bluuebunny/crossref_metadata_2025 huggingface dataset:
Automated Sectioning, Staining, and Imaging of Histologically Preserved Tissues
Authors: Monteith Corey , Parker Kristy , Springer Joey , Farahani Navid , Rhodes Christopher
Published: April 2020

Objective

Tissue microstructure contains a wealth of information with extraordinary value for basic research and therapy discovery. But because histology is manual and tedious, tissue protocols are not set up for highly automated, structured, data‐driven methods that prevail in other areas of life sciences research.

Methods

We adapt block‐face lamination array tomography (B‐FLAT) for automated, high‐throughput histological sectioning. We can section, stain, mount, image, and microdissect paraffin‐embedded tissue sections on tape, allowing for much higher speed, accuracy, and consistency in section capture, as well as automated imaging and image registration.

Results

We successfully captured, deparaffinized, stained, and imaged serial histological sections on tape. Separately, we validated numerous histological and immunohistochemical (IHC) stains relevant to renal pathology and established their equivalence to conventional glass slide methods. We generated a series of at least 30 hematoxylin and eosin (H&E)‐stained mouse kidney sections, digitally imaged and registered at 20X magnification, and reconstructed entire renal glomeruli with nuclear resolution adequate for algorithmic cell counting.

Conclusions

Our automation approach recapitulates the performance of conventional histology practices, but allows for much higher speed, accuracy, and consistency in section capture. This enables future development to dramatically improve section quality, throughput, and sample biases.

Support or Funding Information

Funding provided by NIH grant 1R43DK120281‐01
Dietary Potassium Attenuates the Effects of Dietary Sodium on Vascular Function in Salt‐Resistant Adults
Authors: Smiljanec Katarina , Mbakwe Alexis , Gonzalez Macarena Ramos , Lennon Shannon L.
Published: April 2020

Cardiovascular disease remains a major public health problem in the U.S. and is the result of lifestyle factors, such as nutrition, that play an important role. Excess sodium consumption can increase blood pressure (BP) while consumption of potassium has been shown to have BP lowering properties. While the effect of these two nutrients on BP is fairly well understood, their impact on vascular function, particularly any interactive effects, has received less attention. Endothelial dysfunction, characterized by impaired dilation, is an important non‐traditional risk factor for atherosclerosis. Therefore, the purpose of this study was to determine if dietary potassium can offset the deleterious effect of high sodium on endothelial function as assessed by brachial artery flow‐mediated dilation (FMD). Thirty‐three adults with salt‐resistant BP (16M,17F; 27±1yr) completed 7 days each of the following diets: 65 mmol potassium/50 mmol sodium (MK/LS); 65 mmol potassium/300 mmol sodium (MK/HS); and 120 mmol potassium/300 mmol sodium (HK/HS) in random order. On the last day of each diet, 24‐hour ambulatory BP, 24‐hour urine collection and a blood draw was performed. FMD was assessed in response to reactive hyperemia. 24‐hr BP did not differ between diets confirming that subjects were salt‐resistant. Sodium excretion increased on both HS diets compared to LS/MK (p<0.05) and potassium excretion was increased on the HK diet compared to MK/LS and MK/HS (p<0.05) confirming diet compliance. Aldosterone and plasma renin activity were elevated on MK/LS compared to both HS diets (p<0.05). FMD was impaired on MK/HS (5.4±0.5%) compared to both MK/LS (7.0±0.6%; p<0.05) and HK/HS (6.5±0.5%; p<0.05) while there was no difference between the MK/LS and HK/HS diets (p>0.05). These data suggest that potassium provides vascular protection against the deleterious effects of high sodium on conduit artery function.

Support or Funding Information

This work was supported by NIH NIGMS grant # P20GM113125.
Semaphorin‐3F, a Potential Biomarker for Lymphatic Anomalies and Bone Cell Function
Authors: Solorzano Ernesto , Kelly Michael , Safadi Fayez
Published: April 2020

Lymphatic anomalies (LA) are rare conditions characterized by abnormal vascularization of lymphatic vessels (lymphangiogenesis). Gorham‐Stout Disease (GSD) is an aggressive type of LA which invades cortical bone and makes it “disappear”. There is little evidence elucidating the etiology of GSD and its osteolytic tendencies due to the rarity of the disease. In addition, there are no biomarkers capable of predicting GSD progression or treatment outcome. Normal bone does not have lymphatic vessels. It is believed that in GSD lymphatic endothelial cells (LEC) undergo genetic changes early in development that facilitate their migration and vessel formation in bone. LECs and their secretome content (lymph) are likely both crucial to lymphatic migration into bone and the cortical destruction seen in GSD. In this study, we focused on the LEC secretome with the goal of identifying proteins capable of modifying LEC and bone cell interactions. We initially assessed the effects of normal LEC conditioned media (LEC‐CM) on bone cell differentiation and function in vitro, and found that LEC‐CM stimulates osteoclast differentiation. Next, we subjected LEC‐CM to mass spectrometry and over 500 proteins were identified; we focused specifically on Semaphorin‐3F (SEMA3F) as SEMA3F effectively inhibits lymphangiogenesis in part, by competing with vascular endothelial growth factor‐C (VEGF‐C), for binding to the neuropilin‐2 receptor. We measured mRNA expression of both SEMA3F and VEGF‐C in untreated and Rapamycin treated LECs, a clinically effective therapy for patients with lymphatic anomalies. SEMA3F mRNA expression was upregulated while VEGF‐C was downregulated in Rapamycin treated versus untreated LEC cultures. We next assessed the effects of exogenous SEMA3F on LEC viability and on osteoblast differentiation and function. We found that SEMA3F reduced LEC viability in a dose‐dependent fashion. In addition, SEMA3F inhibited osteoblast differentiation as measured by alkaline‐phosphatase activity and matrix mineralization independent of its effects on osteoblast viability or proliferation. Experiments are currently underway to determine the expression/localization of SEMA3F, VEGF‐C and neuropilin‐2 in lymphatic anomalies (LA) tissues obtained from pediatric patients at our facility. We expect that SEMA3F expression will be downregulated while VEGF‐C will be upregulated in LA compared to normal lymphatics. In summary, our data suggest that SEMA3F might be a possible biomarker for patient with LA conditions or contributes to their pathogenesis.

Support or Funding Information

Lymphatic Malformation Institute, USA | Rebecca D. Considine Institute of Research, Akron Children’s Hospital, Akron, OH, USA| Integrated Pharmaceutical Medicine Program, NEOMED, Rootstown, OH, USA
Muscle sympathetic nerve activity in altitude acclimatized lowlanders during leg cycling exercise
Authors: Moralez Gilbert , Hansen Alexander B. , Amin Sachin B. , Hofstatter Florian , Tymko Michael M. , Ainslie Philip N. , Hearon Christopher M. , Lawley Justin S.
Published: April 2020

Acute normobaric hypoxia augments muscle sympathetic nerve activity (MSNA) during dynamic cycling exercise; however, the effects of chronic hypobaric hypoxia exposure on this response remain unknown. The purpose of the present study was to determine MSNA reactivity to moderate intensity cycling exercise in lowlanders at sea‐level (344m; Kelowna, BC, Canada) and after 14–21 days of high‐altitude acclimatization (~4300m; Cerro de Pasco, Peru). MSNA (radial microneurography), mean arterial pressure (brachial artery catheter), and heart rate (electrocardiogram) were measured at rest and during cycling exercise (60% of peak workload) in six healthy male participants. Chronic exposure to hypobaric hypoxia increased resting MSNA burst frequency from 14.0 ± 3.2 burst/min at sea‐level, to 38.2 ± 4.5 burst/min (P<0.001). The increase in MSNA during semi recumbent cycling exercise during chronic hypobaric hypoxia exposure was significantly lower compared to sea‐level (Δ MSNA: sea‐level: 106 ± 46% vs. high‐altitude: 43 ± 15% vs; P<0.001). To our knowledge, this is the first investigation to compare MSNA between normobaric normoxia and chronic hypobaric hypoxia during leg cycling exercise. These data indicate that chronic hypobaric hypoxic exposure blunts the relative increase in MSNA during moderate‐intensity dynamic cycling exercise, potentially due to a sympathetic “ceiling” being reached.

Support or Funding Information

Wilderness Medical Society research in‐training grant (GM)
Increased Interest in Anatomical Donation as a Result of Positive Media Coverage
Authors: Walker Cam , Ostervoss Tamara L.
Published: April 2020

A 99‐year old female anatomical donor at Oregon Health & Science University (OHSU) in 2018 had an unusual anomaly: Situs inversus (SI) with levocardia [left‐facing heart]. Her SI and levocardia went mostly unnoted throughout her long life, despite several surgical interventions and five successful pregnancies. After consultation with Ms. Ostervoss (director of OHSU’s body donation program), and the donor family, a poster presentation highlighting this donor was prepared and shown at last year’s American Association of Anatomy meeting at Experimental Biology in Orlando, FL, in April 2019 (Walker and Hankin 2019). Additionally, a press release to coincide with the poster presentation was prepared at OHSU for local media consumption. The donor’s story was distributed by print, radio, and television, worldwide.

The benefit to the OHSU BDP of publicizing this unusual case of SI can be best seen in Figure 1, which documents the number of consent forms registered with the BDP between 2014–2019. Between 2014 and 2018, annual preregistration vacillated between 350 and 400 individuals. However, after the widespread publication of the case of SI with levocardia in 2019, the number of preregistered donors surged to approximately 460 individuals, with the majority of additional registrations taking place after publication of the findings in widely viewed media sources. This is particularly apparent in Figure 2, where preregistration numbers are compared month to month between years 2017, 2018, and 2019. For example, in June, 21 persons (2017), and 29 persons (2018) preregistered to be anatomical donors. In June 2019, 48 individuals. The disparity continued in July, when 25 individuals preregistered in 2017, 34 in 2018, and 50 in 2019. The OHSU BDP undertakes other forms of ongoing outreach as well, though none were performed shortly before or after the SI press release in 2019.

Health professionals have long known the benefits of anatomical donation. Transparency in body donation is important as well. In this case, nascent medical students saw unique, but not pathological, anatomical variation that will stick in their memories. The family of the donor, after being contacted by Ms. Ostervoss, received closure and a great sense of satisfaction that their loved one contributed to the education of medical students and the public as a whole. Living individuals with SI may also benefit from a future study about the causes of SI.
Exploring the Kinetics and Cleavage Sites of Cysteine Cathepsins for Vaccine Design
Authors: Matson Shayna , Rivera Kristian , Becker Tynan , Kuhn Thomas
Published: April 2020

Cysteine proteases are key players in antigen processing, with roles in both pathogen digestion and major histocompatibility complex class II (MHCII) formation. A selection of the cleaved peptide chains are presented to CD4+ T cells to initiate an adaptive immune response. Those peptides that elicit the strongest T cell activation are termed immunodominant.

In this project, the kinetics and cleavage preference of cathepsins S and L individually and S, B & H in combination were examined using hemagglutinin (HA) protein from the A/New Caledonia/20/99 H1N1 strain of influenza as a substrate. HA was digested in solution with cathepsins at specific time points both at pH 5.0 and 6.4. Post‐digestion, we separated the products by mass and characterized the peptide fragments using mass spectroscopy.

We found that specific core peptides remained intact for up to 16 hours and correspond to a sub‐set of the immunodominant or subdominant HA peptides regardless of how many enzymes were applied. The kinetics of the digestion of cathepsins S, B & H in combination differed by pH with the higher pH showing slower kinetics. Both the individual cathepsins S and L displayed kinetics more similar that of pH 6.4 than 5.0 when the mixture of 3 enzymes were applied. This provides confirmation that the peptide repertoire can be impacted by regulation of the activity of individual cathepsins in addition to the pH of the endosomes where processing occurs.

Information on the cleavage sites and kinetics of cathepsins increases in‐silico predictive capacity when designing vaccines against novel pathogens.

Support or Funding Information

Research funded by NIH grant R01AI118888‐01A1 and NIH grants UL1GM118991, TL4GM118992, or RL5GM118990.
Isolation stress during adolescence increases conditioned place preference to cocaine in female rats
Authors: Rivero Carlos J. , Freire Jaime A. , Santiago Ivan , Delgado Cristian , Segarra Annabell C.
Published: April 2020

Adolescence is a developmental period characterized by neuronal growth. Environmental factors such as maternal neglect, social stress and drugs of abuse have a direct impact on future behavioral patterns. Because rats are highly social animals, we decided to investigate if isolation stress during adolescence increases conditioned place preference (CPP) to drugs of abuse. Since lower D2 receptor expression in the mesocorticolimbic system is known to increase cocaine conditioning in adult animals, and because adolescents and adults differ in dopaminergic connectivity and D2 receptor populations, we investigated if D2 receptors are altered during these conditions. Female rats were weaned at postnatal day 23 and housed singly or in pairs. At day 34 rats were tested in an open field, at day 35 in an elevated plus maze and at day 36 they were tested for CPP to cocaine (15mg/kg). At post‐natal day 48, rats were euthanized, brains collected and stored at −80°C. Single‐housed rats showed greater conditioning to cocaine when compared to group‐housed rats. This suggest that housing rats singly increases the rewarding properties of cocaine. Single‐housed rats also show lower D2 receptor immunoreactivity in the Nucleus accumbens (NAc) and the Prefrontal cortex (PFC) when compared to group‐housed rats. This correlates with the greater conditioning observed in single housed rats. Isolation stress had the effect of lowering D2 receptor populations in the NAc and PFC thus augmenting the rewarding properties of cocaine in adolescent female rats. Chronic stress during adolescence thus can lower D2 receptors and could be a determining factor in substance abuse and relapse.

Support or Funding Information

Financial assistance was provided by the Office of International Science and Engineering (OISE) of NSF through the Partnerships for Research and Education (PIRE) Program (OISE‐#1545803).
A rare finding of suspected synovial chondromatosis
Authors: Beger Aaron W. , Dudzik Beatrix , Millard Jonathan A. , Ford Stephen , Thompson Brent J.
Published: April 2020

Introduction

Human gross anatomy laboratory dissections provide a rich learning environment, sometimes including rare anatomical variation or pathological findings. Here we describe a rare knee joint abnormality, which provided an opportunity to review pathological processes unique to this region and expand the learning experience beyond the standard curriculum.

Findings

During a routine lower limb dissection of an 88‐year‐old formalin‐fixed Caucasian male donor, first‐year medical students discovered an enlarged semimembranosus bursa in the superomedial corner of the right popliteal fossa. Opening of the bursa revealed nine irregular‐shaped osteochondral bodies bathed in synovial fluid, with an average length of 1.76 cm and an average width of 1.25 cm. The donor exhibited no obvious trauma, however bilateral knee joint dissections revealed advanced osteoarthritis. Glenohumeral and acetabulofemoral joint cavities and their surrounding bursae were explored bilaterally, with the only significant finding being bilateral glenohumeral osteoarthritis. Histological studies supplemented by a review of the literature suggests a diagnosis of secondary synovial chondromatosis.

Discussion

Synovial chondromatosis outlines a rare, benign process characterized by the metaplasia of synovial tissue into chondrocytes within joint cavities, tendinous sheaths, or bursae. It can be further classified into one of two categories: primary, where the proliferation of hyaline cartilage in synovial membranes is seemingly spontaneous, or secondary, in which osteocartilaginous bodies are generated as a result of trauma or inflammatory joint pathologies. The condition most often affects males, has a monoarticular presentation, with a proclivity for the knee joint, followed by the shoulder and hip. Cases in the ankle, elbow, wrist, and temporomandibular joints have also been described. Although it is classified as benign and non‐metastatic, pain, swelling, and functional deficits have been reported. Treatment most often consists of surgical excavation of loose bodies with open arthrotomy and synovectomy.

Conclusion

Anomalies discovered through anatomical dissection provide unique opportunities to research pathological processes that may not otherwise be covered in standard anatomical curricula.
The Microbiomes of Sea Turtle Nests in Venice Beach, Florida
Authors: Mota Mario Jorge , Heidel Greg , Mahounou Agossou , Hoover Shelby
Published: April 2020

Sea turtles nest throughout the world’s beaches and the fate of their clutches depends on many factors including the biotic and abiotic properties of the sand.

It is well documented how a multitude of sand properties can impact the nest microenvironment and consequentially have a positive or negative effect on incubation. The present study looks at the microbiotic diversity in the sand of four beach areas of varying human traffic on Venice Beach, Florida. Our goal was to document bacterial species presence, relate them to human usage and extrapolate the potential role these bacteria species have on their respective sea turtle nest nest microbiomes.

This study was conducted during the sea turtle summer nesting season and samples were collected at four locations in Venice Beach every week from July until October. The locations were a pubic beach, a private beach, a dog beach and an isolated beach. This study design documents changes within the same beach and nesting site. Samples were also collected along the dune and high tide line and at three different depths in the same location to document microbiotic differences across and at different depths of the beach planform, respectively. Sargassum and other macroalgae were collected at random locations and its bacterial fauna identified to see if they could serve as possible bacterial sources for beach sand.

Sand samples were incubated in trypic soy broth and then plated in agar that selected for gram positive and negative bacteria. After individual bacterial colonies were isolated, DNA was extracted using standard procedures. The 16s ribosome was target with a primer, DNA was amplified using a PCR, and the DNA was sent for Sanger sequencing.

Bacteria species were identified by comparing their DNA sequences against several online databases such as BLAST. Data collected from several biochemical tests (oxidase test, glucose/fructose/lactose fermentation, anaerobic test, MacConkey and Mannitol Salt media, etc) done prior to DNA isolation were used to confirm the bacteria species identified by the database.

Our results show that there is diversity in bacterial fauna in the sand samples from the different sea turtle nesting areas of Venice Beach. Although some bacteria species (i.e. Pseudomonas), were found more prevalent in sand from hatched than unhatched clutches we cannot, at this time, state they played a significant role in the fate of the clutch. However, species diversity could not be explained in relation to sand properties and we believe it to be correlated to the level of anthropogenic impact. Further research is needed to test the direct influence these bacterial species have on the incubation of sea turtle eggs.

Support or Funding Information

Study was funded by National University’s Undergraduate Research Opportunity Program
Association between a non‐invasive assessment of frailty and vascular dysfunction in old mice
Authors: Cole Jazmin A. , Henson Grant D. , Walker Ashley E.
Published: April 2020

Advancing age is characterized by not only an increased risk for cardiovascular diseases (CVDs), but also a decline in functional reserve and impaired adaptive capacity across multiple physiologic systems, also known as frailty. Impaired vascular function is a known contributor to CVDs and potentially has a role in increased frailty. In patients with overt disease, measures of frailty are related to vascular endothelial cell dysfunction. However, the relation between vascular endothelial function and frailty in a non‐disease population is unknown. It is also unknown if dysfunction of a particular vascular bed is more closely related to frailty.

To examine the relation between vascular function and frailty in the context of similar genetics and environment, we studied wildtype C57BL/6 mice. In young (9 mo, n=7) and old (23–30 mo, n=27) male and female mice, we assessed endothelial‐dependent dilation (EDD) ex vivo in isolated, pressurized mesentery arteries and middle cerebral arteries (MCAs) by measuring the dilation to increasing doses of acetylcholine (ACh, 10−9 to 10−4 M) after pre‐constriction (by phenylephrine, 2 μM). Endothelium‐independent dilation (EID) was assessed by the dose response to sodium nitroprusside (SNP, 10−10 to 10−4 M) after pre‐constriction.

Mouse frailty was assessed using a previously established non‐invasive 31‐item frailty index based on clinical signs of deterioration. Clinical assessments included evaluation of the integumentary, musculoskeletal, vestibulocochlear/auditory, ocular, nasal, digestive, urogenital, and respiratory systems as well as body weight, body surface temperature, and signs of discomfort. The severity of each deficit was assessed by two independent observers and assigned a value between 0–1, with a higher score indicating more severe frailty.

Frailty index was ~6 fold greater in old compared with young mice (p<0.001). Maximal EDD was 16.1% lower in mesentery arteries (p=0.04) and 11.3% lower in MCAs (p=0.12) for old compared with young mice. Among the old mice, frailty index was correlated with mesentery artery maximal EDD (r=−0.53 p=0.002), and remained significant after controlling for age (partial correlation: r=−0.38, p=0.03). In contrast, there was no correlation between frailty index and MCA maximal EDD among old mice (r=0.16, p=0.21). Among old mice, there was no correlation between frailty index and maximal EID in mesentery arteries or MCAs (p>0.05).

This study demonstrates that frailty, independent of age, is associated with mesentery artery endothelial dysfunction in wildtype, non‐diseased mice. However, it remains unknown whether the mesentery artery dysfunction is a cause or consequence of the greater frailty. Interestingly, we found a lack of association between frailty and cerebral artery endothelial function. Future research is needed to determine the mechanisms linking frailty and mesentery artery dysfunction.

Support or Funding Information

Funded by NIH R56AG064016
The Impact of Zebrafish Research on Student Development at a Small Liberal Arts University
Authors: Lee Hyuna , Graham Lauren K. , Cline Noah M. , Darwazeh Amelia , Shapiro Atley , Stieglitz Grace R. , Sun Stephanie , Ramirez Danny A.
Published: April 2020

Executing high‐level research at a small, liberal arts college in southern California is seen as a daunting task due to limited resources and funds. Unlike larger institutions that have teams of paid lab managers and personnel, Dr. Hyuna Lee’s zebrafish lab at Biola University is run solely by undergraduate students. The team performs zebrafish husbandry and conducts in‐depth experiments throughout the academic year, attaining hands‐on experience that cultivates their ability to utilize creativity and critical thinking in a laboratory setting. The zebrafish lab also collaborates with other departments within the School of Science Technology and Health at Biola University. For instance, students work with the Chemistry, Physics, and Engineering (CPE) department and the Biology department’s microbiology and parasitology laboratory courses. The students in the CPE department previously utilized their state‐of‐the‐art fabrication lab to design and construct customized research devices‐‐such as an optokinetic drum, multiple imaging devices, and a brine shrimp hatchery. The Biology department’s microbiology and parasitology laboratories investigated water samples from the aquatic housing system to identify potential threats to the health of the zebrafish population. To better understand how participation in the lab impacts individual development, the team generated a pilot survey to collect data on student perception and experience with research at Biola University. The data demonstrates a strong correlation between participation in directed research and positive experience with research and course labs. The study also suggests that directed research students have more personal and professional development in the sciences than students who have not participated in research. Working in an independent, student‐run facility with inter‐departmental collaboration seems to be an efficacious method to enhance student satisfaction, research appreciation, and department success within the undergraduate population.

Support or Funding Information

None.
Cellular Encapsulation and Immunostaining Using a Photo Polymerized Hydrogel
Authors: Din Shabana , White Laura , Saito Kan , Desmond Lisa , MilliporeSigma Jeffrey Turner
Published: April 2020

Rationale

Immunocytochemistry is a valuable technique used to visualize protein expression, localization, and cellular structural changes by microscopic imaging. However, immunofluorescence of suspension cells, such as primary immune cells or rare circulating tumor cells, can be technically challenging requiring either expensive equipment and or mechanical manipulation. Furthermore, low amount of cellular sample can prevent proper assessment of experimental results and reduce throughput.

Objective

Here we introduce a photo polymerized hydrogel, Cell Capture, which uniformly encapsulates non‐adherent cells and permits immuno labeling and visualization by fluorescence imaging.

Methods and Results

Cell Capture reagent was formulated and optimized for maximal permeability to antibodies while ensuring cellular retention within the hydrogel. Preparation of samples for immunoassaying in microtiter plates led to decreased workflow times, compared to antibody staining in bulk processing due to the removal of centrifugation steps. Immortalized T lymphocyte Jurkat cells were plated in varying cell numbers, ranging from 4×105 to 5×103, followed by staining with CD3 antibody. Cell Capture demonstrated minimal cell loss with phenotypic membrane staining of CD3 and additional T cell targets, including multiplex tri color detection by microscopy. Cell Capture was further compared to traditional method of non‐adherent cell immobilization by extra cellular matrix coating of surfaces. Superior cellular retention properties were observed when using Cell Capture, demonstrating the advantages of this technology. Image cytometry is a common method to image suspension cells but requires a significant financial investment. To demonstrate Cell Capture as an alternative to image cytometry, cells were fixed into Cell Capture hydrogel following flow cytometry evaluation. Imaging by fluorescence microscopy, showed clear and distinct staining of measured immune targets in Jurkat cells, verifying Cell Capture as a valid alternative to image cytometry.

Conclusion

Cell Capture reagent allows for convenient immune labeling and detection of nonadherent cells in a high throughput fashion, demonstrating the utility and flexibility over traditional suspension cell immunocytochemistry methods.
The Effect of Inspired Hypoxia and Hyperoxia on Respiratory Sinus Arrhythmia Reactivity
Authors: Lazar Brendan , Nguyen Stephanie , Betton Matthew , Olar Evan , Day Trevor A.
Published: April 2020

Respiratory sinus arrhythmia (RSA) is the normal fluctuation in heart rate (HR) in phase with the respiratory cycle, whereby HR increases during inspiration and decreases during expiration. The underlying mechanisms and potential utility remain elusive. RSA magnitude is thought to be affected by autonomic balance, whereby a dominant parasympathetic nervous system (NS) at rest facilitates RSA, and sympathetic NS activation attenuates its magnitude. Carotid body stimulation via acute hypoxia elicits sympathetic activation, whereas hyperoxia facilitates sympathetic withdrawal. In addition, RSA has been hypothesized to facilitate improvements in ventilation/perfusion (V/Q) matching, improving oxygenation when hypoxic. We hypothesized that (a) acute hypoxia and hyperoxia would increase and decrease the magnitude of RSA reactivity, respectively and (b) when hypoxic, those with larger RSA magnitude during deep breathing would have larger improvements in oxygen saturation (SpO2). Healthy participants (n=13) were instrumented with a pneumotachometer and coached to breathe at three percentages (30, 40 and 50%) of their forced vital capacity (FVC) using visual cues while inspiring room air (21% FIO2), hypoxia (13.5% FIO2), hyperoxia (100% FIO2). RSA was quantified via the peak‐valley approach, and RSA reactivity (RSAR) was quantified via linear regression to calculate the slope of changes in RSA magnitude across all three FVC levels. RSA magnitude increased linearly with increases in inspired tidal volume in all three gases (R2>0.98). No significant differences were found in RSAR slopes between room air (0.32±0.2 ΔBPM/%FVC), hypoxia (0.33±0.26 ΔBPM/%FVC), and hyperoxia (0.34±0.26 ΔBPM/%FVC; P=0.97). There was also no correlation between RSA magnitude during deep breathing (50%FVC) and the associated improvements in SpO2 in hypoxia (r=−0.11, P=0.72). Given that RSAR is not affected by oxygen availability, our data provides evidence that hypoxic stimulation of the carotid bodies elicits activation of both arms of the autonomic nervous system, maintaining autonomic balance and normal heart rate variability. In addition, RSA magnitude does not appear to bestow advantages on V/Q matching in the context of acute hypoxia.

Support or Funding Information

MRU Faculty of Science and Technology
The interaction between p53 and Mdm2 is independent of MEG3–p53 association
Authors: Bauer Nicholas C. , Yang Anli , Wang Xin , Zhou Yunli , Klibanski Anne , Soberman Roy J.
Published: April 2020

The ability of the long noncoding RNA MEG3 to suppress cell proliferation led to its recognition as a tumor suppressor. MEG3 has previously been shown to bind to p53 in vitro, which led us to hypothesize that MEG3 functions by disrupting the interaction of p53 and its E3 ubiquitin ligase Mdm2. To test this hypothesis in vivo, we built a cross‐nearest neighbor/Monte Carlo analytical method based on two color direct stochastic optical reconstruction microscopy (dSTORM), a single‐molecule localization microscopy (SMLM) technique. Our data support the interaction of MEG3 and p53. Surprisingly, this association had no effect on the binding of p53 and Mdm2, distinct from the most commonly proposed model for the mechanism of MEG3 action. Additionally, our mathematical approach to analyzing SMLM data has general applicability to assessing molecular interactions in a native cellular context.

Support or Funding Information

RJS: NIH R01CA193520, NIH R01DK062472, NIH S10RR027931; NCB: NIH T32DK007540; AK and YZ: NIH R01CA193520; and The Jarislowsky Foundation
Endothelial Cells Express Activin A which may serve as an antagonist to TGFβ binding to Activin‐Like Kinase 1 Receptor
Authors: Hawker James , Woods Griffin , Morrissey Nancy , Patel Mintoo , Duffy Iain
Published: April 2020

Transforming growth factor (TGFβ) family and their receptors have been shown to be involved in angiogenesis as well as tumor cells. Members of the family that have been reported to act on endothelial cells are TGFβ, Bone Morphogenetic protein 9 and 10 (BMP‐9, BMP‐10), and Activin A. They all can signal through the type I receptor Activin‐Like Kinase 1 (ALK‐1). We have screened endothelial cells vs tumor cells for ALK‐1, Endoglin, and potential ligands BMP‐10 and Activin A to determine which ligands and receptors may be active in signaling in angiogenesis. We show that endothelial cells express ALK‐1 and endoglin as well as BMP‐10 and Activin A by quantitative RT‐PCR. Transfecting Human Umbilical Vein Endothelial Cells (HUVEC) compared to human 293 cells and Hela cells with an ALK‐1 plasmid resulted in increased expression of ALK‐1 in all cell lines. ALK‐1 overexpression did not affect Smad 1 or Smad 5 expression but increased BMP‐10 and Activin A expression in HUVEC. This data suggests these factors may function as endogenous ligands to control endothelial function and angiogenesis. TGFβ can only bind ALK‐1 on endothelial cells at 37°C but not 23 or 4° C. But our results indicate that endogenous Activin A is bound to ALK‐1 and can be displaced by added TGFβ. This supports the finding above that HUVEC express Activin A. Thus Activin A may function as an endogenous regulator of TGFβ function in endothelial cells.

Support or Funding Information

Research support was from an SFSC Innovation Grant (MP), Portz National Honors Fellowships (GW and NM), and funds from Saint Leo University (ID).
Teaching coupled reactions in the lab class
Authors: Park Chad K.
Published: April 2020

We present some preliminary protocols for coupled reaction assays. Students are asked to move from direct activity assays using alkaline phosphatase (AP) and para‐nitrophenolphosphate into coupled reaction systems. We study phosphofructokinase (PFK) with a coupled reaction assay. To segue from AP to PFK, we spend one lab period trying to force AP to catalyze the reverse reaction and discuss energetics and Le Chatelier’s principle. The concept of reversibility of a reaction and the role of the enzyme as a catalyst for both the forward and reverse reaction is discussed. This lays the groundwork for using the popular pyruvate kinase, lactate dehydrogenase coupled reaction system for measuring the rate of ATP hydrolysis for many enzyme systems including that of our final project, PFK.
Serial interval and generation interval for respectively the imported and local infectors estimated using reported contact-tracing data of COVID-19 in China
Authors: Li Menghui , Liu Kai , Song Yukun , Wang Ming , Wu Jinshan
Published: April 2020

Backgrounds

The emerging virus, COVID-19, has caused a massive out-break worldwide. Based on the publicly available contact-tracing data, we identified 337 transmission chains from 10 provinces in China and estimated the serial interval (SI) and generation interval (GI) of COVID-19 in China.

Methods

Inspired by possibly different values of the time-varying reproduction number for the imported cases and the local cases in China, we divided all transmission events into three subsets: imported (the zeroth generation) infecting 1st-generation locals, 1st-generation locals infecting 2nd-generation locals, and others transmissions among 2+ generations. The corresponding SI (GI) is respec-tively denoted as , and . A Bayesian approach with doubly interval-censored likelihood is employed to fit the lognormal, gamma, and Weibull distribution function of the SI and GI using the identified 337 transmission chains.

Findings

It is found that the estimated , and , thus overall both SI and GI decrease when generation increases.
Antibody testing for COVID-19: A report from the National COVID Scientific Advisory Panel
Authors:   National COVID Testing Scientific Advisory Panel, Adams Emily R , Ainsworth Mark , Anand Rekha , Andersson Monique I , Auckland Kathryn , Baillie J Kenneth , Barnes Eleanor , Beer Sally , Bell John , Berry Tamsin , Bibi Sagida , Carroll Miles , Chinnakannan Senthil , Clutterbuck Elizabeth , Cornall Richard J , Crook Derrick W , De Silva Thushan , Dejnirattisai Wanwisa , Dingle Kate E , Dold Christina , Espinosa Alexis , Eyre David W , Farmer Helen , Fernandez Mendoza Maria , Georgiou Dominique , Hoosdally Sarah J , Hunter Alistair , Jeffrey Katie , Klenerman Paul , Knight Julian , Knowles Clarice , Kwok Andrew J , Leuschner Ullrich , Levin Robert , Liu Chang , Lopez-Camacho Cesar , Martinez Garrido Jose Carlos , Matthews Philippa C , McGivern Hannah , Mentzer Alexander J , Milton Jonathan , Mongkolsapaya Juthathip , Moore Shona C , Oliveira Marta S , Pereira Fiona , Perez Lopez Elena , Peto Timothy , Ploeg Rutger J , Pollard Andrew , Prince Tessa , Roberts David J , Rudkin Justine K , Sanchez Veronica , Screaton Gavin R , Semple Malcolm G , Skelly Donal T , Slon-Campos Jose , Smith Elliot Nathan , Sobrino Diaz Alberto Jose , Staves Julie , Stuart David , Supasa Piyada , Surik Tomas , Thraves Hannah , Tsang Pat , Turtle Lance , Walker A Sarah , Wang Beibei , Washington Charlotte , Watkins Nicholas , Whitehouse James
Published: April 2020

Background

The COVID-19 pandemic caused >1 million infections during January-March 2020. There is an urgent need for reliable antibody detection approaches to support diagnosis, vaccine development, safe release of individuals from quarantine, and population lock-down exit strategies. We set out to evaluate the performance of ELISA and lateral flow immunoassay (LFIA) devices.

Methods

We tested plasma for COVID (SARS-CoV-2) IgM and IgG antibodies by ELISA and using nine different LFIA devices. We used a panel of plasma samples from individuals who have had confirmed COVID infection based on a PCR result (n=40), and pre-pandemic negative control samples banked in the UK prior to December-2019 (n=142).

Results

ELISA detected IgM or IgG in 34/40 individuals with a confirmed history of COVID infection (sensitivity 85%, 95%CI 70-94%), vs. 0/50 pre-pandemic controls (specificity 100% [95%CI 93-100%]). IgG levels were detected in 31/31 COVID-positive individuals tested ≥10 days after symptom onset (sensitivity 100%, 95%CI 89-100%). IgG titres rose during the 3 weeks post symptom onset and began to fall by 8 weeks, but remained above the detection threshold. Point estimates for the sensitivity of LFIA devices ranged from 55-70% versus RT-PCR and 65-85% versus ELISA, with specificity 95-100% and 93-100% respectively. Within the limits of the study size, the performance of most LFIA devices was similar.

Conclusions

Currently available commercial LFIA devices do not perform sufficiently well for individual patient applications. However, ELISA can be calibrated to be specific for detecting and quantifying SARS-CoV-2 IgM and IgG and is highly sensitive for IgG from 10 days following first symptoms.
Variation in Aerosol Production Across Oxygen Delivery Devices in Spontaneously Breathing Human Subjects
Authors: Iwashyna Theodore J. , Boehman Andre , Capecelatro Jesse , Cohn Amy M. , Cooke James M. , Costa Deena Kelly , Eakin Richard M. , Prescott Hallie C. , Woolridge Margaret S.
Published: April 2020

We sought to assess whether HHFNC results in greater production of aerosolized particles than 6 liters per minute nasal cannula, using state-of-the-art techniques of aerosol measurement, in spontaneously breathing human volunteers in a simulated hospital room.

For each volunteer, we first measured background aerosol levels in the room immediately prior to testing. We then measured aerosol levels while the healthy volunteer laid in bed - - with the head of bed at 30 degrees - - wearing the following oxygen delivery devices: (a) 6L/min nasal canula (NC) with humidification; (b) non-re-breather mask (NRB) with 15L/min gas flow, non-humidified; (c) HHFNC with 30L/min gas flow; (d) HHFNC with 60L/min gas flow. Two scanning mobility particle sizing (SMPS) systems (TSI 3080/3030, TSI 3080/3750) were used to measure aerosols 10 to 500 nanometer (nm) in size for each of the oxygen delivery devices.

There was no variation in aerosol level within patients between room air, 6 L/min NC, 15 L/min NRB, 30 L/min HHFNC, and 60 L/min HHFNC, regardless of coughing.
Why lockdown? Simplified arithmetic tools for decision-makers, health professionals, journalists and the general public to explore containment options for the novel coronavirus
Authors: Killeen Gerry F. , Kiware Samson S
Published: April 2020

Half the world’s population is already under lock-down and the remainder will have to follow if the ongoing novel coronavirus 2019 (COVID-19) virus pandemic is to be contained. Faced with such brutally difficult decisions, it is essential that as many people as possible understand (1) why lock-down interventions represent the only realistic way for individual countries to contain their national-level epidemics before they turn into public health catastrophes, (2) why these need to be implemented so early, so aggressively and for such extended periods, and (3) why international co-operation to conditionally re-open trade and travel between countries that have successfully eliminated local transmission represents the only way to contain the pandemic at global level. Here we present simplified arithmetic models of COVID-19 transmission, control and elimination in user-friendly Shiny and Excel formats that allow non-specialists to explore, query, critique and understand the containment decisions facing their country and the world at large. Based on parameter values representative of the United Republic of Tanzania, which is still early enough in its epidemic cycle and response to avert a national catastrophe, national containment and elimination with less than 10 deaths is predicted for highly rigorous lock down within 5 weeks of the first confirmed cases and maintained for 15 weeks. However, elimination may only be sustained if case importation from outside the country is comprehensively contained by isolating for three weeks all incoming travellers, except those from countries certified as COVID-free in the future. Any substantive relaxation of these assumptions, specifically shortening the lock-down period, less rigorous lock-down or imperfect importation containment, may facilitate epidemic re-initiation, resulting in over half a million deaths unless rigorously contained a second time. Removing contact tracing and isolation has minimal impact on successful containment trajectories because high incidence of similar mild symptoms caused by other common pathogens attenuates detection success of COVID-19 testing. Nevertheless, contact tracing is recommended as an invaluable epidemiological surveillance platform for monitoring and characterizing the epidemic, and for understanding the influence of interventions on transmission dynamics.
Knowledge and practice of preventive measures against COVID-19 infection among pregnant women in a low-resource African setting
Authors: Nwafor Johnbosco Ifunanya , Aniukwu Joseph Kenechi , Anozie Bonaventure Okechukwu , Ikeotuonye Arinze Chidiebere
Published: April 2020

Background

Coronavirus disease pandemic has resulted in death of thousands of people across several countries. Several preventive measures have been recommended to halt the spread of the disease and its associated mortality. However, the level knowledge and practice of these preventive measures against COVID-19 infection among pregnant women, which constitute vulnerable groups, are yet to be evaluated.

Aim

To determine the knowledge and practice of preventive measures against COVID-19 infection among pregnant women in Abakaliki.

Materials and Methods

This was a self-administered questionnaire-based cross-sectional study conducted from February 1, 2020 to March 31, 2020 among 284 antenatal clinic attendees at Alex Ekwueme Federal University Teaching Hospital, Abakaliki, Ebonyi State. A pretested and validated questionnaire was used to collect the data. Data analysis was done using SPSS version 22.

Results

Of 284 participants, 60.9% (n=173) had adequate knowledge of the preventive measures against COVID-19 infection. However, the overall practice of these preventive measures among the participants were poor as 69.7% of the participants were not practicing the preventive measures against the coronavirus. The determinants of poor practice of the preventive measures among the participants were being in age group 31-40 years (AOR=2.04, 95%CI: 1.26 - 5.37, p=0.022), married (AOR=2.99, 95%CI: 1.40 - 6.33, p=0.035) grandmultiparous (AOR=3.11, 95%CI: 1.32 - 6.56, p=0.021), residing in rural area (AOR=2.08, 95%CI: 1.32 - 4.05, p=0.031), and having no formal education (AOR=6.73, 95%CI: 2.66 - 18.34, p=0.002).

Conclusion

The study showed that most of the participants had adequate knowledge of preventive measures against COVID-19 infection but the practice of these preventive measures were poor among the participants.
Community Memory of COVID-19 Infections Post Lockdown as a Surrogate for Incubation Time
Authors: Prakash Meher K.
Published: April 2020

If the knowledge of the incubation time is helpful in designing non-pharmaceutical interventions such as quarantine measures, can one use the number of cases arising after a lockdown to check (a) if our assumptions of the incubation time were correct and (b) if the quarantine measures were as successful as they could theoretically be. These are the two questions we raise by studying the number of new cases arising after lockdowns in a few European countries. The analysis which purely relies on the publicly available data of the numbers of new infections, rather than extensive contact tracing of individual patients, suggests a “memory” of the infections in the community with a median of 13.3 days. This distribution of the memory of infections which may even be considered as a surrogate of the incubation time in a perfect lockdown, suggests that even a perfect quarantine of 30 days is only 90% complete.
Estimating SARS-CoV-2 seroprevalence and epidemiological parameters with uncertainty from serological surveys
Authors: Larremore Daniel B. , Fosdick Bailey K. , Bubar Kate M. , Zhang Sam , Kissler Stephen M. , Metcalf C. Jessica E. , Buckee Caroline O. , Grad Yonatan H.
Published: April 2020

Establishing how many people have already been infected by SARS-CoV-2 is an urgent priority for controlling the COVID-19 pandemic. Patchy virological testing has hampered interpretation of confirmed case counts, and unknown rates of asymptomatic and mild infections make it challenging to develop evidence-based public health policies. Serological tests that identify past infection can be used to estimate cumulative incidence, but the relative accuracy and robustness of various sampling strategies has been unclear. Here, we used a flexible framework that integrates uncertainty from test characteristics, sample size, and heterogeneity in seroprevalence across tested subpopulations to compare estimates from sampling schemes. Using the same framework and making the assumption that serological positivity indicates immune protection, we propagated these estimates and uncertainty through dynamical models to assess the uncertainty in the epidemiological parameters needed to evaluate public health interventions. We examined the relative accuracy of convenience samples versus structured surveys to estimate population seroprevalence and found that sampling schemes informed by demographics and contact networks outperform uniform sampling. The framework can be adapted to optimize the design of serological surveys given particular test characteristics and capacity, population demography, sampling strategy, and modeling approach, and can be tailored to support decision-making around introducing or removing interventions.