Qian Wang

Pathological ocular neovascularization remains a leading cause of irreversible vision loss. This study investigates the role of Polo-like kinase 2 (PLK2) in coordinating the transition of endothelial cells into pathological phenotypes. Single-cell transcriptomic and clinical tissue analysis revealed that PLK2 is specifically enriched in the vascular component of human proliferative membranes and is markedly upregulated in human fibrovascular membrane tissue. PLK2 endothelial knockdown (eKD) inhibited vascular branching and tip cell filopodia formation in postnatal mice. Analysis of murine oxygen-induced retinopathy (OIR) single cell sequencing datasets identified PLK2 as a specific marker for actively sprouting tip cells and proliferative endothelial subpopulations during pathological neovascularization. PLK2 eKD or conditional knockout (eCKO) significantly attenuated pathological neovascularization in OIR. Additionally, PLK2 eKD inhibited laser-induced choroidal neovascularization (CNV) in adult mice. In vitro assays confirmed that PLK2 depletion via shRNA or CRISPR/Cas9 markedly inhibited endothelial cell proliferation, migration, and tube formation across various microvascular cells, whereas ectopic PLK2 overexpression promoted a robust pro-angiogenic phenotype. Mechanistically, PLK2 is a critical regulator of mitochondrial bioenergetics; genetic depletion led to a reduction of mitochondrial Complex I activity, decreased ATP levels, and impaired oxygen consumption, accompanied by mitochondrial membrane depolarization and the accumulation of reactive oxygen species (ROS). The anti-angiogenic effects and apoptotic induction caused by PLK2 deficiency were significantly rescued by glucose supplementation or antioxidant N-acetylcysteine (NAC) treatment. Thus, by maintaining mitochondrial bioenergetic efficiency and mitigating oxidative stress, PLK2 sustains the high-energy demands of pathological endothelial cell activation and sprouting. PLK2 represents a promising therapeutic strategy for pathological ocular angiogenesis.

Introduction: Adequate sedation is essential for paediatric patients undergoing invasive haematologic-oncologic procedures. The combination of propofol and remifentanil is commonly used yet is associated with respiratory depression. Esketamine has anaesthetic, analgesic and sympathomimetic properties and is known to cause less respiratory depression than other sedatives. This study aims to assess esketamine versus remifentanil in combination with propofol for invasive haematological-oncological procedures for paediatric patients. Methods and analysis: This prospective, randomised, double-blind, two-period crossover trial will include 80 paediatric patients aged 6–12 years, with American Society of Anesthesiologists Physical Status II to III, who are scheduled to undergo repeated invasive procedures including bone marrow aspirates and lumbar punctures under sedation. Participants will be randomised to two sequences: AB or BA. In sequence AB, children will receive propofol (3 mg/kg) and esketamine (0.5 mg/kg) in period 1 and propofol (3 mg/kg) and remifentanil (1 µg/kg) in period 2. In sequence BA, the order is reversed. The primary endpoint of this trial is the incidence of desaturation events, defined as SpO ₂ <90% during the sedation and recovery. The secondary endpoints include: (1) the interruption rate of the procedure; (2) total dose of propofol; (3) the number of times esketamine or remifentanil is added; (4) recovery time; (5) distress behaviour; (6) a composite of sedation-related adverse events, including psychotomimetic effects (hallucinations, nightmares, fatigue, dizziness and headache), nausea, vomiting, bradycardia and hypotension; (7) operator satisfaction; (8) patient satisfaction and (9) anaesthetist satisfaction. Ethics and dissemination: The trial was approved by the Ethics Committee of the Children’s Hospital of Soochow University (Approval No. 2025008). The results of this trial will be submitted for peer review and publication in a scientific journal. Trial registration number: ChiCTR2500098533.

Background: Flexible fiberoptic bronchoscopy is being increasingly carried out under general anesthesia in pediatric patients. However, the optimal anesthetic protocol still remains unclear. This study aims to test the hypothesis that the addition of low-dose rocuronium during laryngeal mask airway-assisted flexible fiberoptic bronchoscopy in children can enhance procedural outcomes and improve overall management. Methods: In this randomized, double-blind, placebo-controlled trial, 140 children aged 1–6 years scheduled for flexible fiberoptic bronchoscopy under general anesthesia will be randomly allocated in a 1:1 ratio to either the rocuronium group or the normal saline group ( n = 70 in each group). All children will receive intravenous propofol at a dose of 3 mg⋅kg ^–1 and remifentanil at 3 μg⋅kg ^–1 for anesthesia induction. This will be followed by either intravenous rocuronium at 0.3 mg⋅kg ^–1 or an equivalent volume of normal saline. Sevoflurane will be titrated to achieve the target anesthesia depth, while remifentanil will be initiated at a rate of 0.05 μg⋅kg ^–1 ⋅min ^–1 and subsequently adjusted as required. Mechanical ventilation will be performed via laryngeal mask airway and the bronchofiberscope is inserted through an adapter connected to the laryngeal mask airway. The primary endpoint, reflecting effectiveness of anesthesia, is the frequency of coughing episodes during the operation. Secondary endpoints include: (1) cumulative remifentanil dosage; (2) failure rate of flexible fiberoptic bronchoscopy operation; (3) hemodynamic changes; (4) cough scores in the post-anesthesia care unit; (5) procedure time; (6) discharge time; (7) adverse events; (8) satisfaction scores of bronchoscopist; (9) satisfaction scores of anesthesiologist; and (10) satisfaction scores of parents 24 h after the operation. Discussion: We hypothesize that the addition of low-dose rocuronium during laryngeal mask airway-assisted flexible fiberoptic bronchoscopy under general anesthesia may optimize operation conditions, reduce coughing frequency, lower remifentanil dosage and the failure rate, stabilize hemodynamic fluctuations, minimize adverse events and enhance satisfaction, thereby providing valuable reference for clinical anesthesia practice in these procedures. Study Protocol Registration: https://www.chictr.org.cn/showprojEN.html?proj=277486 , identifier [ChiCTR2500105407].

Mitochondria are central to endothelial cell activation and angiogenesis, with the RNA polymerase mitochondrial (POLRMT) serving as a key protein in regulating mitochondrial transcription and oxidative phosphorylation. In our study, we examined the impact of POLRMT on angiogenesis and found that its silencing or knockout (KO) in human umbilical vein endothelial cells (HUVECs) and other endothelial cells resulted in robust anti-angiogenic effects, impeding cell proliferation, migration, and capillary tube formation. Depletion of POLRMT led to impaired mitochondrial function, characterized by mitochondrial depolarization, oxidative stress, lipid oxidation, DNA damage, and reduced ATP production, along with significant apoptosis activation. Conversely, overexpressing POLRMT promoted angiogenic activity in the endothelial cells. In vivo experiments demonstrated that endothelial knockdown of POLRMT, by intravitreous injection of endothelial specific POLRMT shRNA adeno-associated virus, inhibited retinal angiogenesis. In addition, inhibiting POLRMT with a first-in-class inhibitor IMT1 exerted significant anti-angiogenic impact in vitro and in vivo. Significantly elevated expression of POLRMT was observed in the retinal tissues of streptozotocin-induced diabetic retinopathy (DR) mice. POLRMT endothelial knockdown inhibited pathological retinal angiogenesis and mitigated retinal ganglion cell (RGC) degeneration in DR mice. At last, POLRMT expression exhibited a substantial increase in the retinal proliferative membrane tissues of human DR patients. These findings collectively establish the indispensable role of POLRMT in angiogenesis, both in vitro and in vivo.

Irritable bowel syndrome (IBS) is a common functional bowel disorder characterized by abdominal pain and visceral hypersensitivity. Reducing visceral hypersensitivity is the key to effectively relieving abdominal pain in IBS. Increasing evidence has confirmed that the thalamic nucleus reuniens (Re) and 5-hydroxytryptamine (5-HT) neurotransmitter system play an important role in the development of colorectal visceral pain, whereas the exact mechanisms remain largely unclear. In this study, we found that high expression of the 5-HT_2B receptors in the Re glutamatergic neurons promoted colorectal visceral pain. Specifically, we found that neonatal maternal deprivation (NMD) mice exhibited visceral hyperalgesia and enhanced spontaneous synaptic transmission in the Re brain region. Colorectal distension (CRD) stimulation induced a large amount of c-Fos expression in the Re brain region of NMD mice, predominantly in glutamatergic neurons. Furthermore, optogenetic manipulation of glutamatergic neuronal activity in the Re altered colorectal visceral pain responses in CON and NMD mice. In addition, we demonstrated that 5-HT_2B receptor expression on the Re glutamatergic neurons was upregulated and ultimately promoted colorectal visceral pain in NMD mice. These findings suggest a critical role of the 5HT_2B receptors on the Re glutamatergic neurons in the regulation of colorectal visceral pain.

Background and purpose: Acute kidney injury (AKI) is a common serious complication in sepsis patients with a high mortality rate. This study aimed to develop and validate a predictive model for sepsis associated acute kidney injury (SA-AKI). Methods: In our study, we retrospectively constructed a development cohort comprising 733 septic patients admitted to eight Grade-A tertiary hospitals in Shanghai from January 2021 to October 2022. Additionally, we established an external validation cohort consisting of 336 septic patients admitted to our hospital from January 2017 to December 2019. Risk predictors were selected by LASSO regression, and a corresponding nomogram was constructed. We evaluated the model's discrimination, precision and clinical benefit through receiver operating characteristic (ROC) curves, calibration plots, decision curve analysis (DCA) and clinical impact curves (CIC) in both internal and external validation. Results: AKI incidence was 53.2% in the development cohort and 48.2% in the external validation cohort. The model included five independent indicators: chronic kidney disease stages 1 to 3, blood urea nitrogen, procalcitonin, D-dimer and creatine kinase isoenzyme. The AUC of the model in the development and validation cohorts was 0.914 (95% CI, 0.894-0.934) and 0.923 (95% CI, 0.895-0.952), respectively. The calibration plot, DCA, and CIC demonstrated the model's favorable clinical applicability. Conclusion: We developed and validated a robust nomogram model, which might identify patients at risk of SA-AKI and promising for clinical applications.

The mitochondrial integrity and function in endothelial cells are essential for angiogenesis. TIMM44 (translocase of inner mitochondrial membrane 44) is essential for integrity and function of mitochondria. Here we explored the potential function and the possible mechanisms of TIMM44 in angiogenesis. In HUVECs, human retinal microvascular endothelial cells and hCMEC/D3 brain endothelial cells, silence of TIMM44 by targeted shRNA largely inhibited cell proliferation, migration and in vitro capillary tube formation. TIMM44 silencing disrupted mitochondrial functions in endothelial cells, causing mitochondrial protein input arrest, ATP reduction, ROS production, and mitochondrial depolarization, and leading to apoptosis activation. TIMM44 knockout, by Cas9-sgRNA strategy, also disrupted mitochondrial functions and inhibited endothelial cell proliferation, migration and in vitro capillary tube formation. Moreover, treatment with MB-10 (“MitoBloCK-10”), a TIMM44 blocker, similarly induced mitochondrial dysfunction and suppressed angiogenic activity in endothelial cells. Contrarily, ectopic overexpression of TIMM44 increased ATP contents and augmented endothelial cell proliferation, migration and in vitro capillary tube formation. In adult mouse retinas, endothelial knockdown of TIMM44, by intravitreous injection of endothelial specific TIMM44 shRNA adenovirus, inhibited retinal angiogenesis, causing vascular leakage, acellular capillary growth, and retinal ganglion cells degeneration. Significant oxidative stress was detected in TIMM44-silenced retinal tissues. Moreover, intravitreous injection of MB-10 similarly induced oxidative injury and inhibited retinal angiogenesis in vivo. Together, the mitochondrial protein TIMM44 is important for angiogenesis in vitro and in vivo, representing as a novel and promising therapeutic target of diseases with abnormal angiogenesis.

Mounting evidence indicates that microRNAs (miRNAs) play critical roles in various pathophysiological conditions and diseases, but the physiological roles of extracellular miRNAs on the disease-related ion channels remain largely unknown. Here, we showed that miR-1306-3p evoked action potentials and induced inward currents of the acutely isolated rat dorsal root ganglion (DRG) neurons. The miR-1306-3p–induced effects were significantly inhibited by A317491, a potent inhibitor of the P2X3 receptor (P2X3R), or disappeared after the knockdown of P2X3Rs in DRG neurons. We further identified R180, K315, and R52 as the miR-1306-3p interaction sites on the extracellular domain of P2X3Rs, which were distinct from the orthosteric ATP-binding sites. Intrathecal injection of miR-1306-3p produced visceral pain but not somatic pain in normal control rats. Conversely, intrathecal application of a miR-1306-3p antagomir and A317491 significantly alleviated visceral pain in a rat model of chronic visceral pain. Together, our findings suggest that miR-1306-3p might function as an endogenous ligand to activate P2X3Rs, eventually leading to chronic visceral pain.

Background Melanoma is an aggressive form of skin cancer worldwide. Phosphoinositide-3-kinase regulatory subunit 2 (PIK3R2) exerts carcinogenic roles in various tumors. So far, the function and mechanism of PIK3R2 in melanoma are not been fully clarified. Objective We aimed to clarify the role of PIK3R2 in melanoma. Methods PIK3R2 expressions in melanoma clinical tissues and melanoma cells were measured using quantitative real-time PCR and Western blot. In addition, PIK3R2 expressions in different tumor stages of melanoma were determined by immunohistochemistry assay. Meanwhile, PIK3R2 function was evaluated using loss or gain-of-function assays, Cell Counting Kit-8 assay, flow cytometry, and Transwell analysis. Furthermore, PIK3R2 mechanism in melanoma was assessed by a series of rescue experiments. Results PIK3R2 was highly expressed in melanoma tissues and cells, and PIK3R2 expressions were the highest in Stage IV. Functionally, PIK3R2 knockdown repressed melanoma cell proliferation, invasion, epithelial-mesenchymal transition, and facilitated cell apoptosis. Also, PIK3R2 overexpression produced an opposite trend. Mechanistically, PIK3R2 facilitated melanoma progression by activating PI3K/AKT/NF-κB pathway. Furthermore, PIK3R2 knockdown restrained the melanoma tumor growth in vivo. Conclusions PIK3R2 aggravated melanoma by activating PI3K/AKT/NF-κB pathway, prompting that PIK3R2 might be a therapeutic target for melanoma.