Grading of ductal breast carcinoma by cytomorphology and image morphometry with histologic correlation

OBJECTIVE: To investigate the relevance of image analysis for grading breast carcinoma. STUDY DESIGN: Twenty-five ductal breast carcinoma cases were chosen randomly from routine fine needle aspiration clinics. The results of cytomorphologic grading and image morphometry were correlated with those of histologic grading. The five image morphometric parameters studied were nuclear diameter, nuclear area, nuclear roundness, nuclear perimeter and grey level to compare with chromatin texture. RESULTS: Cytologic grading alone had a high correlation with histologic grading. The lowest correlation was found in grade 2 tumors. When cytologic grading was supplemented with image morphometric parameters, the correlation was higher than that of cytologic grading alone. CONCLUSION: Cytologic grading has a high correlation with histologic grading. The correlation improves further on supplementation with image morphometric parameters.     

Nitric oxide induces apoptosis in cutaneous T cell lymphoma (HuT-78) by downregulating constitutive NF-κB

Constitutive active NF-κB have been shown to protect cutaneous T cell lymphoma (CTCL) cells from apoptosis. In the present study, we have studied the cytotoxic potential of nitric oxide generating compound, sodium nitroprusside (SNP) on CTCL cell line, HuT-78. Treatment of cells with SNP resulted in decrease in mitochondrial membrane potential, cytochrome c release, activation of caspase-3 and poly (ADP ribose) polymerase cleavage. SNP treatment inhibited activation of NF-κB in a concentration dependent manner. SNP increased the expression of IκBα without affecting the phosphorylation of IκBα. Downregulation of NF-κB by SNP decreased p65 nuclear translocation as evident by confocal fluorescence microscopy. Further it was found that SNP treatment caused downregulation of Bcl-2 family member (Bcl-xl) in HuT-78 cells. Thus, we have provided evidence that SNP induces apoptosis in CTCL cell line, HuT-78 by downregulating constitutive NF-κB and thereby Bcl-xl expression.     

Identification and association of ISSR markers for thermal stress in polyvoltine silkworm Bombyx mori

Evaluation of genetic resources is an essential prerequisite for their effective utilization. In India, the tropical climate prevails in most of the sericultural belts, where temperature goes beyond the ambient during summer, adversely affecting the silkworm rearing. Although polyvoltine silkworms are poor in silk content, they are mostly tolerant to tropical conditions and diseases. With an aim to identify potential silkworm races/breeds specific to thermo-tolerance for their effective utilization in breeding programme, 15 selected polyvoltine silkworm races were studied for their thermo-tolerance behaviour. Their genomic DNA samples were analyzed for ISSR-PCR using 15 selected primers. The UPGMA analysis based on Nei and Li algorithm has clustered the 15 silkworm races into five groups and one isolate. ALSCAL-multidimensional scaling has not only supported the information generated by the dendrogram, but it has made the genetic distance among races more clear and substantiating their status in terms of thermal stress where pupation rate was taken as indicator. Further, discriminant function analysis (DFA) was done with three groups of silkworms classified for thermal stress viz. susceptible, moderately tolerant and tolerant. The canonical correlation value was estimated to be 0.987 (Wilk's lambda = 0.004; chi2 = 36.044, p < 0.05). DFA clearly discriminated the above three groups. Beta statistics with t value and its significance for the markers identified through stepwise multiple regression analysis (MRA) revealed a total of five bands (807(1300), 808(3000), 808(4000), 834(4000), and 834(3000)) showing correlation with pupation rate after thermal treatment. Out of them, marker 8083000 showed maximum and highly significant correlation (r = 0.757, p < 0.001, t = 4.182) with pupation rate among the silkworm races. The identified putative markers are being used to develop DNA marker to be used in evolving thermo-tolerant silkworm breeds using marker assisted selection programme.     

Cytomorphologic and morphometric limitations of the assessment of atypia in fibroadenoma of the breast

OBJECTIVE: To investigate the relevance of nuclear morphometry in separating the categories of "fibroadenoma" and "fibroadenoma with atypia." STUDY DESIGN: Thirty consecutive breast lumps, on which a fine needle aspiration (FNA) diagnosis of fibroadenoma was followed by excision and histopathologic confirmation of the diagnosis, were included. Atypia on cytology was based on cell overlap, nuclear enlargement and cell dyscohesion. Nuclear morphometric comparison was carried out between the categories of fibroadenoma, fibroadenoma with atypia and grade 1 ductal carcinoma cases that formed part of an earlier study. The parameters employed were area, roundness, diameter, perimeter and grey level. RESULTS: Among the 30 cases of fibroadenoma reported on FNA, an additional component of atypia was noted in 5. On subsequent histopathology, atypia was not confirmed in any of the cases. On morphometric analysis, a significant difference was noted between fibroadenoma and fibroadenoma with atypia categories, as between fibroadenoma and grade 1 ductal carcinoma cases. However, no significant difference was noted between fibroadenoma with atypia and grade 1 ductal carcinoma cases. CONCLUSION: FNA assessment of atypia in cases of fibroadenoma is difficult. Even conventional nuclear morphometry, though supporting the initial impression of atypia, does not help with this assessment. Also, based on morphometry alone, there may be difficulty separating fibroadenomas with atypia and grade 1 ductal carcinomas. Larger studies, employing other morphometric parameters, such as chromatin texture and fractal dimension, may shed further light on the subject.     

Exploitation of marine bacteria for production of gold nanoparticles

Background

Gold nanoparticles (AuNPs) have found wide range of applications in electronics, biomedical engineering, and chemistry owing to their exceptional opto-electrical properties. Biological synthesis of gold nanoparticles by using plant extracts and microbes have received profound interest in recent times owing to their potential to produce nanoparticles with varied shape, size and morphology. Marine microorganisms are unique to tolerate high salt concentration and can evade toxicity of different metal ions. However, these marine microbes are not sufficiently explored for their capability of metal nanoparticle synthesis. Although, marine water is one of the richest sources of gold in the nature, however, there is no significant publication regarding utilization of marine micro-organisms to produce gold nanoparticles. Therefore, there might be a possibility of exploring marine bacteria as nanofactories for AuNP biosynthesis.

Results

In the present study, marine bacteria are exploited towards their capability of gold nanoparticles (AuNPs) production. Stable, monodisperse AuNP formation with around 10?nm dimension occur upon exposure of HAuCl₄ solution to whole cells of a novel strain of Marinobacter pelagius, as characterized by polyphasic taxonomy. Nanoparticles synthesized are characterized by Transmission electron microscopy, Dynamic light scattering and UV-visible spectroscopy.

Conclusion

The potential of marine organisms in biosynthesis of AuNPs are still relatively unexplored. Although, there are few reports of gold nanoparticles production using marine sponges and sea weeds however, there is no report on the production of gold nanoparticles using marine bacteria. The present work highlighted the possibility of using the marine bacterial strain of Marinobacter pelagius to achieve a fast rate of nanoparticles synthesis which may be of high interest for future process development of AuNPs. This is the first report of AuNP synthesis by marine bacteria.     

Genetic Diversity and Population Structure of Indian Golden Silkmoth (Antheraea assama)

Background

The Indian golden saturniid silkmoth (Antheraea assama), popularly known as muga silkmoth, is a semi-domesticated silk producing insect confined to a narrow habitat range of the northeastern region of India. Owing to the prevailing socio-political problems, the muga silkworm habitats in the northeastern region have not been accessible hampering the phylogeography studies of this rare silkmoth. Recently, we have been successful in our attempt to collect muga cocoon samples, although to a limited extent, from their natural habitats. Out of 87 microsatellite markers developed previously for A. assama, 13 informative markers were employed to genotype 97 individuals from six populations and analyzed their population structure and genetic variation.

Methodology/Principal Findings

We observed highly significant genetic diversity in one of the populations (WWS-1, a population derived from West Garo Hills region of Meghalaya state). Further analysis with and without WWS-1 population revealed that dramatic genetic differentiation (global F_ST = 0.301) was due to high genetic diversity contributed by WWS-1 population. Analysis of the remaining five populations (excluding WWS-1) showed a marked reduction in the number of alleles at all the employed loci. Structure analysis showed the presence of only two clusters: one formed by WWS-1 population and the other included the remaining five populations, inferring that there is no significant genetic diversity within and between these five populations, and suggesting that these five populations are probably derived from a single population. Patterns of recent population bottlenecks were not evident in any of the six populations studied.

Conclusions/Significance

A. assama inhabiting the WWS-1 region revealed very high genetic diversity, and was genetically divergent from the five populations studied. The efforts should be continued to identify and study such populations from this region as well as other muga silkworm habitats. The information generated will be very useful in conservation of dwindling muga culture in Northeast India.     

Design, synthesis and biological activity of sphingosine kinase 2 selective inhibitors

Sphingosine kinase (SphK) has emerged as an attractive target for cancer therapeutics due to its role in cell survival. SphK phosphorylates sphingosine to form sphingosine 1-phosphate (S1P), which has been implicated in cancer growth and survival. SphK exists as two different isotypes, namely SphK1 and SphK2, which play different roles inside the cell. In this report, we describe SphK inhibitors based on the immunomodulatory drug, FTY720, which is phosphorylated by SphK2 to generate a S1P mimic. Structural modification of FTY720 provided a template for synthesizing new inhibitors. A diversity-oriented synthesis generated a library of SphK inhibitors with a novel scaffold and headgroup. We have discovered subtype selective inhibitors with K(i)'s in the low micromolar range. This is the first report describing quaternary ammonium salts as SphK inhibitors.     

Association of A/T rich microsatellites with responses to artificial selection for larval developmental duration in the silkworm Bombyx mori

Simple sequence repeats (SSRs) and interSSR (ISSR) marker systems were used in this study to reveal genetic changes induced by artificial selection for short/long larval duration in the tropical strain Nistari of the silkworm Bombyx mori. Artificial selection separated longer larval duration (LLD) (29.428 (+/-) 0.723 days) and shorter larval duration (SLD) (22.573 (+/-) 0.839 days) lines from a base, inbred population of Nistari (larval span of 23.143 (+/-) 0.35 days). SSR polymorphism was observed between the LLD and SLD lines at one microsatellite locus, Bmsat106 (CA7) and at two loci of 1074 bp and 823 bp generated with the ISSR primer UBC873. Each of these loci was present only in the LLD line. The loci segregated in the third generation of selection and were fixed in opposite directions. In the F2 generation of the LLD x SLD lines, the alleles of Bmsat106 and UBC8731074bp segregated in a 1:1 ratio and the loci were present only in the LLD individuals. UBC873823bp was homozygous. Single factor ANOVA showed a significant association between the segregating loci and longer larval duration. Together, the two alleles contributed to an 18% increase in larval duration. The nucleotide sequences of the UBC8731074bp and UBC873823bp loci had 67% A/T content and consisted of direct, reverse, complementary and palindromic repeats. The repeats appeared to be "nested" (59%) in larger repeats or as clustered elements adjacent to other repeats. Of 203 microsatellites identified, dinucleotides (67.8%) predominated and were rich in A/T and T/A motifs. The sequences of the UBC8731074bp and UBC873823bp loci showed similarity (E = 0.0) to contigs located in Scaffold 010774 and Scaffold 000139, respectively, of the B. mori genome. BLASTN analysis of the UBC8731074bp sequence showed significant homology of (nt.) 45-122 with upstream region of three exons from Bombyx. The complete sequence of this locus showed approximately 49% nucleotide conservation with transposon 412 of Drosophila melanogaster and the Ikirara insertions of Anopheles gambiae. The A + T richness and lack of coding potential of these small loci, and their absence in the SLD line, reflect the active process of genetic change associated with the switch to short larval duration as an adaptation to the tropics.     

COBioSIFTER – A CORBA-Based Distributed Multi-Agent Biological Information Management System

In this paper, we describe the COBioSIFTER project that aims at constructing a framework for managing information from the biological domain. It uses a distributed multi-agent approach based on the CORBA middleware and contains an architecture made up of functionally heterogeneous agents. A series of experiments has been carried out with a prototype of COBioSIFTER to illustrate its advantages such as high performance, scalability, heterogeneity, and fault tolerance. The results of these experiments demonstrate the effectiveness of such an agent-based biological information framework.     

Cell blocks from scraping of cytology smear: comparison with conventional cell block

OBJECTIVE: To compare cytomorphology preservation and immunohistochemistry results between conventional cell blocks (CCB) and cytoscrape cell blocks (SCB). STUDY DESIGN: Fine needle aspiration (FNAC) was done in 17 consecutive cases. Air-dried smears for May-Grünwald-Giemsa stain and wet-fixed smear for hematoxylin-eosin (H-E) stain were prepared. Simultaneously another pass was made in each case for preparation of material for CCB. One of the H-E-stained smears was spared for SCB. SCB was compared with CCB for cell morphology. Immunostaining was performed both cell blocks, as well as on FNA smears in 8 cases. Results were evaluated for intensity of staining and percentage of cells showing positivity. RESULTS: CCB and SCB sections showed adequate cellularity in all cases. Morphologic preservation was good in SCB sections. There was good architectural and nuclear preservation in all cases of SCB. Immunostaining results showed better and clear intensity of staining with little background in all cell block cases. CONCLUSION: SCB is a valuable technique in cell blocks from stained FNA smears. The cytomorphologic details are equally good in SCB and CCB. Additional panels of immunostaining can be done on SCB for better diagnosis and classification, particularly in cases in which repeat FNA is not possible.