Studies on the Santa Lucia (El Salvador) strain of Plasmodium falciparum in Aotus trivirgatus monkeys.

The Santa Lucia strain of Plasmodium falciparum was isolated from El Salvador, Central America, and established in Aotus trivirgatus monkeys. Transmission from monkey to monkey via the bites of infected Anopheles freeborni, A. maculatus, and A, albimanus mosquitoes was obtained in 20 of 27 attempts. Prepatent periods in the monkeys ranged from 17 to 46 days with a mean of 24.3 days. Parasitemias and mortality were higher following sporozoite inoculation into animals which had been previously infected with P. vivax than in those with no previous malaria experience. Monkeys previously infected with P. vivax and P. cynomolgi had lower maximum parasitemias than those previously infected with P. vivax only.     

Development of acetosyringone-inducible Gateway® and Golden Gate expression vectors for heterologous gene expression in Agrobacterium tumefaciens

In Vitro Cellular & Developmental Biology - Plant - As a plant genetic engineer, Agrobacterium tumefaciens utilizes phenolic compounds, such as acetosyringone, to activate its virulence genes...     

Mouse mast cell secretory granules can function as intracellular ionic oscillators

Fluorescent Ca2+ probes and digital photo-sectioning techniques were used to directly study the dynamics of Ca2+ in isolated mast cell granules of normal (CB/J) and beige (Bg(j)/Bg(j)) mice. The resting intraluminal free Ca2+ concentration ([Ca2+]L) is 25 +/- 4.2 microM (mean +/- SD, n = 68). Exposure to 3 microM inositol 1,4,5-trisphosphate (InsP3) induced periodic oscillations of luminal Ca2+ ([Ca2+]L) of approximately 10 microM amplitude and a period around 8-10 s. The [Ca2+]L oscillations were accompanied by a corresponding oscillatory release of [Ca2+]L to the extraluminal space. Control experiments using ruthenium red (2 microM) and thapsigargin (100 nM) ruled out artifacts derived from the eventual presence of mitochondria or endoplasmic reticulum in the isolated granule preparation. Oscillations of [Ca2+]L and Ca2+ release result from a Ca2+/K+ exchange process whereby bound Ca is displaced from the heparin polyanionic matrix by inflow of K+ into the granular lumen via an apamin-sensitive Ca2+-sensitive K+ channel (ASK(Ca)), whereas Ca2+ release takes place via an InsP3-receptor-Ca2+ (InsP3-R) channel. These results are consistent with previous observations of [Ca2+]L oscillations and release in/from the endoplasmic reticulum and mucin granules, and suggest that a highly conserved common mechanism might be responsible for [Ca2+]L oscillations and quantal periodic Ca2+ release in/from intracellular Ca2+ storage compartments.     

The positive impact of cytological specimens for EGFR mutation testing in non‐small cell lung cancer: a single South East Asian laboratory’s analysis of 670 cases

B. Pang, D. Matthias, C.W. Ong, A.N. Dhewar, S. Gupta, G.L. Lim, M.‐E. Nga, J.E. Seet, A. Qasim, T.‐M. Chin, R. Soo, R. Soong and M. Salto‐Tellez The positive impact of cytological specimens for EGFR mutation testing in non‐small cell lung cancer: a single South East Asian laboratory’s analysis of 670 cases Objectives: To compare the rejection rates of non‐small cell lung cancer (NSCLC) samples obtained by differing sampling methods for testing by Sanger sequencing for epidermal growth factor receptor (EGFR) mutations. To assess the association between unsatisfactory outcomes and the quantity of DNA extracted from cytological versus histological samples. Methods: Six hundred and seventy NSCLC samples referred to our centre from 2008 to 2010 were reviewed as a consequence of sample rejection, presence of EGFR mutations, cytological versus histological sampling methods, DNA quantity and the unsatisfactory genotyping rate. Results: Eighty samples were rejected for testing in similar proportions of histological and cytological samples (11.9% versus 10.9%) usually (n = 75) because the amount of cellular material was judged insufficient in small biopsies or cytology samples. The remaining 590 samples on which EGFR testing was attempted yielded 51 (8.6%) unsatisfactory test outcomes caused by failure of the polymerase chain reaction (PCR) (n = 47 cases), uninterpretable Sanger chromatograms (n = 3 cases) and insufficient DNA extracted for PCR (n = 1 case). The difference in rates of unsatisfactory outcomes between cytological samples (seven of 147 samples or 4.7%) versus tissue samples (44 of 443 samples or 9.9%) was clinically relevant but not statistically significant (Mann–Whitney test; P < 0.081). There was no association between the concentration of DNA extracted and the likelihood of an unsatisfactory analysis; which was similar in all types of sections (large and small) while 0% of 37 cytology slides were unsatisfactory. Conclusions: Utilizing cytology samples for EGFR testing avoids unnecessary patient re‐biopsing and yields a clinically superior satisfactory rate to the overall satisfactory rate of tissue biopsies of NSCLC. The quality rather than quantity of DNA extracted may be a more important determinant of a satisfactory result.     

Airway gene transfer using cationic emulsion as a mucosal gene carrier

BACKGROUND: Delivery of genes to airway mucosa would be a very valuable method for gene therapy and vaccination. However, there have been few reports on suitable gene delivery systems for administration. In this study, we use a cationic emulsion system, which is physically stable and facilitates the transfer of genes in the presence of up to 90% serum, as a mucosal gene carrier. METHODS AND RESULTS: Cationic lipid emulsion was formulated with squalene and 1,2-dioleoyl-sn-glycero-3-trimethylammoniumpropane (DOTAP) as major components. Emulsions formed stable complexes with DNA and protected and transferred DNA to target cells against DNase I digestion in the presence of mucosal destabilizers such as heparin sulfate (a polysaccharide of the glycosaminoglycan family in mucosa) and Newfectan (a natural lung extract of bovine) in an in vitro system. In contrast, commercial liposomes and counter liposomes, made with an identical lipid composition of emulsions, failed. After in vivo intranasal instillation, the cationic emulsion showed at least 200 times better transfection activity than the liposomal carriers in both nasal tissue and lung. CONCLUSIONS: These findings show that cationic emulsions can mediate gene transfection into airway epithelium, making it a good choice for transferring therapeutic genes and for genetic vaccination against an pathogenic infection via an airway route.