Supernumerary chromosomal elements in lymphocytes cultured from phenotypically normal human adult males

In a group of phenotypically normal men there were approximately 0.24% of metaphase lymphocytes with extra chromosomal elements along with the regular 46 chromosomes. They ranged in size from small acrocentric-acentric elements to elements longer than any chromosome arm. These elements have been referred to as supernumerary chromosomal elements. No significant effects due to donor's age, smoking history, season, storage of blood samples prior to culture, or culture medium, were found either in the frequency of supernumerary elements per cell or in the frequency of cells with supernumerary elements. Furthermore, the same subject did not consistently exhibit supernumerary elements. Furthermore, the same subject did not consistently exhibit supernumerary elements when sampled during four successive quarters of the year. Some of these elements in pairs were identified by G-banding technique as translocation chromosomes bearing long arms of chromosome number 2 and presumptive short arms of chromosome 8, acentric long arms of chromosome 4, and iso-acentric chromosomes for the long arms of chromosome 5. Presumably, more than one type of cytogenetic event occurred in their formation. Circumstantial evidence has been presented to show that the means of elimination of these supernumerary elements is a process of chromosomal disintegration.     

Anti-immunoglobulin-induced proliferation of B cells : Parallelism in the inhibition by chloroquine, monensin and immunoglobulin

Proliferation of rabbit lymphocytes was induced with goat anti-rabbit immunoglobulin. Chloroquine and monensin, known to inhibit internalization-related events, yielded inhibition of proliferation that paralleled the inhibition by a specific competitive ligand, rabbit immunoglobulin (IgG), whereas inhibition by puromycin did not. Moreover, virtually all of the cells that can be activated in freshly isolated populations adhered to anti-immunoglobulin-coated Petri plates, whereas all of the activatable population was recovered in the non-adherent fraction after a brief incubation of the cells with anti-immunoglobulin to induce internalization of surface membrane immunoglobulin. Using immunofluorescence it was further observed that monensin and Chloroquine inhibit the reappearance of surface immunoglobulins on the cell surface to some extent subsequent to their removal induced by anti-immunoglobulin.     

Poster Sessions BP02: Oligodendrocytes and Schwann Cells. RAS activation regulates angiogenic and anti‐angiogenic factor expression in NF1‐derived Schwann cells

Neurofibromatosis Type 1 tumors are highly vascularized and contain Schwann cells with hyperactivated Ras. In vitro, the NF1‐derived neurofibromin deficient Schwann cells have an angiogenic profile, which favors angiogenesis and sustains the growth of the NF1‐derived tumors. This study examined the relationship of the activation state of Ras as it related to the expression of angiogenic and antiangiogenic factors in both cultured NF1‐derived Schwann cells and normal human Schwann cells. Western blot analysis of normal human Schwann cells revealed low expression of angiogenic vascular endothelial growth factor (VEGF) as well as low expression of the antiangiogenic pigment epithelium derived factor (PEDF). Relative to normal human Schwann cells, NF1‐derived Schwann cells have increased RAS activity and a three‐fold increase in VEGF expression. Surprisingly, PEDF was also expressed in the NF1‐derived Schwann cells at approximately the same level as VEGF expression. Using a retroviral construct, we introduced the GAP‐related domain of neurofibromin into the NF1‐derived Schwann cells to reduce the level of activated Ras. Relative to the untreated NF1‐derived Schwann cells the Schwann cells expressing the GAP‐related domain expressed about one‐half the VEGF but twice the PEDF. We conclude that decreasing the Ras activity in NF1‐drived Schwann cells will not only decrease proliferation, but also slow tumor angiogenesis due to the decreased expression of angiogenic and increased expression of antiangiogenic factors.     

Discovery of QTLs for water mining and water use efficiency traits in rice under water-limited condition through association mapping

Developing trait introgressed rice cultivars is essential to sustain yield under aerobic conditions. Here, we report DNA markers governing variability in root traits, water use efficiency (WUE) and other biometric traits like total leaf area by association mapping. A set of 173 diverse rice germplasm accessions were phenotyped for root traits in specially designed root structures and WUE using carbon isotope discrimination (Δ¹³C) during the monsoon season (July to October) of two consecutive years (2007 and 2008). The panel was genotyped using 291 SSR markers spanning the entire genome of rice. Root biomass varied between 1.8 and 16.3 g plant^?1 while root length between 22 and 78 cm representing significant genetic variability. Similarly, Δ¹³C varied from 18 to 23 ‰. The SSR markers showed extensive polymorphism with around 73 % of all the markers revealing polymorphism information content values more than 0.5. Model-based structure analysis using the squared-allele frequency correlations revealed six subgroups among the panel with an average LD decay of about 10–20 cM. The Benjamini–Hochberg analysis was carried out to compute the false discovery rate combined with the analysis of effective LD. A total of 82 markers were involved in 175 significant (corrected P values and Q values <0.05) marker–trait associations (MTAs) across experiment 1 and experiment 2 and for the pooled data. Out of these, 22 markers were found to be associated with more than one trait. Common markers with significant associations were discovered for root biomass, total leaf area and total biomass suggesting the interdependency of these traits. Finally, 12 markers showed significant and stable MTAs across the experiments for different traits. An in silico analysis indicated that 45 % of the MTAs overlapped with previously reported QTLs and can be used for QTL introgression through breeding.     

Experimental determination of sarcomere force–length relationship in type-I human skeletal muscle fibers

The objectives of this study were to measure the active and passive force–length (F–L) relationships in type-I human single muscle fibers and to compare the results to predictions from the sliding filament model (the “standard model”). We measured isometric forces in chemically skinned fibers at different sarcomere lengths (SLs) in separate maximal activations. The experimental tolerance interval for optimal SL was calculated to be (2.37, 2.95 μm), which included the prediction by the standard model (2.64, 2.81 μm). Average passive slack length was 2.22±0.08 μm, and the passive F–L relationship was well described by an exponential function. Best fit lines were used to estimate the ascending and descending limbs from the active F–L data using the average SL obtained from a digital image of the fiber. The experimental descending limb was also estimated using the shortest SL to address the possible effects of sarcomere inhomogeneity (SI). The experimental slopes of the ascending and descending limbs, 0.42 F_o/μm and ?0.52 F_o/μm (vs. ?0.55 F_o/μm with the shortest SL) respectively, F_o being the maximal isometric force, were significantly less in magnitude than those from the standard model. These results suggested that the difference between experimental and standard models was not fully explained by SI and other factors could be important. The broader experimental F–L curve compared to the standard model implies that human muscle has functionally a wider operating length range where its force-generating capacity is not compromised.     

Cytokinetics and sce baselines in rat and human lymphocytes during successive divisions in the presence of different culture media

Summary Peripheral blood cultures were set up from male rats and humans in TC199, RPMI 1640, and minimal essential medium in the presence of 5-bromo, 2-deoxyuridine and harvested at 48, 72, and 96 h. Mitotic indices were compared in the different media at all three harvest times, but cytokinetic patterns and baseline sister chromatid exchange (SCE) frequencies were evaluated exclusively in the 72- and 96-h cultures. In general, lymphocyte division kinetics, as determined by average lymphocyte division (ALD) numbers, were comparable between rat and human lymphocytes cultured in any of the three media and harvested at either of the culture times. The numbers of SCEs were distributed between and within the two systems independent of ALD numbers or the harvest time. Overall, no influence of media was detected on the distribution of SCEs. Despite a number of similarities in growth characteristics between rat and human lymphocytes, the rat lymphocyte test system has distinct advantages over that of the human because of the easy access to rat blood samples and the absence of the many restrictions applicable to humans.     

Effect of food and water deprivation on the parameters of the mouse bone marrow micronucleus test

This study reports the influence of acute starvation on spontaneous and cyclophosphamide (CP) induced micronucleus (MN) frequencies in the bone marrow polychromatic erythrocytes (PCE) of CD-1 mice. Groups of mice (5/sex) were deprived of either food alone or food and water for 0, 24, 48 and 72 h prior to sacrifice. Although there was no evidence of a significant increase in MN-PCE frequencies among the starved groups, a significant depressant effect of starvation on erythropoietic activity was observed. Peak levels of CP-induced (40 mg/kg b.w.) MN-PCE's appeared later in male mice deprived of food and water after treatment compared to mice given food and water ad libitum. The results indicate that starvation is detrimental to bone marrow erythropoiesis and that starvation may alter the response of mice to clastogens.     

Anchor-dependent lipofection with non-glycerol based cytofectins containing single 2-hydroxyethyl head groups

Detailed structure-activity investigations aimed at probing the anchor chain length dependency for glycerol-based lipofectins have been reported previously. Herein, we report on the first detailed investigation on the anchor-dependent transfection biology of non-glycerol based simple monocationic cytofectins containing single 2-hydroxyethyl head group functionality using 11 new structural analogs of our previously published first generation of non-glycerol based transfection lipids (lipids 1-11). The C-14 and C-16 analogs of DOMHAC (lipids 4 and 5, respectively) were found to be remarkably efficient in transfecting COS-1 cells. In addition, the present anchor-dependency investigation also revealed that the C-14 analog of DOHEMAB (lipid 10) is significantly efficient in transfecting both COS-1 and NIH3T3 cells. Our results also indicate that too strong lipid-DNA interactions might result in weaker transfection for non-glycerol based cationic lipids. In summary, the anchor-dependence investigations presented here convincingly demonstrate that non-glycerol based cationic lipids containing a single hydroxyethyl head group and hydrophobic C-14 or C-16 anchors are promising non-toxic cationic transfection lipids for future use in liposomal gene delivery.