Detection of suspicious cells and rejection of artefacts in cervical cytology using the Leyden Television Analysis System.

In slide based automation of cervical cytology the first stage of analysis involves finding possibly suspicious cells, or areas on the slide with these types of cells. By using a television based system such as the Leyden Television Analysis System (LEYTAS), a number of detection methods can be applied to rapidly screen a large number of fields automatically for suspicious cells. In this paper, results using a parameter based on increased nuclear DNA content of cells are given and a second detection method based on a chromatin pattern feature, called chromatin contrast, is discussed. Two blind trials on 41 positive and 22 negative cervical slides, using the Leyden Television Analysis System to detect suspicious cells with an increased nuclear DNA content, were promising. In 1 of the 41 positive cases no suspicious cells were found. In the negative specimens, suspicious cells were detected in 1 of 9 cases and 1 of 13 cases, with the two detection parameters investigated. These findings are discussed and some automatic artefact rejection procedures with preliminary results are given.     

Synthesis,structure, antimycobacterial and anticancer evaluation of new pyrrolo-phenanthroline derivatives

A study concerning design, synthesis, structure and in vitro antimycobacterial and anticancer evaluation of new fused derivatives with pyrrolo[2,1-c][4,7]phenanthroline skeleton is described. The strategy adopted for synthesis involves a [3?+?2] dipolar cycloaddition of several in situ generated 4,7-phenanthrolin-4-ium ylides to different substituted alkynes and alkenes. Stereo- and regiochemistry of cycloaddition reactions were discussed. The structure of the new compounds was proven unambiguously, single-crystal X-ray diffraction studies including. The antimycobacterial and anticancer activity of a selection of new synthesized compounds was evaluated against Mycobacterium tuberculosis H37Rv under aerobic conditions and 60 human tumour cell line panel, respectively. Five of the tested compounds possess a moderate antimycobacterial activity, while two of the compounds have a significant antitumor activity against renal cancer and breast cancer.     

Exploring the role of microRNAs in axolotl regeneration

The axolotl, Ambystoma mexicanum, is used extensively for research in developmental biology, particularly for its ability to regenerate and restore lost organs, including in the nervous system, to full functionality. Regeneration in mammals typically depends on the healing process and scar formation with limited replacement of lost tissue. Other organisms, such as spiny mice (Acomys cahirinus), salamanders, and zebrafish, are able to regenerate some damaged body components. Blastema is a tissue that is formed after tissue injury in such organisms and is composed of progenitor cells or dedifferentiated cells that differentiate into various cell types during regeneration. Thus, identifying the molecules responsible for initiation of blastema formation is an important aspect for understanding regeneration. Introns, a major source of noncoding RNAs (ncRNAs), have characteristic sizes in the axolotl, particularly in genes associated with development. These ncRNAs, particularly microRNAs (miRNAs), exhibit dynamic regulation during regeneration. These miRNAs play an essential role in timing and control of gene expression to order and organize processes necessary for blastema creation. Master keys or molecules that underlie the remarkable regenerative abilities of the axolotl remain to be fully explored and exploited. Further and ongoing research on regeneration promises new knowledge that may allow improved repair and renewal of human tissues.     

Nucleosides,XL1 Synthesis and Properties of lin-Naphthamidazole-Ribonucleosides

Abstract

The fusion reaction between 1-trimethylsilyl-naphth[2,3-d]imidazole (3) and its 2-methyl derivative (4) with 2, 3, 5-tri-O-benzoyl-1-bromo-D-ribofuranose (6) leads to anomeric mixtures of the corresponding 2', 3', 5'-tri-O-benzoyl-1α- and β-D-ribofuranosylnaphth[2,3-d]imidazoles (7, 11 and 13). Separation of the anomers was achieved by chromatographical means and debenzoylation yielded the corresponding nucleosides (8, 12 and 10, 14). Structural proofs are based on elementary analysis, UV- and ¹H-NMR spectra.     

Anopheles gambiae cadherin AgCad1 binds the Cry4Ba toxin of Bacillus thuringiensis israelensis and a fragment of AgCad1 synergizes toxicity

A midgut cadherin AgCad1 cDNA was cloned from Anopheles gambiae larvae and analyzed for its possible role as a receptor for the Cry4Ba toxin of Bacillus thuringiensis strain israelensis. The AgCad1 cadherin encodes a putative 1735-residue protein organized into an extracellular region of 11 cadherin repeats (CR) and a membrane-proximal extracellular domain (MPED). AgCad1 mRNA was detected in midgut of larvae by polymerase chain reaction (PCR). The AgCad1 protein was localized, by immunochemistry of sectioned larvae, predominately to the microvilli in posterior midgut. The localization of Cry4Ba binding was determined by the same technique, and toxin bound microvilli in posterior midgut. The AgCad1 protein was present in brush border membrane fractions prepared from larvae, and Cry4Ba toxin bound the same-sized protein on blots of those fractions. The AgCad1 protein was expressed transiently in Drosophila melanogaster Schneider 2 (S2) cells. 125I-Cry4Ba toxin bound AgCad1 from S2 cells in a competitive manner. Cry4Ba bound to beads extracted 200 kDa AgCad1 and a 29 kDa fragment of AgCad1 from S2 cells. A peptide containing the AgCad1 region proximal to the cell (CR11-MPED) was expressed in Escherichia coli. Although Cry4Ba showed limited binding to CR11-MPED, the peptide synergized the toxicity of Cry4Ba to larvae. AgCad1 in the larval brush border is a binding protein for Cry4Ba toxin. On the basis of binding results and CR11-MPED synergism of Cry4Ba toxicity, AgCad1 is probably a Cry4Ba receptor.     

Expression analysis of the nucleocytoplasmic lectin 'Orysata' from rice in Pichia pastoris

The Oryza sativa lectin, abbreviated Orysata, is a mannose-specific, jacalin-related lectin expressed in rice plants after exposure to certain stress conditions. Expression of a fusion construct containing the rice lectin sequence linked to enhanced green fluorescent protein in Bright Yellow 2 tobacco cells revealed that Orysata is located in the nucleus and the cytoplasm of the plant cell, indicating that it belongs to the class of nucleocytoplasmic jacalin-related lectins. Since the expression level of Orysata in rice tissues is very low the lectin was expressed in the methylotrophic yeast Pichia pastoris with the Saccharomyces α-factor sequence to direct the recombinant protein into the secretory pathway and express the protein into the medium. Approximately 12 mg of recombinant lectin was purified per liter medium. SDS/PAGE and western blot analysis showed that the recombinant lectin exists in two molecular forms. Far western blot analysis revealed that the 23 kDa lectin polypeptide contains an N-glycan which is absent in the 18.5 kDa polypeptide. Characterization of the glycans present in the recombinant Orysata revealed high-mannose structures, Man9-11 glycans being the most abundant. Glycan array analysis showed that Orysata interacts with high-mannose as well as with more complex N-glycan structures. Orysata has potent anti-human immunodeficiency virus and anti-respiratory syncytial virus activity in cell culture compared with other jacalin-related lectins.     

Morphology of the stomach of the tropical house gecko Hemidactylus mabouia (Squamata: Gekkonidae)

Rodrigues Sartori, S. S., Nogueira, K. O. P. C., Rocha, A. S. and Neves, C. A. 2011. Morphology of the stomach of the tropical house gecko Hemidactylus mabouia (Squamata: Gekkonidae). —Acta Zoologica (Stockholm) 92 : 179–186. Hemidactylus mabouia is a common species in Brazil, which facilitates its use in research in several areas and allows display it as a benchmark for studies with reptiles. To study the morphology of the stomach of H. mabouia, we carried out anatomical, histological and histochemical analysis. The stomach of H. mabouia is ‘J’ shaped and can be divided into oral fundic (OF), aboral fundic (AF) and pyloric regions. The surface epithelium is composed of mucosecretory cells (MC) containing neutral mucins. In the lamina propria of the OF region, are large ramified tubulo‐acinar glands, which become smaller, less ramified and more tubular towards the AF region, and are simple tubular and short in the pyloric region. The fundic glands contain differentiated neck and pit. The neck is composed of MC containing neutral mucins and the pit is made of oxyntopeptic cells (OC). The OC of the OF region contained many zymogen granules, while those of the AF region contained few zymogen granules and many mitochondria, which suggests the existence of a gradient of pepsinogen and hydrochloric acid secretion. Pyloric glands consisted of MC containing neutral mucins and both argyrophil and argentaffin endocrine cells.     

High-Copy Suppressor Analysis Reveals a Physical Interaction between Sec34p and Sec35p, a Protein Implicated in Vesicle Docking

A temperature-sensitive mutant, sec34-2, is defective in the late stages of endoplasmic reticulum (ER)-to-Golgi transport. A high-copy suppressor screen that uses the sec34-2 mutant has resulted in the identification of the SEC34 structural gene and a novel gene called GRP1. GRP1 encodes a previously unidentified hydrophilic yeast protein related to the mammalian Golgi protein golgin-160. Although GRP1 is not essential for growth, the grp1Delta mutation displays synthetic lethal interactions with several mutations that result in ER accumulation and a block in the late stages of ER-to-Golgi transport, but not with those that block the budding of vesicles from the ER. Our findings suggest that Grp1p may facilitate membrane traffic indirectly, possibly by maintaining Golgi function. In an effort to identify genes whose products physically interact with Sec34p, we also tested the ability of overexpressed SEC34 to suppress known secretory mutations that block vesicular traffic between the ER and the Golgi. This screen revealed that SEC34 specifically suppresses sec35-1. SEC34 encodes a hydrophilic protein of approximately 100 kDa. Like Sec35p, which has been implicated in the tethering of ER-derived vesicles to the Golgi, Sec34p is predominantly soluble. Sec34p and Sec35p stably associate with each other to form a multiprotein complex of approximately 480 kDa. These data indicate that Sec34p acts in conjunction with Sec35p to mediate a common step in vesicular traffic.