Extended Exposure Topotecan Significantly Improves Long-Term Drug Sensitivity by Decreasing Malignant Cell Heterogeneity and by Preventing Epithelial-Mesenchymal Transition.

Maximum tolerable dosing (MTD) of chemotherapeutics has long been the gold standard for aggressive malignancies. Recently, alternative dosing strategies have gained traction for their improved toxicity profiles and unique mechanisms of action, such as inhibition of angiogenesis and stimulation of immunity. In this article, we investigated whether extended exposure (EE) topotecan could improve long-term drug sensitivity by preventing drug resistance. To achieve significantly longer exposure times, we used a spheroidal model system of castration-resistant prostate cancer. We also used state-of-the-art transcriptomic analysis to further elucidate any underlying phenotypic changes that occurred in the malignant population following each treatment. We determined that EE topotecan had a much higher barrier to resistance relative to MTD topotecan and was able to maintain consistent efficacy throughout the study period (EE IC50 of 54.4 nM (Week 6) vs. MTD IC50 of 2200 nM (Week 6) vs. 83.8 nM IC50 for control (Week 6) vs. 37.8 nM IC50 for control (Week 0)). As a possible explanation for these results, we determined that MTD topotecan stimulated epithelial-mesenchymal transition (EMT), upregulated efflux pumps, and produced altered topoisomerases relative to EE topotecan. Overall, EE topotecan resulted in a more sustained treatment response and maintained a less aggressive malignant phenotype relative to MTD topotecan.

Aviation and the Microbiome.

INTRODUCTION: Although the impact of microorganisms on their hosts has been investigated for decades, recent technological advances have permitted high-throughput studies of the collective microbial genomes colonizing a host or habitat, also known as the microbiome. This literature review presents an overview of microbiome research, with an emphasis on topics that have the potential for future applications to aviation safety. In humans, research is beginning to suggest relationships of the microbiome with physical disorders, including type 1 and type 2 diabetes mellitus, cardiovascular disease, and respiratory disease. The microbiome also has been associated with psychological health, including depression, anxiety, and the social complications that arise in autism spectrum disorders. Pharmaceuticals can alter microbiome diversity, and may lead to unintended consequences both short and long-term. As research strengthens understanding of the connections between the microbiota and human health, several potential applications for aerospace medicine and aviation safety emerge. For example, information derived from tests of the microbiota has potential future relevance for medical certification of pilots, accident investigation, and evaluation of fitness for duty in aerospace operations. Moreover, air travel may impact the microbiome of passengers and crew, including potential impacts on the spread of disease nationally and internationally. Construction, maintenance, and cleaning regimens that consider the potential for microbial colonization in airports and cabin environments may promote the health of travelers. Altogether, the mounting knowledge of microbiome effects on health presents several opportunities for future research into how and whether microbiome-based insights could be used to improve aviation safety.Davis JT, Uyhelji HA. Aviation and the microbiome. Aerosp Med Hum Perform. 2020; 91(8):651-661.

U.S. Statement of Demonstrated Ability Aeromedical Waivers.

INTRODUCTION: The Statement of Demonstrated Ability (SODA) is a type of U.S. aeromedical waiver used for disqualifying conditions that are not expected to change. About 21,000 (2%) U.S. pilots possess a SODA waiver.METHODS: We matched all pilot medical exams from the FAA's medical certification database from 2002 through 2011 to their respective accidents in the National Transportation Safety Board accident database. The association of SODA waivers and SODA conditions with the odds of an accident were explored using logistic regression techniques.RESULTS: For 3rd class flight exams, the presence of a SODA waiver was not associated with the odds of an accident. For the 1st and 2nd class exams, the accident odds ratio (OR = 1.45) was statistically significant. Crop dusting operations accounted for 17 of the 40 accidents where SODAs were present and returned a significant accident OR = 1.68. SODAs were not associated with the odds of accidents during other commercial operations. Six SODA conditions (amputation, internal eye, external eye, visual fields, bone and joint, and miscellaneous) were also found to have elevated ORs but were based on very small accident counts. NTSB investigators and the authors reviewed all accidents and none thought the SODA condition to be contributory.DISCUSSION: SODA waivers were not associated with increased accident odds except for crop dusting operations. Six specific SODA conditions also had elevated odds of an accident, but there was no evidence they contributed to the accidents. Overall, U.S. pilots with SODA waivers appear to have a satisfactory safety record.Mills WD, Davis JT. U.S. Statement of Demonstrated Ability aeromedical waivers. Aerosp Med Hum Perform. 2019; 90(12):1034-1040.

The U.S. Experience with Special Issuance Waivers.

INTRODUCTION: The special issuance (SI) waiver is the primary mechanism for U.S. pilots who do not meet FAA standards to obtain a medical certificate. About 34,000 pilots possess an SI waiver, but there is a large gap in knowledge of the relationship of SI waivers to aviation safety. METHODS: All FAA pilot medical exams from 2002 through 2011 were matched to the National Transportation Safety Board accident database. The association of an SI waiver with accidents was explored using logistic regression models. Accident rates were also calculated using a novel technique based on pilots' reported flight times on their applications for medical certification. RESULTS: For third-class flight exams overall, the presence of an SI waiver is associated with 8.7% lower odds of an accident than regular issuance exams. The calculated overall accident rate was 6.6 per 100,000 h. For the first and second-class exams, there was no significant association of SI waivers with safety for the overall group. The oldest and youngest pilots in the third-class group and the younger pilots in the first and second-class groups had somewhat elevated accident odds. The significance of these higher odds is uncertain, especially for the younger pilots who have a probable flight time bias. DISCUSSION: The overall FAA program of special issuance waivers shows no detrimental effect on aviation accidents and enables a large number of pilots to safely continue their aviation pursuits in spite of failure to meet specific regulatory medical standards.Mills WD, Davis JT. The U.S. experience with special issuance waivers. Aerosp Med Hum Perform. 2018; 89(10):905-911.

Substrate Stiffness Influences the Time Dependence of CTGF Protein Expression in Müller Cells.

Following ocular trauma and retinal detachment, gliotic changes in the retina may develop over the subsequent month, a process known as PVR (proliferative vitreoretinopathy). There have been no successful therapeutic interventions to inhibit PVR. The protein CTGF (Connective Tissue Growth Factor) has been associated with retinal PVR and other fibrotic diseases of the retina in clinical studies but the mechanistic link between different pathologies and retinal gliosis has not been determined. In addition, CTGF has been previously noted to be associated, in some cases, with YAP/TAZ (Yes-associated protein and Tafazzin protein complex), transcriptional regulatory proteins that change subcellular localization in response to mechanical cues, such as the stiffness of the underlying material. We have previously shown that the mRNA for CTGF is markedly (100-fold) upregulated in retinal Müller cells grown on soft substrates. In order to evaluate if the mechanism by which mechanotransduction modulating CTGF production in retinal Müller cells involves the YAP/TAZ complex, this study tests the influence of substrate stiffness on the time dependence of CTGF protein expression, as well as subcellular localization of YAP/TAZ using a conditionally-immortalized mouse retinal Müller cell line plated on laminin-coated, polyacrylamide substrates of varying elastic modulus. Changes were assayed using immunohistochemistry and ELISA (Enzyme-Linked ImmunoSorbent Assay). In retinal Müller cells, the relationship between elastic modulus and the pattern of CTGF protein expression was bimodal, with CTGF levels rising more rapidly for cells on hard substrates and more slowly for cells grown on soft substrates. In addition, nuclear localization of YAP/TAZ corresponded directly to the maximum CTGF expression.

Physical defects in basement membrane-mimicking collagen-IV matrices trigger cellular EMT and invasion.

In fibrosis and cancer, degradation of basement membrane (BM) and cell invasion are considered as key outcomes of a cellular transformation called epithelial-mesenchymal transition (EMT). Here, we pose a converse question - can preexisting physical defects in the BM matrix cause EMT in normal epithelial cells? On a BM-mimicking matrix of collagen-IV-coated polyacrylamide (PA) gel, we have discovered a reverse phenomenon in which preexisting defects trigger EMT in normal epithelial cells. Through spatiotemporal measurements and simulations in silico, we demonstrate that the EMT precedes cellular mechanoactivation on defective matrices, but they occur concurrently on stiff matrices. The defect-dependent EMT caused cell invasion though a stroma-mimicking collagen-I layer, which could be disabled through MMP9 inhibition. Our findings reveal that the known BM degradation caused by cellular EMT and invasion is not a one-way process. Instead, normal epithelial cells can exploit physical defects in the BM matrix to undergo disease-like cellular transformations.

PHARMACOKINETIC STUDY OF ORAL ε-AMINOCAPROIC ACID IN THE NORTHERN ELEPHANT SEAL (MIROUNGA ANGUSTIROSTRIS).

ε-Aminocaproic acid (EACA) is a lysine analogue antifibrinolytic drug used to treat bleeding disorders in humans and domestic animals. Its use in zoological medicine is rare, and dosage is anecdotal. One possible application of EACA is to treat bleeding associated with prepatent Otostrongylus arteritis in Northern elephant seals ( Mirounga angustirostris ) presenting to wildlife rehabilitation centers. This study used an in vitro model of hyperfibrinolysis and a thromboelastograph-based assay to estimate the therapeutic plasma concentration of EACA in elephant seals (85 µg/ml, 95% confidence interval = 73.8-96.8 µg/ml). A concurrent pharmacokinetic study of orally administered, single-dose EACA found that doses of 75 and 100 mg/kg achieved therapeutic plasma concentrations (>85 µg/ml), but the drug was rapidly eliminated and remained in the therapeutic range for only 0.4 and 1.5 hr, respectively. Models of repeated oral dosing at 100 mg/kg every 6 hr predict that therapeutic plasma concentration will be maintained for 31.7% (7.6 hr) of a 24-hr period. More frequent dosing would be required to maintain continuous therapeutic concentrations but would be impractical in a wildlife rehabilitation setting. Further pharmacodynamic studies to evaluate the duration of action of EACA in elephant seals and a prospective, placebo-controlled study are needed to determine if EACA is effective in decreasing bleeding associated with prepatent Otostrongylus arteritis and other bleeding disorders in this species.

The influence of substrate elastic modulus on retinal pigment epithelial cell phagocytosis.

To better understand if a complex process such as phagocytosis is influenced by substrate stiffness, we investigated the influence of substrate elastic modulus on phagocytosis in the retinal pigment epithelial (RPE) cell line ARPE-19. RPE cells lie on Bruch's membrane, directly under the retina, and phagocytose the shed photoreceptor outer segments. Bruch's membrane is known to increase in stiffness by an order of magnitude with age and thus, this study has potential relevance in explaining retinal changes in age-related macular degeneration. ARPE-19 cells were plated on laminin-coated polyacrylamide substrates of varying elastic modulus. After 14 days in culture, a solution of latex fluorescent beads suspended in PBS was placed in each well. After an incubation time of 4h, flow cytometry was performed to determine the number of cells that phagocytosed a bead. The number of ARPE-19 cells that phagocytosed a bead decreased continuously as a function of increasing substrate elastic modulus (p=0.0135), and this was found to be a linear relationship (slope=-0.03305 ± 0.01104, R2=0.4726 per 10,000 cells). Our results suggest that RPE cells display decreased phagocytosis when grown on firmer substrates, and thus, RPE cells in older eyes, in which Bruch's membrane is stiffer, may demonstrate decreased phagocytosis. Impaired phagocytosis by RPE cells may contribute to impaired metabolism of photoreceptor outer segments and to development of macular degeneration. Material stiffness may be a critical parameter in the development of neural therapies, including retinal prosthetics and stem cell therapies.

Partner communication, discordant fertility goals, and contraceptive use in urban Kenya.

This study assesses the degree of partner communication and perceived partner concordance and their association with contraceptive use among 2,891 women and 1,362 men in urban Kenya. Twenty-three percent of men and 30% of women report never discussing FP with their partner. Approximately 70% of participants perceive their partner to have concordant fertility desires. Multivariate analyses revealed that both male (AOR = 7.7 [95% CI = 5.5-10.7]) and female (AOR = 2.8 [95% CI = 2.3-3.3]) participants were more likely to use contraception if they report discussing FP with their partner. Participants who perceive that their partner wants fewer children also were more likely to use contraception (AOR, females = 1.8 [95% CI = 1.2-2.8]), (AOR, males = 1.9 [95% CI = 1.2-3.1]). Discussion of FP is a key determinant of contraceptive use; in couples with discordant fertility goals, pro-natalist males do not always dictate contraceptive behavior in urban Kenya.

Timing and circumstances of first sex among female and male youth from select urban areas of Nigeria, Kenya, and Senegal.

PURPOSE: To examine the timing and circumstances of first sex among urban female and male youth in Kenya, Nigeria, and Senegal. METHODS: Recently collected data are used to examine youth sexual behaviors in Kenya, Nigeria, and Senegal. In each country, a large, representative sample of women (ages 15-49) and men (ages 15-59) was collected from multiple cities. Data from youth (ages 15-24) are used for the analyses of age at sexual initiation, whether first sex was premarital, and modern family planning use at first sex. Cox proportional hazard models and logistic regression analyses are performed to determine factors associated with these outcomes. RESULTS: Across all three countries, a greater percentage of male youth than female youth report initiating sex with a nonmarital partner. More educated youth are less likely to have initiated sex at each age. In Nigeria and Senegal, poor female youth report earlier first sex than wealthier female youth. In Kenya, richer female youth are more likely to have premarital first sex and to use contraception/condom at first sex than their poorer counterparts. Older age at first sex and youth who report that first sex was premarital are significantly more likely to use a method of contraception (including condom) at first sex. City-specific distinctions are found and discussed for each outcome. CONCLUSIONS: Programs seeking to reduce HIV and unintended pregnancy risk among urban youth need to undertake needs assessments to understand the local context that influences the timing and circumstances of first sex in each city/country-specific context.

Muller cell expression of genes implicated in proliferative vitreoretinopathy is influenced by substrate elastic modulus.

PURPOSE: Matrix stiffness is recognized increasingly as a significant factor in cell and tissue function. To understand better the mechanosensitivity of Müller cells and its association with vitreoretinal disorders, we examined morphology, propagation, and expression of genes in Müller cells that were cultured on substrates of varying elastic moduli. METHODS: A conditionally immortalized mouse Müller cell line was cultured on laminin-coated polyacrylamide substrates with calibrated Young's moduli. Glass was used as a control. Phase contrast, fluorescence, and atomic force microscopy were used to study cell morphology and propagation. Expression of extracellular matrix (ECM) genes was analyzed using quantitative reverse-transcription PCR. RESULTS: The adherent area, stiffness, and propagation of Müller cells all are affected by matrix stiffness, but to different extents and with different ranges of sensitivity. Of 85 ECM genes tested 11 showed a continuous >4-fold increase or decrease in mRNA expression as a function of the substrate elastic modulus. The changes were statistically significant in four genes: connective tissue growth factor (Ctgf, P = 0.04), tenascin C (Tnc, P = 0.035), Collagen Iα1 (Col1a1, P = 0.0001), and Collagen IVα3 (Col4a3, P = 0.05), with all showing increased expression on softer substrates. CONCLUSIONS: There are significant changes in morphology, cytoskeletal integrity, and gene regulation in Müller cells as a function of the stiffness of the substrate. Changes in local tissue elastic modulus may have a role in vitreoretinal disorders. These findings also may have implications for strategies for improved integration of retinal prosthetics, and for stem cell therapies, particularly targeting the transcriptional regulators YAP and TAZ.