Nuclear Diameter Research Articles

Purification and production of Plasmodium falciparum zygotes from in vitro culture using magnetic column and Percoll density gradient

BackgroundPlasmodium falciparum zygotes develop in the mosquito midgut after an infectious blood meal containing mature male and female gametocytes. Studies of mosquito-produced P. falciparum zygotes to elucidate their biology and development have been hampered by high levels of contaminating mosquito proteins and macromolecules present in zygote preparations. Thus, no zygote-specific surface markers have been identified to date. Here, a methodology is developed to obtain large quantities of highly purified zygotes using in vitro culture, including purification methods that include magnetic column cell separation (MACS) followed by Percoll density gradient centrifugation. This straightforward and effective approach provides ample material for studies to enhance understanding of zygote biology and identify novel zygote surface marker candidates that can be tested as transmission blocking vaccine (TBV) candidates.MethodsPlasmodium falciparum gametocyte cultures were established and maintained from asexual cultures. Gametocytes were matured for 14 days, then transferred into zygote media for 6 h at 27 ± 2 °C to promote gamete formation and fertilization. Zygotes were then purified using a combination of MACS column separation and Percoll density gradient centrifugation. Purity of the zygotes was determined through morphological studies: the parasite body and nuclear diameter were measured, and zygotes were further transformed into ookinetes. Immunofluorescence assays (IFA) were also performed using the ookinete surface marker, Pfs28.ResultsAfter stimulation, the culture consisted of transformed zygotes and a large number of uninfected red blood cells (RBCs), as well as infected RBCs with parasites at earlier developmental stages, including gametes, gametocytes, and asexual stages. The use of two MACS columns removed the vast majority of the RBCs and gametocytes. Subsequent use of two Percoll density gradients enabled isolation of a pure population of zygotes. These zygotes transformed into viable ookinetes that expressed Pfs28.ConclusionThe combined approach of using two MACS columns and two Percoll density gradients yielded zygotes with very high purity (45-fold enrichment and a pure population of zygotes [approximately 100%]) that was devoid of contamination by other parasite stages and uninfected RBCs. These enriched zygotes, free from earlier parasites stages and mosquito-derived macromolecules, can be used to further elucidate the biology and developmental processes of Plasmodium.

Nesprin-2 accumulates at the front of the nucleus during confined cell migration.

The mechanisms by which cells exert forces on their nuclei to migrate through openings smaller than the nuclear diameter remain unclear. We use CRISPR/Cas9 to fluorescently label nesprin-2 giant, which links the cytoskeleton to the nuclear interior. We demonstrate that nesprin-2 accumulates at the front of the nucleus during nuclear deformation through narrow constrictions, independently of the nuclear lamina. We find that nesprins are mobile at time scales similar to the accumulation. Using artificial constructs, we show that the actin-binding domain of nesprin-2 is necessary and sufficient for this accumulation. Actin filaments are organized in a barrel structure around the nucleusin the direction of movement. Using two-photon ablation andcytoskeleton-inhibiting drugs, we demonstrate an actomyosin-dependent pulling force on the nucleus from the front of the cell. The elastic recoil upon ablation is dampened when nesprins are reduced at the nuclear envelope. We thus show that actin redistributes nesprin-2 giant toward the front of the nucleus and contributes to pulling the nucleus through narrow constrictions, in concert with myosin.

High percentages of larval tetraploids in the yellowtail tetra Astyanax altiparanae induced by heat-shock: The first case in Neotropical characins

Tetraploid fish are an important source of diploid gametes for polyploid production, and they can be induced by inhibition of the first mitotic cell division. Dispite its potential, very few protocols regarding tetraploidization and subsequent application to mass production of triploid fish for aquaculture exist. In the yellowtail tetra Astyanax altiparanae, triploids present sterility and increased carcass yield, suggesting the generation of tetraploid for large-scale production of such triploids. In this study, the efficacy of heat shock treatments (40° for 2 min) at 16, 18, 20, 22, 24, and 26 min post-fertilization (mpf) at an incubation temperature of 22 °C were compared on tetraploid induction of A. altiparanae. Ploidy status was confirmed by flow cytometry, karyotyping, and nuclear diameter of erythrocytes. Afterwards, sperm parameters and fertility capacity of tetraploid males were evaluated. The results showed that heat shock decreased the hatching rate, especially at 22 (19.1 ± 9.6%), 24 (16.0 ± 9.1%), and 26 mpf (10.3 ± 6.2%) in comparison with the control group (62.2 ± 7.7%). The flow cytometric analysis showed that the control group and the treatments with heat shock at 16 and 18 mpf presented only diploid individuals. However, from 20 mpf onwards, tetraploid individuals were observed, and the highest percentage arose at 26 mpf (94.55%). When mature, tetraploid males are capable of producing diploid spermatozoa with fertilization capacity, generating a 100% triploid offspring. The results indicate that the heat shock treatment at 26 mpf (40 °C for 2 min) is optimum for tetraploid induction in the A. altiparanae. Additionally, tetraploid mature males generated diploid spermatozoa and can be used to mass production of triploid fish. The current data are the first report of tetraploids in family Characidae, presenting an application in basic and applied sciences.

Comparative analysis of nucleomorphometric parameters in methyl green-pyronin-stained sections of oral epithelial dysplasia, oral submucous fibrosis and oral squamous cell carcinoma.

Context:The diagnosis and grading of epithelial dysplasia is based on a combination of architectural and cytological changes. A gradual increase in quantitative DNA aberrations has been found to correlate with increasing degree of dysplasia in oral mucous membranes.Aims:The aim of this study is to assess nuclear parameters in potentially malignant and malignant lesions of the oral cavity and to assess cytomorphometric changes in the nucleus and nucleolus in oral epithelial dysplasia (OED), oral submucous fibrosis (OSMF), oral squamous cell carcinoma (OSCC) and normal oral mucosa using methyl green-pyronin staining to determine its suitability for detecting potentially malignant lesions and the stage of carcinogenesis.Methods:Forty-five archival sections of OED, OSMF and OSCC and 5 cases of normal oral mucosa as the control group were stained according to methyl green-pyronin-staining protocol. Cytomorphometric parameters such as nuclear diameter, nucleolar diameter, number of nucleoli and cytoplasmic RNA were assessed.Statistical Analysis Used:The study was subjected to statistical analysis to evaluate the association between morphometric parameters using analysis of variance test, followed by Bonferroni's post hoc analysis.Results:A progressive increase in the nuclear parameters as well as cytoplasmic RNA content was observed between normal mucosa through dysplasia and OSMF to OSCC.Conclusion:This study serves as an effective diagnostic aid in assessing nuclear parameters in potentially malignant and malignant epithelial lesions affecting oral cavity.

Morphometric Analysis of Suprabasal Cell Layer in Oral Epithelial Dysplasia: A Computer-assisted Microscopic Study.

ABSTRACTBackground:Oral leukoplakia is the most common precancerous lesion. Various grading systems based on histological findings have been proposed for dysplasia. Recently, computer-assisted morphometric analysis has been established to reduce the interobserver and intraobserver variability during the histological grading of epithelial dysplasia. This study was undertaken to establish the morphometric changes in the suprabasal cell layer of different grades of oral epithelial dysplasia.Materials and Methods:Forty paraffin-embedded tissue sections (10 normal mucosa, 10 mild dysplasia, 10 moderate dysplasia, and 10 severe dysplasia cases) were stained using hematoxylin and eosin stain, and analyzed for cellular and nuclear morphometry using binocular microscope.Results:Our results showed that values of nuclear area, nuclear perimeter, nuclear volume density, and nuclear/cytoplasmic (N/C) ratio were increased gradually in dysplasia compared to control groups and the values were statistically significant (P = 0.001). Nuclear diameter and cellular area were increased in dysplasia when compared to control group (P = 0.001). Mild and moderate dysplasia showed decreased value of nuclear form factor compared to control group, whereas severe dysplasia showed highest value. A fair correlation was found when comparing histological grading and grouping based on nuclear area, nuclear perimeter, N/C ratio, and nuclear volume density.Conclusion:Nuclear features reflect cell behavior, and its morphometric analysis can be considered as a reliable tool for differentiating various grades of epithelial dysplasia.

Cytological and Cytomorphometric Study of Exfoliated Cells of the Oral Mucosa in Diabetic Patients.

ABSTRACTBackground:Systemic disorder like diabetes mellitus is on the rise in developing countries like India and in many parts of the world. Early diagnosis of this disease will help to prevent the complications due to uncontrolled diabetes mellitus. Dentist helps in the early diagnosis of this disease as patients manifest several oral manifestations.Aims and Objectives:The aim of this study was to study the cytological and cytomorphometric changes of exfoliated cells of oral mucosa in diabetic patients.Materials and Methods:Thirty diabetic patients and 15 healthy controls were included in the study. From the buccal mucosa, smears were collected and Papanicolaou stain was used for staining. Cytological and cytomorphometric study was carried out. The cell diameter and nuclear diameter were measured on these smears and were compared with the control group with the help of “unpaired Student t test.”Results:Buccal mucosa exfoliated cells’ mean nuclear diameter of diabetic group and control group was 11.198 and 9.494 µm, respectively and the difference was highly significant.Conclusion:Our study suggests significant cellular changes in the diabetic patients, which may aid us in early detection of diabetes.

Histological Classification of Thymoma

Background: Many thymoma classifications have been followed and have been updated by newer or alternative schemes. Many classifications were based on the morphology and histogenesis of normal thymus as the backbone, while other classifications have followed a more simplified scheme, whereby thymomas were grouped based on biological behavior. The WHO classification is currently the advocated one, which is based on “organotypical” features (i.e. histological characteristics mimicking those observed in the normal thymus) including cytoarchitecture (encapsulation and a “lobular architecture”) and the cellular composition, mostly the nuclear morphology is generally appreciated.
 Objectives: This study aims to re-classify thymomas by establishing certain morphometric parameters to evaluate the epithelial cells nuclei. An appraisal of thymoma classification as cortical/ lymphocytic/ type B1 and B2 or medullary/ spindle cells/ type A will be attempted as objective re-evaluation of thymoma.
 Patients: This study is a retrospective evaluation of 50 cases of thymoma, 20 cases of thymic hyperplasia and 10 cases of normal thymus (control group). Using a 5 µm formalin-fixed paraffin embeded tissue sections, stained with hematoxylin-eosin stain, these cases were previously classified histologically into lymphocytic (6 cases), lymphoepithelial (mixed) (28 cases) and epithelial (16 cases), including 2 cases of spindle cell thymoma.
 Methods: Computer-assisted morphometry was performed for 80 cases. This involves digitization of the histopathological features and application of morphometric analysis through software. The morphometric parameters used are nuclear area, maximum nuclear diameter and Form factor of epithelial cells nuclei. Ten normal thymus glands from the archived cases were also evaluated as a control groups.
 Results: The results showed that epithelial thymomas possess significantly different nuclear areas from that of a normal thymus, the maximum nuclear diameter (D-Max) follows the same pattern and adds no further outcomes. The numerical morphometric analysis showed no significant differences between lymphocytic predominant thymoma and those classified as cortical thymoma (Type B2). Thus it does not support such a re-classification. Form factor is an indication of pleomorphism, but it should be cautiously used when spindle cells are present since it may give a false indication of pleomorphism.
 Conclusion: Computer-Assisted morphometric analysis provides an objective, reproducible and comparable results for thymoma histological classification.
 Keywords: Thymoma, Histological classification, Grading, Morphometry, Computer-assistedmorphometry.

PTEN-Induced Putative Kinase 1 (PINK1)/Parkin-Mediated Mitophagy Protects PC12 Cells Against Cisplatin-Induced Neurotoxicity.

BackgroundThe pathogenesis of chemotherapy-induced neuropathy, a dose-dependent adverse effect of cisplatin, involves mitochondrial dysfunction. PTEN-induced putative kinase 1 (PINK1)/Parkin-mediated mitophagy removes damaged mitochondria under various pathological conditions. The objective of this study was to determine mitophagy status and its effects on mitochondrial function and neuronal cell damage after cisplatin treatment using an in vitro model of cisplatin-induced neurotoxicity.Material/MethodsPC12 cells were transfected with Parkin or Parkin siRNA using lentiviral particles and Lipofectamine 3000™, respectively, and then were exposed to 10 μM cisplatin. The expression of autophagic proteins was measured by Western blot analysis. Mitophagy in PC12 cells was detected by confocal microscopy analysis of mitochondria-lysosomes colocalization and autophagic flux. The effects of PINK1/Parkin-mediated mitophagy on cisplatin-induced neurotoxicity were assessed via mitochondrial function, neuritic length, nuclear diameter, and apoptosis.ResultsCisplatin activated PINK1/Parkin-mediated mitophagy in PC12 cells. Autophagic flux analysis revealed that cisplatin inhibits the late stage of the autophagic process. The knockdown of Parkin suppressed cisplatin-induced mitophagy, aggravating cisplatin-induced depolarization of mitochondria, cellular ATP deficits, reactive oxygen species outburst, neuritic shortening, nuclear diameter reduction, and apoptosis, while Parkin overexpression enhanced mitophagy and reversed these effects.ConclusionsPINK1/Parkin-regulated mitophagy can protect against cisplatin-related neurotoxicity, suggesting therapeutic enhancement of mitophagy as a potential intervention for cisplatin-induced peripheral neuropathies. The interference of cisplatin with autophagosome-lysosome fusion may be partly responsible for cisplatin-induced neurotoxicity.

Comparison of nuclear size in mature and hypermature cataract

Comparison of nuclear size in mature and hypermature cataract - IJCEO- Print ISSN No: - 2395-1443 Online ISSN No:- 2395-1451 Article DOI No:- 10.18231/j.ijceo.2019.103, Indian Journal of Clinical and Experimental Ophthalmology-Indian J Clin Exp Ophthalmol

Cellular Alteration of the Oral Mucosa due to Orthodontic Appliances and Tobacco Smoking: A Cytomorphometric Study

Introduction: Fixed orthodontic treatment causes chronic low grade irritation on the oral mucosa. Tobacco smoking is considered to be a risk factor in the development of oral cancer. This study involves a cytomorphometric analysis on the buccal mucosal cells in patients undergoing fixed orthodontic treatment who are tobacco smokers.
 Methods: Exfoliated buccal mucosal cells were obtained from orthodontic patients (smokers & non-smokers) as well as smokers and non-smokers who are not undergoing any orthodontic treatment. Smears were stained with rapid PAP stain and digital images of 50 cells were analysed using the Image J software.
 Results: Statistical analysis revealed that there were significant increase in the values of nuclear and cell diameter between non-smokers undergoing orthodontic treatment and smokers (p<0.001 & p=0.014 respectively), and between non-smokers undergoing orthodontic treatment and smokers undergoing orthodontic treatment (p<0.001 & p=0.015 respectively). Also, nuclear area and cell area showed significant differences between many of the study groups.
 Conclusion: It can be concluded that there are nuclear and cellular alterations in patients undergoing fixed orthodontic treatment who are smokers

Evaluation of the relation between Lens Opacities Classification System III grading and nuclear size by direct measurement.

CONTEXT:Although relation between Lens Opacities Classification System III (LOCS III) and nuclear density is established, no data are available about nuclear size at different LOCS III grades.AIMS:The aim of this study is to evaluate the relation between LOCS III grading of nuclear opacity obtained preoperatively and the size of the nucleus obtained during cataract surgery.SETTINGS AND DESIGN:This was a prospective observational study carried out in a hospital attached to medical college.MATERIALS AND METHODS:Patients who underwent manual small-incision cataract surgery or extra-large temporal tunnel cataract extraction and gave consent were included in this study. Institutional Ethics Committee clearance was taken for the study. Preoperative LOCS III grading was obtained at slit-lamp biomicroscope. Ocular dimensions were obtained by preoperative immersion biometry. The thickness and diameter of the nucleus obtained by extraction were measured up to 10 μ accuracy. Data were analyzed for the change in nuclear thickness, nuclear diameter, age, lens thickness, and anterior chamber depth in relation to the LOCS III grade of the nucleus.STATISTICAL ANALYSIS USED:Statistical analysis used in this study was one-way ANOVA, mean, and range.RESULTS:There was a significant increase (P < 0.05) in nuclear thickness, nuclear diameter, and age with increasing LOCS III grade of the nucleus. The change in nuclear size was linear between Grades 1 and 4. The nuclear size did not increase between Grades 4 and 5. It increased steeply from Grade 5 to Grade 6.9.CONCLUSION:LOCS III grading of the nucleus can be utilized for determining the nuclear thickness and diameter preoperatively. These data can be helpful in adjusting machine parameters during phacoemulsification.

Actin Accumulates Nesprin-2 at the Front of the Nucleus During Confined Cell Migration

The mechanisms by which cells exert forces on their nuclei to migrate through openings smaller than the nuclear diameter remain unclear. In microfluidic devices, the hourglass shape of the nucleus and its strain patterns as it translocates through narrow constrictions suggest pulling forces. We use CRISPR/Cas9 to label nesprin-2 giant, a protein that links the cytoskeleton to the interior of the nucleus. We demonstrate that nesprin-2 giant accumulates at the front of the nucleus during nuclear deformation through narrow constrictions, independently of the nuclear lamina. We find that nesprins are more mobile than lamin A/C, and hypothesize that nesprin accumulation at the front is a consequence of forward pulling by the cytoskeleton. Using artificial constructs, we show indeed that the actin-binding domain of nesprin-2 is necessary and sufficient to generate this accumulation, and that microtubules are not involved. Actin filaments are organized in a barrel structure around the moving nucleus in the direction of movement. This and estimates of nesprin elongation suggest that actin pulling forces on nesprins are distributed around the deformed nucleus towards the front.

Abstract 2670: Spatial potassium buffering among clustered glioblastoma cells is proposed to enable survival scavenging of nutrients released by interior necrotic cells

Abstract Glioblastoma cells are known for surviving harsh environments and for forming secondary structures of Scherer (cell clusters around neurons/axons, blood vessels, regions of coagulative necrosis, &amp; in the subpia). The astrocytic lineage of glioblastomas suggests recapitulation of astrocytic-type syncytia that buffer potassium released from neurons/axons. Here, multiple glioblastoma cell lines were stressed by nutrient withdrawal to observe cell behavior in view of their astrocytic lineage &amp; known genome sequence of 1 cell line. Methods: Single cell suspensions of 4 glioblastoma cell lines (U87, LN229, T98G &amp; A172) were added to a settling chamber (Neuro Probe, Gaithersburg, MD). After 0.5 hr of gradual nutrient withdrawal, slides (Diff Quik stain) were analyzed, 2 replicate wells per cell line. U87 cells with cytoplasmic vacuoles were quantified to indicate the degeneration in cell clusters. When vacuoles collectively had a diameter at least 1/3 of an ave. nuclear diameter in the same 100X microscopic field, the vacuolated cell was deemed +. Potassium ion channel genes potentially used by glioblastoma cells per the Online Mendelian Inheritance in Man (OMIM) database were checked in genome sequence results for U87 cells (Clark MJ, et al. PLoS Genet 2010 6(1): e10000832). Results: All glioblastoma cell lines displayed cell adherence &amp; wrapping around single swollen, often heavily vacuolated degenerating cells of the same lineage after nutrient withdrawal. As clustered U87 cells (putative syncytia) increased in cell number, the number of constituent cells with vacuoles decreased (Pearson regressions, p=.04, p=.006) in 2 replicate wells. Whereas single cells were +, 72% and 51%, vacuolated cells fell to 18% and 11% in putative syncytia of over 20 cells in 100X microscopic fields across the width (7 mm) of each well. Thus, within increasing-sized putative syncytia, which offer increasing potassium buffering capacity, significantly fewer constituent cells displayed vacuoles. Presumably this was due to available nutrients from one to few necrotic cells in their midst during nutrient withdrawal. Potassium ion channeling within astrocytic-type syncytia, termed "spatial buffering" explains how viable glioblastoma cells adhere to degenerating cells despite high potassium levels. Genome sequencing of U87 cells revealed that only KCNMB2 &amp; KCNJ16 of 47 potassium channeling genes pertinent to brain, were negatively affected by genetic defects. Functional losses of KCNMB2 as an enhancer for KCNMA1 (intact), and KCNJ16 as a co-assembler with other family members (intact), are unlikely to prevent potassium buffering. Preservation of nearly all potassium channeling genes in U87 suggests that maintenance of spatial buffering in astrocytic-type syncytia permits viable cells to surround necrotic cells and salvage released nutrients as a survival strategy. Citation Format: Marie Beckner. Spatial potassium buffering among clustered glioblastoma cells is proposed to enable survival scavenging of nutrients released by interior necrotic cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2670.

The Distribution of Kidney Tumors in Our Hospital, The Relationship Between Pathological Stage, Nuclear Grade and Tumor Diameter: Analysis of 140 Cases

Objective: Most of the renal neoplasms are the epithelial and malign ones. The renal cell carcinoma (RCC), which is the most frequently seen malign renal tumor, consists of 2-4% of all the malign tumors seen among adult individuals and 80-90% of all the malign renal tumors. RCCs peak at sixties and seventies and its prevalence among males is two folds of its prevalence among females. RCCs have bad prognosis. The recurrence is seen in 40% of the cases, and the metastasis despite the early diagnosis was detected in 50% of cases. In present study, it was aimed to distribution of renal tumors in our hospital, discuss the prognostic factors such as the, pathological stage, nuclear grade and tumor diameter accompanied by literature information. Patients and Method: By retrospectively scanning the records of Pathology Department of Medical Faculty, Firat University, for the period between 1988 and 2009, 140 renal tumor cases were determined and the previous diagnoses, histological degrees, pathological stages of these cases were evaluated and the final diagnoses were taken as base in the present study. In pathologically staging (pT) of RCCs, TNM classification was used, whereas Fuhrman nuclear grading (FNG) was used in nuclear grading procedure. The patients were divided into different groups based on their genders and ages. The statistical analyses were performed in SPSS-12.0 software by using Chi-Square and Correlation tests. Results: When compared to the literature in terms of the mean age, the present study involves relatively young population (mean age 57), and the majority was male (male/ female:1.1). The distributions of renal tumours, malign and benign subtypes, and RCC subtypes show similarities with the general literature. When examining the RCCs statistically, no significant relationship was found between diagnosis and age, and between nuclear grade and tumor diameter (p>0.05). In the present study, the RCCs with largest diameter were papillary type (9,78cm) ones, whereas the RCCs with lowest diameter values were classic type. A statistically significant relationship was observed between the benign, borderline and malign tumor diagnoses and diameters (p= 0.023). It was determined that approximately one-four of RCC cases were in highly pathological stage (21,6%, pT3a-3b). A direct proportional and statistically significant relationship was found between nuclear grade and diameter(p=0,002). Conclusion: RCCs have poor prognosis and their frequency is increasing. Pathological stage, nuclear grade and diameter are the leading factors affecting the prognosis. As in our study, in retrospective studies involving old ages and not enough sampling, tumor diameter may be used as an alternative prognostic factor for pathological staging in tumors with a stage above pT2.

PB1690 ACUTE MYELOID LEUKEMIA WITH “CUP‐LIKE” BLASTS – ASSOCIATION WITH NPM1 AND FLT3 MUTATIONS AND IMMUNOPHOTYPIC FEATURES

Background:The diagnosis and classification of acute myeloid leukemias (AMLs) are based on morphologic, cytogenetic and molecular findings. AMLs with “cup‐like” (CL) morphology have been associated with a high frequency of FLT3‐ITD and NPM1 mutations.Aims:The aim of this study was to evaluate the relation of CL morphology with the most important prognostic markers for AML and immunophenotypic features.Methods:74 patients [M/F = 1, median age 60.5 years (17‐88)] consecutively diagnosed with non‐FAB‐M3, M4 or M5 AMLs that had available FLT3 and NPM1 mutational status were included. CL morphology was defined as a dim colored invagination, resembling a cup or pocket shaped, measuring ≥25% of the nuclear diameter. Two independent examiners, after counting 200 blasts on peripheral blood or bone marrow slides, determined the % of CL blasts (discrepant counts were settled with the aid of a third examiner). The results were then compared with clinical, laboratory, immunophenotypic, cytogenetic [G Band Karyotype(KT)] and molecular (FLT3‐ITD and NPM1) findings obtained at diagnosis. CL % were classified into 3 groups: A(&lt;5%), B(5‐9%), C(≥10%). Statistical analysis with T‐Student, X2 and Fisher exact tests were performed.Results:CLs varied from 0‐30%; 63 patients (85%) had &lt;5% (group A); 4 (5,4%) were B and 7 (9%) had ≥10% (C). Group C patients were mostly female (5 of 7). Median white blood counts were higher in group C (69x109/L, range 1.9‐202x109/L) vs B (22.7 × 109/L, range 12‐28.4 × 109/L) vs A (12.8 × 109/L, range 1.2 ‐212 × 109/L) (CxA p = 0.002). Immunophenotyping studies showed that cases with higher CL percentages were less likely CD34 positive: [C (1 of 7) vs B (100%, 4/4) vs A (77%, 48/62) (CxA, p &lt; 0.002)] and that they also tended to be more likely HLA‐DR negative [C (3/7) vs B (4/4) vs A (47/54), (CxA, p = 0.06)]. KTs were normal in 16/45 of group A, 2/4 of group B and in 1/3 of group C. FLT3‐ITD was present in 15% (11/74) of this cohort. These mutations were most frequent in group C [(3/7, 43%) vs. B (1/4, 25%) vs A (7/63, 11%) (CxA p = 0.055)]. NPM1 mutations were identified in 12/70 (17%) of the cases. These mutations were also more frequent in group C [6/7; 85.7%) vs B (0/4) vs A (6/59, 10%), (CxA/B p &lt; 0.001)].Summary/Conclusion:It is important to rapidly identify features related to cytogenetic and molecular abnormalities in patients with AML. Thus, a morphological evaluation is an important step of acute leukemia diagnosis. Previous studies had reported the association of CL morphology with higher blood counts, a more mature immunophenotype (negativity for CD34 and HLA‐DR), normal KT and FLT3‐ITD/NPM1 mutations. Notably in our study, these associations were only statistically significant when CLs were &gt;10% of the blasts, with the exception of normal KT (probably due to the small number of evaluable cases). Curiously, at least in 3 other studies, CL morphology was also more frequent in females. Recent studies using electron‐microscopy have demonstrated mitochondrial accumulation at the CL region which could be related to the pathogenesis of theses AMLs. In summary, identifying CL morphology is not difficult, and our study can validate a method to predict phenotypic features and NPM1 mutations with a high degree of confidence.

The orbit and size-frequency distribution of long period comets observed by Pan-STARRS1

We introduce a new technique to estimate the comet nuclear size frequency distribution (SFD) that combines a cometary activity model with a survey simulation and apply it to 150 long period comets (LPC) detected by the Pan-STARRS1 near-Earth object survey. The debiased LPC size-frequency distribution is in agreement with previous estimates for large comets with nuclear diameter ≳1 km but we measure a significant drop in the SFD slope for small objects with diameters <1 km and approaching only 100 m diameter. Large objects have a slope αbig = 0.72 ± 0.09(stat.) ± 0.15(sys.) while small objects behave as αsmall = 0.07 ± 0.03(stat.) ± 0.09(sys.) where the SFD is ∝10αHN and HN represents the cometary nuclear absolute magnitude. The total number of LPCs that are >1 km diameter and have perihelia q < 10 au is 0.46 ± 0.15 × 109 while there are only 2.4 ± 0.5(stat.) ± 2(sys.) × 109 objects with diameters >100 m due to the shallow slope of the SFD for diameters <1 km. We estimate that the total number of ‘potentially active’ objects with diameters ≥1 km in the Oort cloud, objects that would be defined as LPCs if their perihelia evolved to <10 au, is (1.5 ± 1) × 1012 with a combined mass of 1.3 ± 0.9 M⊕. The debiased LPC orbit distribution is broadly in agreement with expectations from contemporary dynamical models but there are discrepancies that could point towards a future ability to disentangle the relative importance of stellar perturbations and galactic tides in producing the LPC population.

Modeling meiotic chromosome pairing: a tug of war between telomere forces and a pairing-based Brownian ratchet leads to increased pairing fidelity

Meiotic homolog pairing involves associations between homologous DNA regions scattered along the length of a chromosome. When homologs associate, they tend to do so by a processive zippering process, which apparently results from avidity effects. Using a computational model, we show that this avidity-driven processive zippering reduces the selectivity of pairing. When active random forces are applied to telomeres, this drop in selectivity is eliminated in a force-dependent manner. Further simulations suggest that active telomere forces are engaged in a tug-of-war against zippering, which can be interpreted as a Brownian ratchet with a stall force that depends on the dissociation constant of pairing. When perfectly homologous regions of high affinity compete with homeologous regions of lower affinity, the affinity difference can be amplified through this tug of war effect provided the telomere force acts in a range that is strong enough to oppose zippering of homeologs while still permitting zippering of correct homologs. The degree of unzippering depends on the radius of the nucleus, such that complete unzippering of homeologous regions can only take place if the nucleus is large enough to pull the two chromosomes completely apart. A picture of meiotic pairing thus emerges that is fundamentally mechanical in nature, possibly explaining the purpose of active telomere forces, increased nuclear diameter, and the presence of ‘Maverick’ chromosomes in meiosis.

Cytomorphometric Study of Exfoliated Cells of Buccal Mucosa in Type II-Diabetic Patient

India is the country with highest prevalence of diabetes mellitus in the world. Other condition like premalignant lesions and condition, oral exfoliative cytology may be more appropriate as the non-invasive techniques for early diagnosis. So, cytology technique may help to diagnose diabetes disease and help to prevent the complications. Aims: The present study is conducted to evaluate the cytological and cytomorphometric changes of exfoliated cells of buccal mucosa in type-II diabetic patients. Methods and Material: The diabetic group comprised of 30 diabetic patients, the age of 30-60 years suffering from diabetes mellitus at least from one year. Control group comprised of 15 healthy volunteers above the age of 30 years. Smears were taken from the buccal mucosa and stained with papanicolaou stain. The stained smears were measured for the cellular diameter (CD) and nuclear diameter (ND) along the longest axis of the cell using software digital camera with axiovision of version 2.0. Collected data were analyzed using ‘Unpaired student t test’. Results: The results showed the mean nuclear diameter of the exfoliated cells of diabetic group was 11.198µm and that of the control group was 9.494µm and the difference showed very highly significance (P<0.001). The mean cell diameter of the exfoliated cells of diabetic group showed 56.52µm and the control group showed 53.148µm and the difference between these group was significant (P<0.01). Conclusions: This study suggests that there were significant cellular and nuclear changes in the diabetic patients. The knowledge of which can be helpful for the cytopathologist in the early recognition of the cellular changes in the diabetic patients.

Toward early diagnosis of oral cancer: Diagnostic utility of cytomorphological features, a pilot study.

Introduction:Early detection of oral cancer is one of the most efficient ways to reduce the high mortality from this disease because of the ready accessibility of the oral cavity. We need to devise urgent diagnostic tools to detect early oral premalignant and malignant lesions.Aim:The aim of the present study was to grade the oral lesions in an attempt toward developing a novel cytological grading system. Further, morphometric analysis of cellular parameters was also performed to compare their significance in differentiating benign from malignant lesions.Materials and Methods:The present study was conducted at a tertiary care hospital catering to the low socioeconomic population. Patients presenting in the various Out Patient Departments with suspicious oral lesions were evaluated by cytology in the Department of Pathology.Results:A total of 72 patients were evaluated with a mean age of 43.54 ± 10.35 years. The involvement of the buccal mucosa was the most common site of oral lesions. Cytologically, the lesions were graded according to the oral/oro-pharyngeal cytology grading system into grades A to F. Cyto-morphometric analysis showed an increasing trend in mean nuclear diameter from benign to malignant cases while the mean cytoplasmic diameter decreased, value of P < 0.05 was observed indicating a statistically significant difference between the two groups.Conclusions:Cytological features of pleomorphism are a unique feature in oral carcinoma reflecting intracellular alterations in cells. Grading of lesions according to cytological characteristics can be helpful in standardizing the reporting of the oral lesion. However, our study was restricted by limited data; we emphasize more extensive studies to assess the usefulness and applicability of such a grading system. We also conclude that the use of cytomorphometry can improve the diagnostic reliability of exfoliative cytology.

The value of rapid semi-automated morphometric analysis system in predicting malignancy in thyroid follicular neoplasm

Aims: To constuct and evaluate a rapid semiautomated system in discriminating malignant from benign follicular thyroid neoplasms. Methods: Sixty formalin fixed paraffin embeded blocks of follicular thyroidneoplasms were retreived and slides were prepared and H and E stained. A morphometric system was constructed to be as rapid as possible and was tested.Results: No signficant difference was found in ratio of largest nuclear to smallest nuclear diameter between follicualr adenoma and follicualr carcinoma groups.Conclusion: Rapid semiautomated morphometric analysis is of no help in predicting malignancy in follicular thyroid neoplasm.