Differential proteomics of circulating extracellular vesicles of placental origin isolated from women with early-onset preeclampsia reveal aberrant innate immune and hemostasis processes.

PROBLEM: Early-onset preeclampsia (EOPE) is a severe gestational hypertensive disorder with significant feto-maternal morbidity and mortality due to uteroplacental insufficiency. Circulating extracellular vesicles of placental origin (EV-P) are known to be involved in the pathophysiology of EOPE and might serve as an ideal reservoir for its specific biomarkers. Therefore, we aimed to characterize and perform comparative proteomics of circulating EV-P from healthy pregnant and EOPE women before delivery. METHOD OF STUDY: The EV-P from both groups were isolated using immunoaffinity and were characterized using transmission electron microscopy, dynamic light scattering, nanoparticle tracking analysis, and immunoblotting. Following IgG albumin depletion, the pooled proteins that were isolated from EV-P of both groups were subjected to quantitative TMT proteomics. RESULTS: Circulating term EV-P isolated from both groups revealed ∼150 nm spherical vesicles containing CD9 and CD63 along with placental PLAP and HLA-G proteins. Additionally, the concentration of EOPE-derived EV-P was significantly increased. A total of 208 proteins were identified, with 26 among them being differentially abundant in EV-P of EOPE women. This study linked the pathophysiology of EOPE to 19 known and seven novel proteins associated with innate immune responses such as complement and TLR signaling along with hemostasis and oxygen homeostasis. CONCLUSION: The theory suggesting circulating EVs of placental origin could mimic molecular information from the parent organ-"the placenta"-is strengthened by this study. The findings pave the way for possible discovery of novel prognostic and predictive biomarkers as well as provide insight into the mechanisms driving the pathogenesis of EOPE.

Isolation of serum-derived placental/amnio-chorionic extracellular vesicles across pregnancy by immunoaffinity using PLAP and HLA-G.

In brief: Circulating extracellular vesicles of placental/amniochorionic origin carry placental/amniochorionic proteins and nucleic acids with the potential to facilitate non-invasive diagnosis of pregnancy-related disorders. The study reports an improvised method for the enriched isolation of extracellular vesicles of placental/amniochorionic origin using the two markers, PLAP and HLA-G. Abstract: Extracellular vesicles (EVs) are membrane-bound nanovesicles secreted from the cells into extracellular space and body fluids. They are considered 'fingerprints of parent cells', which can reflect their physiological and functional states. During pregnancy, EVs are produced by the syncytiotrophoblasts and extravillous trophoblasts and are released into the maternal bloodstream. In the present study, placental alkaline phosphatase (PLAP)-specific extracellular vesicles were isolated from maternal serum-derived EVs (SDE) across pregnancy. Transmission electron microscopy and dynamic light scattering analysis showed that the isolated EVs exhibited a spherical morphology with ~30-150 nm size range. Nanoparticle tracking analysis indicated that the concentration of PLAP+ serum-derived EVs (PLAP+-SDE) increased across the gestation. PLAP+-SDE contained DNA with LINE1 promoter methylation pattern. C19 miRNA cluster miRNAs (miR 515-5p, 519e and 520f) were present in PLAP+-SDE along with other miRNAs (miR-133-3p, miR210-3p and miR-223-3p). PLAP+-SDE confirmed the presence of EV markers (CD63 and CD9), along with placental proteins (PLAP and cullin 7). A modified novel strategy to extract an enriched population of circulating placental/amniochorionic EVs was devised employing an additional marker of extravillous trophoblasts, human leukocyte antigen G (HLA-G), along with PLAP. The isolated pooled placental/amniochorionic (PLAP+&HLA-G+) serum-derived EVs (PP-SDE) showed ~two-fold increased protein levels of HLA-G in the third-trimester pregnant women compared to the non-pregnant controls. Future studies will be focused on validation of this novel strategy to isolate an enriched population of placental/amniochorionic EVs to facilitate a better understanding of placental physiology and pathophysiology.

Evidence-based interventions to reduce maternal malnutrition in low and middle-income countries: a systematic review.

Introduction: Despite remarkable strides in global efforts to reduce maternal mortality, low-and middle-income countries (LMICs) continue to grapple with a disproportionate burden of maternal mortality, with malnutrition emerging as a significant contributing factor to this enduring challenge. Shockingly, malnourished women face a mortality risk that is twice as high as their well-nourished counterparts, and a staggering 95% of maternal deaths in 2020 occurred within LMICs. The critical importance of addressing maternal malnutrition in resource-constrained settings cannot be overstated, as compelling research studies have demonstrated that such efforts could potentially save thousands of lives. However, the landscape is marred by a scarcity of evidence-based interventions (EBIs) specifically tailored for pregnant individuals aimed at combatting maternal malnutrition and reducing mortality rates. It is against this backdrop that our study endeavors to dissect the feasibility, adoption, sustainability, and cost-effectiveness of EBIs designed to combat maternal malnutrition. Methods: Our comprehensive search encompassed eight prominent databases covering the period from 2003 to 2022 in LMICs. We began our study with a comprehensive search across multiple databases, yielding a total of 149 studies. From this initial pool, we eliminated duplicate entries and the remaining studies underwent a thorough screening process resulting in the identification of 63 full-text articles that aligned with our predefined inclusion criteria. Results: The meticulous full-text review left us with a core selection of six articles that shed light on interventions primarily centered around supplementation. They underscored a critical issue -the limited understanding of effective implementation in these countries, primarily attributed to inadequate monitoring and evaluation of interventions and insufficient training of healthcare professionals. Moreover, our findings emphasize the pivotal role of contextual factors, such as cultural nuances, public trust in healthcare, the prevalence of misinformation, and concerns regarding potential adverse effects of interventions, which profoundly influence the successful implementation of these programs. Discussion: While the EBIs have shown promise in reducing maternal malnutrition, their true potential for feasibility, adoption, cost-effectiveness, and sustainability hinges on their integration into comprehensive programs addressing broader issues like food insecurity and the prevention of both communicable and non-communicable diseases.

Early prediction of pre-eclampsia using circulating placental exosomes: Newer insights.

Pre-eclampsia (PE), a multifactorial de novo hypertensive pregnancy disorder, is one of the leading causes of foeto-maternal morbidity and mortality. Currently, antihypertensive drugs are the first-line therapy for PE and evidence suggests that low-dose aspirin initiated early in high risk pregnancies may reduce the risk of development or severity of PE. However, an early prediction of this disorder remains an unmet clinical challenge. Several potential serum biomarkers associated with maternal immunoregulation and placental angiogenesis have been evaluated but are ineffective and inconsistent for early prediction. Although placental biomarkers would be more specific and sensitive in predicting the risk of PE, accessing the placenta during pregnancy is not feasible. Circulating placental exosomes (pEXO), originating from foeto-maternal interface, are being evaluated as the placenta's surrogate and the best source of non-invasive placental biomarkers. pEXO appear in the maternal circulation starting from six weeks of gestation and its dynamic biological cargo across pregnancy is associated with successful pregnancy outcomes. Therefore, monitoring changes in pEXO expression profiles could provide new insights into the prediction, diagnosis and treatment of PE. This narrative review comprehensively summarizes the available literature on the candidate predictive circulating biomarkers evaluated for PE to date. In particular, the review elucidates the current knowledge of distinct molecular signatures emanating from pEXO in pre-eclamptic women to support the discovery of novel early predictive biomarkers for effective intervention and management of the disease.

Prediction of maternal and foetal outcomes among patients with preeclampsia using circulatory biomarkers (MMP-9 & ST2) - A prospective cohort study.

INTRODUCTION: Preeclampsia is one of the major causes of maternal and foetal morbidity and mortality worldwide. Its complications include but are not limited to eclampsia, intracerebral haemorrhage and cardiovascular diseases in the later stages of life. The combination of clinical and risk variables and a panel of multiple biomarkers will help clinicians in risk stratification and prognostication of clinical outcomes among preeclamptic women. We evaluated MMP-9 (matrix metalloproteinase - 9) and ST2 (suppression of tumorigenicity 2) for utility as biomarkers and for predicting maternal and foetal outcomes in women with preeclampsia. METHODS: This prospective cohort study involved 49 preeclamptic women and 80 healthy controls. Biomarkers were measured in plasma using ELISA. The patients were followed up to assess maternal and foetal outcomes. RESULTS: The mean value of MMP-9 was 2.42 ng/mL in the preeclamptic group and 2.67 ng/mL in controls. The mean value of ST2 (1937.4 ± 747.81) in the preeclamptic group was high compared to the control group (1005.7 ± 683.6) and the difference was significant (P = 0.0001). The study population was divided into those with high and low MMP-9 and those with high and low ST2. Lower levels of MMP-9 seemed to be related to both early and late onset preeclampsia. The ROC (Receiver Operating Characteristic) curve did not show the ability to predict maternal and foetal outcomes. DISCUSSION: Our study demonstrated that women with preeclampsia had low MMP-9 and high ST2 compared to healthy pregnant women. But neither of the biomarkers could predict complications of preeclampsia.

A New Model of Learning: Environmental Health in a Global World.

INTRODUCTION: Environmental Health in a Global World at New York University was re-designed as a class participatory effort, challenging undergraduate students to understand environmental hazards and the resultant adverse health outcomes by embracing the inherent complexity of environmental risks and proposing solutions. METHODS: Following introductory lectures, students are placed into teams and assigned a specific perspective, or avatar, which includes learning to see the challenge from the perspective of a technical expert such as a biologist, an engineer, or an anthropologist. The teams then design specific systems maps to visualize the complex interactions that lead to adverse health outcomes after a given environmental exposure. The maps highlight potential leverage points where relatively minor interventions can provide a disproportionate benefit in health outcomes. The teams then explore potential interventions and identify the potential unintended consequences of those actions, develop and advocate for innovative new strategies to mitigate risk and improve outcomes. RESULTS AND DISCUSSION: Over the past 5 years, we have taught this methodology to over 680 students with strong, student-oriented results. The teams created and presented more than 100 strategies, addressing a diverse set of environmental challenges that include water contamination, gun violence, air pollution, environmental justice, health security, and climate change. Developing the strategies helped the students understand environmental threats in a more holistic way, provided them with some agency in finding solutions, and offered an opportunity for them to improve their presentation skills. The responses in course evaluations have been enthusiastic, with many students reporting a deep impact on their college experience.

USE OF OPTICAL COHERENCE TOMOGRAPHY IN DETECTING RETINAL TEARS IN ACUTE, SYMPTOMATIC POSTERIOR VITREOUS DETACHMENT.

PURPOSE: To evaluate the association of posterior vitreous opacities (PVOs) on optical coherence tomography with retinal tears identified on examination in patients with acute, symptomatic posterior vitreous detachment (PVD). METHODS: Data were retrospectively collected from the medical records of 388 patients with acute, symptomatic PVD between January 1, 2021, and June 30, 2021. Included patients had received a primary diagnosis of PVD and presented with flashes and/or floaters. Optical coherence tomography scans were reviewed by two separate readers for the presence of PVOs. The primary outcome was the presence of retinal tear on fundus photograph and on examination. RESULTS: Of 388 patients who presented with acute PVD symptoms, 90 (23.2%) were found to have a retinal tear on dilated fundus examination. Among these patients, 78 (86.7%) were found to have PVOs on optical coherence tomography. Statistical analysis demonstrated a significant relationship between the presence of PVOs and retinal tear ( P < 0.01). The sensitivity and specificity of this finding was 86.7% and 72.5%, respectively. Further analysis included area under the curve from receiver operating characteristic curve which was found to be 0.80. CONCLUSION: The presence of PVOs on optical coherence tomography is suggestive of a retinal tear in patients with acute, symptomatic PVD.

Clinical Proteomics for Meningioma: An Integrated Workflow for Quantitative Proteomics and Biomarker Validation in Formalin-Fixed Paraffin-Embedded Tissue Samples.

Clinical proteomics is a rapidly emerging frontier in laboratory medicine. High-throughput proteomic investigations of biopsy tissues provide mechanistic insights into complex human diseases. For large-scale proteomics, formalin-fixed and paraffin-embedded (FFPE) tissue samples offer a viable alternative to fresh-frozen (FF) tissues that have restricted availability. In this context, meningioma is one of the most common primary brain tumors where innovation in diagnostics and therapeutic targets can benefit from clinical proteomics. We present here an integrated workflow for quantitative proteomics and biomarker validation of meningioma FFPE tissues. Applying label-free quantitative (LFQ) proteomics, we reproducibly (Pearson's correlation: 0.84-0.91) obtained an in-depth proteome coverage (nearly 4000 proteins per sample) from 120 min gradient of single unfractionated mass spectrometry run. Furthermore, building upon LFQ data and literature curated set of meningioma-associated proteins, we validated VIM, AHNAK, and CLU from FFPE tissues using selected reaction monitoring (SRM) assay and compared its performance with FF tissues. This study illustrates how knowledge from label-free proteomics can be integrated for selecting peptides for targeted validation and suggests that FFPE tissues are comparable to FF tissues for SRM assays. This quantitative clinical proteomics workflow is scalable for large-scale clinical diagnostics studies in the future, for example, utilizing the global repository of FFPE tissues in meningioma and possibly in other cancers.

Characteristics, safety and cost-effectiveness analysis of self-administered outpatient parenteral antibiotic therapy via a disposable elastomeric continuous infusion pump at two county hospitals in Houston, Texas, United States.

WHAT IS KNOWN AND OBJECTIVE: Outpatient parenteral antibiotic therapy (OPAT) is an attractive option for patients who require parenteral antimicrobials as outpatients. Few OPAT studies have assessed the impact of IV antibiotic therapy via elastomeric continuous pumps, with most having been conducted outside the United States and few in county hospitals. The OPAT program in Harris Health system, the county hospital system of Houston, Texas, United States, has implemented a disposable elastomeric continuous infusion pump (eCIP) for self-administered intravenous antibiotics (s-OPAT) since December 2018. Our goal was to describe the clinical characteristics of patients discharged with an eCIP, as well as the safety and cost-effectiveness of this pump. METHODS: We retrospectively analysed patients discharged from Harris Health hospitals between 12/2018 and 02/2021 with s-OPAT via eCIP at home. We extracted various patient characteristics and outcomes related to OPAT. RESULTS AND DISCUSSION: Among 481 OPAT patients during the study period, 91 patients received s-OPAT via eCIP. A total of 1925 days of s-OPAT were administered at home, with a median duration of 12 days. Eighty-three patients (93.4%) achieved a cure from infection, six patients (6.6%) had side effects, and nine patients (9.9%) experienced 30-day hospital readmission. Twenty-two patients (24.2%) presented to the ED during s-OPAT, with 13 patients (14.3%) presenting with PICC line concerns. We estimated that s-OPAT via eCIP saved $2,360,500 to $3,503,900 compared to inpatient-only therapy. WHAT IS NEW AND CONCLUSION: Our study showed that patients with s-OPAT via eCIP had a high cure rate with a relatively low incidence of side effects and 30-day hospital readmission. ED visits during therapy were relatively high, which indicates the necessity of close patient monitoring via the OPAT program. eCIP appears to be a good option to facilitate an early disposition of patients in county hospitals.

Real World Long-term Assessment of The Efficacy of Tocilizumab in Patients with COVID-19: Results From A Large De-identified Multicenter Electronic Health Record Dataset in the United States.

BACKGROUND: Studies have shown conflicting results on the efficacy of tocilizumab (TCZ) for patients with COVID-19, with many confounders of clinical status and limited duration of the observation. Here, we evaluate the real-world long-term efficacy of TCZ in COVID-19 patients. METHODS: We conducted a retrospective study of hospitalized adult patients with COVID-19 using a large US-based multicenter COVID-19 database (Cerner Real-World Data; updated in September, 2020). The TCZ group was defined as patients who received at least one dose of the drug. Matching weight (MW) and a propensity score weighting method were used to balance confounding factors. RESULTS: A total of 20,399 patients were identified. 1,510 and 18,899 were in the TCZ and control groups, respectively. After MW adjustment, no statistically significant differences in all-cause mortality were found for the TCZ vs. control group (Hazard Ratio [HR]:0.76, p=0.06). Survival curves suggested a better trend in short-term observation, driven from a subgroup of patients requiring oxygen masks, BIPAP or CPAP. CONCLUSION: We observed a temporal (early) benefit of TCZ, especially in patients on non-invasive high-flow supplemental oxygen. However, the benefit effects faded with longer observation. The long-term benefits and risks of TCZ should be carefully evaluated with follow-up studies.

Application of 2D-DIGE and iTRAQ Workflows to Analyze CSF in Gliomas.

Proteomics is an indispensable tool for disease biomarker discovery. It is widely used for the analysis of biological fluids such as cerebrospinal fluid (CSF), blood, and saliva, which further aids in our understanding of disease incidence and progression. CSF is often the biospecimen of choice in case of intracranial tumors, as rapid changes in the tumor microenvironment can be easily assessed due to its close proximity to the brain. On the contrary studies comprising of serum or plasma samples do not truly reflect the underlying molecular alterations due to the presence of protective blood-brain barrier. We have described in here the detailed workflows for two advanced proteomics techniques, namely, 2D-DIGE (two-dimensional difference in-gel electrophoresis) and iTRAQ (isobaric tag for relative and absolute quantitation), for CSF analysis. Both of these techniques are very sensitive and widely used for quantitative proteomics analysis.

Proteomic analysis of Streptomyces coelicolor in response to Ciprofloxacin challenge.

Multi-drug tolerance is an important phenotypic property that complicates treatment of infectious diseases and reshapes drug discovery. Hence a systematic study of the origins and mechanisms of resistance shown by microorganisms is imperative. Since soil-dwelling bacteria are constantly challenged with a myriad of antibiotics, they are potential reservoirs of resistance determinants that can be mobilized into pathogens over a period of time. Elucidating the resistance mechanisms in such bacteria could help future antibiotic discoveries. This research is a preliminary study conducted to determine the effects of ciprofloxacin (CIP) on the intrinsically resistant Gram-positive soil bacterium Streptomyces coelicolor. The effect was investigated by performing 2-DE on total protein extracts of cells exposed to sub-lethal concentrations of ciprofloxacin as compared to the controls. Protein identification by MALDI-TOF/TOF revealed 24 unique differentially expressed proteins, which were statistically significant. The down-regulation of proteins involved in carbohydrate metabolism indicated a shift in the cell physiology towards a state of metabolic shutdown. Furthermore, the observed decline in protein levels involved in transcription and translation machinery, along with depletion of enzymes involved in amino acid biosynthesis and protein folding could be a cellular response to DNA damage caused by CIP, thereby minimizing the effect of defective and energetically wasteful metabolic processes. This could be crucial for the initial survival of the cells before gene level changes could come into play to ensure survival under prolonged adverse conditions. These results are a first attempt towards profiling the proteome of S. coelicolor in response to antibiotic stress. This article is part of a Special Issue entitled: Trends in Microbial Proteomics. BIOLOGICAL SIGNIFICANCE: Soil-dwelling bacteria could serve as a reservoir of resistance determinants for clinically important bacteria. In this work, we investigated, for the first time, the differential proteomic profile of S. coelicolor cells in response to sub-inhibitory concentrations of Ciprofloxacin using 2-DE. Results indicate a shift in the cell physiology towards a state of metabolic shutdown, possibly to counter the DNA damage by ciprofloxacin. Further, up-regulation of GAPDH, RNA pol mRNA and Translation IF2 protein indicates a reprogramming of the cell for long-term survival. This study could serve as a basis for further investigations to elucidate the general mechanism by which soil bacteria exhibit resistance to fluroquinolones. This may help in developing new drug protocols and inventing novel drugs to counter resistance to this class of antibiotics in pathogenic bacteria.