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91.
Summary The purpose of this study was to develop a nonenzymatic method of isolating adult islets using atrophied pancreata from copper-deficient rats and to analyze their morphologic characteristics and behavior in culture. This unusual model of isolation was studied because islets remain intact in the course of dietary copper deficiency while the acinar glandular component of the pancreas undergoes selective atrophy and lipomatosis. Small fragments containing islets were readily microdissected from atrophied glands and placed in culture. Within 24 h the fragments congealed into small irregular- to spherical-shaped masses within which the darker profile of islets could be distinguished. Within a period of 3 to 5 d, islet tissue began to bud from the lipocytic mass until by Day 7 spherical aggregates of intact islet tissue separated from the residual fragments. Subsequent to further in vitro treatment, these islets could be maintained as free viable spherical masses if periodically agitated, as attached stationary islets which developed monolayer growth if left undisturbed and as aggregated masses of islet tissue forming megaislets if combined in small groups. Grouped islets treated with actinomycin D and cycloheximide did not exhibit aggregation when incubated with these inhibitors. This suggests that megaislet formation was an active process requiring protein-RNA synthesis rather than passive clumping or aggregation that can accompany metabolically altered or dying islets undergoing cellular shedding and adhesion. Immunohistochemical localization demonstrated that insulin, glucagon, somatostatin, and pancreatic polypeptide-immunoreactive cell types were present within the islets derived from this technique. The cellular topography of these islets was not unlike that described by others for islets cultured from enzymatic isolation. This culture model may serve as a resource for mature, viable islets isolated without mechanical or enzymatic disaggregation which can have attenuating effects on islet function. This work was supported by a research grant from the Diabetes Research and Education Foundation.  相似文献   
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Background

Influenza pandemic remains a serious threat to human health. Viruses of avian origin, H5N1, H7N7 and H9N2, have repeatedly crossed the species barrier to infect humans. Recently, a novel strain originated from swine has evolved to a pandemic. This study aims at improving our understanding on the pathogenic mechanism of influenza viruses, in particular the role of non-structural (NS1) protein in inducing pro-inflammatory and apoptotic responses.

Methods

Human lung epithelial cells (NCI-H292) was used as an in-vitro model to study cytokine/chemokine production and apoptosis induced by transfection of NS1 mRNA encoded by seven infleunza subtypes (seasonal and pandemic H1, H2, H3, H5, H7, and H9), respectively.

Results

The results showed that CXCL-10/IP10 was most prominently induced (> 1000 folds) and IL-6 was slightly induced (< 10 folds) by all subtypes. A subtype-dependent pattern was observed for CCL-2/MCP-1, CCL3/MIP-1α, CCL-5/RANTES and CXCL-9/MIG; where induction by H5N1 was much higher than all other subtypes examined. All subtypes induced a similar temporal profile of apoptosis following transfection. The level of apoptosis induced by H5N1 was remarkably higher than all others. The cytokine/chemokine and apoptosis inducing ability of the 2009 pandemic H1N1 was similar to previous seasonal strains.

Conclusions

In conclusion, the NS1 protein encoded by H5N1 carries a remarkably different property as compared to other avian and human subtypes, and is one of the keys to its high pathogenicity. NCI-H292 cells system proves to be a good in-vitro model to delineate the property of NS1 proteins.
  相似文献   
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Andropogon virginicus L. (broomsedge) is a perennial bunchgrass of temperate origin which was introduced to the Hawaiian Islands early in the 20th Century. Since its introduction broomsedge has become the dominant species of savanna vegetation on the windward sides of Oahu and Hawaii (the Big Island). Despite its ecological success, the phenology of broomsedge-especially the timing of its periods of active growth in the summer and dormancy (or semi-rest) in the winter-is not synchronized with the summer-dry, winter-wet moisture seasonality of its range in Hawaii. Rather, this species responds to a shortening-daylength cue by flowering in late autumn, and the onset of dormancy and the progression of subsequent phenophases seems to be mostly under endogenous control following the flowering response. The lack of synchronization between the onset of active growth by broomsedge and the onset of the rainy season does not conform to the specific assumption that the ecological dominance of the perennial bunchgrass life-form in savanna vegetation results from a coupling of phenological strategy to moisture seasonality. It also brings into question the more general assumption that a perfect coupling of phenology to the prevailing climatic conditions is a prerequisite for ecological success.  相似文献   
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Three samples of silage taken from the surface of a silo and from depths of 20 and 45 cm in the silo were studied for identification of the potential agents causing symptoms of organic dust toxic syndrome. The samples were examined by dilution plating before and after aerosolization in an acoustical dust generator. Aerosol samples were collected by liquid impinger and filter cassettes. The samples were examined for total aerobic bacteria, anaerobic bacteria, gram-negative bacteria, lactobacilli, listeriae, thermophilic actinomycetes, fungi, and endotoxin. Very high levels of total aerobic bacteria and fungi were found in the surface sample (up to 10(9) CFU/g in the bulk sample and up to 10(9) CFU/m3 after aerosolization), whereas the corresponding values from the deepest site were 100 to 50,000 times lower. Aspergillus fumigatus predominated among the fungi, whereas Bacillus and gram-negative organisms (Pseudomonas, Alcaligenes, Citrobacter, and Klebsiella species) prevailed among bacteria. Thermophilic actinomycetes occurred in numbers up to 10(7) CFU/g in the bulk samples, whereas anaerobic bacteria, lactobacilli, and listeriae were only few or absent. The concentration of endotoxin was high in the surface sample (up to 211.4 Endotoxin Units/mg) and about 200-fold lower in the sample from the deepest site. The results show that contact with dust from the surface of silage carries the risk of exposure to high concentrations of microorganisms, of which A. fumigatus and endotoxin-producing bacteria are the most probable disease agents.  相似文献   
100.
A growing variety of “genotype-by-sequencing” (GBS) methods use restriction enzymes and high throughput DNA sequencing to generate data for a subset of genomic loci, allowing the simultaneous discovery and genotyping of thousands of polymorphisms in a set of multiplexed samples. We evaluated a “double-digest” restriction-site associated DNA sequencing (ddRAD-seq) protocol by 1) comparing results for a zebra finch (Taeniopygia guttata) sample with in silico predictions from the zebra finch reference genome; 2) assessing data quality for a population sample of indigobirds (Vidua spp.); and 3) testing for consistent recovery of loci across multiple samples and sequencing runs. Comparison with in silico predictions revealed that 1) over 90% of predicted, single-copy loci in our targeted size range (178–328 bp) were recovered; 2) short restriction fragments (38–178 bp) were carried through the size selection step and sequenced at appreciable depth, generating unexpected but nonetheless useful data; 3) amplification bias favored shorter, GC-rich fragments, contributing to among locus variation in sequencing depth that was strongly correlated across samples; 4) our use of restriction enzymes with a GC-rich recognition sequence resulted in an up to four-fold overrepresentation of GC-rich portions of the genome; and 5) star activity (i.e., non-specific cutting) resulted in thousands of “extra” loci sequenced at low depth. Results for three species of indigobirds show that a common set of thousands of loci can be consistently recovered across both individual samples and sequencing runs. In a run with 46 samples, we genotyped 5,996 loci in all individuals and 9,833 loci in 42 or more individuals, resulting in <1% missing data for the larger data set. We compare our approach to similar methods and discuss the range of factors (fragment library preparation, natural genetic variation, bioinformatics) influencing the recovery of a consistent set of loci among samples.  相似文献   
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