Ustilago maydis Mating Hyphae Orient Their Growth toward Pheromone Sources

Snetselaar, K. M., Bolker, M., and Kahmann, R. 1996. Ustilago maydis mating hyphae orient their growth toward pheromone sources. Fungal Genetics and Biology 20, 299-312. When small drops of Ustilago maydis sporidia were placed 100-200 μm apart on agar surfaces and covered with paraffin oil, sporidia from one drop formed thin hyphae that grew in a zig-zag fashion toward the other drop if it contained sporidia making the appropriate pheromone. For example, a2b2 mating hyphae grew toward a1b1 and a1b2 mating hyphae, and the filaments eventually fused tip to tip. Time-lapse photography indicated that the mating hyphae can rapidly change orientation in response to nearby compatible sporidia. When exposed to pheromone produced by cells in an adjacent drop, haploid sporidia with the a2 allele began elongating before sporidia with the a1 allele. Sporidia without functional pheromone genes responded to pheromone although they did not induce a response, and sporidia without pheromone receptors induced formation of mating hyphae although they did not form mating hyphae. Diploid sporidia heterozygous at b but not at a formed straight, rigid, aerial filaments when exposed to pheromone produced by the appropriate haploid sporidia. Again, the a2a2b1b2 strain formed filaments more quickly than the a1a1b1b2 strain. Taken together, these results suggest that the a2 pheromone diffuses less readily or is degraded more quickly than the a1 pheromone.     

Probits of mixtures

The tolerances of individuals (insects, parasites) in a population have a frequency or probability distribution called a tolerance distribution. Many tolerance distributions in bioassay studies can be the result of a rather heterogeneous population of individuals and can often be modelled as a mixture of a number of standard unimodal distributions. A probit analysis can be generalized to the case where the tolerance distribution is a mixture of location and scale parameter distributions. In this article, the existence and determination of the maximum likelihood estimates are investigated. An expectation-maximization (EM) algorithm for probits of mixtures is developed and it is shown that by application of the EM algorithm, the problem of probits of mixtures can be separated into a series of probits of individual component tolerance distributions.     

Micro-neutralization test for mumps virus using the 96-well tissue culture plate and PAP (peroxidase-antiperoxidase) staining technique

A rapid micro-test method for mumps virus neutralization was developed. In this method, a 96-well tissue culture plate was used for preparation of cell monolayers and the PAP staining technique was used for visualization of mumps virus infected cells. Clusters of infected cells were observed as a focus and the numbers of foci could be counted by the naked eye 2 days after the infection. A linear relationship between virus dilutions and focus numbers was observed. When neutralizing antibodies in sera from cases of natural mumps infection were assayed, a good correlation was observed between those obtained by the focus reduction method applying the micromethod and those obtained by the ordinary plaque method. Our results indicate that this micromethod is useful in mumps virus neutralization tests and it has many advantages over other methods previously reported.     

The association between frequency of thiabendazole treatment and the development of resistance in field isolates of Ostertagia spp. of sheep

Martin P. J., Anderson N., Lwin T., Nelson G. and Morgan T. E. 1984. The association between frequency of thiabendazole treatment and the development of resistance in field isolates of Ostertagia spp. of sheep. International Journal for Parasitology14: 177–181. Isolates of Ostertagia spp. were obtained from grazing sheep 3,4 and 5 years after nil, planned (five per year) and regular (3-weekly) treatments with thiabendazole (TBZ). Levels of resistance to TBZ were measured by an in vitro egg hatch assay and a controlled anthelmintic efficiency assay. Isolates from planned treatment groups showed an increase in the level of resistance; the lethal concentrations of TBZ to 50% of eggs (LC₅₀s) were 3, 3 and 6 times the LC₅₀s of isolates from nil treatment groups for years 3, 4 and 5 respectively. The LC₅₀s of isolates from regular treatment groups were 14 times higher than those from nil treatment groups in each year. To assess the potential for an increase in level of resistance, additional egg assays were done 14 days after treatment with 44 mg kg^t?1 of TBZ on sheep infected with the planned group isolates for each year. This treatment raised the LC₅₀S for years 3, 4 and 5 respectively by 3, 2 and 1–5 times the LC₅₀s of the same isolates which had not been exposed to additional TBZ treatment. The controlled anthelmintic efficiency assay using 44 mg kg^t?1 of TBZ produced a significant reduction in the number of adult and immature worms from the nil isolate but failed significantly to reduce the number of worms from the planned and regular isolates. A three component analysis resolved the nonlinear trends of the log dose-probit plots in egg hatch assays for isolates from planned treatment groups into subpopulations of susceptible, hybrid and resistant individuals each with different levels of resistance. The proportions of these subpopulations changed in accordance with the level of resistance observed in each year.     

Influence of shape-memory stent grafts on local aortic compliance

The effect of repair techniques on the biomechanics of the aorta is poorly understood, resulting in significant levels of postoperative complications for patients worldwide. This study presents a computational analysis of the influence of Nitinol-based devices on the biomechanical performance of a healthy patient-specific human aorta. Simulations reveal that Nitinol stent-grafts stretch the artery wall so that collagen is stretched to a straightened high-stiffness configuration. The high-compliance regime (HCR) associated with low diastolic lumen pressure is eliminated, and the artery operates in a low-compliance regime (LCR) throughout the entire cardiac cycle. The slope of the lumen pressure–area curve for the LCR post-implantation is almost identical to that of the native vessel during systole. This negligible change from the native LCR slope occurs because the stent-graft increases its diameter from the crimped configuration during deployment so that it reaches a low-stiffness unloading plateau. The effective radial stiffness of the implant along this unloading plateau is negligible compared to the stiffness of the artery wall. Provided the Nitinol device unloads sufficiently during deployment to the unloading plateau, the degree of oversizing has a negligible effect on the pressure–area response of the vessel, as each device exerts approximately the same radial force, the slope of which is negligible compared to the LCR slope of the native artery. We show that 10% oversizing based on the observed diastolic diameter in the mid descending thoracic aorta results in a complete loss of contact between the device and the wall during systole, which could lead to an endoleak and stent migration. 20% oversizing reaches the Dacron enforced area limit (DEAL) during the pulse pressure and results in an effective zero-compliance in the later portion of systole.

     

Osteoclasts in Multiple Myeloma Are Derived from Gr-1+CD11b+Myeloid-Derived Suppressor Cells

Osteoclasts play a key role in the development of cancer-associated osteolytic lesions. The number and activity of osteoclasts are often enhanced by tumors. However, the origin of osteoclasts is unknown. Myeloid-derived suppressor cells (MDSCs) are one of the pre-metastatic niche components that are induced to expand by tumor cells. Here we show that the MDSCs can differentiate into mature and functional osteoclasts in vitro and in vivo. Inoculation of 5TGM1-GFP myeloma cells into C57BL6/KaLwRij mice led to a significant expansion of MDSCs in blood, spleen, and bone marrow over time. When grown in osteoclastogenic media in vitro, MDSCs from tumor-challenged mice displayed 14 times greater potential to differentiate into mature and functional osteoclasts than those from non-tumor controls. Importantly, MDSCs from tumor-challenged LacZ transgenic mice differentiated into LacZ+osteoclasts in vivo. Furthermore, a significant increase in tumor burden and bone loss accompanied by increased number of osteoclasts was observed in mice co-inoculated with tumor-challenged MDSCs and 5TGM1 cells compared to the control animals received 5TGM1 cells alone. Finally, treatment of MDSCs from myeloma-challenged mice with Zoledronic acid (ZA), a potent inhibitor of bone resorption, inhibited the number of osteoclasts formed in MDSC cultures and the expansion of MDSCs and bone lesions in mice. Collectively, these data provide in vitro and in vivo evidence that tumor-induced MDSCs exacerbate cancer-associated bone destruction by directly serving as osteoclast precursors.     

Effect of Compounds from Moringa oleifera Lam. on in Vitro Non-Alcoholic Fatty Liver Disease (NAFLD) Model System

Non-alcoholic fatty liver disease (NAFLD) is currently the most common chronic liver disease in the world, with a prevalence of 25 % in many countries. To date, no drug has been approved to treat NAFLD, therefore, the use of phytochemicals to prevent this disease is meaningful. In this study, we focused on the effects of Moringa oleifera Lam. on diabetes, attempted to isolate compounds that regulate NAFLD. Compounds 1 and 2 were isolated from the ethyl acetate fraction of M. oleifera. Spectral data revealed that they were 1-hydroxy-3-phenylpropan-2-yl benzoate ( 1 ) and benzyl benzylcarbamate ( 2 ), respectively. The three-dimensional structure of compound 1 was determined by single crystal X-ray structural analysis. Neither compound was toxic to HepG2 cells, and compound 1 was found to have a concentration-dependent inhibitory effect on intracellular lipid accumulation induced by stimulation of linoleic acid (LA). As a result of measuring the effects of compound 1 on the intracellular lipid production-related protein, it was found that compound 1 enhanced protein expression that promotes lipolysis. On the other hand, since the action of compound 1 was similar to that of PPARα agonists, it is deduced that compound 1 enhanced the activity of PPARα and further enhanced the expression of lipolytic proteins, which is related to the suppression of intracellular lipid accumulation. Furthermore, as the result of docking simulation, compound 1 had a higher binding affinity to the ligand binding site of PPARα than fenofibrate, which is a PPARα agonist, and thus compound 1 was considered to be promising as an agonist of PPARα.     

Relationship between Marteilioides chungmuensis infection and reproduction in the Pacific oyster, Crassostrea gigas

Marteilioides chungmuensis, a protozoan paramyxean parasite, infects the oocytes of the Pacific oyster, Crassostrea gigas. The effects of infection on the reproductive cycle of C. gigas were investigated over two consecutive years at Okayama Prefecture, Japan. In male oysters, gonadal development began during February/March, maturity was achieved in June and spawning activity extended from July to September. In November and December, male oysters were not seen, probably because their gonads regressed to connective tissue and they transformed into undifferentiated oysters. By contrast, female oysters, in which parasite spore formation occurred, were still carrying oocytes until the following March and the spawning process of female oysters took 5 months longer than that of males in epizootic areas. The prevalence of M. chungmuensis infection increased from July to September, when most female oysters had their spawning period, and declined from October to the following April when oysters were at the spent stage. The prevalence of infection increased again in May of the following year and high prevalence was observed in the following July. When prevalence was compared between oysters of different age classes, higher prevalence was detected in older than in younger oysters. Histological examination showed that infected oysters produced oocytes continuously and spawned repeatedly from October to March, during which period healthy oysters were reproductively inactive. Parasites can infect the oocytes of infected oysters throughout the longer spawning period. These observations suggest that M. chungmuensis extends the reproductive period of infected oysters for its own reproductive benefit.     

A fluid salt-bridging cluster and the stabilization of p53

p53 is a homotetrameric tumor suppressor protein that is found to be mutated in most human cancers. Some of these mutations, particularly mutations to R337, fall in the tetramerization domain and cause defects in tetramer formation leading to loss of function. Mutation to His at this site has been found to destabilize the tetramer in a pH-dependent fashion. In structures of the tetramerization domain determined by crystallography, R337 from one monomer makes a salt bridge with D352 from another monomer, apparently helping to stabilize the tetramer. Here we present molecular dynamics simulations of wild-type p53 and the R337His mutant at several different pH and salt conditions. We find that the 337-352 salt bridge is joined by two other charged side chains, R333 and E349. These four residues do not settle into a fixed pattern of salt bridging, but continue to exchange salt-bridging partners on the nanosecond time scale throughout the simulation. This unusual system of fluid salt bridging may explain the previous finding from alanine scanning experiments that R333 contributes significantly to protein stability, even though in the crystal structure it is extended outward into solvent. This extended conformation of R333 appears to be the result of a specific crystal contact and, this contact being absent in the simulation, R333 turns inward to join its interaction partners. When R337 is mutated to His but remains positively charged, it maintains the original interaction with D352, but the newly observed interaction with E349 is weakened, accounting for the reduced stability of R337H even under mildly acidic conditions. When this His is deprotonated, the interaction with D352 is also lost, accounting for the further destabilization observed under mildly alkaline conditions. Simulations were carried out using both explicit and implicit solvent models, and both displayed similar behavior of the fluid salt-bridging cluster, suggesting that implicit solvent models can capture at least the qualitative features of this phenomenon as well as explicit solvent. Simulations under strongly acidic conditions in implicit solvent displayed the beginnings of the unfolding process, a destabilization of the hydrophobic dimer-dimer interface. Computational alanine scanning using the molecular mechanics Poisson-Boltzmann surface area method showed significant correlation to experimental unfolding data for charged and polar residues, but much weaker correlation for hydrophobic residues.