Vascular Patterning as Integrative Readout of Complex Molecular and Physiological Signaling by VESsel GENeration Analysis.

The molecular signaling cascades that regulate angiogenesis and microvascular remodeling are fundamental to normal development, healthy physiology, and pathologies such as inflammation and cancer. Yet quantifying such complex, fractally branching vascular patterns remains difficult. We review application of NASA's globally available, freely downloadable VESsel GENeration (VESGEN) Analysis software to numerous examples of 2D vascular trees, networks, and tree-network composites. Upon input of a binary vascular image, automated output includes informative vascular maps and quantification of parameters such as tortuosity, fractal dimension, vessel diameter, area, length, number, and branch point. Previous research has demonstrated that cytokines and therapeutics such as vascular endothelial growth factor, basic fibroblast growth factor (fibroblast growth factor-2), transforming growth factor-beta-1, and steroid triamcinolone acetonide specify unique "fingerprint" or "biomarker" vascular patterns that integrate dominant signaling with physiological response. In vivo experimental examples described here include vascular response to keratinocyte growth factor, a novel vessel tortuosity factor; angiogenic inhibition in humanized tumor xenografts by the anti-angiogenesis drug leronlimab; intestinal vascular inflammation with probiotic protection by Saccharomyces boulardii, and a workflow programming of vascular architecture for 3D bioprinting of regenerative tissues from 2D images. Microvascular remodeling in the human retina is described for astronaut risks in microgravity, vessel tortuosity in diabetic retinopathy, and venous occlusive disease.

Use of VESsel GENeration with Optical Coherence Tomography Angiography and Fluorescein Angiography for Detection and Quantification of Vascular Changes in Mild and Moderate Diabetic Retinopathy.

(1) Background: Previously, VESsel GENeration (VESGEN) software was used to map and quantify vascular changes observed on fluorescein angiography (FA) in subjects (n = 15 eyes) with retinal pathology ranging from mild non-proliferative diabetic retinopathy (NPDR) to proliferative diabetic retinopathy (PDR). In the current study, we used VESGEN for the assessment of individuals with early-stage NPDR imaged by FA (Cohort 1) and by optical coherence tomography angiography (OCTA; Cohort 2). (2) Methods: Cohort 1 included type 2 diabetics (T2D), represented 21 eyes (ranging from no DR to moderate DR), and also included nondiabetic controls (NDC; n = 15 eyes). Cohort 2 consisted of 23 eyes from T2D subjects (including no DR subjects and moderate DR subjects) and NDC (n = 18 eyes). (3) Results: In the FA-VESGEN study, total tortuosity (Tv) of microvessels (G ≥ 6) increased in T2D with mild DR compared to the controls. In contrast, the VESGEN analysis of OCTA images showed that vessel length (characterized as density) was lower in T2D subjects before the diagnosis of DR and following the diagnosis of DR when compared to the controls. Additionally, T2D showed a significant decrease in vessel area (density). (4) Conclusions: FA elucidated the vessel morphology of small-generation microvessels to a greater degree than OCTA; however, OCTA identified changes in vessel density better than FA. VESGEN analysis can be used with both standard FA and OCTA to facilitate our understanding of early events in DR, including before the clinical diagnosis of DR.

Remediation Programs for Regulated Health Care Professionals: A Scoping Review.

PURPOSE: Clinical competence is essential for providing safe, competent care and is regularly assessed to ensure health care practitioners maintain competence. When deficiencies in competence are identified, practitioners may undergo remediation. However, there is limited evidence regarding the effectiveness of remediation programs. The purpose of this review is to examine the purpose, format, and outcomes of remediation programs for regulated health care practitioners. METHODS: All six stages of the scoping review process as recommended by Levac et al were undertaken. A search was conducted within MEDLINE, Embase, CINAHL, ERIC, gray literature databases, and websites of Canadian provincial regulatory bodies. Emails were sent to Registrars of Canadian regulatory bodies to supplement data gathered from their websites. RESULTS: A total of 14 programs were identified, primarily for physicians (n = 8). Reasons for remediation varied widely, with some programs identifying multiple reasons for referral such as deficiencies in recordkeeping (n = 7) and clinical skills (n = 6). Most programs (n = 9) were individualized to address specific deficiencies in competence. The process of remediation followed three stages: (1) assessment, (2) active remediation, and (3) reassessment. Most programs (n = 12) reported that remediation was effective in improving competence. CONCLUSIONS: Regulatory bodies should consider implementing individualized remediation programs to ensure that clinicians' deficiencies in competence are addressed effectively. Further research is indicated, using reliable and valid outcome measures to assess competence immediately after remediation programs and beyond.

A Framework for Developing Antiracist Medical Educators and Practitioner-Scholars.

With an increasing awareness of the disparate impact of COVID-19 on historically marginalized populations and acts of violence on Black communities in 2020, academic health centers across the United States have been prioritizing antiracism strategies. Often, medical students and residents have been educated in the concepts of equity and antiracism and are ready to tackle these issues in practice. However, faculty are not prepared to respond to or integrate antiracism topics into the curriculum. Leaders in faculty affairs, education, diversity, and other departments are seeking tools, frameworks, expertise, and programs that are best suited to meet this imminent faculty development need. In response to these demands for guidance, the authors came together to explore best practices, common competencies, and frameworks related to antiracism education. The focus of their work was preparing faculty to foster antiracist learning environments at traditionally predominantly White medical schools. In this Scholarly Perspective, the authors describe their collaborative work to define racism and antiracism education; propose a framework for antiracism education for faculty development; and outline key elements to successfully build faculty capacity in providing antiracism education. The proposed framework highlights the interplay between individual learning and growth and the systemic and institutional changes needed to advance antiracist policies and practices. The key elements of the framework include building foundational awareness, expanding foundational knowledge on antiracism, embedding antiracism education into practice, and dismantling oppressive structures and measuring progress. The authors list considerations for program planning and provide examples of current work from their institutions. The proposed strategies aim to support all faculty and enable them to learn, work, and educate others in an antiracist learning environment.

Can Targeting Hypoxia-Mediated Acidification of the Bone Marrow Microenvironment Kill Myeloma Tumor Cells?

Multiple myeloma (MM) is an incurable cancer arising from malignant plasma cells that engraft in the bone marrow (BM). The physiology of these cancer cells within the BM microenvironment (TME) plays a critical role in MM development. These processes may be similar to what has been observed in the TME of other (non-hematological) solid tumors. It has been long reported that within the BM, vascular endothelial growth factor (VEGF), increased angiogenesis and microvessel density, and activation of hypoxia-induced transcription factors (HIF) are correlated with MM progression but despite a great deal of effort and some modest preclinical success the overall clinical efficacy of using anti-angiogenic and hypoxia-targeting strategies, has been limited. This review will explore the hypothesis that the TME of MM engrafted in the BM is distinctly different from non-hematological-derived solid tumors calling into question how effective these strategies may be against MM. We further identify other hypoxia-mediated effectors, such as hypoxia-mediated acidification of the TME, oxygen-dependent metabolic changes, and the generation of reactive oxygen species (ROS), that may prove to be more effective targets against MM.

Opposite response of blood vessels in the retina to 6° head-down tilt and long-duration microgravity.

The Spaceflight Associated Neuro-ocular Syndrome (SANS), associated with the headward fluid shifts incurred in microgravity during long-duration missions, remains a high-priority health and performance risk for human space exploration. To help characterize the pathophysiology of SANS, NASA's VESsel GENeration Analysis (VESGEN) software was used to map and quantify vascular adaptations in the retina before and after 70 days of bed rest at 6-degree Head-Down Tilt (HDT), a well-studied microgravity analog. Results were compared to the retinal vascular response of astronauts following 6-month missions to the International Space Station (ISS). By mixed effects modeling, the trends of vascular response were opposite. Vascular density decreased significantly in the 16 retinas of eight astronauts and in contrast, increased slightly in the ten retinas of five subjects after HDT (although with limited significance). The one astronaut retina diagnosed with SANS displayed the greatest vascular loss. Results suggest that microgravity is a major variable in the retinal mediation of fluid shifts that is not reproduced in this HDT bed rest model.

Determinants of serum magnesium abnormalities and outcome among admissions to the intensive care unit.

Serum magnesium is a frequently measured and treated electrolyte. However, few studies have examined magnesium level abnormalities and outcome in critically ill patients. Our objective was to determine the epidemiology and outcome of magnesium abnormalities among patients admitted to intensive care units (ICU). A retrospective cohort including adult patients admitted to three ICUs in southeast Queensland was assembled. Magnesium levels < 0.7, 0.7-1.0, and > 1.0 mmol/L were classified as low, normal, and high, respectively. Among 14,101 patients, the median age was 59.3 (interquartile range; IQR, 45.1-70.5), 7493 (56.4%) were male, and the median APACHE III score was 48 (IQR, 34-66). At admission, 3357 (23.8%) patients were classified as having hypomagnesemia, 1682 (11.9%) hypermagnesemia, 165 (1.2%) mixed, and 8897 (63.1%) as normal. Patients with magnesium abnormalities were more likely to be underweight and to have higher APACHE III scores. The overall 30-day case fatality was 8.2% (1155/14,101). Compared to those with normal levels, patients with hypermagnesemia at admission were at two-fold increased crude risk for death (relative risk; RR, 2.09; 95% confidence interval; CI, 1.83-2.39; p < 0.0001). After controlling for confounding variables in logistic regression analysis, neither admission hypo- nor hypermagnesemia was associated with death. However, development of ICU acquired hypermagnesemia among those with normal (odds ratio; OR, 1.34; 95% CI, 1.02-1.77; p = 0.034) and low (OR, 1.67; 95% CI, 1.15-2.41; p = 0.006) admission magnesium levels increased the risk for death. Magnesium abnormalities are common among patients managed in ICUs. The determinants of ICU-acquired hypermagnesemia and its adverse effect on outcome warrants further investigation.

Decreased Vascular Patterning in the Retinas of Astronaut Crew Members as New Measure of Ocular Damage in Spaceflight-Associated Neuro-ocular Syndrome.

Purpose: Ocular structural and functional changes, collectively termed spaceflight-associated neuro-ocular syndrome (SANS), have been described in astronauts undergoing long-duration missions in the microgravity environment of the International Space Station. We tested the hypothesis that retinal vascular remodeling, particularly by smaller vessels, mediates the chronic headward fluid shifts associated with SANS. Methods: As a retrospective study, arterial and venous patterns extracted from 30° infrared Heidelberg Spectralis retinal images of eight crew members acquired before and after six-month missions were analyzed with NASA's recently released VESsel GENeration Analysis (VESGEN) software. Output parameters included the fractal dimension and overall vessel length density that was further classified into large and small vascular branching generations. Vascular results were compared with SANS-associated clinical ocular measures. Results: Significant postflight decreases in Df, Lv, and in smaller but not larger vessels were quantified in 11 of 16 retinas for arteries and veins (P value for Df, Lv, and smaller vessels in all 16 retinas were ≤0.033). The greatest vascular decreases occurred in the only retina displaying clinical evidence of SANS by choroidal folds and optic disc edema. In the remaining 15 retinas, decreases in vascular density from Df and Lv ranged from minimal to high by a custom Subclinical Vascular Pathology Index. Conclusions: Together with VESGEN, the Subclinical Vascular Pathology Index may represent a new, useful SANS biomarker for advancing the understanding of SANS etiology and developing successful countermeasures for long duration space exploration in microgravity, although further research is required to better characterize retinal microvascular adaptations.

Self-Monitoring of Blood Glucose in Youth-Onset Type 2 Diabetes: Results From the TODAY Study.

OBJECTIVE: To determine whether self-monitoring of blood glucose (SMBG) is associated with lower HbA1c in youth with type 2 diabetes taking oral medications only or after starting insulin for persistently elevated HbA1c. RESEARCH DESIGN AND METHODS: Treatment Options for Type 2 Diabetes in Adolescents and Youth (TODAY) study participants (n = 699) taking oral medications were asked to perform SMBG twice daily. After reaching primary outcome (PO) (HbA1c ≥8% [64 mmol/mol]) over 6 months or an inability to wean from temporary insulin because of metabolic decompensation), insulin glargine was started. HbA1c and percent of SMBG (SMBG%) (percent days when the meter was used one or more times) before and after PO were analyzed. RESULTS: SMBG declined over time and was inversely related to HbA1c (P < 0.0001). Of 298 youth who reached PO and started insulin, 282 had SMBG data. At PO, mean ± SD age was 15.8 ± 2.3 years, BMI 35.5 ± 7.9 kg/m2, and HbA1c 9.6 ± 2.0% (81 ± 21.9 mmol/mol); 65.3% were female. Median SMBG% was 40% at PO, which increased to 49% after 6 months and fell to 41% after 1 year on insulin. At PO, 22% of youth checked ≥80% of days, which increased to 25% and fell to 19% after 6 and 12 months using insulin, respectively. At PO, compared with those who checked <80%, youth who checked ≥80% were younger and with a lower BMI, HbA1c, and blood pressure. SMBG ≥80% was associated with ≥1% reduction in HbA1c at 6 and 12 months after insulin initiation. CONCLUSIONS: Low SMBG adherence was common and associated with higher HbA1c. Optimal SMBG frequency in youth using or not using insulin, and whether less frequent SMBG is a marker for overall worse self-care, require further study.

Trends in the evolution of the prodynorphin gene in teleosts: cloning of eel and tilapia prodynorphin cDNAs.

The detection of the prodynorphin gene in anuran amphibians and lungfishes may indicate that this gene arose as a result of the duplication of the proenkephalin gene early during the divergence of the Sarcopterygii, or that this gene may predate the divergence of the ray-finned fish and the lobe-finned fish. The cloning of prodynorphin-related genes from the pufferfish and zebrafish supports the latter hypothesis. This study analyzes trends in the radiation of the prodynorphin gene in teleosts. Prodynorphin cDNAs were cloned from the brain of the eel Anguilla rostrata and the Nile tilapia, Oreochromis niloticus. These teleost prodynorphin sequences have distinct alpha-neoendorphin, dynorphin A, and dynorphin B sequences, and a novel opioid sequence, YGGFI. The relationship of these teleost prodynorphin sequences to other actinopterygian and sarcopterygian prodynorphin sequences will be discussed.

Using whole mount in situ hybridization to link molecular and organismal biology.

Whole mount in situ hybridization (WISH) is a common technique in molecular biology laboratories used to study gene expression through the localization of specific mRNA transcripts within whole mount specimen. This technique (adapted from Albertson and Yelick, 2005) was used in an upper level undergraduate Comparative Vertebrate Biology laboratory classroom at Syracuse University. The first two thirds of the Comparative Vertebrate Biology lab course gave students the opportunity to study the embryology and gross anatomy of several organisms representing various chordate taxa primarily via traditional dissections and the use of models. The final portion of the course involved an innovative approach to teaching anatomy through observation of vertebrate development employing molecular techniques in which WISH was performed on zebrafish embryos. A heterozygous fibroblast growth factor 8 a (fgf8a) mutant line, ace, was used. Due to Mendelian inheritance, ace intercrosses produced wild type, heterozygous, and homozygous ace/fgf8a mutants in a 1:2:1 ratio. RNA probes with known expression patterns in the midline and in developing anatomical structures such as the heart, somites, tailbud, myotome, and brain were used. WISH was performed using zebrafish at the 13 somite and prim-6 stages, with students performing the staining reaction in class. The study of zebrafish embryos at different stages of development gave students the ability to observe how these anatomical structures changed over ontogeny. In addition, some ace/fgf8a mutants displayed improper heart looping, and defects in somite and brain development. The students in this lab observed the normal development of various organ systems using both external anatomy as well as gene expression patterns. They also identified and described embryos displaying improper anatomical development and gene expression (i.e., putative mutants). For instructors at institutions that do not already own the necessary equipment or where funds for lab and curricular innovation are limited, the financial cost of the reagents and apparatus may be a factor to consider, as will the time and effort required on the part of the instructor regardless of the setting. Nevertheless, we contend that the use of WISH in this type of classroom laboratory setting can provide an important link between developmental genetics and anatomy. As technology advances and the ability to study organismal development at the molecular level becomes easier, cheaper, and increasingly popular, many evolutionary biologists, ecologists, and physiologists are turning to research strategies in the field of molecular biology. Using WISH in a Comparative Vertebrate Biology laboratory classroom is one example of how molecules and anatomy can converge within a single course. This gives upper level college students the opportunity to practice modern biological research techniques, leading to a more diversified education and the promotion of future interdisciplinary scientific research.

Stimulation of Xenopus laevis oocyte maturation by methyl-beta-cyclodextrin.

The cholesterol-depleting drug methyl-beta-cyclodextrin (Me-beta-CD) was tested for its effects on amphibian oocyte maturation, cholesterol depletion, and low-density membrane recovery. Progesterone-induced oocyte maturation was accelerated by pretreatment of cells with 5-50 mM Me-beta-CD in a dose-dependent manner. Treatment of oocytes with 50 mM Me-beta-CD alone was sufficient to induce germinal vesicle breakdown, stimulate formation of meiotic spindles, and stimulate phosphorylation of mitogen-activated protein kinase over time courses longer than those observed after progesterone treatment. After short-term (30 min) labeling of oocytes with [(3)H]cholesterol, 30-90 min of treatment with 5-50 mM Me-beta-CD removed 50%-70% of cell- associated label, and cholesterol depletion was not observed with alpha-cyclodextrin. After long-term (20-23 h) labeling of oocytes with [(3)H]cholesterol, Me-beta-CD treatment resulted in dose- dependent cholesterol depletion in the 5-50 mM range, and 50 mM Me-beta-CD removed approximately 50% of cell-associated label after 9 h. Treatment of oocytes with 5-50 mM Me-beta-CD also decreased recovery of low-density membrane by detergent-free sucrose gradient centrifugation. These results implicate cholesterol and low-density membrane domains in the signaling mechanisms leading to germinal vesicle breakdown in amphibian oocytes.