bluuebunny/crossref_metadata_2025 huggingface dataset:
Coping with the COVID-19 crisis: an overview of service adaptation and challenges encountered by a rural Psychiatry of Later Life (POLL) team
Authors: Usman M. , Fahy S.
Published: July 2020

The COVID-19 pandemic has posed many challenges in maintaining standards of care and treatment for patients while managing the increased anxieties of patients, carers and the public in general. This paper highlights several clinical, administrative, medicolegal and IT implications of COVID-19 on the delivery of mental healthcare to an elderly vulnerable patient cohort due to recommended social distancing measures. Our Psychiatry of Later Life team has adapted to this by restricting face-to-face consultation, while continuing to provide telephone support. We have modified our documentation standard and have improved some aspects of our team working by facilitating flexible working arrangement and relevant training for staff as well as by embracing new technology. Notwithstanding the challenges therefore, this exceptional time has also opened avenues for new and innovative opportunities that can be further explored even when the current crisis eventually passes.
An approach to teaching psychiatry to medical students in the time of Covid-19
Authors: Guerandel A. , McCarthy N. , McCarthy J. , Mulligan D. , Lane A. , Malone K.
Published: July 2020

In this time of Covid-19, life in healthcare has changed immeasurably. It has rapidly been injected with an ‘all hands-on deck’ approach, to facilitate the necessary adaptations required to reduce the spread of the virus and deliver frontline clinical care. Inevitably aspects of these changes have disrupted the delivery of medical education, notably clinical placements have been cancelled and social distancing guidelines prohibit face-to-face teaching. The training of future doctors is an essential part of this effort. Indeed, the emergence of a global health threat has underlined its continued importance. For medical educators and students alike, we have been presented with a challenge. Concurrently, this presents us with an impetus and opportunity for innovation. For some time now, a transformation in medical education has been called for, with an increasing recognition of the need to prepare students for the changing landscape of healthcare systems. This has included a focus on the use of technology-enhanced and self-directed learning. As a team of educators and clinicians in psychiatry, working in the School of Medicine and Medical Sciences (SMMS) in University College Dublin (UCD), we will share how we have responded. We outline the adaptations made to our ‘Psychiatry’ module and consider the influence this may have on its future delivery. These changes were informed by direct student input.
The Plan of the Saxon Shore Fort at Richborough
Authors: Wilmott Tony , Smither Philip
Published: July 2020

Recent excavation and coring of the collapsed east wall of the Saxon Shore fort of Richborough has revealed the manner in which the wall collapsed. This led to a re-evaluation of the original siting of the wall, which must have lain to the west of where it is usually depicted. Reassessment of previous excavations, including the examination of original records from the J.P. Bushe-Fox excavations of the 1920s and 1930s leads to the conclusion that the so-called ‘unfinished’ or ‘abandoned’ east wall foundation was in fact the base of the built east wall, from which the collapse derives. A revised fort plan based upon this conclusion is suggested. Supplementary material is available online (https://doi.org/10.1017/S0068113X20000379), and includes additional backing tables and illustrations referenced in the text.
Clinical and socio-economic predictors of work participation in adult CHD patients
Authors: Sarno Lauren A. , Cortright Lindsay , Stanley Tiara , Tumin Dmitry , Li Jennifer S. , Sang Charlie J.
Published: July 2020

Background:

Adults with CHD have reduced work participation rates compared to adults without CHD. We aimed to quantify employment rate among adult CHD patients in a population-based registry and to describe factors and barriers associated with work participation.

Methods:

We retrospectively identified adults with employment information in the North Carolina Congenital Heart Defects Surveillance Network. Employment was defined as any paid work in a given year. Logistic regression was used to examine patients’ employment status during each year.

Results:

The registry included 1,208 adult CHD patients with a health care encounter between 2009 and 2013, of whom 1,078 had ≥1 year of data with known employment status. Overall, 401 patients (37%) were employed in their most recent registry year. On multivariable analysis, the odds of employment decreased with older age and were lower for Black as compared to White patients (odds ratio = 0.78; 95% confidence interval: 0.62, 0.98; p = 0.030), and single as compared to married patients (odds ratio = 0.50; 95% confidence interval: 0.39, 0.63; p < 0.001).

Conclusion:

In a registry where employment status was routinely captured, only 37% of adult CHD patients aged 18–64 years were employed, with older patients, Black patients, and single patients being less likely to be employed. Further work is needed to consider how enhancing cardiology follow-up for adults with CHD can integrate support for employment.
Cardiac workup and monitoring in hospitalised children with COVID- 19
Authors: Fremed Michael A. , Lytrivi Irene D. , Liberman Leonardo , Anderson Brett R. , Barry Oliver M. , Choudhury Tarif A. , Chrisomalis-Dring Sophia , Ferris Anne , Glickstein Julie S. , Krishnan Usha , Levasseur Stéphanie , Rosenzweig Erika B. , Shah Amee , Silver Eric S. , Suh Sanghee , Turner Mariel E. , Weller Rachel , Woo Joyce , Starc Thomas J.
Published: July 2020

Approximately, 1.7 million individuals in the United States have been infected with SARS-CoV-2, the virus responsible for the novel coronavirus disease-2019 (COVID-19). This has disproportionately impacted adults, but many children have been infected and hospitalised as well. To date, there is not much information published addressing the cardiac workup and monitoring of children with COVID-19. Here, we share the approach to the cardiac workup and monitoring utilised at a large congenital heart centre in New York City, the epicentre of the COVID-19 pandemic in the United States.
Measuring overweight and obesity in Chinese American children using US, international and ethnic-specific growth charts
Authors: Lau Jennifer D , Elbaar Laminasti , Chao Eda , Zhong Olivia , Yu Chihang Ray , Tse Raymond , Au Loretta
Published: July 2020

Objective:

The aim is to determine the disparity between the overweight and obesity prevalence of Chinese American school-aged children and adolescents as measured by the Centers for Disease Control and Prevention (CDC) growth reference and the prevalence as measured by international and ethnic-specific-growth references.

Design:

This retrospective, cross-sectional study measured overweight and obesity prevalence among a paediatric population using the CDC, International Obesity Task Force (IOTF), World Health Organization (WHO) and an ethnic Chinese growth curve.

Setting:

A community health centre in New York City, USA.

Participants:

Chinese American children aged 6–17 years in 2017 (N 9160).

Results:

The overweight prevalence was 24 % (CDC), 23 % (IOTF), 30 % (WHO) and 31 % (China). The obesity prevalence was 10 % (CDC), 5 % (IOTF), 10 % (WHO) and 10 % (China). When disaggregated by age and sex, the difference was the most prominent in girls; using the China reference compared with using the CDC reference almost doubles the overweight prevalence (school-aged: 31 v. 17 %, P < 0·001, adolescent: 27 v. 14 %, P < 0·001) and the obesity prevalence (school-aged: 11 v. 5 %, P < 0·001, adolescent: 7 v. 4 %, P < 0·001).

Conclusions:

Use of the CDC reference compared with the Chinese ethnic-specific reference results in lower overweight and obesity prevalence in Chinese American girls. Almost half of the girls who were overweight and half of the girls who were obese were not identified using the CDC reference. Using ethnic-specific references or ethnic-specific cut-points may help improve overweight identification for Chinese American children.
Validation of a new software eAT24 used to assess dietary intake in the adult Portuguese population
Authors: Goios Ana CL , Severo Milton , Lloyd Amanda J , Magalhães Vânia PL , Lopes Carla , Torres Duarte PM
Published: July 2020

Objective:

The aim of the current study was to evaluate the accuracy of the new software eAT24 used to assess dietary intake in the National Food, Nutrition and Physical Activity Survey (IAN-AF) against urinary biomarkers: N (nitrogen), K (potassium) and Na (sodium).

Design:

We conducted a cross-sectional study. Two non-consecutive 24-h dietary recalls (24-HDR) were applied, and a 24-h urine sample was collected. We examined differences between estimates from dietary and urine measures, Pearson correlation coefficients were calculated and the Bland–Altman plots were drawn. Multiple linear regression was used to evaluate the factors associated with the difference between estimates.

Setting:

Sub-sample from the Portuguese IAN-AF sampling frame.

Participants:

Ninety-five adults (men and women) aged 18–84 years.

Results:

The estimated intake calculated using the dietary recall data was lower than that estimated from urinary excretion for the three biomarkers studied (protein 94·3 v. 100·4 g/d, K 3212 v. 3416 mg/d and Na 3489 v. 4003 mg/d). Considering 2 d of recall, the deattenuated correlation coefficients were 0·33, 0·64 and 0·26 for protein, K and Na, respectively. For protein, differences between dietary and urinary estimates varied according to BMI (β = −1·96, P = 0·017). The energy intake and 24-h urine volume were significantly associated with the difference between estimates for protein (β = 0·03, P < 0·001 and β = −0·02, P = 0·002, respectively), K (β = 0·71, P < 0·001 and β = −0·42, P = 0·040, respectively) and Na (β = 1·55, P < 0·001 and β = −0·81, P = 0·011, respectively).

Conclusions:

The new software eAT24 performed well in estimating protein and K intakes, but lesser so in estimating Na intake, using two non-consecutive 24-HDR.
Ovarian Carcinomas: Insights into Origins Using Confocal Microscopy and Fluorescence in Situ Hybridization on Intact Paraffin Sections
Authors: Bailey GW , Dimlich RVW , Alexander KB , McCarthy JJ , Pretlow TP , Wolf N G , Abdul-Karim F W , Schork N J , Schwartz S
Published: July 2020

Although ovarian carcinomas are the most lethal gynecologic tumors, their origins remain unclear. Do they develop from malignant transformation of benign neoplasms through a multistep process of tumor progression, or do they arise de novo? The histologically benign and/or low malignant potential (LMP) components in heterogeneous ovarian carcinomas have been considered as evidence supporting the theory of tumor progression. These components are interpretted as the remnants of pre-existing neoplasms that underwent malignant transformation. In two other possible interpretations, however, such components may be clones which developed independently (de novo hypothesis) or they may represent malignant epithelium which underwent focal maturation (maturation hypothesis).

To evaluate genetic relationships of the histological components in heterogeneous ovarian carcinomas, 10 such neoplasms and 5 normal ovary controls were examined using fluorescence in situhybridization (FISH) on intact paraffin sections (6 μm).The retention of tissue architecture allowed direct correlation of detectable genetic aberrations with histology, and comparison of malignant components with adjacent histologically benign or LMP components.
Giant Human T-Lymphocytes (Jurkat Cells) Expressing HIV-1 Tat-72: LM and TEM.
Authors: Gilloteaux J , Koch J , Jamison J M , Authelet M , Sowick C , Summers JL
Published: July 2020

The human immunodeficiency virus (HIV-1) is dependent upon the synthesis of TAT for efficient transcriptional activation and replication. Following translocation to the nucleus TAT interacts with a cis-acting TAT-responsive element located between nucleotides +1 and +42 within the HIV-1 long terminal repeat. This region contains an RNA stem loop structure TAR which interacts with TAT and cellular proteins to increase transcription 1000 fold We have transfected human T-lymphocytes with an expression vector containing the HIV-1 LTR driving the synthesis of TAT-72. A stable cell line was generated by selection with G418. Cells were grown on round glass coverslips coated with a solution of fibronectin overnight (at 37 °C). In addition to light microscopy (LM) observations, cells on coverslips were fixed with a 3.5% glutaraldehyde buffered solution (0.1 M Na cacodylate) for 5 min at room temperature and gently centrifliged in a microfuge (400 rpm, 3min) to obtain a cellular pellet.
Quantitative Laser Scanning Confocal Autofluorescence Microscopy (Lscam) of Normal and Premalignant Colonic Tissues
Authors: Wang Hsing-Wen , Willis Joseph , Sivak Michael V , Izatt Joseph A
Published: July 2020

Laser-induced fluorescence (LIF) spectroscopy of tissues has been proposed as an adjunct to endoscopy for in vivo gastrointestinal (GI) diagnosis of premalignant lesions. In previous studies, under ultraviolet excitation (350-370 nm), sources of LIF signal differences in normal, and premalignant colonie tissues have been identified including 1) changes in gross tissue morphology, 2) increased hemoglobin absorption in adenomatous tissues, and 3) increased red fluorescence in dysplastic cells. However, many questions remain regarding the specific origins and biochemical correlates of GI tissue autofluorescence.

We report on a study of quantitative ultraviolet LIF spectral microscopy in human colonic tissues using a laser scanning confocal microscope with argon laser excitation 351-364 nm. Frozen sections (6μm) of normal colon (n=10) and tubular adenoma (n=8) were prepared from fresh surgical resection specimens using a previously published protocol. To identify histological components accurately, LSCAM slides were alcohol fixed and H&E histologically stained following confocal imaging.
Nanogold-Labeled Amyloid β Protein Targets 20s Proteasome
Authors: Hainfeld James F , Gregori Luisa , Simon Martha N , Goldgaber Dmitry
Published: July 2020

Proteasomes are multicatalytic structures found in cells that play a major role in the proteolytic degradation of proteins. Before entering the proteasome, proteins are marked like trees to be cut by attachment of ubiquitin. Neurodegenerative disorders of aging, including Alzheimer's, show abnormal accumulation of plaques and neurofibrillary tangles (NT). The latter are composed of paired helical filaments containing ubiquitinated tau protein. The fact that ubiquitinated tau is not degraded by the proteasomes may be a key to the formation of NT. Proteasomes, like other cellular pathways, are controlled by various mechanisms. It was found that amyloid β protein (Aβ) inhibits chymotrypsin-like proteasomal activity. Additionally, Aβ was found in elevated quantity in NTs in Alzheimer's diseased brain tissue. Understanding the mechanism of action of Aβ therefore becomes important.

In order to track the regulatory Aβ protein (Aβ1-40), a variant was synthesized with a terminal cysteine (Aβ1-40C40) that could be gold labeled.
Effects of Soil Removal and Flooding on Leaf Cell Structure of Quercus Falcata Var. Pagodaefolia and Qurecus Nigra
Authors: Ning Zhu Hua , Abdollahi Kamran K
Published: July 2020

Soil removal and flooding are serious problems for many urban trees. The main purpose of this paper is to outline some changes which occur in leaves of cherrybark oak (Quercus falcata var. pagodaefolia) and water oak (Quercus nigra) during soil removal and flooding treatments. There has not been any prior documentation of leaf tissue of trees under these conditions. Two-year-old saplings of cherrybark oaks and water oaks were randomly planted in twenty-four experimental plots. Trees were randomly subjected to a combination of soil removal and flooding treatments for one year. Leaf tissues were collected from trees in controlled and treated plots. Tissues were fixed overnight at 4° C in 4% glutaraldehyde buffered with 0.07M cacodylate at pH of 7.2 and post-fixed for one hour in 1% osmium tetroxide in the same buffer. After washing thoroughly in the buffer, tissues were dehydrated in a graded ethanol series and embedded in Eponate 12 resin. Thin sections of 75nm thick were cut with a MT-6000 ultratome and placed on 300 mesh copper grids. Sections were then positive stained with uranyl acetate followed by lead citrate. All examinations of the cell structure were made using a Zeiss EM 10C transmission electron microscope.
Localization of Nitric Oxide Synthase in the Salivary Gland Ducts of Feeding Female Dermacentor Variabilis (SAY) and the Effect of Endogenous Nitric Oxide on Isolated Salivary Gland Ducts
Authors: Lamoreaux William J , Fitzgerald Malinda E C , Needham Glen R , Frase Sharon , Bayakly Nabil , Coons Lewis B
Published: July 2020

The tick Dermacentor variabilis returns much of the water and ions from the bloodmeal back into the host as saliva produced in paired salivary glands. These glands are made up of acini and a system of ducts (Fig. 1). Bloodsucking alternates with salivation, but saliva is constantly produced by the acini and expelled into the ducts where it accumulates during bloodsucking. Ducts consist of a layer of epithelial cells, a cuticular coil, and an epicuticular lining of the lumen (Figs. 2, 3). Actin-like filaments run from the coil into the epithelial cells as shown by TEM and CLSM (Figs. 3, 4). These filaments are absent in cytochalasin D treated ducts.

Nitric oxide synthase (=NADPH diaphorase) is present in duct epithelial cells as shown by histochemistry (Fig. 5). We measured nitric oxide (NO) indirectly by the conversion of L-arginine to L-citrulline. NO production and production of saliva is dopamine dependent. NO activates guanylyl cyclase as measured by an RIA for cGMP. The lumen width in dopamine stimulated isolated ducts increased by 19.7% as measured optically. Ducts pre-treated with 10 μM L-NAME, an NOS specific inhibitor, showed no significant increase in width. The addition of 10 μM L-arginine to these ducts partially restored the increase in lumen diameter.
Detection of Concannavalin a Receptors in Trophozoites and Cysts of Acanthamoeba sp. (W4) Philippine Isolate
Authors: Argayosa AM , Natividad FF , Matias RR , Enriquez GL
Published: July 2020

Distribution of glucose and mannose moieties of Acanthamoeba sp. (W4) Philippine isolate was detected using fluorescem isothiocyanate (FITC)- labeled Concannavalin A (Con A) lectin. Green fluorescent patches around the plasma membrane of agglutinated trophozoites (Fig.1) were observed. Isolated Acanthamoeba cyst exhibited strong fluorescence on the cyst wall Brighter fluorescence was detected on the site of adherence between the Acanthamoeba (W4) cysts and trophozoites (Fig. 1,3). These lectin receptors were concentrated at the uroidal region of the trophozoite. The fluorescence, however, was absent in the newly forming hyaline cap (Fig.4). Upon addition of α -methyl-mannoside (0.5 M), Con A binding to sugar moieties in cyst and trophozoites was blocked and no fluorescence was observed.

The binding specificity of Con A-FITC and Acanthamoeba cell surface mannose moieties demonstrate topographical distribution of lectin receptor sites. Ultrastructurally, ferritin-labeled Con A at cell adhesion sites showed clustering of lectin receptors. Occurrence of fluorescence in Naegleria sp. using Con A-FITC has been shown to concentrate at the uroidal region but no fluorescence was seen at the anterior of newly formed pseudopodia.
Endosome Fusion and Tubule Formation in Cells Overexpressing GFP-Rab5 Fusion Proteins
Authors: Roberts RL , Barbieri MA , Stahl PD
Published: July 2020

Previous studies have demonstrated that GTPgS and rab5:Q79L, a constitutively active rab5 mutant, stimulate endosome fusion in in vitro assays [Cell Reg. 1:113-124 (1989)]. Additionally, electron microscopy has shown that under these conditions, endosomes form incompletely fused couplets [Cell Reg. 1:113-124 (1989); Arch. Biochem. Biophys. 326:64-72 (1996)]. CHO and BHK cells that overexpress rab5:Q79L develop ‘giant’ cytoplasmic vesicles that exhibit many characteristics of early endosomes including immunoreactivity for rab 5 and transferrin receptor (data not shown). The formation of these vesicles has been analyzed by time-lapse video microscopy which shows the enlarged endosomes arise primarily by fusion of smaller vesicles. These endosome fusion events occur by two distinct mechanisms that differ in temporal and spatial characteristics of the membrane merger process and we refer to these processes as “explosive” versus “bridge” fusion. “Explosive” fusion is a rapid process in which the initial opening that forms between contacting vesicles expands rapidly and permits an abrupt, complete coalescence of membranes (Fig. 1).
Ultrastructural Evidence for Proteoglycans Mediating an Attachment of Type VI Collagen to Banded Collagen Fibrils
Authors: R. Keene Douglas , C. Ridgway Catherine , V. Iozzo Renato
Published: July 2020

Immunolocalizaton studies of type VI collagen in skin have previously demonstrated that type VI collagen forms a flexible network that anchors large interstitial structures such as nerves, blood vessels, and collagen fibers into the surrounding connective tissues matrix. The purpose of this study is to determine if individual type VI collagen microfilaments might be connected to banded collagen fibrils, thereby stabilizing the network.

Solid phase binding assays suggest a specific, high affinity interaction between the core protein of the dermatan sulfate proteoglycan decorin and type VI collagen, and immunocytochemical studies in fetal and neonate rabbit cornea suggest an association of decorin with type VI microfilaments. Other studies in skin and perichondrium have localized decorin to a region between the d and e bands of banded collagen fibrils. However, no direct documentation has demonstrated a specific structural interaction between type VI microfilaments and banded collagen fibrils. We, therefore, sought to determine if type VI microfilaments cross banded collagen fibrils between the “d” and “e” bands.
Rat Reactive Astrocyte Marker (IFAP-70/280KD) is Expressed in Rat Spinal Cord Motor Neurons Following Transection of Sciatic Nerve.
Authors: Kriho V , Pappas GD , Lieska N , Wu C-M , Yang H-Y
Published: July 2020

Following injury to peripheral nerves, processes involved in regeneration must be activated, restoring the original architecture and synaptic connections of the neuron. This is essential for the efficient operation of the sophisticated communications network of the nervous system. In order to accomplish these tasks, complex changes occur in gene expression. Regenerating neurons shift into a growth mode wherein large amounts of cytoskeletal proteins and other growth-associated proteins are produced. These materials, which are synthesized and produced in the neuronal cell body, are then transferred to the axon via axonal transport systems. Among the cytoskeletal and associated proteins upregulated following injury to the CNS are actin, tubulin and the intermediate filament-associated protein, IFAP-70/280kD. The latter is the subject of this investigation.

Intermediate filaments (IF) are a major constituent of the cytoskeleton of most eukaryotic cells. The IF cytoskeleton is a highly dynamic structure that reorganizes continuously as the cell divides and changes shape in response to its environment.
Standardization of Routine Immunohistochemistry (IHC) by Antigen Retrieval (AR): is it Possible?
Authors: Shi S-R , Chaiwun B , Cote RJ , Young L , Chen T , Taylor CR
Published: July 2020

The standardization of IHC has been a critical issue since 1977. The most significant challenge for the standardization of routine IHC may be the adverse influence of formalin, which is in part due to variable periods of formalin-fixation ranging from 12 hours to days or weeks, resulting in variable intensity of immunostaining for formalin sensitive antigens. The world-wide use of the AR technique for IHC in both clinical and research histopathology has demonstrated that the AR is a breakthrough in pathology. Based on numerous studies of AR-IHC with excellent results for a variety of interesting markers used in diagnostic pathology, the possibility has been raised that AR technique may be useful in the standardization of IHC. The purpose of this study is to demonstrate the possibility of standardization of IHC based on the ‘test battery‘ approach for establishing an optimal protocol of AR in order to achieve a state of ‘maximal retrieval’, showing a equal comparability of AR-IHC staining of archival paraffin-embedded tissues under varying periods of formalin-fixation.
A New Attractive Approach of Three- Dimensional(3D) Reconstruction of the Relationships Between Hormone Secreating Cells and its Microvessel Network Patterns in Rat Pituitary Gland by Confocal Laser Scanning Miroscopy(CLSM) With Combined Fluorescent and Non-Fluorescent Probes.
Authors: Itoh J , Takumi I , Sanno N , Kawai K , Serizawa A , Teramoto A , Osamura RY
Published: July 2020

There have been considerable interests in the relationship between hormone secreting endocrine cells and microvessel network patterns in human pituitary gland. However, its 3D microcirculatory networks have not been studied extensively. Therefore, this study was designed to visualize hormone secreting endcrine cells and microvessels in 3D by CLSM and to reveal relationship between hormone secretion and microvessel networks in the rat pituitary gland.

CLSM has been an exciting new apparatus because of its more powerful resolution than conventional light microscopy and the high-performance on 3D optical sectioning. The images taken with CLSM can be digitized and made susceptible to image analysis manipulation and computer-assisted 3D reconstructions.

The female normal adult Wistar rats were used in this study. To demonstrate microvessels, fluorescent probe( 30 mg/ml)-conjugated gelatin solution was injected to left ventricle of the heart After complete perfusion, the pituitary glands were resected and immediately fixed in cold graded PFA(0.5- 8 %) containing 15 % picric acid solutions (4 °C. GH, ACTH cells or S100 protein positive fcfliculostellate (FS) cells were demonstrated by indirect TRTTC-labeled or HRP labeled antibody methods.
Light and Electron Microscope Immunohistochemical Localization of Decorin and Biglycan in Osteoblasts and Early Bone Matrix of Developing Rat Bone.
Authors: Yamada T , Ono Y , Lee I , Ikeda T , Takagi M
Published: July 2020

Decorin and biglycan are the predominant proteoglycans (PGs) isolated from bone of several animal species. Previous light microscope (LM) immunohistochemical studies of fetal human bone demonstrated that osteoid and osteoblasts reacted with both biglycan and decorin antibodies, but min-eralized matrix did not; however, the precise distribution of immunostaining was not examined at the electron microscope (EM) level. The present study examines LM and EM immunolocalization of decorin and biglycan in osteoblasts and early bone matrix of developing mandible of fetal rats, using polyclonal antibodies (LF113, LF106) directed against synthetic peptides corresponding to nonho-mologous regions of the two core proteins.

Fetuses were collected at embryonic day 15-18 from pregnant Wistar rats. After aldehyde fixation, developing jaws with and without osmium post-fixation were dehydrated, and embedded in paraffin, Spurr's resin, or LR gold resin for morphological and immunohistochemical observations. Sections cut from paraffin- or LR gold resin-embedded specimens were immunostained with LF113 specific for rat decorin or LF106 specific for rat biglycan, which were kindly provided by Dr. L.W. Fisher, National Institute of Dental Research, NIH, Maryland.
The Nature and Distribution of Large Proteoglycans in Developing Rat Bone.
Authors: Lee I , Ono Y , Kubushiro N , Ikeda T , Takagi M
Published: July 2020

The nature and distribution of large hyaluronate-binding proteoglycans (PGs) in early bone formation bf developing rat mandible were biochemically and immunohistochemically investigated with a monoclonal antibody (MAb) 5D5, which is derived from bovine sciera and specifically recognizes the core protein of the large PGs such as versican, neurocan and brevican, but not that of aggrecan.

The first sign of mandibular bone formation is evident in the embryonic day 15 (Day 15). At the site of initiation of bone formation, a center of ossification was seen in the close proximity to Meckel’s cartilage. At the light microscopic level, these centers contained fine strips of the intercellular matrix surrounded by osteoblasts (Fig. 1). At the EM level, these matrices contained very fine fibrillar material, numerous, osteoblastic cell processes, and vesicular structures, but did not show any typical mineralization pattern. In Day 16, clusters of needle-like crystals or bone nodules, which varied considerably in size and shape, were seen in the intercellular matrix among osteoblasts (Fig. 2).
Light and Electron Microscope Immunohistochemical and Biochemical Studies of Vitronectin in Developing Rat Bone.
Authors: Takagi M , Kumagai T , Lee I , Ono Y , Maeno M
Published: July 2020

Recently, Seiffert using light microscope (LM) immunohistochemical methods, has localized vitronectin (Vn) in the mineralized bone matrix of mature mouse long bone after demineralization with 10 % nitric acid, indicating that Vn is a specific component of bone tissue. This raises the possibility that Vn is involved in regulation of bone metabolism. The present study extends previous studies to the electron microscope (EM) level and utilizes biochemical methods to determine the distribution and nature of Vn in early bone formation of developing rat mandible with rabbit antimurine Vn IgG (antiVn).

Developing jaws of fetuses were collected at embryonic day 15-18 from pregnant Wistar rats. After al-dehyde fixation, specimens with and without osmium post-fixation were dehydrated, and embedded in paraffin, Spurr's resin, or LR gold resin for morphological and immunohistochemical observations. Sections cut from paraffin- or LR gold resin-embedded specimens were immunostained with antiVn, which was kindly provided by Dr. D. Seiffert, The Scripps Research Institute, La Jolla, California.
Spectral Karyotyping
Authors: Ried Thomas , Macville Merryn , Manoir Stan du , Veldman Tim , Weaver Zoe , Liyanage Marek , Schröck Evelin
Published: July 2020

In human leukemia and solid tumors cytogenetic analysis provides critical information of diagnostic and prognostic importance. Frequently however, the unambiguous characterization of all aberrant chromosomes is difficult using conventional chromosome banding techniques alone. We have developed a novel approach, termed spectral karyotyping (SKY), based on the hybridization of 24 differentially labeled human chromosome painting probes that allows the simultaneous color identification of all human chromosomes. This genome scanning method combines Fourier spectroscopy, CCD-imaging, and optical microscopy to visualize the hybridization of differentially labeled chromosomes painting probes. One of the most important analysis algorithms is the spectral-based classification algorithm that enables multiple different spectra in the image to be identified and highlighted in classification-colors. This allows assignment of a specific classification-color to all human chromosomes based on their spectra. This algorithm assumes that the (reference) spectrum of each chromosome has been measured and stored in a reference library in the computer
Actin Dynamics and Force Generation in the Motility of Listeria Monocytogenes
Authors: Fung David C , Theriot Julie A
Published: July 2020

The gram-positive bacterium Listeria monocytogenes is one of several intracellular pathogens which can move about within its host's cytoplasm using a form of actin-based motility. This motility plays an important role in the virulence of the microbe, which can cause serious disease in humans. Actin is a host-cell protein whose polymerization is required for the locomotion of animal cells. L. monocytogenes exploits this normal cellular machinery for its own movement by creating a “comet” tail of cross-linked actin filaments behind it. Actin polymerization occurs at the bacterial surface and is required to drive the bacterium forward.

We are interested in how L. monocytogenes induces the polymerization of actin and how this polymerization can lead to the generation of force. To study this, we have used time-lapse videomicroscopy and a cell-free system derived from Xenopus egg extracts. We have observed rare instances of a saltatory movement of L. monocytogenes in these extracts.
Probing The Mechanisms Underlying Kinetochore Behavior In Vertebate Cells Using Combinations of Advanced Light and 3-D Electron Microscopy
Authors: McEwen B F , Heagle AB , Rieder CL
Published: July 2020

For daughter cells to receive equal copies of the genome during mitosis, the replicated chromosomes must attach to and move bi-directionally on the mitotic spindle. A chromosome becomes attached to the spindle via a pair specialized structures, known as kinetochores, that are positioned on opposite sides of its primary constriction (one on each of the two chromatids). In addition to being the spindle attachment site, kinetochores are also involved in producing and/or transmitting the forces for chromosome motion. In vertebrates the kinetochore closest to a spindle pole at the time of nuclear envelope breakdown usually is the first to attach to the spindle. As a result of this attachment the now “monooriented” chromosome moves toward the closest pole where its only attached kinetochore initiates oscillatory motions toward and away from that pole until the unattached sister kinetochore acquires microtubules (Mts) from the opposite spindle pole.
Electron Tomography of Rat-Liver Mitochondria: Structure of Cristae and Interactions With Endoplasmic Reticulum
Authors: Mannella CA , Buttle K , Tessitore K , Pixley S , Rath B , D’Arcangelis D , Marko M
Published: July 2020

We are using electron tomography to investigate the internal organization of mitochondria and the interactions of these organelles with other cellular components. Two-axis tilt-series are recorded from 0.2-1.0-μm sections using either an AEI EM7 high-voltage or JEOL 4000 intermediate-voltage electron microscope. Reconstructions are computed using programs in SPIDER and visualized with the Sterecon system.

Three-dimensional reconstructions of conventionally fixed and plastic-embedded rat-liver mitochondria (isolated and in situ) indicate that the cristae, the infoldings of the inner membrane, are pleiomorphic, defining compartments that are connected to each other and to the surface of the inner membrane by narrow (30-40-nm diameter) tubular regions (Fig. 1). This is contrary to the “baffle” model commonly shown in textbooks and has important implications for the diffusion of ions, metabolites, and macromolecules inside the organelle. The same basic organization is also observed in reconstructions of mitochondria in rat-liver sections prepared by the cryo-fixation method of Tokuyasu(Fig. 2).
Three-Dimensional Structure of the Gap Junction Connexon and Intercellular Channel
Authors: Perkins Guy , Goodenough Dan , Sosinsky Gina
Published: July 2020

Gap junctions are specialized cell-cell contact areas by which cells communication with each other. Within these contact areas are tens to thousands of membrane channels. A gap junction membrane channel (also referred to as an intercellular channel) is unique among membrane channels in that it is composed of two oligomers with each of two adjacent tissue cells contributing one oligomer (called a connexon or hemichannel). The pore of the intercellular channel controls the passage of small molecules and ions from one cell to another.

We are interested in how the structure and surface topology of the gap junction connexon at its extracellular surface influences the docking and formation of an intercellular communicating channel. It has been demonstrated that connexons made from some connexins will dock and form functional channels with some but not all connexons made from other isoforms. This selectivity is surprising considering that the primary sequences of the docking domains are highly conserved.
Scanning Electron Microscopic Study of Erythroblastic Islands Obtained by “Spleen-Stamp Culture” Method
Authors: Fujii Y , Terada N , Ueda H , Baba T , Ohno S
Published: July 2020

During hematopoiesis of mammalian animals, an erythroblastic island in the bone marrow has been reported to contain a central macrophage surrounded by many erythroblasts. Fetal mouse livers and spleens, which are treated with collagenase, have been used to get the erythroblastic island in tissue cultures. In this study, we have cultured some stamped cells obtained from spleens of anemic mice and examined their structures by scanning electron microscopy.

Hemolytic anemia of female BALB/c mice was induced by acetylphenylhydrazine (4mg/g body weight) injection for 5 days. Spleens were resected on the second day after the last injection and a “spleen-stamp culture“ method was performed. Briefly, they were cut with razor blades (Fig. 1a) and splenic cells on the cut tissue surface were stamped on collagen-coated coverslips (Fig.1b). The stamped splenic cells were covered with dialysis membranes (Fig. 1c). They were cultured with 55% IMDM, 20% L-15 and 25% FBS, containing 2U/ml erythropoietin (EPO) in humidified atmosphere of 5% CO2 at 37°C for 1-3 days.
Microcontact Printed Substrates for Growing Central Nervous System Neurons and Glia
Authors: Craighead H G , Isaacson M , John P St , Davis R , Banker G , Esch T , Kam L , Shain W , Turner J N
Published: July 2020

The application of nano- and microfabrication to biology is an emerging area linking engineering and biology. We are using microfabrication techniques to make patterns of specific bioactive molecules on glass and silicon surfaces to study central nervous system neurons and glia. Cell adhesion and tissue reaction to implanted silicon based electronic prosthetic devices are being studied, and the methods expanded to interface neurons with integrated circuits to study their electrical properties. The number, type and distribution of cells adhering to particular portions of a substrate can be influenced by patterning amine groups, proteins or polypeptides. We are using microcontact printing as an alternative to lithography for chemical patterning.

Primary cultures of rat cortical astrocytes and hippocampal neurons, and continuous cultures of transformed astrocytes were prepared on microcontact printed patterns produced by an elastomer stamp made from a silicon master. Fig. 1 shows two such patterns prepared by depositing self-assembled monolayers of N1[3-(Trimethoxylsily)propyl] diethylenetriamine (DETA) and octadecyltrichlorosilane (OTS).
How Multiphoton Excitation can Illuminate Biophysics
Authors: Bailey GW , Dimlich RVW , Alexander KB , McCarthy JJ , Pretlow TP , Webb W W
Published: July 2020

Multiphoton molecular excitation by the strongly focused femtosecond pulses of infrared light generated as an 80 MHZ pulse train by a mode locked laser provides intrinsic submicron three dimensional spatial resolution of fluorescence excitation and photochemistry for laser scanning fluorescence microscopy. Because two-photon excitation requires simultaneous (∼10-16 seconds), absorption of two-photons focused laser intensities of about 1022 photons/cm2s are required. Since the rate of absorption is proportional to the square of the intensity, excitation is limited to the focal volume and is negligible elsewhere along the double cone of the focused illumination. Therefore, out of focus photodamage and fluorescence are generally negligible and laser scanning fluorescence microscopy with multiphoton excitation is intrinsically three dimensionally resolved with no out of focus background. Since the appropriate wave lengths are infrared for multiphoton excitation of ultraviolet or visible absorbing molecules, out of focus photodamage is eliminated. This allows imaging of useful ultraviolet absorbing indicators, vital DNA stains and autofluorescence in living cells with minimal, but not necessarily negligible, photodamage.
Quantitative Analysis of Coronary Atherosclerosis by Intra Vascular Ultrasound
Authors: Vince D G , Shekhar R , Cothren R M , Nissen S E , Tuzcu E M , Cornhill J F
Published: July 2020

Traditional methods for studying human coronary artery disease have significant limitations. Angiography allows evaluation only of the geometry of the remaining lumen; it cannot provide information on the structural or cellular composition of the arterial wall, which is essential to understand the processes involved in the progression of atherosclerosis. Intravascular ultrasound (IVUS) imaging is a new technology that permits tomographical visualization of a cross section through the vessel wall (Fig 1 A.) The development and refinement of IVUS has provided a powerful in vivo method to assess plaque morphology. Recent clinical studies have documented its sensitivity in detecting atherosclerosis, and it is increasingly employed to assist in selecting an appropriate therapeutic intervention. Perhaps more importantly, the potential of IVUS to quantify the structure and geometry of normal and atherosclerotic coronary arteries will allow one to characterize specific lesions and to differentiate the plaques that lead to various clinical syndromes. This study presents a three-dimensional (3D) reconstruction technique suitable for clinical use that accurately preserves 3D geometric information throughout the cardiac cycle without requiring mechanical IVUS catheter withdrawal aids.
Application of Multiphoton Imaging to Study of the Vasculature.
Authors: Zipfel Warren R , Williams Rebecca M , Watt W
Published: July 2020

Nonlinear or multiphoton fluorescence microscopy utilizes the simultaneous absorption of two (or three) longer wavelength photons to excite fluorophores (Denk et.al., 1990). For example, UV absorbing fluorophores such as DAPI or lndo-1 are excited using 700 nm near infrared light rather than 350 nm UV excitation. This relatively new form of laser scanning fluorescence microscopy has excellent optical sectioning capabilities in thick, highly scattering tissues. The high degree of intrinsic optical sectioning arises from the spatial nature of the excitation dependence, rather than from a confocal aperture or deconvolution algorithm. The fluorescence arising from any point in the specimen depends on the second or third power of the intensity (i.e. two and three photon excitation, respectively). This squaring (or cubing) of the illumination point spread function effectively confines the excitation to a tenth of a femtoliter optical volume when using a high NA lens. 680 to 1100 nm mode-locked lasers producing pulses of 100 femtosecond duration at a repetition rate of 80 Mhz are used to efficiently produce fluorescence excitation at average powers that are usually in the 0.5 to 10 mW range at the sample.
Quantitative Evaluation of a Standard for Immunogold Labeling of Collagen Type IV
Authors: Springe MJ , Hann CR , Johnson DH
Published: July 2020

Quantitative gold labeling studies allow the comparison of tissue antigens only if labeling conditions are the same. When labeling conditions deviate slightly, the labeling efficiency can also shift. Labeling efficiency can be determined on a standard containing known concentrations of the target antigen. When labeling this standard, the labeling density (LD) divided by the antigen concentration (C) gives the labeling efficiency (LE).(l) If the assumption is accepted that labeling efficiency is the same over the standard and tissue, then the antigen concentration in the tissue can be calculated from the labeling density. In this study, a labeling standard was constructed that contained collagen type IV in five concentrations and a blank agarose gel. Our design criteria specified that these six gels measure less than 2 mm across and the zones between protein concentrations be clearly delineated.

The labeling standard was constructed by diluting lyophilyzed collagen type IV in water to final concentrations of 1,2,3,4, and 5 mg/ml.
Exhaustive Photon Reassignment™: A Method Offering Enhanced Sensitivity and Quantitative Accuracy for High Resolution Fluorescence Microscopy
Authors: Kramer Jennifer A , Moorthy Ramkumar K , Casavan William N , Hitrys David C
Published: July 2020

Confocal microscopy is a technique that allows researchers to obtain a highly resolved, high-contrast image of a focal plane in a fluorescent specimen by excluding or rejecting light emanating from out-of-focus planes. However, the basic design of confocals results in limitations for many biologists. It is often difficult to avoid photobleaching of specimens, to visualize fine or faintly-labeled structures, or to acquire high-quality images using short exposure times. The photomultiplier tubes used as the amplification detectors in these systems are restricted to a dynamic range of 8 bits (255 intensity levels), produce noise, and are not quantitatively linear detectors.

Members of the Biomedical Imaging Group at the University of Massachusetts Medical School in Worcester, MA have spent the last fifteen years developing and perfecting a digital imaging system that helps scientists overcome these problems. Scanalytics is the exclusive worldwide licensee of this patented technology which allows researchers to obtain high-resolution, quantitatively accurate three-dimensional images of fluorescent specimens
Structure-Property Relationships in Sputtered Magneto-Optic Multilayers
Authors: Sinclair R , Bertero G A , Visokay MR
Published: July 2020

One technology for high-density information storage employs magneto-optic thin films. Key material properties include a large magneto-optic Kerr rotation of polarized light (especially in the blue wavelength range), high perpendicular magnetic anisotropy, good reflectivity and a large magnetic coercivity. These can be achieved in nano-scale magnetic multilayers (e.g., Co-Pt) in which one of the constituent layers is ferromagnetic. Naturally the magnetic properties can be manipulated to a great extent by the preparation conditions, and the microstructure is most readily revealed by cross-section high-resolution electron microscopy (HREM). In our work, extremely high quality multilayers were fabricated by sputtering using a variety of inert gases, and most notably the influence of interface sharpness on properties was assessed. Intermetallic compounds, which are essentially “natural” multilayers, were also extensively studied.

Figure 1 shows magneto-optic Kerr rotation hysteresis loops for multilayers sputtered in 5 mtorr of either argon or the heavier xenon gas.
HRTEM Study of Nanostructure of Co/Au Multilayers
Authors: Liu Y , Robertson BW , Shan ZS , Sellmyer D J
Published: July 2020

Multilayer thin films of Co/Au with large magnetic anisotropy are attracting strong interest. Among the factors affecting the film properties are the layer thicknesses of the Au layer and the Co layer. This paper reports the detailed HRTEM study of the nanostructure evolution of Co/Au films as a function of the Au thickness. Two DC-magnetron sputtered films with the same Co thickness 5Å but different Au thicknesses (Au60Å/Co5Å)x9/Au400Å (denoted Au60) and (Au10Å/Co5Å)x50/Au400Å (denoted Au 10) were chosen for detailed study. The Au60 film shows a well defined perpendicular magnetization loop with loop squareness S=l and an anisotropy value of 3xl06erg/cm3 while the Au 10 film shows an irregular perpendicular magnetization loop with S=0 and an anisotropy value of 1.5xl06erg/cm3.

For cross sectional TEM samples, bright field TEM images show that the grain size of the Au underlayer is about 20-80 nm. Slight convex curvature of the multilayers is observed.