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Tumor acidosis is one of the hallmarks indicating the initiation and progression of various cancers. Here, we present a protocol for preparing a hyperpolarized (HP) 13C-bicarbonate tissue pH MRI imaging contrast agent to detect aggressive tumors. We describe the steps for the formulation and polarization of a precursor molecule 13C-glycerol carbonate (13C-GLC), the post-dissolution reaction, and converting HP 13C-GLC to an injectable HP 13C-bicarbonate solution. We then detail procedures for MRI data acquisition to generate tumor pH maps for assessing tumor aggressiveness. For complete details on the use and execution of this protocol, please refer to Mu et al.1.

BACKGROUND: Mechanical ventilation is a common life-saving procedure but can lead to serious complications, including ARDS and oxygen toxicity. Nonadherence to lung-protective ventilation guidelines is common. We hypothesized that a respiratory therapist-driven mechanical ventilation bundle could increase adherence to lung-protective ventilation and decrease the incidence of pulmonary complications in the ICU. METHODS: A respiratory therapist-driven protocol was implemented on August 1, 2018, in all adult ICUs of a Midwestern academic tertiary center. The protocol targeted low tidal volume, adequate PEEP, limiting oxygen, adequate breathing frequency, and head of the bed elevation. Adherence to lung-protective guidelines and clinical outcomes were retrospectively observed in adult subjects admitted to the ICU and on ventilation for ≥ 24 h between January 2011 and December 2019. RESULTS: We included 666 subjects; 68.5% were in the pre-intervention group and 31.5% were in the post-intervention group. After adjusting for body mass index and intubation indication, a significant increase in overall adherence to lung-protective ventilation guidelines was observed in the post-intervention period (adjusted odds ratio 2.48, 95% CI 1.73-3.56). Fewer subjects were diagnosed with ARDS in the post-intervention group (adjusted odds ratio 0.22, 95% CI 0.08-0.65) than in the pre-intervention group. There was no difference in the incidence of ventilator-associated pneumonia, ventilator-free days, ICU mortality, or death within 1 month of ICU discharge. CONCLUSIONS: A respiratory therapist-driven protocol increased adherence to lung-protective mechanical ventilation guidelines in the ICU and was associated with decreased ARDS incidence. [ABSTRACT FROM AUTHOR]

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Importance: Orthognathic surgery is utilized to rectify facial deformities, but it can lead to neurosensory alterations. Electroacupuncture has been shown to enhance sensitivity and motor functions in patients post-surgery. However, its application in traumatic facial injuries remains inadequately researched. Objective: To investigate the effects of electroacupuncture on sensitivity and orofacial function in patients undergoing orthognathic surgery.Design, Setting, and Population: A randomized clinical trial involving patients undergoing bimaxillary orthognathic surgery and genioplasty who are randomly allocated to either a physiotherapy (PT) or physiotherapy plus electroacupuncture group (PTEA). Exposures: Participants will be allocated to their respective therapies for a duration of six weeks. The PT group will receive a 50-minute physical therapy session along with 30 minutes of simulated procedure. The PTEA group will receive 50 minutes of physical therapy followed by 30 minutes of electroacupuncture. Main Outcomes and Measures: Before treatment, sensitivity will be assessed using the SMILE Sensitivity Test-BAURU kit, edema will be evaluated using the MD Anderson Cancer Center Head and Neck Lymphedema protocol, range of motion will be measured using a digital caliper, and muscle pain and fatigue will be gauged using numerical scales. The chewing function will be evaluated using the Chewing Quality Assessment Questionnaire. Results: All assessments will be repeated at three and six months following the initiation of treatment. Conclusions and Relevance: This study may provide reliable and high-quality clinical evidence regarding the impact of electroacupuncture on restoring altered sensation and motor function in patients undergoing orthognathic surgery.

OBJECTIVE A risk stratified sedation weaning protocol improved patient outcomes in a pediatric intensive care unit (PICU). We sought to determine the protocol effect on medication costs. METHODS This was a retrospective observational cohort study in an academic tertiary care children’s hospital PICU (2018–2020) comparing the cost when weaning benzodiazepine, alpha agonist, and/or opioid infusions in intubated children <18 years of age. RESULTS There were 84 total sedation weaning instances (pre-protocol n = 41 and post-protocol n = 41); 2 patients had 2 encounters, 1 in each phase. The total cost (in 2022 United States Dollars) of sedation weaning was $400,328.87 ($15,994.44/kg) pre-protocol compared with $170,458.85 ($11,227.52/kg) post-protocol. The median cost of sedation wean per patient for pre-protocol patients was $3197.42 (IQR: $322.66–$12,643.29) and post-protocol patients was $1851.44 (IQR: $425.05–$5355.85; p = 0.275). A linear regression model estimated the expected cost of sedation wean for post-protocol patients to be $5173.20 lower than for pre-protocol patients of the same weight and overall drug risk (p = 0.036). The proportion of withdrawal symptoms in the pre-protocol patients (16%) was not significantly different from the proportion in the post-protocol patients (14%; p = 0.435). CONCLUSIONS Implementation of a PICU sedation weaning protocol in a single-center conferred cost benefit without negatively impacting patient outcomes. A larger multicenter study would provide insight to the applicability to PICUs in varied settings with differing patient populations. [ABSTRACT FROM AUTHOR]

Aim: To determine the tensile load capacity (TLC) and the tearing characteristics for interrupted and vertical mattress sutures with different insertion points from the wound margin, and the effect of the bite size when using vertical mattress sutures. Materials and Methods: A total of 120 gingiva and lining mucosa samples obtained from pig jaws were divided into groups according to the suturing technique (interrupted and vertical mattress sutures), distance of the insertion points from the wound margin (margin, 1, 3, and 5 mm) and bite size (1, 3, and 5 mm). The TLC of the suture and the tearing characteristics were evaluated using a tensile tester device. Results: The TLC was significantly higher for vertical mattress sutures than for interrupted sutures regardless of the distance of the insertion points from the wound margin (intergroup p <.001). This distance significantly influenced the TLC for vertical mattress sutures (p <.05) but not for interrupted sutures (p >.05). Testing the tearing characteristics revealed that no tissue tearing occurred in groups when the insertion points were more than 3 mm from the wound margin. Conclusion: The TLC is higher for vertical mattress sutures than for interrupted sutures, and it increases when the insertion points are farther from the wound margin. [ABSTRACT FROM AUTHOR]

This review addresses the current understanding of In Vitro Maturation (IVM) treatment, including indications and effective treatment protocols influencing oocyte developmental competence. A comprehensive literature search was performed to gather relevant studies, clinical trials, and reviews related to IVM. Databases such as PubMed, MEDLINE, and pertinent medical journals were searched. The selected literature was analyzed and synthesized to offer a comprehensive overview. IVM has emerged as a promising technique for inducing maturation in immature oocytes across various developmental stages. Its applications extend to areas utilizing In Vitro Fertilization (IVF), gaining traction as a treatment option for Polycystic Ovary Syndrome (PCOS) and fertility preservation in cancer patients. Recent advancements have led to improved global pregnancy rates, resulting in successful births. IVM also holds potential in reducing risks associated with conventional IVF, including ovarian hyperstimulation syndrome and multiple pregnancies. Despite these advantages, IVM adoption in clinical practice remains limited. Ongoing research aims to refine therapeutic protocols and expand clinical indications. IVM holds promise in assisted reproductive technology, spanning applications from cancer patient fertility preservation to addressing PCOS. Enhanced pregnancy rates highlight efficacy, while risk reduction compared to IVF underscores its importance. Further research is needed for optimal use across patient groups, emphasizing protocol refinement and expanded applications. [ABSTRACT FROM AUTHOR]

Currently, there is no established system for quantifying patterns of ocular ductions. This poses challenges in tracking the onset and evolution of ocular motility disorders, as current clinical methodologies rely on subjective observations of individual movements. We propose a protocol that integrates image processing, a statistical framework of summary indices, and criteria for evaluating both cross-sectional and longitudinal differences in ductions to address this methodological gap. We demonstrate that our protocol reliably transforms objective estimates of ocular rotations into normative patterns of total movement area and movement symmetry. This is a critical step towards clinical application in which our protocol could first diagnose and then track the progression and resolution of ocular motility disorders over time. [ABSTRACT FROM AUTHOR]

Noninvasive imaging of endogenous retinal fluorophores, including vitamin A derivatives, is vital to developing new treatments for retinal diseases. Here, we present a protocol for obtaining in vivo two-photon excited fluorescence images of the fundus in the human eye. We describe steps for laser characterization, system alignment, positioning human subjects, and data registration. We detail data processing and demonstrate analysis with example datasets. This technique allays safety concerns by allowing for the acquisition of informative images at low laser exposure. For complete details on the use and execution of this protocol, please refer to Bogusławski et al. (2022).1.

ABSTRACT Objective: To describe the adherence to the sepsis protocol by obstetric nurses in the obstetric triage of a high-risk maternity reference center. Methods: This was a quantitative, documental, and retrospective study involving 105 pregnant women treated in obstetric triage under sepsis criteria. Data were collected through electronic medical records using structured forms and were organized into tables employing descriptive statistics. This research adhered to ethical principles concerning human studies. Results: Of the checklists for initiating the SEPSIS protocol by obstetric nurses, 105 were identified. Regarding the protocol steps performed, lactate was collected in 97.1% of cases and blood cultures in 98.1%, antibiotic therapy was administered in 94.3%, and hydration was carried out in 51.4% of the cases. Conclusion: The initiation of the sepsis protocol for all women meeting the criteria was confirmed. However, the steps were not fully implemented as recommended by the institutional protocol, and the recommended broad-spectrum antibiotic was not administered.

Summary: Tumor acidosis is one of the hallmarks indicating the initiation and progression of various cancers. Here, we present a protocol for preparing a hyperpolarized (HP) 13C-bicarbonate tissue pH MRI imaging contrast agent to detect aggressive tumors. We describe the steps for the formulation and polarization of a precursor molecule 13C-glycerol carbonate (13C-GLC), the post-dissolution reaction, and converting HP 13C-GLC to an injectable HP 13C-bicarbonate solution. We then detail procedures for MRI data acquisition to generate tumor pH maps for assessing tumor aggressiveness.For complete details on the use and execution of this protocol, please refer to Mu et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Summary: Drug resistance is currently the biggest challenge in cancer chemotherapy. Here, we present a protocol to develop a chemotherapy drug screening process by constructing a cancer prognostic model (PM) using public databases. We describe steps for downloading code and data, preparing the expression matrix and metadata for analysis, screening modeling genes, and constructing a PM. We then detail procedures for constructing predictive websites for cancer patients’ survival based on their age, tumor stage, gene expression levels, and risk scores.For complete details on the use and execution of this protocol, please refer to Bai et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Summary: The emergence of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Alpha variant in 2020 demonstrated the need for reanalysis of diagnostic tests to ensure detection of emerging variants. Here, we present a protocol for creating and characterizing SARS-CoV-2 variant testing panels using remnant clinical samples for diagnostic assay testing. We describe steps for characterizing SARS-CoV-2 remnant clinical samples and preparing them into pools and their use in preparing varying quantities of virus. We then detail procedures for verifying variant detection using the resulting sample panel.For complete details on the use and execution of this protocol, please refer to Rao et al.1,2 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Yujin Choi,1,&ast; Na-Yeon Ha,2,&ast; Ae-Ran Kim,3 Haein Jeong,2 Ojin Kwon,1 Ki-Sun Park,1 Jieun Kim,1 Jinsung Kim,2 Hyungjun Kim1 1KM Science Research Division, Korea Institute of Oriental Medicine, Daejeon, Republic of Korea; 2Division of Digestive Diseases, Department of Korean Internal Medicine, Kyung Hee University College of Korean Medicine, Kyung Hee University Korean Medicine Hospital, Seoul, Republic of Korea; 3R&D Strategy Division, Korea Institute of Oriental Medicine, Daejeon, Republic of Korea&ast;These authors contributed equally to this workCorrespondence: Jinsung Kim, Department of Digestive Diseases, College of Korean Medicine, Kyung Hee University, 26 Kyungheedae-ro, Dongdaemun-gu, Seoul, 02447, Republic of Korea, Tel +82 2 958 8895, Email oridoc@khu.ac.kr Hyungjun Kim, Division of KM Science Research, Korea Institute of Oriental Medicine, Daejeon, 34054, Republic of Korea, Tel +82 42 869 2796, Email heyjoon73@kiom.re.krIntroduction: Irritable bowel syndrome (IBS) is a chronic condition characterized by recurrent abdominal pain associated with bowel movements. Modified Gwakjeongtang (MGT), an herbal prescription rooted in traditional East Asian medicine, consists of thirteen botanical drugs known for their potential to enhance intestinal barrier function, regulate gastrointestinal motility, and exhibit anti-inflammatory and antioxidant properties. Despite a few previous clinical trials highlighting MGT’s potential for IBS symptom management, limited evidence exists with placebo control.Methods and Analysis: In this pilot randomized clinical trial protocol, we aim to exploratively evaluate the efficacy and safety of MGT in patients with diarrhea-predominant IBS (IBS-D) by comparing it with a placebo. A total of 60 IBS-D patients will be enrolled, and eligible participants will be randomly allocated to either the MGT or placebo groups. Over a 4-week period, they will receive MGT or placebo granules three times a day. The primary endpoint will be the overall response rate post-treatment, determined through daily assessments of abdominal pain intensity and stool consistency.Ethics and Dissemination: This clinical trial protocol has received approval from the Korean Ministry of Food and Drug Safety for an investigational new drug application and Institutional Review Board of the Kyung Hee University Korean Medicine Hospital. The research findings will be submitted and published in international peer-reviewed journal.Trial Registration: Clinical research information service (registration number: KCT0008523).Keywords: irritable bowel syndrome, herbal medicine, Gwakhyang-Jeonggi-San, Huoxiang-Zhengqi-San, Kkako-Shoki-San, clinical protocol

Background: Various protocols have been approved to improve the response rate leading to successful fertilizationin poor ovarian responders (PORs). The application of double ovarian stimulation (DuoStim) in the follicular andluteal phases of the same ovarian cycle has been shown as an intriguing option to achieve more oocyte retrievals inthe shortest time. The aim of the current study, is to compare the outcomes of different protocols, minimal stimulation(MS) and Duostim.Materials and Methods: This randomized clinical trial was performed on 42 in vitro fertilization (IVF) candidateswith POR diagnosis. Patients were classified into two equal groups and treated with the DuoStim protocol and MSprotocol. The IVF outcomes, including retrieved follicles, oocytes, metaphase II (MII) oocytes and embryos, werecompared between these groups.Results: The patients’ characteristics including age, anti-mullerian hormone (AMH), follicle-stimulating hormone(FSH), luteinizing hormone (LH), and antral follicle count (AFC) were collected and compared. It showed there wasno significant difference between the two groups' baseline characteristics (P>0.05). We observed that the DuoStimprotocol resulted in a significantly higher score in comparison with the MS protocols, including the number of follicles(6.23 ± 2.93 vs. 1.77 ± 1.66, P

Summary: We present a protocol to generate a liver tumor murine model utilizing orthotopic ultrasound-guided intrahepatic injections of tumor cells and a partial hepatectomy. We describe cell and animal preparation and ultrasound-guided intrahepatic injections. We then depict how to conduct a partial hepatectomy addressing solutions to potential problems. This approach introduces a rapid and reproducible orthotopic implantation model of liver tumor model in a minimally invasive manner, allowing the surgical procedure to be optimized with minimal intra-abdominal injury.For complete details on the use and execution of this protocol, please refer to Brown et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Summary: Cancer cells with homologous recombination deficiency (HRD) exhibit a distinctive vulnerability to poly(ADP-ribose) polymerase inhibitors (PARPis). To address the limitations of existing methodologies incapable of providing real-time insights into homologous recombination (HR) status, we present an adenovirus-based functional assay designed to quantify cellular HR activity. Here, we delineate the step-by-step procedure for producing the adenovirus harboring an HR reporter, processing primary cells, and assessing HR activity in primary ovarian cancer cells.For complete details on the use and execution of this protocol, please refer to Lee et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

ObjectivesSpine surgery is associated with early impairment of gastrointestinal motility, with postoperative ileus rates of 5-12%. A standardized postoperative medication regimen aimed at early restoration of bowel function can reduce morbidity and cost, and its study should be prioritized.MethodsA standardized postoperative bowel medication protocol was implemented for all elective spine surgeries performed by a single neurosurgeon from March 1, 2022 to June 30, 2022 at a metropolitan Veterans Affairs medical center. Daily bowel function was tracked and medications were advanced using the protocol. Clinical, surgical, and length of stay data are reported.ResultsAcross 20 consecutive surgeries in 19 patients, mean age was 68.9 years [standard deviation (SD) = 10; range 40-84]. Seventy-four percent reported preoperative constipation. Surgeries consisted of 45% fusion and 55% decompression; lumbar retroperitoneal approaches constituted 30% (10% anterior, 20% lateral). Two patients were discharged in good condition prior to bowel movement after meeting institutional discharge criteria; the other 18 cases all had return of bowel function by postoperative day (POD) 3 (mean = 1.8-days, SD = 0.7). There were no inpatient or 30-day complications. Mean discharge occurred 3.3-days post-surgery (SD = 1.5; range 1-6; home 95%, skilled nursing facility 5%). Estimated cumulative cost of the bowel regimen was $17 on POD 3.ConclusionsCareful monitoring of return of bowel function after elective spine surgery is important for preventing ileus, reducing healthcare cost, and ensuring quality. Our standardized postoperative bowel regimen was associated with return of bowel function within 3 days and low costs. These findings can be utilized in quality-of-care pathways.

Background: Various protocols have been approved to improve the response rate leading to successful fertilization in poor ovarian responders (PORs). The application of double ovarian stimulation (DuoStim) in the follicular and luteal phases of the same ovarian cycle has been shown as an intriguing option to achieve more oocyte retrievals in the shortest time. The aim of the current study is to compare the outcomes of different protocols, minimal stimulation (MS) and Duostim. Materials and Methods: This randomized clinical trial was performed on 42 in vitro fertilization (IVF) candidates with POR diagnosis. Patients were classified into two equal groups and treated with the DuoStim protocol and MS protocol. The IVF outcomes, including retrieved follicles, oocytes, metaphase II (MII) oocytes and embryos, were compared between these groups. Results: The patients' characteristics including age, anti-mullerian hormone (AMH), follicle stimulating hormone (FSH), luteinizing hormone (LH), and antral follicle count (AFC) were collected and compared. It showed there was no significant difference between the two groups baseline characteristics (P>0.05). We observed that the DuoStim protocol resulted in a significantly higher score in comparison with the MS protocols, including the number of follicles (6.23 ± 2.93 vs. 1.77 ± 1.66, P<0.001), retrieved oocytes (3.86 ± 2.57 vs. 1.68 ± 1.58, P=0.002), MII oocytes (3.36 ± 2.42 vs. 1.27 ± 1.27, P=0.001) and obtained embryos (2.04 ± 1.64 vs. 0.77 ± 0.86, P=0.003). Conclusion: The DuoStim protocol is a favourable and time saving plan that is associated with more oocytes in a single stimulation cycle. The DuoStim protocol significantly can result in more frequent MII oocytes and embryos. We figured that the higher number of oocytes and embryos might have led to a higher rate of pregnancy (registration number: IRCT20200804048303N1). [ABSTRACT FROM AUTHOR]

Introduction: Avoidant/restrictive food intake disorder (ARFID) is an eating disorder characterised by a pattern of eating that leads to failure to meet appropriate nutritional and/or energy needs. Method: In the absence of evidence-based inpatient guidelines for adolescents with ARFID, we set out to develop and pilot an inpatient protocol for adolescents with ARFID. Identification of the key differences between managing inpatients with ARFID and anorexia nervosa (AN) led to modification of an existing AN protocol with the goals of better meeting patient needs, enhancing alignment with outpatient care, and improving outcomes. A case report of an adolescent with ARFID who had three hospital admissions is presented to highlight these changes. Interviews with this patient and her family were undertaken, together with key staff, to explore the challenges of the AN protocol for this patient and the perceived benefits and any limitations of the ARFID protocol for this patient and others. Results: The new ARFID protocol supports greater choice of meals, without the need for rest periods after meals and bathroom supervision. The similarities with the AN protocol reflect the need to promote timely weight gain through meal support, including a staged approach to nutritional supplementation. The protocol appears to have been well accepted by the patient and her family, as well as by staff, and continues to be used in cases of ARFID. Conclusion: Further evaluation would help identify how well this protocol meets the needs of different adolescents with ARFID. [ABSTRACT FROM AUTHOR]

Currently, there is no established system for quantifying patterns of ocular ductions. This poses challenges in tracking the onset and evolution of ocular motility disorders, as current clinical methodologies rely on subjective observations of individual movements. We propose a protocol that integrates image processing, a statistical framework of summary indices, and criteria for evaluating both cross-sectional and longitudinal differences in ductions to address this methodological gap. We demonstrate that our protocol reliably transforms objective estimates of ocular rotations into normative patterns of total movement area and movement symmetry. This is a critical step towards clinical application in which our protocol could first diagnose and then track the progression and resolution of ocular motility disorders over time.

Summary: Due to anatomical and biological similarities with humans, pigs are increasingly used for inflammation- and immune-related studies in biomedical research, including the field of osteonecrosis and osteoimmunology. Here, we present a protocol for rib extraction, isolation of the bone marrow by centrifugation, and processing to obtain bone-marrow-derived macrophages (BMDMs). Then, we describe the procedures of in vitro experiments to evaluate the cell phenotype.For complete details on the use and execution of this protocol, please refer to Andre et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Summary: The limitations associated with distinguishing serum Fe2+ and Fe3+ hinder the widespread application of ferroptosis, beyond laboratory settings. Here, we present a protocol for deep mining the correlation between acute pancreatitis and ferroptosis using the MIMIC-III database and STATA software. We describe steps for using Cox regression, decision curve analysis (DCA), and receiver operating characteristic (ROC) approaches to establish the relationship between them and determine the relevant factors. This protocol has potential application in establishing novel research models that integrate both fundamental and clinical methodologies.For complete details on the use and execution of this protocol, please refer to Yueling Deng et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Summary: Phage therapy has re-emerged as a promising treatment for non-resolving infections. Given the lack of approved phage treatments, there is a need to establish a compassionate use pipeline. Here, we present a protocol for phage matching, treatment, and monitoring for compassionate bacteriophage use in non-resolving infections. We describe steps for consultation and request implementation, evaluating and comparing different aspects of phage activity, and phage production. We then detail procedures for multidisciplinary meetings, ethics approvals, phage therapy, and follow-up.For complete details on the use and execution of this protocol, please refer to Onallah et al.1,2 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Summary: Here, we present a workflow for analyzing multi-omics data of plasma samples in patients with post-COVID condition (PCC). Applicable to various diseases, we outline steps for data preprocessing and integrating diverse assay datasets. Then, we detail statistical analysis to unveil plasma profile changes and identify biomarker-clinical variable associations. The last two steps discuss machine learning techniques for unsupervised clustering of patients based on their inherent molecular similarities and feature selection to identify predictive biomarkers.For complete details on the use and execution of this protocol, please refer to Wang et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Summary: Investigating the spatially distributed information contained in fMRI data is essential for understanding brain functions. Here, we present a protocol to dynamically predict short-term changes in neural patterns using trial-by-trial blood-oxygen-level-dependent (BOLD) activity of a seed region. We describe steps for setting fMRI data acquisition parameters and quantification of changes in multivariate patterns. We then detail procedures for defining seed regions and identifying brain areas in which changes in multivariate patterns can be predicted by BOLD activity of the seed region.For complete details on the use and execution of this protocol, please refer to Möhring et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Abstract Background Life expectancy of patients with metastatic castration-resistant prostate cancer (mCRPC) is still limited despite several systemic treatments. Within five years after diagnosis of primary prostate cancer, 10–20% of the patients have mCRPC and curation is not an option. Radionuclide therapy (RNT) targeted against prostate-specific membrane antigen (PSMA) emerged as a new treatment option and showed effective results in patients with mCRPC. Survival benefit after [177Lu]Lu-PSMA RNT has already been demonstrated in several clinical trials. However, [225Ac]Ac-PSMA (225Ac-PSMA) appears to be an even more promising radiopharmaceutical for the treatment of mCRPC. The use of alpha emitting radionuclides offers advantages over beta emitting radionuclides due to the high linear energy transfer effective for killing tumor cells and the limited range to reduce the radiation effects on the healthy tissue. However, these results are based on retrospective data and safety data of 225Ac-PSMA are still limited. Therefore, a prospective trial is needed to determine the optimal amount of activity that can be administered. Methods The 225Ac-PSMA-Imaging & Therapy (I&T) trial is an investigator-initiated phase I, single-center, open label, repeated dose-escalation and expansion trial. Patient with PSMA-positive mCRPC after at least one line of chemotherapy and/or one line of nonsteroidal antiandrogen will be treated with 225Ac-PSMA-I&T in increasing amount of activity per cycle. Dose-escalation following an accelerated 3 + 3 design which allows to open the next dose-level cohort in the absence of dose limiting toxicity while the previous one is still ongoing. Up to 4 treatment cohorts will be explored including up to 3 dose-escalation cohorts and one expansion cohort where patients will be administered with the recommended dose. A total of up to 30 patients will be enrolled in this trial. All patients will be evaluated for safety. Additionally, dosimetry was performed for the patients in the dose-escalation cohorts after the first 225Ac-PSMA-I&T administration. Discussion This trial will assess the safety and tolerability of 225Ac-PSMA-I&T in patients with mCRPC to recommend the optimal dose for the phase II trial. Trial registration ClinicalTrials.gov, (NCT05902247). Retrospectively registered 13 June 2023.

Background: Life expectancy of patients with metastatic castration-resistant prostate cancer (mCRPC) is still limited despite several systemic treatments. Within five years after diagnosis of primary prostate cancer, 10–20% of the patients have mCRPC and curation is not an option. Radionuclide therapy (RNT) targeted against prostate-specific membrane antigen (PSMA) emerged as a new treatment option and showed effective results in patients with mCRPC. Survival benefit after [177Lu]Lu-PSMA RNT has already been demonstrated in several clinical trials. However, [225Ac]Ac-PSMA (225Ac-PSMA) appears to be an even more promising radiopharmaceutical for the treatment of mCRPC. The use of alpha emitting radionuclides offers advantages over beta emitting radionuclides due to the high linear energy transfer effective for killing tumor cells and the limited range to reduce the radiation effects on the healthy tissue. However, these results are based on retrospective data and safety data of 225Ac-PSMA are still limited. Therefore, a prospective trial is needed to determine the optimal amount of activity that can be administered. Methods: The 225Ac-PSMA-Imaging & Therapy (I&T) trial is an investigator-initiated phase I, single-center, open label, repeated dose-escalation and expansion trial. Patient with PSMA-positive mCRPC after at least one line of chemotherapy and/or one line of nonsteroidal antiandrogen will be treated with 225Ac-PSMA-I&T in increasing amount of activity per cycle. Dose-escalation following an accelerated 3 + 3 design which allows to open the next dose-level cohort in the absence of dose limiting toxicity while the previous one is still ongoing. Up to 4 treatment cohorts will be explored including up to 3 dose-escalation cohorts and one expansion cohort where patients will be administered with the recommended dose. A total of up to 30 patients will be enrolled in this trial. All patients will be evaluated for safety. Additionally, dosimetry was performed for the patients in the dose-escalation cohorts after the first 225Ac-PSMA-I&T administration. Discussion: This trial will assess the safety and tolerability of 225Ac-PSMA-I&T in patients with mCRPC to recommend the optimal dose for the phase II trial. Trial registration: ClinicalTrials.gov, (NCT05902247). Retrospectively registered 13 June 2023. [ABSTRACT FROM AUTHOR]

Summary: Apoptosis-associated speck-like protein containing a c-terminal caspase activation and recruitment domain (ASC) specks are elevated in the cerebrospinal fluid (CSF) of Alzheimer’s disease and related dementias (AD/ADRDs) patients. Here, we present a flow cytometry protocol to quantify ASC specks. We describe steps for fluorescently labeling ASC specks using antibody technology, visualizing with imaging flow cytometry, and gating based on physical characteristics. CSF ASC specks levels positively correlate with phosphorylated tau (Thr181) and negatively correlate with amyloid β ratio (42/40), thus serving as a neuroinflammatory biomarker for diagnosing AD/ADRDs.For complete details on the use and execution of this protocol, please refer to Jiang et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Summary: Despite optimal multimodal treatment including surgical resection, 50%–80% of high-grade soft tissue sarcoma (STS) patients metastasize. Here, we present a protocol for the generation and use of post-surgical minimal residual disease models to investigate metastatic relapse in STS patient-derived xenografts. We describe steps for orthotopic engraftment of high-grade STS patient-derived tumor tissue. We then detail procedures for primary tumor resection with broad, negative resection margins and follow-up until metastases using MRI.For complete details on the use and execution of this protocol, please refer to Fischer et al. (2023).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Summary: Here, we present a protocol for programing a degree-to-careers dashboard in R using Posit and Shiny. We describe steps for installing software, obtaining datasets, and munging and joining data. We then detail procedures for programing and publishing the dashboard in Shiny web application that includes a filtered data table. The resulting dashboard links academic programs with careers and income data and may be useful to inform decision-making by higher education leaders and policymakers.For complete details on the use and execution of this protocol, please refer to Perkins and Carrier (2023).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Summary: Cancer registry data on pediatric gliomas come with inherent limitations as inclusion criteria and registration practices of these tumors differ between registries due to specific guidelines that are lacking. These limitations can lead to biased estimates in incidence and survival outcomes. Here, we present a protocol to investigate data quality and comparability for retrospective population-based pediatric glioma studies. We describe steps for obtaining institutional permissions, dealing with data quality issues, regrouping tumors, and reporting tumors in a clinically relevant manner.For complete details on the use and execution of this protocol, please refer to Hoogendijk et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Summary: Immunophenotyping of out-of-hospital cardiac arrest (OHCA) patients is of increasing interest but has challenges. Here, we describe steps for the design of the clinical cohort, planning patient enrollment and sample collection, and ethical review of the study protocol. We detail procedures for blood sample collection and cryopreservation of peripheral blood mononuclear cells (PBMCs). We detail steps to modulate immune checkpoints in OHCA PBMC ex vivo. This protocol also has relevance for immunophenotyping other types of critical illness.For complete details on the use and execution of this protocol, please refer to Tamura et al. (2023).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Summary: We present a protocol for the rapid postmortem bedside procurement of selected tissue samples using an endoscopic endonasal surgical technique that we adapted from skull base surgery. We describe steps for the postmortem collection of blood, cerebrospinal fluid, a nasopharyngeal swab, and tissue samples; the clean-up procedure; and the initial processing and storage of the samples. This protocol was validated with tissue samples procured postmortem from COVID-19 patients and can be applied in another emerging infectious disease.For complete details on the use and execution of this protocol, please refer to Khan et al. (2021)1 and Khan et al. (2022).2 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Summary: Total antioxidant capacity (TAC), representative of the capacity to combat oxidative stress, is closely linked to numerous diseases. Here, we present a protocol for measuring TAC using minimal samples that are stable across varying pH levels and at room temperature. We describe steps for preparing and loading samples and working solutions and conducting and analyzing the colorimetric reaction. Sample sources include aqueous humor, vitreous, tears, and plasma, which allow the protocol to be used in various clinical diagnostic settings.For complete details on the use and execution of this protocol, please refer to publications by Tsao et al. (2022).1,2 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Summary: Immunodynamics in the tumor microenvironment can be precisely examined by using multiple antigen identification approaches. Here, we present a protocol for capturing expression levels of multiple target proteins in the same specimen at single-cell resolution using a tyramide signal amplification-based immunofluorescent multiplexing system. We describe steps for tumor tissue microarray preparation, multiplex immunohistochemistry staining, image acquisition, and quantification. This protocol can quantify immune cells in tissues from patients or experimental disease models at a protein level.For complete details on the use and execution of this protocol, please refer to Chung et al. (2023),1 Tang et al. (2022),2 and Tang et al. (2022).3 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Summary: Federated learning is a cooperative learning approach that has emerged as an effective way to address privacy concerns. Here, we present a protocol for training MERGE: a federated multi-input neural network (NN) for COVID-19 prognosis. We describe steps for collecting and preprocessing datasets. We then detail the process of training a multi-input NN. This protocol can be adapted for use with datasets containing both image- and table-based input sources.For complete details on the use and execution of this protocol, please refer to Casella et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Abstract Background Postpartum hemorrhage is the leading cause of maternal death and severe maternal morbidity worldwide. Previous studies have reported the importance of multidisciplinary treatment approaches for postpartum hemorrhage; however, only a few studies have shown a clear improvement in maternal outcomes. Therefore, this study aimed to investigate the efficacy of a call system for postpartum hemorrhage in a tertiary emergency facility for rapid multidisciplinary treatment and its effect on maternal outcomes. Methods This single-center retrospective cohort study included patients transferred to our hospital due to postpartum hemorrhage between April 1, 2013, and March 31, 2019. The primary outcome was mortality, and the secondary outcomes were morbidity (duration of hospital stay, duration of intensive care unit stay, admission to the intensive care unit, respirator use, duration of ventilator support, acute kidney injury, transfusion-associated circulatory overload/transfusion-related acute lung injury, hysterectomy, composite adverse events, blood transfusion initiation time, blood transfusion volume, and treatment for postpartum hemorrhage). An in-hospital call system implementation commenced on April 1, 2016. The study outcomes were compared 3 years before and after implementing the call system. Results The blood transfusion initiation time and duration of hospital stay were significantly shortened after implementing the call system for postpartum hemorrhage. No maternal deaths were observed after implementing the system. Conclusions Implementing call systems specialized for postpartum hemorrhage in tertiary emergency facilities may improve maternal outcomes.

ObjectivesTo determine the impact of a criteria-led discharge initiative (CLD) on the hospital length of stay of patients undergoing a robotic-assisted laparoscopic prostatectomy (RALP). MethodsThis is a cohort study of prospectively collected data completed at a major tertiary hospital from December 2017 to August 2020. The CLD initiative consists of 4 criteria: clinical haemodynamic stability (heart rate < 100 beats/minute, systolic blood pressure > 100mmHg), a drain output of less than 50 mL, flatulence or bowel movement, and the ability to tolerate an oral diet. The primary outcome was hospital length of stay for patients before and after the introduction of CLD. ResultsOne hundred men undergoing RALP before the implementation of the CLD initiative were compared to 118 men undergoing RALP following the implementation of CLD. The patients had similar baseline demographic features. There was a significant difference found in hospital LOS with the pre-CLD group LOS (mean = 1.8 days, SE = 0.12) being longer than the LOS in the post-CLD group (mean = 1.4 days, SE = 0.09, P = 0.015). There were no significant between-group differences in the proportion of patients discharged on the first postoperative day and the 30-day readmission rate. ConclusionWithin our study population, we have demonstrated that the introduction of CLD was associated with reduced hospital LOS with no increase in adverse events. These findings support the need for the development of CLD in other conditions.

Summary: Infection with Helicobacter suis, which causes many cases of gastric disease, is not reliably diagnosed. Here, we present a protocol for detecting H. suis infection. We describe steps for collecting gastric biopsies and sera from patients, preparing DNA for PCR, and targeting the H. suis-specific gene. We then define procedures for inoculating biopsies onto primary agar plates and transferring colonies to secondary agar plates. Finally, we detail whole-genome sequencing of bacteria and assess H. suis infection in sera with ELISA.For complete details on the use and execution of these protocols, please refer to Matsui et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Summary: Immune dysregulation and inflammation by hepatic-resident leukocytes is considered a key step in disease progression of non-alcoholic fatty liver disease and non-alcoholic steatohepatitis toward cirrhosis and hepatocellular carcinoma. Here, we provide a protocol for isolation and characterization of liver-resident immune cells from fine-needle biopsies obtained from a rodent model and humans. We describe steps for isolating leukocytes, cell sorting, and RNA extraction and sequencing. We then detail procedures for low-input mRNA sequencing analyses. : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Summary: Plasma extracellular vesicles (EVs) represent a potential resource for biomarkers of multiple diseases. Here, we present a protocol for obtaining EVs from human plasma using asymmetrical flow field-flow fractionation technology. We describe steps for using tandem mass tags to perform comparative proteomic studies of a large clinical cohort. We then detail targeted quantitative analysis of differential proteins based on a parallel reaction monitoring technique.For complete details on the use and execution of this protocol, please refer to Wu et al. (2020)1 and Li et al. (2023).2 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Summary: Data quality issues have been acknowledged as one of the greatest obstacles in medical artificial intelligence research. Here, we present DeepFundus, which employs deep learning techniques to perform multidimensional classification of fundus image quality and provide real-time guidance for on-site image acquisition. We describe steps for data preparation, model training, model inference, model evaluation, and the visualization of results using heatmaps. This protocol can be implemented in Python using either the suggested dataset or a customized dataset.For complete details on the use and execution of this protocol, please refer to Liu et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Summary: The lymph node (LN) is a critical biological site for immune maturation after vaccination as it includes several cell populations critical for priming the antibody response. Here, we present a protocol for sampling the LN and isolating cell populations to evaluate immunogens targeting germline cells. We describe steps for media and tube preparation and sample collection using an ultrasound-guided LN fine-needle aspiration procedure. This protocol is safe, quick, low-cost, and less invasive than excisional biopsy.For complete details on the use and execution of this protocol, please refer to Leggat et al. (2022).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Background: Postpartum hemorrhage is the leading cause of maternal death and severe maternal morbidity worldwide. Previous studies have reported the importance of multidisciplinary treatment approaches for postpartum hemorrhage; however, only a few studies have shown a clear improvement in maternal outcomes. Therefore, this study aimed to investigate the efficacy of a call system for postpartum hemorrhage in a tertiary emergency facility for rapid multidisciplinary treatment and its effect on maternal outcomes. Methods: This single-center retrospective cohort study included patients transferred to our hospital due to postpartum hemorrhage between April 1, 2013, and March 31, 2019. The primary outcome was mortality, and the secondary outcomes were morbidity (duration of hospital stay, duration of intensive care unit stay, admission to the intensive care unit, respirator use, duration of ventilator support, acute kidney injury, transfusion-associated circulatory overload/transfusion-related acute lung injury, hysterectomy, composite adverse events, blood transfusion initiation time, blood transfusion volume, and treatment for postpartum hemorrhage). An in-hospital call system implementation commenced on April 1, 2016. The study outcomes were compared 3 years before and after implementing the call system. Results: The blood transfusion initiation time and duration of hospital stay were significantly shortened after implementing the call system for postpartum hemorrhage. No maternal deaths were observed after implementing the system. Conclusions: Implementing call systems specialized for postpartum hemorrhage in tertiary emergency facilities may improve maternal outcomes. [ABSTRACT FROM AUTHOR]

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Combination strategies to improve immunotherapy response in microsatellite stable metastatic colorectal cancer (MSS mCRC) remain an unmet need. Several single-arm clinical trials have shown promising synergistic effects between regorafenib and ICIs; however, some contradictory results have also been reported. Randomized controlled trials are needed to further validate the combination of regorafenib with ICIs. In addition, low-dose radiotherapy has been demonstrated to induce local immune responses by reprogramming the tumor microenvironment when combined with high-dose radiotherapy and ICIs. In this study, we designed a prospective, randomized, controlled phase II trial to investigate the efficacy and safety of regorafenib in combination with high/lowdose radiotherapy plus toripalimab in MSS mCRC compared to regorafenib alone. Patients with MSS metastatic adenocarcinoma of the colon or rectum will be enrolled and randomly assigned into two arms: a control arm and an experimental arm. Patients in the control arm will receive regorafenib monotherapy (120 mg once daily on days 1-21 of each 28 days cycle). Patients in the experimental arm will first receive one cycle of regorafenib (80 mg once daily on days 1-21 of each 28 days cycle) and toripalimab (240mg, q3w), followed by high-dose (4-8 fractions of 8-12Gy) and low-dose (1-10Gy at 0.5-2Gy/fraction) radiotherapy, and then continue regorafenib and toripalimab treatment. The primary endpoint is the objective response rate, and the secondary endpoints are disease control rate, duration of remission, median progress-free survival, median overall survival, and adverse events. Recruitment started in August 2023 and is ongoing. [ABSTRACT FROM AUTHOR]

Background: In 2022 the Academy of Breastfeeding Medicine (ABM) published Clinical Protocol #36: The Mastitis Spectrum, which aims to update clinical approaches to management of benign lactation-related breast inflammation. The protocol has been timely because of the exponential increase in knowledge about the human milk microbiome over the past decade. This Commentary aims to continue respectful debate amongst clinicians and researchers within the Academy of Breastfeeding Medicine and more broadly, confident that we share a fundamental commitment to promote breastfeeding and support the well-being of lactating women, their infants and their families. Analysis: Although Clinical Protocol #36 offers advances, it does not fulfil the principles of best practice implementation science for translation of evidence into clinical guidelines. Clinical Protocol #36 inaccurately represents studies; misrepresents theoretical models as proven aetiologies; does not consistently attribute sources; does not reliably apply the SORT taxonomy; and relies upon single case reports. As a result, various recommendations in Clinical Protocol #36 lack an evidence-base or credible underlying theoretical model. This includes recommendations to use 'lymphatic drainage' massage, therapeutic ultrasound, and oral lecithin. Similarly, based on a contestable theoretical model which is presented as fact, Clinical Protocol #36 makes the recommendation to either reduce frequency of milk removal or to maintain current frequency of milk removal during an episode of breast inflammation. Although Clinical Protocol #36 limits this advice to cases of 'hyperlactation', the diagnosis 'hyperlactation' itself is undefinable. As a result, this recommendation may put breastfeeding women who present with breast inflammation at risk of worsened inflammation and decreased breast milk production. Conclusion: Clinical Protocol #36 offers some advances in the management of breast inflammation. However, Clinical Protocol #36 also exposes clinicians to two international trends in healthcare which undermine health system sustainability: overdiagnosis, including by over-definition, which increases risk of overtreatment; and antibiotic over-use, which worsens the crisis of global antimicrobial resistance. Clinical Protocol #36 also recommends unnecessary or ineffective interventions which may be accessed by affluent patients within advanced economies but are difficult to access for the global majority. The Academy of Breastfeeding Medicine may benefit from a review of processes for development of Clinical Protocols. [ABSTRACT FROM AUTHOR]

There is a lack of clinical protocols for re-irradiation in paediatric central nervous system (CNS) tumours. To fill this void, the Swedish Workgroup of Paediatric Radiotherapy (SBRTG) compiled national guidelines on re-irradiation in paediatric CNS tumours (diffuse intrinsic pontine glioma, ependymoma, germinoma and medulloblastoma). These have been in clinical practice since 2019 in all paediatric radiotherapy centres in Sweden. Since the implementation, the guidelines have been complemented with a yearly review on clinical outcome and toxicities in all paediatric patients treated according to the guidelines. This article presents the Swedish national guidelines on re-irradiation in paediatric CNS tumours. • Re-irradiation of CNS tumours in paediatric patients is becoming more common. • Clinical guidelines for re-irradiation of paediatric CNS tumours are lacking. • We present Swedish national guidelines on re-irradiation in paediatric CNS tumours. [ABSTRACT FROM AUTHOR]