Amino Acid Sequence of Oligopeptide Causes Marked Difference in DNA Compaction and Transcription.

Compaction of T4 phage DNA (166 kbp) by short oligopeptide octamers composed of two types of amino acids, four cationic lysine (K), and four polar nonionic serine (S) having different sequence order was studied by single-molecule fluorescent microscopy. We found that efficient DNA compaction by oligopeptide octamers depends on the geometrical match between phosphate groups of DNA and cationic amines. The amino acid sequence order in octamers dramatically affects the mechanism of DNA compaction, which changes from a discrete all-or-nothing coil-globule transition induced by a less efficient (K4S4) octamer to a continuous compaction transition induced by a (KS)4 octamer with a stronger DNA-binding character. This difference in the DNA compaction mechanism dramatically changes the packaging density, and the morphology of T4 DNA condensates: DNA is folded into ordered toroidal or rod morphologies during all-or-nothing compaction, whereas disordered DNA condensates are formed as a result of the continuous DNA compaction. Furthermore, the difference in DNA compaction mechanism has a certain effect on the inhibition scenario of the DNA transcription activity, which is gradual for the continuous DNA compaction and abrupt for the all-or-nothing DNA collapse.

B-type natriuretic peptide response and reverse left ventricular remodeling after surgical correction of functional mitral regurgitation in patients with advanced cardiomyopathy.

BACKGROUND: Restrictive mitral annuloplasty (RMA) can reverse left ventricular (LV) remodeling and reduce plasma B-type natriuretic peptide (BNP), a surrogate biomarker of heart failure. However, the relationship between reverse LV remodeling and plasma BNP changes after RMA is poorly defined. We explored the main hemodynamic factors contributing to change in plasma BNP after RMA in patients with functional mitral regurgitation (MR). METHODS: Twenty-four patients with moderate to severe functional MR secondary to LV systolic dysfunction [ejection fraction (EF) <40%] underwent 64-row multidetector computed tomography (MDCT) before and 1.4 months after RMA. LV end-diastolic volume index (EDVI), end-systolic volume index (ESVI), LVEF, and regional and global end-systolic wall stress (ESS) were calculated from 3-dimensional MDCT images, with blood samples for plasma BNP measurement collected the same day. RESULTS: After RMA, LV volumes and global ESS were decreased, while LVEF improved (all p<0.01). There were significant correlations between changes in LVEDVI and LVESVI (r=0.90, p<0.0001), LVESVI and global ESS (r=0.54, p=0.006), and global ESS and LVEF (r=-0.60, p=0.002). The median value for the plasma BNP also decreased from 597 pg/ml [interquartile range (IQR), 360-934 pg/ml] to 207 pg/ml (IQR, 124-271 pg/ml), in association with changes in LVEDVI (r=0.47, p=0.019), LVESVI (r=0.56, p=0.004), LVEF (r=-0.60, p=0.002), and global ESS (r=0.74, p<0.0001). Multivariate regression analysis showed that global ESS change was the strongest contributor to change in natural-log-transformed plasma BNP (standardized partial regression coefficient=0.59, p=0.004), indicating a strong association between decrease in LV afterload and reduction in plasma BNP level after RMA. CONCLUSIONS: There may be a significant association between LV reverse remodeling and plasma BNP change after RMA. Furthermore, LV end-systolic myocardial stress may be the key mechanical stimulus influencing plasma BNP after surgical correction for functional MR. Whether these favorable BNP responses and reverse remodeling can predict improved survival requires further study.

Restrictive mitral annuloplasty with or without surgical ventricular restoration in ischemic dilated cardiomyopathy with severe mitral regurgitation.

BACKGROUND: We assessed changes in left ventricular (LV) volume and function and in regional myocardial wall stress in noninfarcted segments after restrictive mitral annuloplasty (RMA) with or without surgical ventricular restoration (SVR). METHODS AND RESULTS: Thirty-nine patients with ischemic cardiomyopathy (ejection fraction ≤ 0.35) and severe mitral regurgitation (≥ 3) were studied before and 2.8 months after surgery with cine-angiographic multidetector computed tomography (cine-MDCT). Eighteen underwent RMA alone (RMA group) and 21 underwent RMA and SVR (RMA+SVR group). In addition to measuring conventional parameters (LV end-diastolic volume index [LVEDVI], LV end-systolic volume index [LVESVI], and LV ejection fraction), we evaluated the regional circumferential end-systolic wall stress and mean circumferential fiber shortening in both the basal and mid-LV regions using 3-dimensional cine-MDCT images. LV end-diastolic and end-systolic volume indexes were significantly greater in the RMA+SVR group than in the RMA group preoperatively, but these values did not differ significantly postoperatively. LV end-diastolic and end-systolic volume indexes decreased significantly, by 21% and 27% after RMA and by 35% and 42% after RMA and SVR, and the percent reductions in LV end-diastolic and end-systolic volume indexes were significantly larger in the RMA+SVR group. Regional end-systolic wall stress decreased and circumferential fiber shortening increased significantly in the noninfarcted regions after RMA with or without SVR. CONCLUSIONS: RMA plus SVR showed a potentially greater reduction of LV end-diastolic and end-systolic volume indexes than RMA alone. In selected patients with more advanced LV remodeling, concomitant SVR may favorably affect the LV reverse-remodeling process induced by RMA.

Collagenous colitis appeared after 6-year administration of lansoprazole.

Collagenous colitis (CC) is one of the causes of undefined watery diarrhea, which is histologically accompanied by thickening of the subepithelial collagen layer. CC associated with lansoprazole normally occurs within several weeks after initial administration, but no case presenting after long-term administration of lansoprazole has yet been reported. A 77-year-old male with 6-year history of administration of lansoprazole complained of watery diarrhea and weight loss. Colonoscopy revealed disappearance of vascular networks and red spots in the sigmoid colon. Biopsy specimen showed erosion and collagen bands thickened, so the patient was diagnosed as CC. After lansoprazole discontinuation, the watery diarrhea disappeared and histological abnormalities improved.

Reduction in adverse reactions to platelets by the removal of plasma supernatant and resuspension in a new additive solution (M-sol).

BACKGROUND: Leukodepletion reduces but does not eliminate adverse reactions to platelet concentrate (PC). As an alternative strategy, plasma reduction or washing of platelets should be considered. However, the efficacy of this strategy is still unclear. STUDY DESIGN AND METHODS: A total of 12 patients who experienced adverse reactions at a 29 to 100 percent reaction rate for plasma-PC were enrolled. The reactions were allergic reactions and nonhemolytic transfusion reactions, such as chills. Plasma-removed PC (W/R-PC), which was suspended in a recently developed additive solution (M-sol) containing less than 20 mL plasma, was prepared. W/R-PCs in M-sol were then transfused into patients after an overnight storage period; the occurrence of adverse reactions was monitored and 1- and 24-hour corrected count increment (CCI) values were evaluated. RESULTS: Although plasma-PC caused reaction in 12 patients, W/R-PC prevented reactions in 11 of 12 patients, with 1 patient having one minor allergic reaction of 15 transfusions. There was a significant difference in the incidence of reaction (p < 0.0001, Fisher's exact test). On a per-transfusion basis, the reaction rate for W/R-PC (1/156, 0.64%; 95% confidence interval [CI], 0.02%-3.5%) was reduced significantly compared to that for plasma-PC (117/276, 42%; 95% CI, 36%-48%; p < 0.0001). W/R-PC gave findings of satisfactory CCI at 1 hour (22,400 +/- 8,000/microL) and 24 hours (15,400 +/- 8,000/microL). No clinically evident bleeding episodes were recorded. CONCLUSIONS: W/R-PC suspended in M-sol in the presence of less than 20 mL plasma can be transfused safely and eliminate a wide range of adverse reactions to plasma-PC.

Tilt control in optical tweezers.

Laser trapping of micrometer-sized objects floating in water is investigated through the use of a tilted laser beam. With a change in the tilt direction, the orientation of the trapped object can be easily controlled when the object has an asymmetric body or nonuniform refractive index, such as nanowires, living cells, and so on. The method enables efficient orientation control under laser trapping through a simple setup. This method for tilt control may be useful for high-performance laser trapping in bioengineering and microsurgery in single living cells.

Pleiotrophic functions of Epstein-Barr virus nuclear antigen-1 (EBNA-1) and oriP differentially contribute to the efficiency of transfection/expression of exogenous gene in mammalian cells.

The EBNA1 gene and oriP sequence, originally derived from the EBV genome, provide plasmid vectors with artificial chromosome (AC)-like characteristics, including cytoplasm-to-nuclear transport, nuclear retention, replication and segregation of the DNA, while transcriptional up-regulation has been suggested as another activity of the EBNA1/oriP. Transfection as well as expression rates of various nonviral delivery vehicles are highly improved by inserting these genetic elements into plasmid DNA constructs. Here we differentially analyzed the contribution of each function of the EBNA1/oriP to the efficacy of electroporation-mediated genetic delivery and expression in mammalian cells. It was found that the EBNA1/oriP-mediated acceleration of genetic delivery and expression was predominantly due to the promotion of cytoplasm-to-nuclear recruitment as well as enhancement of transcription, while the episomal replication of the EBV-AC was not essentially involved.

In situ analysis of the higher-order genome structure in a single Escherichia coli cell.

We developed a novel method for the in situ analysis of the higher-order structure of an individual genome from a single Escherichia coli cell using laser tweezers. Initially, condensed DNA was stably grasped by a laser without any chemical modification and without physical attachment to an artificial object such as micro-plastic beads. Under optical transport, the trapped genome gradually unfolded in solution due to viscous friction. Interestingly, the nucleoid DNA from a log-phase cell is almost fully elongated, whereas in the stationary phase, unfolding of the nucleoid is characterized by step-wise elongation of 1.7-5.1 microm, corresponding to a size of 5-15 kbp, and a few tightly packed domains remain along the DNA chain. This suggests the coexistence of tightly packed and swollen domains in the genome in the stationary phase.

Hyaluronic acid and its derivative as a multi-functional gene expression enhancer: protection from non-specific interactions, adhesion to targeted cells, and transcriptional activation.

Hyaluronic acid (HA), a natural anionic mucopolysaccharide, can be deposited onto the cationic surface of DNA/polyethyleneimine (PEI) complexes to recharge the surface potential and reduce nonspecific interactions with proteins. HA can also be used as a ligand to target specific cell receptors. Furthermore, HA-coating enhanced the transcriptional activity of the plasmid/PEI complexes, probably by loosening the tight binding between DNA and PEI, which facilitated the approach of transcription factors. Amphoteric HA derivative having spermine side chains (Spn-HA) with a structure similar to HMG protein showed higher transcription-enhancing activity than HA. Plasmid/PEI/Spn-HA ternary complex exhibited 29-fold higher transgene expression efficiency than naked plasmid/PEI complexes in CHO cells.

Enhancement and inhibition of DNA transcriptional activity by spermine: a marked difference between linear and circular templates.

We compared the transcriptional activities of the circular and linear forms of short (4kbp) and giant (106kbp) DNA molecules in the presence of a polyamine, spermine (4+). With an increase in the spermine concentration, transcriptional activity was enhanced, followed by an inhibitory effect, and complete inhibition was observed in the sole case of long, linear templates. A difference between the transcriptional properties of circular and linear conformations is found for giant DNA molecules. These results are discussed in relation to DNA conformational transitions.

Folding transition into a loosely collapsed state in plasmid DNA as revealed by single-molecule observation.

The conformational transition of a plasmid DNA, pGEG.GL3 (12.5 kbp, circular), induced by spermine(4+) was studied through the observation of individual DNA by fluorescence microscopy. We deduced the change in the hydrodynamic radius R(H) from an analysis of the Brownian motion of single DNA molecules. R(H) decreases in a continuous manner with an increase in spermine(4+), in contrast to the large discrete on/off change for long linear DNA. Just after the transition to the collapsed state, a small number of DNA molecules tend to form an assembly, which disperses in the bulk solution without precipitation.

NTP concentration switches transcriptional activity by changing the large-scale structure of DNA.

It is becoming clearer that genetic activity is closely associated with the intracellular energy state. However, the mechanisms of this association are still unclear. In this study, we focused on large-scale changes in the structure of DNA to examine the effect of the NTP concentration on the transcription reaction with T7 RNA polymerase and compared the results with long duplex DNA to those with a short persistent-length(1) fragment. The transcriptional activity dramatically changed only for long duplex DNA within a narrow range of NTP concentrations associated with changes in the large-scale structure of DNA. This result suggests that the energy state may play an essential role in regulating ON/OFF switching on transcriptional activity.