Jove-Journal of Visualized Experiments最新文献

Symptomatic chronic subdural hematoma is treated by surgical evacuation followed by drain insertion for post-operative drainage. There is no international consensus on the location of the drain (subdural or subperiosteal), the type of drainage (passive or active suction), or the duration of drainage (hours or days). However, a growing body of literature highlights the risk of iatrogenic brain injury during subdural drain insertion, causing increased interest in the subperiosteal drainage technique, which has been suggested to be equally effective. There is no consensus on the optimal subperiosteal drain insertion technique, resulting in numerous technical variations in the published literature. Additionally, drain anchoring is crucial to prevent the drain from displacing away from the burr hole. To address both issues, this article presents a standardized method for subperiosteal drain insertion and a novel anchorage technique for drains. All necessary drain entry-, exit-, and anchorage points are clearly defined and marked prior to placement of local anesthesia and skin incision. The stepwise insertion and anchorage of the drain are thoroughly described and illustrated, as well as the removal of the drain after post-operative drainage is complete.

Intracerebral hemorrhage (ICH) can compromise the gastrointestinal tract and trigger intestinal injury, contributing to poor clinical outcomes. This study investigated the effects of lansoprazole on intestinal injury and neurological function in mice with ICH. Mice were stereotactically injected with collagenase to establish an ICH model. Neurological function, brain water content, blood-brain barrier permeability, intestinal injury, and intestinal permeability were evaluated. Levels of LPS and IL-1β in the blood and brain were also assayed. Lansoprazole treatment was associated with alleviated intestinal injury, reduced levels of LPS and IL-1β in blood and brain, decreased brain water content, and inhibited brain inflammation in ICH mice. While lansoprazole administration did not lead to improved performance in the Garcia, forelimb placing, and rotarod tests (short-term neurological function), it significantly enhanced learning and memory (long-term neurological function). These findings indicated that lansoprazole treatment was associated with improved long-term neurological outcomes and reduced intestinal injury in ICH mice.

Observational studies have reported associations between multiple sclerosis (MS) and hematologic malignancies (HM), but findings remain inconsistent. We conducted a bidirectional two-sample Mendelian randomization (MR) analysis using publicly available genome-wide association summary statistics for MS (n = 115,803) and HM (n = 218,792). Primary causal estimates were obtained with the inverse-variance-weighted (IVW) estimator, complemented by prespecified sensitivity analyses for heterogeneity and pleiotropy. Genetically proxied liability to MS was associated with higher odds of leukemia (unspecified subtype) (odds ratio [OR] 1.311, 95% confidence interval [CI] 1.002-1.716, P = 0.048) and Hodgkin lymphoma (HL) (OR 1.224, 95% CI 1.052-1.425, P = 0.009), with no evidence for other leukemia, lymphoma, or plasma-cell neoplasm subtypes (all P > 0.05). Tests indicated no substantial heterogeneity or directional pleiotropy for the leukemia (unspecified subtype) or HL analyses. These results provide genetic evidence consistent with an elevated risk of select HM subtypes among individuals with higher genetic liability to MS; however, the signals should be interpreted cautiously and validated in larger, multi-ancestry datasets and with additional causal frameworks.

Every year, many people around the world are progressively affected by the devastating conditions of health problems such as heart disease, respiratory infections, neurological dysfunction, cognitive stress, cancer, stroke, diabetes, etc., which lead to severe health complications and associated abnormalities. Thus, early health analytics are crucial, as they enable timely intervention with targeted therapies, potentially providing immediate relief and sustained long-term benefits that may slow disease progression. Due to the complex pathophysiological processes and heterogeneous clinical trials in various health conditions, there is a need for highly sensitive, multimodal biomarkers and effective investigative approaches to accurately detect and monitor patient health outcomes. Therefore, machine learning algorithms with various categories and techniques are considered for predicting outcomes, including prognosis, risk assessment, patient stratification, and disease monitoring. The flow of the proposed work is divided into three stages, as the first stage defines the importance of healthcare with case studies, followed by the traditional Machine Learning (ML) algorithms, traditional Deep Learning (DL) approaches, and modern DL techniques (TabNet and AutoInt) in the second stage. Finally, the experiments are implemented to justify the results. This work highlights the grouping of modalities by integrating molecular protein, chemical, and genetic biomarkers with emerging ML features. The results indicate a significant improvement in predicting the accuracy using the proposed methodology.

Our prior work published a protocol for creating resin-embedded arthropods of medical importance for use in educational and outreach activities. This protocol has been adopted by diverse initiatives aiming to create a mounting technique that minimizes damage to the arthropod, creates a safe product for handling, and preserves the ability to see morphological characteristics for identification. Our protocol has evolved and is currently creating resin-embedded arthropods with improved quality thanks to the purchase of additional equipment and process modifications. We present this improved protocol with steps to minimize bubbles, adapt to different types of arthropods, and create final polished blocks with visibility from six sides. We also outline biosafety concerns from fumes during the resin curing, use of power tools, and from dust generated while sanding and polishing resin blocks. This visualized protocol will facilitate the adoption of clear resin casting of arthropods to a broader community. The list of equipment and consumables we have adopted over several years of trial and error will allow other programs to judge if this protocol is a viable option for adoption.

Aberrant long non-coding RNAs (lncRNAs) expression profoundly influences cellular proliferation of osteosarcoma cells, suggesting that they may serve as potential therapeutic targets. However, functional studies of lncRNAs remain largely theoretical and limited to a few validations. Colony formation assays, which specifically reflect the ultimate proliferative fate of single cells, are regarded as the gold standard for evaluating long-term proliferative potential. Here, this protocol established a reliable experimental protocol using colony formation assays in lncRNA-knockdown cells to evaluate the effect of lncRNA on osteosarcoma cell proliferative capacity. This experimental protocol provides a stable, cost-effective, and efficient assay to assess the regulatory effects of lncRNA on osteosarcoma cell proliferation. This approach is not only suitable for research on osteosarcoma cells but also serves as a feasible tool for lncRNA-regulated cell proliferative capacity in other tumors. This study included a detailed detection protocol of lncRNA-knockdown and colony formation assays, with an example of the small nucleolar RNA host gene 6 (SNHG6) knockdown in osteosarcoma cells (143B). Experimental results confirmed efficient suppression of SNHG6 expression, accompanied by a marked reduction in both the number and size of colonies. These findings suggest that SNHG6 is essential for sustaining the long-term proliferative potential of osteosarcoma cells (143B).

Pulmonary fibrosis is characterized by progressive deposition of fibrotic scar tissue within the lung parenchyma, leading to severely impaired gas exchange. It underlies a spectrum of chronic interstitial lung diseases, notably idiopathic pulmonary fibrosis, a condition associated with an exceedingly poor prognosis. Given the lack of effective therapies, robust mouse models are critical for elucidating underlying pathological mechanisms and evaluating novel antifibrotic interventions. Bleomycin-induced pulmonary fibrosis remains the most extensively utilized experimental model. Common routes of administration in mice include intravenous and intraperitoneal injections, invasive open-tracheal instillation, and noninvasive tracheal dripping. However, invasive surgical methods often cause secondary tissue injury, potentially compromising model reproducibility and stability. Conversely, noninvasive tracheal dripping usually results in uneven bleomycin distribution across lung lobes and poses a risk of asphyxiation, thus reducing reproducibility and increasing technical challenges. To address these limitations, a refined aerosol-based intratracheal delivery method is developed that is operationally simpler, minimally invasive, highly reproducible, and ethically superior by significantly reducing animal distress. Using a small-animal laryngoscope to visualize the rima glottidis directly, a specialized aerosolizing needle is inserted into the trachea, markedly narrower than the mouse tracheal diameter. Bleomycin solution is delivered under precisely controlled pressure, generating a fine aerosol. This ensures uniform and efficient distribution of the agent throughout the lung parenchyma. Moreover, one can selectively target the left or right lung by directing the needle into the appropriate bronchus. This optimized model's dose-response relationship is extensively characterized by systematically monitoring changes in lung function, histopathological manifestations, and lung hydroxyproline content. This refined experimental protocol is anticipated to facilitate laboratory standardization, ultimately accelerating the development and preclinical validation of novel antifibrotic therapeutics.

The role of left ventricular systolic function as a prognostic marker for sepsis patients remains an area of ongoing research and debate. The present investigation exhibited the comprehensive evaluation of the relationship between different levels of left ventricular ejection fraction (LVEF) and mortality outcomes in sepsis-diagnosed patients. A retrospective, single-center longitudinal cohort investigation was conducted involving the intensive care unit (ICU) admitted adults' patients at Beth Israel Deaconess Medical Center who underwent transthoracic echocardiography (TTE) during their hospitalization. Individuals diagnosed with sepsis and who received Doppler echocardiography were included in the analysis if transthoracic echocardiography was performed within seven days of ICU admission. All patients of age below 18 or above 90 years, a prior history of cardiac disease or cardiac surgery, and those for whom echocardiography was performed more than seven days after ICU admission were excluded. Patients were stratified into three distinct groups with relation to their LVEF levels: hyperdynamic (LVEF ≥70%), normal (LVEF 55%-70%), and depressed (LVEF ≤55%). The association between different categories of sepsis affected patients mortality outcome and left ventricular ejection fraction (LVEF) was assessed. Among the 3,363 patients analyzed, comprising 1,175 with decreased LVEF, 2,119 with normal LVEF, and 68 with hyperdynamic LVEF, multivariate Cox regression identified hyperdynamic function as the strongest predictor of 28-day mortality. Specifically, hyperdynamic LVEF was independently linked to a 3.643-fold higher hazard of death relative to the normal LVEF group. A significant link has been identified between hyperdynamic left ventricular function and elevated 28-day mortality rates in the ICU-admitted septic patients. This physiological condition underscores the need for enhanced clinical awareness due to its prognostic significance.

This protocol establishes a comprehensive experimental framework for investigating cortical circuit dynamics during EA (electroacupuncture) stimulation. The methodology integrates three key components: chronic cranial window implantation in Thy1-GCaMP6f transgenic mice, standardized EA stimulation at the ST36 acupoint using 2/100 Hz biphasic pulses, and high-resolution two-photon calcium imaging. This integrated approach enables real-time visualization of neural ensemble activity in the primary somatosensory (S1) cortex with millisecond temporal precision, capturing previously inaccessible details of network-level responses to neuromodulation. The technique successfully reveals distinct layer-specific activation patterns and longitudinal plasticity changes, providing critical insights into the cortical mechanisms underlying the effects of EA. By overcoming the fundamental spatiotemporal limitations inherent in conventional fMRI and electrophysiological techniques, this high-resolution platform offers unprecedented analytical capabilities for mapping dynamic neural circuits. The protocol's robust design and reproducible outcomes make it particularly valuable for optimizing targeted neuromodulation therapies and advancing our understanding of circuit-level plasticity in response to peripheral stimulation.

Although cardiac lymphatic vessels have garnered increasing attention in recent years, the relationship between the donor cardiac lymphatic system and acute rejection in heart transplantation remains to be elucidated. Cardiac electrophysiological activity was assessed via electrocardiography (ECG) in normal mice before and after cardiac lymphatic ablation. Major lymphatic vessels of donor hearts were visualized by Evans Blue injection and subsequently ablated via electrocautery. A heart transplantation model featuring donor cardiac lymphatic dysfunction was established using hearts subjected to lymphatic ablation. Cardiac graft function was assessed via echocardiography, western blotting, and ELISA. The impact of donor lymphatic ablation on acute rejection was assessed by hematoxylin-eosin (HE) staining and monitoring of graft survival time. ECG results indicated no significant changes in cardiac electrophysiology before and after lymphatic ablation, confirming that major cardiac vessels remained undamaged. Evans Blue was injected into the cardiac apex to label the lymphatic vessels. Major lymphatics were then ablated using electrocautery. Following the completion of the ablation procedure, the mice were heparinized, and their hearts were subsequently harvested. The aorta and pulmonary artery were transected, and the superior/inferior vena cava, along with the pulmonary veins, were ligated. The prepared (ablated) donor hearts were then transplanted. Levels of cTnI and ejection fraction demonstrated that lymphatic ablation did not exacerbate cardiac injury. HE staining revealed that ablation of donor cardiac lymphatics alleviated acute rejection, reduced myocardial injury, and prolonged graft survival. We successfully established a murine heart transplantation model with donor cardiac lymphatic dysfunction and demonstrated that ablation of donor cardiac lymphatics can mitigate acute rejection, attenuate myocardial damage, and extend graft survival. These findings provide new insights and a foundation for understanding the role of the cardiac lymphatic system in heart transplantation.