Age-related decrease in mouse T cell progenitors.

Mice were given a lethal dose of whole-body gamma-radiation and injected with a 10(5) or 10(6) marrow cells from 10- to 143-week-old syngeneic donors. Nine days later, colony-forming units (CFU) were counted in the spleens of mice given 10(5) cells, and 15 to 21 days after irradiation thymus weights and in some experiments 3H-thymidine uptake or total thymic cellularity were determined in the recipients of 10(6) cells. It was found that in the majority of mouse strains studied there were no significant changes with age in marrow CFU. In contrast, thymic regeneration was significantly impaired when the recipients received marrow cells from donors 100 weeks of age or older. These observations and results obtained in dose-response and time-course studies are best explained by an age-related decrease in marrow T cell progenitors; however, certain findings suggest that in addition the proliferative capacity of these stem cells may at times be moderately impaired.     

New Insights into the Developing Rabbit Brain Using Diffusion Tensor Tractography and Generalized q-Sampling MRI

The use of modern neuroimaging methods to characterize the complex anatomy of brain development at different stages reveals an enormous wealth of information in understanding this highly ordered process and provides clues to detect neurological and neurobehavioral disorders that have their origin in early structural and functional cerebral maturation. Non-invasive diffusion tensor magnetic resonance imaging (DTI) is able to distinguish cerebral microscopic structures, especially in the white matter regions. However, DTI is unable to resolve the complicated neural structure, i.e., the fiber crossing that is frequently observed during the maturation process. To overcome this limitation, several methods have been proposed. One such method, generalized q-sampling imaging (GQI), can be applied to a variety of datasets, including the single shell, multi-shell or grid sampling schemes that are believed to be able to resolve the complicated crossing fibers. Rabbits have been widely used for neurodevelopment research because they exhibit human-like timing of perinatal brain white matter maturation. Here, we present a longitudinal study using both DTI and GQI to demonstrate the changes in cerebral maturation of in vivo developing rabbit brains over a period of 40 weeks. Fractional anisotropy (FA) of DTI and generalized fractional anisotropy (GFA) of GQI indices demonstrated that the white matter anisotropy increased with age, with GFA exhibiting an increase in the hippocampus as well. Normalized quantitative anisotropy (NQA) of GQI also revealed an increase in the hippocampus, allowing us to observe the changes in gray matter as well. Regional and whole brain DTI tractography also demonstrated refinement in fiber pathway architecture with maturation. We concluded that DTI and GQI results were able to characterize the white matter anisotropy changes, whereas GQI provided further information about the gray matter hippocampus area. This developing rabbit brain DTI and GQI database could also be used for educational purposes and neuroscience investigations.     

Breast cancer cells induce stromal fibroblasts to secrete ADAMTS1 for cancer invasion through an epigenetic change

Microenvironment plays an important role in cancer development. We have reported that the cancer-associated stromal cells exhibit phenotypic and functional changes compared to stromal cells neighboring to normal tissues. However, the molecular mechanisms as well as the maintenance of these changes remain elusive. Here we showed that through co-culture with breast cancer cells for at least three to four passages, breast normal tissue-associated fibroblasts (NAFs) gained persistent activity for promoting cancer cell invasion, partly via up-regulating ADAM metallopeptidase with thrombospondin type 1 motif, 1 (ADAMTS1). Furthermore, we demonstrated that the DNA methylation pattern in the ADAMTS1 promoter has no alteration. Instead, the loss of EZH2 binding to the ADAMTS1 promoter and the resulting decrease of promoter-associated histone H3K27 methylation may account for the up-regulation of ADAMTS1. Importantly, the lack of EZH2 binding and the H3K27 methylation on the ADAMTS1 promoter were sustained in cancer cell-precocultured NAFs after removal of cancer cells. These results suggest that cancer cells are capable of inducing stromal fibroblasts to secrete ADAMTS1 persistently for their invasion and the effect is epigenetically inheritable.     

Proteomic profiling of human pleural effusion using two-dimensional nano liquid chromatography tandem mass spectrometry

Pleural effusion, an accumulation of pleural fluid, contains proteins originated from plasma filtrate and, especially when tissues are damaged, parenchyma interstitial spaces of lungs and/or other organs. This study details protein profiles in human pleural effusion from 43 lung adenocarcinoma patients by a two-dimensional nano-high performance liquid chromatography electrospray ionization tandem mass spectrometry (2D nano-HPLC-ESI-MS/MS) system. The experimental results revealed the identification of 1415 unique proteins from human pleural effusion. Among these 124 proteins identified with higher confidence levels, some proteins have not been reported in plasma and may represent proteins specifically present in pleural effusion. These proteins are valuable for mass identification of differentially expressed proteins involved in proteomics database and screening biomarker to further study in human lung adenocarcinoma. The significance of the use of proteomics analysis of human pleural fluid for the search of new lung cancer marker proteins, and for their simultaneous display and analysis in patients suffering from lung disorders has been examined.     

Allelic polymorphism synergizes with variable gene content to individualize human KIR genotype

Killer Ig-like receptor (KIR) genes are a multigene family on human chromosome 19. KIR genes occur in various combinations on different haplotypes. Additionally, KIR genes are polymorphic. To examine how allelic polymorphism diversifies KIR haplotypes with similar or identical combinations of KIR genes, we devised methods for discriminating alleles of KIR2DL1, -2DL3, -3DL1, and -3DL2. These methods were applied to 143 individuals from 34 families to define 98 independent KIR haplotypes at the allele level. Three novel 3DL2 alleles and a chimeric 3DL1/3DL2 sequence were also identified. Among the A group haplotypes were 22 different combinations of 2DL1, 2DL3, 3DL1, and 3DL2 alleles. Among the B group haplotypes that were unambiguously determined were 15 distinct haplotypes involving 9 different combinations of KIR genes. A and B haplotypes both exhibit strong linkage disequilibrium (LD) between 2DL1 and 2DL3 alleles, and between 3DL1 and 3DL2 alleles. In contrast, there was little LD between the 2DL1/2DL3 and 3DL1/3DL2 pairs that define the two halves of the KIR gene complex. The synergistic combination of allelic polymorphism and variable gene content individualize KIR genotype to an extent where unrelated individuals almost always have different KIR types. This level of diversity likely reflects strong pressure from pathogens on the human NK cell response.     

Effects of inositol, LiCl, and heparin on the antibody responses to SRBC by normal and immunodeficient XID mice

Spleen cells from na?ve adult immunocompetent and immunodeficient XID mice were cultured on agar containing sheep red blood cells (SRBC) with and without myo-inositol, scyllo-inositol, lithium chloride, or heparin, and after 1 or 2 days the number of colonies of antiSRBC antibody-forming cells (PFC) were determined. It was found that myo-inositol and scyllo-inositol at one-tenth the concentration were equally effective in increasing the number of specific PFC. Myo-inositol, scyllo-inositol, and lithium chloride accelerated the appearance of direct foci in cultures of spleen cells from normal and XID mice. When heparin was added to cultures of XID spleen cells, PFC were found to be increased on Day 1; however, PFC and foci were not increased in cultures of spleen cells from competent mice until 1 day later. The addition of combinations of these agents to cultures of spleen cells had no positive or negative effect on the generation of foci or PFC. Normal mice given heparin intraperitoneally with SRBC had increased splenic PFC on Days 3 and 4 but not on Day 7. The results suggest that these agents modulate B-cell responses by increasing the rate of proliferation and/or secretion through a signaling pathway(s) distal to, or more likely, independent of Bruton's tyrosine Kinase (BTK). It is not clear that the mechanism is the same with each agent.     

Effects of H-2 and vitamin A on micrognathia in congenic mice.

Pregnant mice congenic with C57BL/10 (B10.A, B10.BR, B10.D2, B10.A(2R), B10.A(5R), B10.A(15R), B10.A(1R), B10.A(18R), and B10.OL) were fed Purina Mouse Chow or the same diet plus 200 IU of vitamin A daily. The pregnant dams were sacrificed on the eighteenth day of gestation, and the fetuses were sexed and examined for defects in mandibular development. On average, micrognathia occurred five times more frequently in female (1.5%) than male (0.3%) fetuses. The addition of vitamin A to the diet affected only females, reducing the frequency of this defect to that observed in males from dams fed the control diet. Micrognathia was strongly associated with micro- or anophthalmia, but not with defects of the palate. C57BL/10 fetuses had the highest frequency of micrognathia (3.2%) and B10.D2 and B10.A(5R) fetuses had the lowest (0.1%). The results suggest that a locus distal to C4 and perhaps proximal to Qa-1 may exert a moderate influence on mandibular development and a second locus proximal to E beta may have a weak effect.     

Quantitative analysis of sensor for pressure waveform measurement

Background

Arterial pressure waveforms contain important diagnostic and physiological information since their contour depends on a healthy cardiovascular system [1]. A sensor was placed at the measured artery and some contact pressure was used to measure the pressure waveform. However, where is the location of the sensor just about enough to detect a complete pressure waveform for the diagnosis? How much contact pressure is needed over the pulse point? These two problems still remain unresolved.

Method

In this study, we propose a quantitative analysis to evaluate the pressure waveform for locating the position and applying the appropriate force between the sensor and the radial artery. The two-axis mechanism and the modified sensor have been designed to estimate the radial arterial width and detect the contact pressure. The template matching method was used to analyze the pressure waveform. In the X- axis scan, we found that the arterial diameter changed waveform (ADCW) and the pressure waveform would change from small to large and then back to small again when the sensor was moved across the radial artery. In the Z-axis scan, we also found that the ADCW and the pressure waveform would change from small to large and then back to small again when the applied contact pressure continuously increased.

Results

In the X- axis scan, the template correlation coefficients of the left and right boundaries of the radial arterial width were 0.987 ± 0.016 and 0.978 ± 0.028, respectively. In the Z-axis scan, when the excessive contact pressure was more than 100 mm Hg, the template correlation was below 0.983. In applying force, when using the maximum amplitude as the criteria level, the lower contact pressure (r = 0.988 ± 0.004) was better than the higher contact pressure (r = 0.976 ± 0.012).

Conclusions

Although, the optimal detective position has to be close to the middle of the radial arterial, the pressure waveform also has a good completeness with a template correlation coefficient of above 0.99 when the position was within ± 1 mm of the middle of the radial arterial range. In applying force, using the maximum amplitude as the criteria level, the lower contact pressure was better than the higher contact pressure.

     

The Contribution of Antibiotic Resistance Mechanisms in Clinical Burkholderia cepacia Complex Isolates: An Emphasis on Efflux Pump Activity

Due to the limited information of the contribution of various antibiotic resistance mechanisms in clinical Burkholderia cepacia complex isolates, Antibiotic resistance mechanisms, including integron analysis, identification of quinolone resistance-determining region mutations, measurement of efflux pump activity, and sequence analysis of efflux pump regulators, were investigated in 66 clinical B. cepacia complex isolates. Species were identified via recA-RFLP and MALDI-TOF. Four genomovars were identified by recA-RFLP. B. cenocepacia (genomovar III) was the most prevalent genomovar (90.1%). Most isolates (60/66, 90.9%) were correctly identified by MALDI-TOF analysis. Clonal relatedness determined by PFGE analysis revealed 30 pulsotypes, including two major pulsotypes that comprised 22.7% and 18.2% of the isolates, respectively. Seventeen (25.8%) isolates harboured class 1 integron with various combinations of resistance genes. Among six levofloxacin-resistant isolates, five had single-base substitutions in the gyrA gene and three demonstrated efflux pump activities. Among the 42 isolates exhibiting resistance to at least one antimicrobial agent, 94.4% ceftazidime-resistant isolates (17/18) and 72.7% chloramphenicol-resistant isolates (16/22) demonstrated efflux pump activity. Quantitation of efflux pump RNA level and sequence analysis revealed that over-expression of the RND-3 efflux pump was attributable to specific mutations in the RND-3 efflux pump regulator gene. In conclusion, high-level expression of efflux pumps is prevalent in B. cepacia complex isolates. Mutations in the RND-3 efflux pump regulator gene are the major cause of efflux pump activity, resulting in the resistance to antibiotics in clinical B. cepacia complex isolates.