth by upregulating microRNA-26a in renal cancer cells.

Accumulating evidence suggests that metformin, a biguanide class of anti-diabetic drugs, possesses anti-cancer properties and may reduce cancer risk and improve prognosis. However, the mechanism by which metformin affects various cancers, including renal cancer still unknown. MiR-26a induces cell growth, cell cycle and cell apoptosis progression via direct targeting of Bcl-2, clyclin D1 and PTEN in cancer cells. In the present study, we used 786-O human renal cancer cell lines to study the effects and mechanisms of metformin. Metformin treatment inhibited RCC cells proliferation by increasing expression of miR-26a in 786-O cells (P < 0.05). As a result, protein abundance of Bcl-2 and cyclin D1 was decreased and PTEN was increased in cells exposed to metformin. Also over-expression of miR-26a can inhibited cell proliferation by down-regulating Bcl-2, cyclin D1 and up-regulating PTEN expression. Therefore, these data for the first time provide novel evidence for a mechanism that the anticancer activities of metformin are due to upregulation of miR-26a and affect its downstream target gene.

Imidazo[4,5-b]pyridines as corticotropin releasing factor receptor ligands.

A series of high affinity CRF receptor ligands with an imidazo[4,5-b]pyridine core is described. Individual analogues were synthesized and tested in a rat CRF receptor binding assay. The best compounds were further tested in the dog N-in-1 pharmacokinetic model to assess plasma levels at 1mg/kg (po) and in the rat situational anxiety model to assess anxiolytic efficacy at 3mg/kg (po). The structure-activity relationships for good receptor binding affinity are described herein.