Growth feedback as a basis for persister bistability

A small fraction of cells in many bacterial populations, called persisters, are much less sensitive to antibiotic treatment than the majority. Persisters are in a dormant metabolic state, even while remaining genetically identical to the actively growing cells. Toxin and antitoxin modules in bacteria are believed to be one possible cause of persistence. A two-gene operon, HipBA, is one of many chromosomally encoded toxin and antitoxin modules in Escherichia coli and the HipA7 allelic variant was the first validated high-persistence mutant. Here, we present a stochastic model that can generate bistability of the HipBA system, via the reciprocal coupling of free HipA to the cellular growth rate. The actively growing state and the dormant state each correspond to a stable state of this model. Fluctuations enable transitions from one to the other. This model is fully in agreement with experimental data obtained with synthetic promoter constructs.As far back as the 1940s, it was known that a small fraction of a bacterial population can survive even when exposed to prolonged antibiotic treatment (1, 2). This phenomenon is termed persistence and members of the surviving subpopulation are called persisters. It has been estimated that the frequency of persisters in normal wild-type populations is extremely small, perhaps of order (3). Although the number of persisters is tiny, they are often the main obstacle to attempts to completely eradicate infection.Remarkably, there is no apparent change in the persisters’ DNA sequence; i.e., their survival is not due to mutation (4). Already in 1944, Bigger suggested that persisters are phenotypically different, in a dormant state instead of an actively growing state (1). The dormant state is presumably better able to deal with common antibiotics, which typically target only actively growing cells. Bigger’s assumption was confirmed by a later study (3). In this study, Balaban et al. investigated the persistence of a single cell of Escherichia coli by using a microfluidic device. They showed that individual persisters do not always remain in the dormant state. Instead, they stochastically transit into an actively growing state and these newly transited cells are indistinguishable from other normally growing cells. Conversely, normal cells can transit into the persistent state. Thus, bacterial persistence at the population level is governed by a single-cell “phenotypic switch.” The precise workings of this switch have to date remained unclear.In the 1980s, Moyed and Bertrand identified the first high-persistence mutant, HipA7, having a persister frequency that is near 10⁻² (4). The discovery of HipA7 facilitated the study of bacterial persistence due to its relatively high proportion of persisters. It was found that HipA7 is formed by a two-residue substitution in the HipA protein. This protein acts as a toxin in a toxin–antitoxin (TA) module (5, 6), where the hipB gene is coexpressed with hipA and the corresponding protein binds to and neutralizes HipA toxicity. To date, HipA is one of only a few molecules that are validated tolerance factors (7).There have already been several models proposed for the Hip system and its connection to persistence. Two modeling groups have claimed that fluctuations cause the apparent coexistence of these two phenotypes, growing and dormant, even though there may or may not be any formal bistability. They were partially driven to this conclusion by their inability to find bistability in their assumed dynamics. The pioneering model of Rotem et al. (8) did not consider the dimerization of HipB and the repression by the HipB dimer of the hip promoter. In the alternate formulation of Koh and Dunlop (9), the HipA-dependent reduction of the translation rate and the growth rate is not included. Thus, both these works claim that bistable states are not necessarily the mechanism underlying persister formation. However, models with a single stable state invariably predict fast transitions between persisters and normally growing cells. For example, simulations in ref. 9 show that transitions from persisters to normally growing cells typically happen within 1 h. In contrast, a sizeable number of persisters can survive even when the antibiotic treatment is maintained for longer than 1 d. If cells stay in a persister state only for less than 1 h, and the persister becomes fragile when it transits into the normally growing state, they would not survive much longer than the other normal cells. The correct picture must include a long-time duration of the persister state.One model has indeed suggested that bistability is the key to the formation of persisters (10). This model made some assumptions now known to be inaccurate, for example that free HipA undergoes dimerization and that the binding of the HipA-HipB complex to the hip promoter is independent from the binding between the HipB dimer and the hip promoter. (Actually they compete with each other in binding to the same operator sites.) However, this model does explain an interesting observation, that often persisters are formed much more readily in stationary phase and in fact persisters seen in normal exponential phase are often just the remnant of persisters formed at a different growth stage. This pattern has been called type I persistence (3) and is the type seen in the HipA7 mutant. As we will see, this occurs due to the fact that the range of bistability can depend on the growth condition. A different issue is that this model is fully deterministic and hence cannot address stochastic effects such as transitions between the two stable states.The drawbacks of these models have motivated us to construct a more precise and comprehensive stochastic model for the HipBA system. A recent paper revealing the structure of HipA and its binding has helped guide us to correct the assumptions in the previous bistable model (11). We show that our approach can consistently account for different classes of experimental data and hence can form a framework for continuing analysis of this important survival strategy for wide classes of bacteria.     

PIKE is essential for oligodendroglia development and CNS myelination

Oligodendrocyte (OL) differentiation and myelin development are complex events regulated by numerous signal transduction factors. Here, we report that phosphoinositide-3 kinase enhancer L (PIKE-L) is required for OL development and myelination. PIKE-L expression is up-regulated when oligodendrocyte progenitor cells commit to differentiation. Conversely, depleting phosphoinositide-3 kinase enhancer (PIKE) expression by shRNA prevents oligodendrocyte progenitor cell differentiation. In both conventional PIKE knockout (PIKE^−/−) and OL-specific PIKE knockout mice, the number of OLs is reduced in the corpus callosum. PIKE^−/− OLs also display defects when forming myelin sheath on neuronal axons during neonatal development, which is partially rescued when PTEN is ablated. In addition, Akt/mTOR signaling is impaired in OL-enriched tissues of the PIKE^−/− mutant, leading to reduced expression of critical proteins for myelin development and hypomyelination. Moreover, myelin repair of lysolecithin-induced lesions is delayed in PIKE^−/− brain. Thus, PIKE plays pivotal roles to advance OL development and myelinogenesis through Akt/mTOR activation.Efficient propagation of action potentials depends on the presence of myelin sheath that spirals around the axon. As a membrane extension from oligodendrocytes (OLs), the myelin sheath has a unique lipid-rich composition that allows electrical insulation for high-speed conduction and fidelity of signal transfer (1). Generation of OLs is a developmentally regulated process, which involves the proliferation of oligodendrocyte progenitor cells (OPCs) at the germinal subventricular zones (SVZ), migration throughout the CNS, differentiation into mature OLs, and adhesion to the axon to form myelin (2). Although most OPCs first appear in early neonatal brain, maturation and myelination of OLs in rodents occur largely in postnatal life between P10 and P60 (1). The timing of s differentiation and myelin formation requires highly localized signaling mechanisms, which involves the coordinated activation/inactivation of Wnt/β-catenin, Hedgehog/Gli1, Jagged1/Notch, and PI3K/Akt/mTOR cascades (3). Disruption of these pathways via gene manipulation or modulation of their regulators results in defective OL development. For example, PI3K depletion causes reduced myelin expression in the cerebral cortex and striatum (4). On the other hand, mutation of PTEN, the negative regulator of PI3K/Akt cascade, causes thickening and unraveling of the myelin sheath surrounding hypertrophic axons in the corpus callosum (CC) (5).Phosphoinositide 3-kinase enhancer L (PIKE-L) is a CNS-specific GTPase that belongs to the centaurin family (6, 7). It participates in numerous cellular events to regulate neuronal activity and survival. Our previous studies show that PIKE-L interacts with both netrin receptor (UNC5H) and metabotropic glutamate receptor I (mGlu₁) to prevent apoptotic cell death (8, 9). In addition, PIKE-L controls cell-surface trafficking of 2-amino-3-(5-methyl-3-oxo-1,2-oxazol-4-yl) propanoic acid receptor and the formation of long-term potentiation in the postsynaptic neurons (10). Moreover, PIKE controls the neuronal dendritogenesis and survival through maintaining the integrity of the PI3K/Akt pathway (11). Indeed, PIKE is an important molecular switch to control the cellular PI3K/Akt activation as it links extracellular stimuli including netrin, glutamate, and neurotrophins to the intrinsic PI3K/Akt activities (12–14). Nevertheless, the functions of PIKE-L in nonneuronal cells such as OLs and astrocytes still remain unexplored. In this paper, we report that PIKE-L signals through the PI3K/Akt pathway to advance CNS myelinogenesis.     

Systematic determination of absolute absorption cross-section of individual carbon nanotubes

Optical absorption is the most fundamental optical property characterizing light–matter interactions in materials and can be most readily compared with theoretical predictions. However, determination of optical absorption cross-section of individual nanostructures is experimentally challenging due to the small extinction signal using conventional transmission measurements. Recently, dramatic increase of optical contrast from individual carbon nanotubes has been successfully achieved with a polarization-based homodyne microscope, where the scattered light wave from the nanostructure interferes with the optimized reference signal (the reflected/transmitted light). Here we demonstrate high-sensitivity absorption spectroscopy for individual single-walled carbon nanotubes by combining the polarization-based homodyne technique with broadband supercontinuum excitation in transmission configuration. To our knowledge, this is the first time that high-throughput and quantitative determination of nanotube absorption cross-section over broad spectral range at the single-tube level was performed for more than 50 individual chirality-defined single-walled nanotubes. Our data reveal chirality-dependent behaviors of exciton resonances in carbon nanotubes, where the exciton oscillator strength exhibits a universal scaling law with the nanotube diameter and the transition order. The exciton linewidth (characterizing the exciton lifetime) varies strongly in different nanotubes, and on average it increases linearly with the transition energy. In addition, we establish an empirical formula by extrapolating our data to predict the absorption cross-section spectrum for any given nanotube. The quantitative information of absorption cross-section in a broad spectral range and all nanotube species not only provides new insight into the unique photophysics in one-dimensional carbon nanotubes, but also enables absolute determination of optical quantum efficiencies in important photoluminescence and photovoltaic processes.Single-walled carbon nanotubes (SWNTs), a model one-dimensional nanomaterial system, constitute a rich family of structures (1). Each single-walled nanotube structure, uniquely defined by the chiral index (n,m), exhibits distinct electrical and optical properties (2–5). Quantitative information of SWNT absorption cross-section is highly desirable for understanding nanotube electronic structures, for evaluating quantum efficiency of nanotube photoluminescence (5, 6) and photocurrent (7–9), and for investigating the unique many-body effects in 1D systems (10–16). Despite its obvious importance, reliable experimental determination of nanotube absorption cross-section at the single-tube level is still challenging (17). Previous absorption measurements on ensemble nanotube samples only provide averaged behavior (18–20). Recent absorption studies of individual nanotubes, suffering from small absorption signals and/or slow laser-frequency scanning, cannot determine the absolute absorption cross-section and are limited in achievable spectral range (15, 21–23).We demonstrate here a high-sensitivity polarization-based homodyne method to measure nanotube absorption spectra. By manipulating the light polarization, we enhanced the nanotube-induced transmission contrast, ΔI/I, by two orders of magnitude, and this enhanced transmission contrast can be quantitatively related to nanotube absorption cross-section along and perpendicular to the nanotube axis. Using this polarization control together with supercontinuum laser source, we realized high-throughput and broadband absorption measurements at the single-tube level; combined with electron diffraction technique on the same tube, it enables absolute determination of absorption cross-sections of individual chirality-defined nanotubes, to our knowledge for the first time. We obtained quantitative absorption spectra of over 50 SWNTs of different chiralities, and established a phenomenological formula for absorption cross-sections of different nanotubes. The chirality-dependent nanotube absorption spectra reveal unique 1D photophysics in nanotubes, including a universal scaling behavior of exciton oscillator strength and of exciton resonance linewidth.     

Changes in Early Serum Metal Ion Levels and Impact on Liver,Kidney, and Immune Markers Following Metal-on-Metal Total Hip Arthroplasty

We retrospectively studied 32 consecutive patients (32 hips) who underwent THA with a Durom large-head, MOM articulation between January 2008 and December 2010. Of the patients who underwent THA using a Trilogy metal on polyethylene prosthesis during the same period, 32 were chosen to form the Trilogy group. 32 volunteers were chosen to form the control group. At the last follow-up, serum metal ion levels, liver and kidney function and host immunologic immune responses were evaluated. The mean Co and Cr levels in the Durom group were 4.33- and 1.95-fold higher than those in the Trilogy group. CD3 +, CD4 + and CD8 + cell levels in the Durom group were significantly decreased. The INF-γ level in the Durom group was significantly higher than that in the Trilogy and control groups.     

体脂量是绝经后女性股骨近端骨强度的决定因素

正Lower body weight or body mass index(BMI)has been known to be higher risk of developing osteoporosis and low-energy fractures via mechanical loading and other factors in both men and women[1].Recently,there have been new insights into the relationships between body composition and bone health.However,most of the previous studies regarding the relative effect of body composition on bone mass yielded inconsistent     

治疗中期的帕金森病患者辅助练习医疗气功前后认知电位P300变化观察

目的:探讨帕金森病患者在临床治疗前后辅助练习医疗气功认知电位P300的变化。方法:使用丹麦DantecConcertoSEEG-16道电生理仪器以及“听觉靶-非靶刺激序列”为诱发条件,对33例药物治疗中期的帕金森病患者作医疗气功辅助治疗3个月,观察练习医疗气功前后的P300电位的变化。结果:帕金森病组P300与正常对照组存在多项指标差异,P3潜伏期延迟尤甚。帕金森病组功前靶潜伏期P2(197±25)ms,N2(258±25)ms,P3(348±28)ms,正常对照组上述指标分别为(171±31),(235±27)ms,(313±25)ms;帕金森病组功前非靶潜伏期N1(129±27)ms,P2(226±26)ms,正常对照组上述指标分别为(99±12),(200±33)ms;帕金森病组功前靶波幅P2(4.5±1.8)μV,P3(7.8±1.7)μV,正常对照组上述指标分别为(6.2±2.7),(5.3±2.9)μV,统计学处理以上指标差异均达显著性或极显著性意义(P<0.05或<0.01)。练习医疗气功后其N2、P3潜伏期缩短的同时犤靶潜伏期N2正常对照组(235±27)ms,帕金森病组功前(258±25)ms,功后(240±21)ms,F值7.5,P<0.05;靶潜伏期P3正常对照组(313±25)ms,帕金森病组功前(348±28)ms,功后(323±25)ms,F=14.1,P<0.01〗,其临床症状也获得改善。结论:帕金森病患者有认知功能改变,在药物治疗的同时,辅助医疗气功治疗可使其获得改善。