Probing the ultrastructure of the mitotic and meiotic spindle in eggs: An expeditious approach based on semi-thin (0.25 μm) serial sections

1983 ◽  
Vol 41 ◽  
pp. 552-555
Author(s):  
Conly L. Rieder ◽  
S. Bowser ◽  
R. Nowogrodzki ◽  
K. Ross ◽  
G. Sluder
Keyword(s):  
Small Sample Size ◽  
Small Sample ◽  
Meiotic Spindle ◽  
Serial Sections ◽  
Mitotic Apparatus ◽  
Thin Sections ◽  
Cell Cycles ◽  
Ultrastructural Studies

Eggs have long been a favorite material for studying the mechanism of karyokinesis in-vivo and in-vitro. They can be obtained in great numbers and, when fertilized, divide synchronously over many cell cycles. However, they are not considered to be a practical system for ultrastructural studies on the mitotic apparatus (MA) for several reasons, the most obvious of which is that sectioning them is a formidable task: over 1000 ultra-thin sections need to be cut from a single 80-100 μm diameter egg and of these sections only a small percentage will contain the area or structure of interest. Thus it is difficult and time consuming to obtain reliable ultrastructural data concerning the MA of eggs; and when it is obtained it is necessarily based on a small sample size.We have recently developed a procedure which will facilitate many studies concerned with the ultrastructure of the MA in eggs. It is based on the availability of biological HVEM's and on the observation that 0.25 μm thick serial sections can be screened at high resolution for content (after mounting on slot grids and staining with uranyl and lead) by phase contrast light microscopy (LM; Figs 1-2).

scholarly journals In vivo assessment of a single adenine mutation in 5′UTR of Endothelin-1 gene in paediatric cases with severe pulmonary hypertension: an observational study

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Abhishek Kumar ◽  
Minati Choudhury ◽  
Sakshi Dhingra Batra ◽  
Kriti Sikri ◽  
Anushree Gupta
Keyword(s):  
Pulmonary Hypertension ◽  
Congenital Heart ◽  
Small Sample Size ◽  
Small Sample ◽  
Severe Pulmonary Hypertension ◽  
Endothelin 1 ◽  
Circulating Levels ◽  
In Vivo Assessment

Abstract Objective Endothelin-1 plays an important role in the pathogenesis of severe pulmonary hypertension. The + 139 ‘A’, adenine insertion variant in 5′UTR of edn1 gene has been reported to be associated with increased expression of Endothelin-1 in vitro. The aim of present study was to explore the association of this variant with the circulating levels of Endothelin-1 in vivo using archived DNA and plasma samples from 38 paediatric congenital heart disease (cyanotic and acyanotic) patients with severe pulmonary hypertension. Results The plasma Endothelin-1 levels were highly varied ranging from 1.63 to75.16 pg/ml. The + 139 ‘A’ insertion variant in 5′UTR of edn1 was seen in 8 out of 38 cases with only one acyanotic sample demonstrating homozygosity of inserted ‘A’ allele at + 139 site (4A/4A genotype). The plasma Endothelin-1 levels in children with homozygous variant 3A/3A genotype were comparable in cyanotic and acyanotic groups. Lone 4A/4A acyanotic sample had ET-1 levels similar to the median value of ET-1 associated with 3A/3A genotype and was absent in cyanotic group presumably due to deleterious higher ET-1 levels. The discussed observations, limited by the small sample size, are suggestive of homozygous adenine insertion variant posing a risk in cyanotic babies with Severe Pulmonary Hypertension.

scholarly journals Development of AOSpine BOnE (Bone Osteobiologics and Evidence) Classification

2019 ◽  
Vol 10 (7) ◽  
pp. 871-874 ◽  
Author(s):  
Jeffrey C. Wang ◽  
S. Tim Yoon ◽  
Darrel S. Brodke ◽  
Jong-Beom Park ◽  
Patrick Hsieh ◽  
...  
Keyword(s):  
Small Sample Size ◽  
Small Sample ◽  
In Vitro Studies ◽  
Level Of Evidence ◽  
Knowledge Forum ◽  
Levels Of Evidence ◽  
Published Evidence ◽  
Patient Reported

Study Design: Classification development. Objectives: The aim of our study was to develop a 3-tier classification for the levels of evidence for osteobiologics and provide a description of the principles by which osteobiologics can be evaluated. BOnE (Bone Osteobiologics and Evidence) classification evaluates each osteobiologic based on the available evidence, and if the published evidence is based on clinical, in vivo or in vitro studies. Methods: The process of establishing the BOnE classification included 5 face-to-face meetings and 2 web calls among members of the AOSpine Knowledge Forum Degenerative. Results: The 3 levels of evidence were determined based on the type of data on osteobiologics: level A for human studies, level B for animal studies, and level C for in vitro studies, with level A being the highest level of evidence. Each level was organized into 4 subgroups (eg, A1, A2, A3, and A4). Conclusions: The use and the variety of osteobiologics for spine fusion has dramatically increased over the past few decades; however, literature on their effectiveness is inconclusive. Several prior systematic reviews developed by AOSpine Knowledge Forum Degenerative reported low level of evidence primarily due to the high risk of bias, small sample size, lack of control groups, and limited patient-reported outcomes. BOnE classification will provide a universal platform for research studies and journal publications to classify a new or an existing product and will allow for creating decision-making algorithms for surgical planning.

scholarly journals Effectiveness and safety of topical amphotericin B in 30% dimethyl sulfoxide cream versus 30% dimethyl sulfoxide cream for nondermatophyte onychomycosis treatment: A pilot study

2021 ◽  
Vol 0 ◽  
pp. 1-6
Author(s):  
Charussri Leeyaphan ◽  
Bordeesuda Suiwongsa ◽  
Phojana Komesmuneeborirak ◽  
Rungsima Kiratiwongwan ◽  
Supisara Wongdama ◽  
...  
Keyword(s):  
Dimethyl Sulfoxide ◽  
Amphotericin B ◽  
Small Sample Size ◽  
Fungal Species ◽  
Small Sample ◽  
Controlled Study ◽  
Mycological Cure ◽  
Accurate Identification

Background: Although topical amphotericin B cream is effective for the treatment of nondermatophyte mold onychomycosis in vitro, studies of its effectiveness and safety in vivo are limited. Objectives: We studied the effectiveness and safety of topical 0.3% amphotericin B in 30% dimethyl sulfoxide cream (amphotericin B cream) in nondermatophyte mold onychomycosis using the vehicle cream 30% dimethyl sulfoxide cream as control. Methods: This randomized controlled study was conducted between January 2019 and November 2020. Patients diagnosed with nondermatophyte mold onychomycosis were randomly divided into two groups of ten patients each: one treated with amphotericin B cream and the other with the vehicle cream. Clinical and mycological cure as well as safety were evaluated. Results: Ten patients each treated with amphotericin B cream and the vehicle cream were included in the study, but only nine patients in the vehicle cream group were available for follow up. All the 19 evaluable patients had distal lateral subungual onychomycosis and the great toenails were affected in 18 (94.7%) of these. Mycological cure was achieved in 8 (80%) patients treated with amphotericin B cream and in 4 (44.4%) patients using the control (vehicle) cream. Clinical cure was achieved in 7 (70%) patients treated with amphotericin B cream, but only in 2 (22.2%) patients on the control cream. No adverse events were observed. Limitations: The small sample size and the fact that PCR fungal identification that provides accurate identification of fungal species was not performed are limitations of our study. Conclusion: Topical amphotericin B cream was both very effective and safe in the treatment nondermatophyte mold onychomycosis. The control (vehicle) cream containing 30% dimethyl sulfoxide also demonstrated some antifungal activity.

Serum α-Inhibin Levels in Polycystic Ovary Syndrome: Relationship to the Serum Androstenedione Level1

1997 ◽  
Vol 82 (6) ◽  
pp. 1939-1943
Author(s):  
Pascal Pigny ◽  
Rachel Desailloud ◽  
Christine Cortet-Rudelli ◽  
Alain Duhamel ◽  
Delphine Deroubaix-Allard ◽  
...  
Keyword(s):  
Regression Analysis ◽  
Polycystic Ovary Syndrome ◽  
Polycystic Ovary ◽  
Small Sample Size ◽  
Small Sample ◽  
Molecular Forms ◽  
Enzyme Linked Immunosorbent Assay ◽  
Ovary Syndrome

Abstract To date, only one study has demonstrated increased serum inhibin levels in women with polycystic ovary syndrome (PCOS). Moreover, no relationship between serum inhibin and either FSH or androgen levels has been noted. This lack of data could be due to 1) the heterogeneity of PCOS and the small sample size of previous studies, and/or 2) the complexity of circulating inhibin molecular forms, which hinders the precise evaluation of bioactive inhibin. In the present study,α -inhibin levels were assayed in the serum of 61 healthy women and 72 PCOS patients by means of an α-α enzyme-linked immunosorbent assay. Serum α-inhibin levels together with LH and androstenedione (A) levels were significantly increased in PCOS women (mean ± sd, 1.45 ± 0.55 vs. 0.94 ± 0.36 U/mL in controls; P < 0.001). Moreover, simple and partial regression analysis demonstrated that serum A levels were positively and independently correlated to serum α-inhibin (r = 0.32; P < 0.01) and LH levels (r = 0.48; P < 0.001) in PCOS. The respective influences of α-inhibin and LH on A variability were 20% and 80%, as determined by multiple regression analysis. In conclusion, in agreement with recent in vitro data, our in vivo results argue for a role of inhibin in the hyperandrogenism of PCOS together with, but independently from, that of LH. Further studies are needed to determine whether this effect is produced by inhibin A and/or B.

scholarly journals PRESENCE OF A BASE LINE LEVEL OF SISTER CHROMATID EXCHANGES IN SOMATIC CELLS OF DROSOPHILA MELANOGASTER IN VIVO AND IN VITRO

Genetics ◽  
1982 ◽  
Vol 100 (2) ◽  
pp. 259-278
Author(s):  
Hideo Tsuji
Keyword(s):  
Drosophila Melanogaster ◽  
Sister Chromatid ◽  
Ganglion Cells ◽  
Sister Chromatid Exchanges ◽  
Base Line ◽  
Cell Cycles ◽  
Chromatid Exchanges ◽  
Third Instar Larvae

ABSTRACT Sister chromatid exchanges (SCEs) under in vivo and in vitro conditions were examined in ganglion cells of third-instar larvae of Drosophila melanogaster (Oregon-R). In the in vivo experiment, third-instar larvae were fed on synthetic media containing 5-bromo-2′-deoxyuridine (BrdUrd). After two cell cycles, ganglia were dissected and treated with colchicine. In the in vitro experiment, the ganglia were also incubated in media containing BrdUrd for two cell cycles, and treated with colchicine. SCEs were scored in metaphase stained with Hoechst 33258 plus Giemsa. The frequencies of SCEs stayed constant in the range of 25-150 vg/ml and 0.25-2.5 vg/ml of BrdUrd in vivo and in vitro, respectively. SCEs gradually increased at higher concentrations, strongly suggesting that at least a fraction of the detected SCEs are spontaneous. The constant levels of SCE frequency were estimated, on the average, at 0.103 per cell per two cell cycles for females and 0.101 for males in vivo and at 0.096 for females and 0.091 for males in vitro. No difference was found in the SCE frequency between sexes at any of the BrdUrd concentrations. The analysis for the distribution of SCEs within chromosomes revealed an extraordinarily high proportion of the SCEs at the junctions between euchromatin and heterochromatin; the remaining SCEs were preferentially localized in the euchromatic regions of the chromosomes and in the heterochromatic Y chromosome. These results were largely inconsistent with those of Gatti et al. (1979).

scholarly journals 1611. Evaluating Clinical Outcomes and Efficacy of Daptomycin in Combination with a Beta-Lactam for the Treatment of Vancomycin-Resistant Enterococcus (VRE) Bacteremia

2020 ◽  
Vol 7 (Supplement_1) ◽  
pp. S799-S800
Author(s):  
Nerea Irusta ◽  
Ana Vega ◽  
Yoichiro Natori ◽  
Lilian M Abbo ◽  
Lilian M Abbo ◽  
...  
Keyword(s):  
Clinical Outcomes ◽  
Primary Outcome ◽  
Small Sample Size ◽  
Univariate Analysis ◽  
Small Sample ◽  
Significant Difference ◽  
Vancomycin Resistant ◽  
Secondary Outcomes ◽  
Related Mortality

Abstract Background In-vitro studies have shown synergistic bactericidal activity with daptomycin (DAP) plus β-lactam antimicrobials against vancomycin resistant enterococci (VRE). There is a paucity of data regarding clinical outcomes with this combination in VRE bloodstream infections (BSI). The purpose of this study was to assess the efficacy of DAP plus a β-lactam with in-vitro activity vs. other therapies for treatment of VRE BSI. Methods IRB-approved, single-center, retrospective study of patients with VRE BSI from 01/2018-09/2019. Patients were excluded if < 18 years old, pregnant, or incarcerated. The primary outcome was time-to-microbiological clearance. Secondary outcomes included infection-related mortality, 30-day all-cause mortality, and incidence of recurrent BSI within 30 days of index culture. Targeted DAP doses were ≥ 8mg/kg and based on MIC. Factors associated with significance for outcomes, via univariate analysis, were evaluated with multivariable logistic regression (MLR), removed in a backward-step approach. Results A total of 85 patients were included, 23 of which received DAP plus a β-lactam. The comparator arm included linezolid or DAP monotherapy. Patients with combination therapy had significantly higher Charlson Comorbidity Index (CCI) (p=0.013) and numerically higher Pitt Bacteremia scores (PBS) (p=0.087) (Table 1). There was no difference seen in the primary outcome (Table 2). Secondary outcomes are provided in Table 2. The presence of polymicrobial infection and higher PBS were significantly associated with infection-related mortality (p=0.008 and p=0.005, respectively) by MLR. A Mann Whitney U test indicated that presence of infection-related mortality was greater for patients with higher MICS (U=20.5, p=0.06). The presence of an underlying source may be related to recurrence of BSI (p=0.075). Table 1: Patient Characteristics Table 2. Primary and Secondary Outcomes Conclusion We did not find a significant difference in time-to-microbiological clearance, although patients treated with DAP and a β-lactam had higher CCI and PBS. These results are limited by retrospective design, small sample size, and potential selection bias. Prospective randomized studies are needed to further validate these findings. Disclosures All Authors: No reported disclosures

scholarly journals EJE Prize 2013: Regulation of aldosterone secretion: from physiology to disease

2013 ◽  
Vol 168 (6) ◽  
pp. R85-R93 ◽  
Author(s):  
Felix Beuschlein
Keyword(s):  
High Throughput Sequencing ◽  
Secondary Hypertension ◽  
Small Sample ◽  
Cellular Systems ◽  
Common Disease ◽  
Aldosterone Secretion ◽  
Technical Developments ◽  
Starting Point

Arterial hypertension is a major cardiovascular risk factor that affects between 10 and 40% of the population in industrialized countries. Primary aldosteronism (PA) is the most common form of secondary hypertension with an estimated prevalence of around 10% in referral centers and 4% in a primary care setting. Despite its high prevalence until recently, the underlying genetic and molecular basis of this common disease had remained largely obscure. Over the past decade, a number of insights have been achieved that have relied onin vitrocellular systems, wild-type and genetically modifiedin vivomodels, as well as clinical studies in well-characterized patient populations. This progress has been made possible by a number of independent technical developments including that of specific hormone assays that allow measurement in small sample volumes as well as genetic techniques that enable high-throughput sequencing of a large number of samples. Furthermore, animal models have provided important insights into the physiology of aldosterone regulation that have served as a starting point for investigation of mechanisms involved in autonomous aldosterone secretion. Finally, national and international networks that have built up registries and biobanks have been instrumental in fostering translational research endeavors in PA. Therefore, it is to be expected that in the near future, further pathophysiological mechanisms that result in autonomous aldosterone secretion will be unraveled.

Parthenogenesis and cytoskeletal organization in ageing mouse eggs

Development ◽  
1986 ◽  
Vol 95 (1) ◽  
pp. 131-145
Author(s):  
Michelle Webb ◽  
Sarah K. Howlett ◽  
Bernard Maro
Keyword(s):  
Critical Concentration ◽  
Metaphase Plate ◽  
Meiotic Spindle ◽  
Cytoskeletal Organization ◽  
Spindle Poles ◽  
Different Types ◽  
Mouse Egg ◽  
Parthenogenetic Embryos

The cytoskeletal organization of the mouse egg changes during ageing in vivo and in vitro. The earliest change observed is the disappearance of the microfilament-rich area overlying the meiotic spindle. This is followed by the migration of the spindle towards the centre of the egg. Finally the spindle breaks down and the chromosomes are no longer organized on a metaphase plate. This spindle disruption may result from changes in the microtubule nucleating material found at the spindle poles and from an increase in the critical concentration for tubulin polymerization. It is possible to correlate the changes in the cytoskeletal organization of the egg occurring during ageing with the different types of parthenogenetic embryos obtained after ethanol activation. These observations strengthen the hypothesis that the actin-rich cortical area that overlies the meiotic spindle forms a domain to which the meiotic cleavage furrow is restricted and provides some insights into the mechanisms by which different types of parthenogenetic embryos are generated.

Mesenchymal Stem Cell Therapy for Acetaminophen-Related Liver Injury: A Systematic Review and Meta-Analysis of Experimental Studies in Vivo

2021 ◽  
Vol 16 ◽  
Author(s):  
Li Wang ◽  
Yiwen Zhang ◽  
Jiajun Zhong ◽  
Yuan Zhang ◽  
Shuisheng Zhou ◽  
...  
Keyword(s):  
Systematic Review ◽  
Stem Cell ◽  
Liver Injury ◽  
Mesenchymal Stem Cell ◽  
Small Sample Size ◽  
Meta Analysis ◽  
Experimental Studies ◽  
Small Sample ◽  
Control Group

Objective: The efficacy of mesenchymal stem cell (MSC) therapy in acetaminophen-induced liver injury has been investigated in animal experiments, but individual studies with a small sample size cannot be used to draw a clear conclusion. Therefore, we conducted a systematic review and meta-analysis of preclinical studies to explore the potential of using MSCs in acetaminophen-induced liver injury. Methods: Eight databases were searched for studies reporting the effects of MSCs on acetaminophen hepatoxicity. The Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) guidelines were used. SYRCLE’s risk of bias tool for animal studies was applied to assess the methodological quality. A meta-analysis was performed by using RevMan 5.4 and STATA/SE 16.0 software. Results: Eleven studies involving 159 animals were included according to PRISMA statement guidelines. Significant associations were found for MSCs with the levels of alanine transaminase (ALT) (standardized mean difference (SMD) − 2.58, p < 0.0001), aspartate aminotransferase (AST) (SMD − 1.75, p = 0.001), glutathione (GSH) (SMD 3.7, p < 0.0001), superoxide dismutase (SOD) (SMD 1.86, p = 0.022), interleukin 10 (IL-10) (SMD 5.14, p = 0.0002) and tumor necrosis factor-α (TNF-α) (SMD − 4.48, p = 0.011) compared with those in the control group. The subgroup analysis showed that the tissue source of MSCs significantly affected the therapeutic efficacy (p < 0.05). Conclusion: Our meta-analysis results demonstrate that MSCs could be a potential treatment for acetaminophen-related liver injury.

scholarly journals Post-embedding colloidal gold immunolocalization of laminin to the lamina rara interna, lamina densa, and lamina rara externa of renal glomerular basement membranes.

1986 ◽  
Vol 34 (7) ◽  
pp. 847-853 ◽  
Author(s):  
D R Abrahamson
Keyword(s):  
Colloidal Gold ◽  
Basement Membranes ◽  
Lamina Densa ◽  
Thin Sections ◽  
Gold Particles ◽  
Rabbit Igg ◽  
Gold Immunolocalization ◽  
Lowicryl K4m

Ultrastructural distribution of laminin within renal glomerular (GBM) and tubular basement membranes (TBM) was investigated using post-embedding immunolocalization with colloidal gold. Rat kidneys were fixed with 4% formaldehyde and embedded at 4 degrees C in Lowicryl K4M medium. Thin sections were then sequentially treated with affinity-purified rabbit anti-laminin IgG and anti-rabbit IgG conjugated to 10 nm diameter colloidal gold. Gold bound specifically to the GBM and TBM with particle densities of 690/micron2 and 731/micron2, respectively. In the GBM, the number of gold particles bound/micron2 of lamina densa greater than lamina rara externa greater than lamina rara interna. Closely similar binding patterns were found when kidneys were fixed with 0.5% glutaraldehyde plus 3% formaldehyde and embedded at 60 degrees C in L.R. White resin, but slightly less gold bound to sections overall than that seen with formaldehyde alone and Lowicryl. Taken together, these results illustrate that anti-laminin IgG, whether applied to fixed sections in vitro or introduced in vivo, bound to the lamina rara interna, lamina densa, and lamina rara externa of the GBM and throughout the TBM.

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