Importance of catalase in the disposal of hydrogen peroxide within human erythrocytes

The catalase within normal, intact human erythrocytes was completely inactivated with amino triazole. The rate of 14CO2 evolution, when the cells were subsequently incubated with 14C-labeled glucose, provided a measure of the rate at which NADPH was being oxidized by the glutathione peroxidase/reductase system for the disposal of H2O2. This rate was determined in control cells and in catalase-inactivated cells while the cells were exposed to H2O2, which was generated at various constant and predetermined rates by glucose oxidase. The results indicated that catalase handles approximately half of the generated H2O2. The glutathione peroxidase/reductase mechanism accounted for the other half. These results are in agreement with our earlier findings on erythrocytes of a subject with a genetic deficiency of catalase. However, an unexpected result with the present approach was the finding that the increased dependence on the glutathione peroxidase/reductase mechanism did not occur until greater than 98% of the catalase had been inactivated. The latter observation indicates that catalase and the glutathione peroxidase/reductase system function intracellularly in a manner very different from that previously ascribed to them. An explanation of the findings requires that the two methods of H2O2 disposal function in a coordinated way, such as a sequential action in which the glutathione peroxidase/reductase system is the rate-limiting step.     

The origin of secondary microseism Love waves

The interaction of ocean surface waves produces pressure fluctuations at the seafloor capable of generating seismic waves in the solid Earth. The accepted mechanism satisfactorily explains secondary microseisms of the Rayleigh type, but it does not justify the presence of transversely polarized Love waves, nevertheless widely observed. An explanation for two-thirds of the worldwide ambient wave field has been wanting for over a century. Using numerical simulations of global-scale seismic wave propagation at unprecedented high frequency, here we explain the origin of secondary microseism Love waves. A small fraction of those is generated by boundary force-splitting at bathymetric inclines, but the majority is generated by the interaction of the seismic wave field with three-dimensional heterogeneity within the Earth. We present evidence for an ergodic model that explains observed seismic wave partitioning, a requirement for full-wave field ambient-noise tomography to account for realistic source distributions.

The surface of the Earth is continuously subjected to perturbing forces that generate seismic waves. Given that 70% of the surface of our planet is covered by oceans, seismic signals due to ocean storms represent the vast majority of seismic data recorded by seismometers on Earth (1). Such data carry information about the energy exchange between different Earth systems, allowing for probing our changing climate (2–4) as well as imaging the internal structure of the Earth (5). The strongest vibrations are called secondary microseisms, excited in the 0.1 to 0.3 Hz frequency range by nonlinear ocean wave–wave interaction (6, 7). They are predominantly composed of seismic surface waves, and Rayleigh waves dominate the vertical component of microseism records (8).The generation mechanism currently accepted for secondary microseisms explains the Rayleigh wave content of vertical-component noise records (9). Secondary microseisms are produced by pressure-like sources at the surface of the ocean. Rayleigh waves are excited below the seafloor due to constructive interference of P and SV body waves. At the ocean–crust interface, they are called Scholte waves when their phase velocity becomes smaller than the minimum phase velocity of the system (10). While at longer periods, ocean waves can directly couple with the seafloor and generate Love waves (11, 12), the generation mechanism of secondary microseisms cannot explain the presence of Love waves on the horizontal components of microseismic records. Observations of secondary microseism Love waves date back to the early (13) and middle (14) 20th century. A few recent studies based on high-quality digital data focused on quantifying the Love-to-Rayleigh ratio in the secondary microseism frequency range (SI Appendix, Table S1). They found that Love-to-Rayleigh ratios are frequency dependent (15) and show a predominance of Rayleigh waves (16, 17), with few exceptions (18).Hypotheses for the generation of secondary microseism Love waves envisage that they can be generated either in the region where the pressure power spectral density (PSD) is strong or along distinct propagation paths within the Earth. The first hypothesis is supported by the presence of bathymetric inclines in the source regions. Such bathymetry may lead to splitting of the vertical second-order pressure force in a component perpendicular to inclines—responsible for Rayleigh waves—and a component tangent to inclines—responsible for Love waves. The second hypothesis is supported by the presence of lateral heterogeneities within the Earth, which can lead to the generation of Love waves due to scattering and focusing/defocusing effects. Ref. 8 observed Love and Rayleigh waves coming from the same direction, concluding that Love waves do originate in the source region. On the other hand, ref. 19 noted that the greater the distance of propagation of Rayleigh waves, the larger the Love wave energy. In addition to these hypotheses, Love waves may originate from Rayleigh-to-Love wave conversion at the ocean–continent boundary, although early numerical simulations suggest that only a few percent of incident Rayleigh wave energy can be converted to Love wave energy (20). To date, no comprehensive theoretical investigations as to which mechanisms can lead to the observed secondary microseism Love waves have been conducted.     

Infection of Nippostrongylus brasiliensis induces development of mucosal-type but not connective tissue-type mast cells in genetically mast cell-deficient Ws/Ws rats

Ws/Ws rats have a small deletion at the tyrosine kinase domain of the c- kit gene and are deficient in both mucosal mast cells (MMC) and connective tissue-type mast cells (CTMC). The role of the c-kit receptor in the development of MMC and CTMC was investigated by infecting Ws/Ws and control +/+ rats with Nippostrongylus brasiliensis (NB), which induces T-cell-dependent mast cell proliferation. Although mast cells did not develop in the skin of Ws/Ws rats, a significant number of mast cells developed in the jejunum after NB infection. These mast cells had the MMC protease phenotype (rat mast cell protease [RMCP] I-/II+) and lacked heparin because they were not stained with berberine sulfate. Globule leukocytes were also detected in the mucosal epithelium of these rats. However, the number of MMC and the serum concentration of RMCP II in NB-infected Ws/Ws rats were only 13% and 7% of those of NB-infected +/+ rats, respectively. A small number of mast cells also developed in the lung, liver, and mesenteric lymph nodes of Ws/Ws rats after NB infection. Although mast cells in these tissues had the MMC phenotype throughout the observation period, the increased mast cells in the lung and liver of +/+ rats acquired a CTMC-like phenotype and were RMCP I+/II+, berberine sulfate+, and formalin resistant. These results indicate that the need for the stimulus through the c-kit receptor appears to be greater in the development of CTMC in the skin as well as for CTMC-like mast cells in the lung and liver than for the development of MMC.     

Mast cells are critical mediators of vaccine-induced Helicobacter clearance in the mouse model

BACKGROUND & AIMS: Despite the proven ability of immunization to prevent Helicobacter infection in mouse models, the precise mechanism of protection has remained elusive. METHODS: We explored the cellular events associated with Helicobacter clearance from the stomach following vaccination by flow cytometry analysis and histological and molecular studies. RESULTS: Kinetic studies showed that the infection is undetectable in vaccinated mice at day 5 postbacterial challenge. Flow cytometry analysis showed that the percentages of mast cells (CD3 - CD117 + ) increased in the lymphoid cells isolated from the stomach at day 4 postchallenge in urease + cholera toxin (CT)-vaccinated mice in comparison with mice administered with CT alone (9.4% +/- 4.4% and 3.1% +/- 1%, respectively, for vaccinated and CT administered, n = 5; P < .01). Quantitative PCR analysis showed an increased messenger RNA (mRNA) expression of the mast cell proteases 1 and 2 at day 5 postchallenge in the stomach of vaccinated mice. In contrast to wild-type mice, mast cell-deficient mice (W/W v mice) were not protected from H felis colonization after vaccination. Indeed only 1 out of 12 vaccinated W/W v mice showed a negative urease test. Remarkably, vaccinated W/W v mice reconstituted with cultured bone marrow-derived mast cells recovered the ability to clear the infection after vaccination (8 out of 10 mast cell-reconstituted mice showed negative urease tests [ P < .006 as compared with wild-type mice]). CONCLUSIONS: These experiments show that mast cells are, unexpectedly, critical mediators of anti- Helicobacter vaccination.     

A relaxation model for memory with high storage density.

We present a relaxation model for memory based on a generalized coulomb potential. The model has arbitrarily large storage capacity and, in addition, well-defined basins of attraction about stored memory states. The model is compared with the Hopfield relaxation model.     

Biomechanical comparison of sagittal-parallel versus non-parallel pedicle screw placement

Background

While convergent placement of pedicle screws in the axial plane is known to be more advantageous biomechanically, surgeons intuitively aim toward a parallel placement of screws in the sagittal plane. It is however not clear whether parallel placement of screws in the sagittal plane is biomechanically superior to a non-parallel construct. The hypothesis of this study is that sagittal non-parallel pedicle screws do not have an inferior initial pull-out strength compared to parallel placed screws.

Methods

The established lumbar calf spine model was used for determination of pull-out strength in parallel and non-parallel intersegmental pedicle screw constructs. Each of six lumbar calf spines (L1-L6) was divided into three levels: L1/L2, L3/L4 and L5/L6. Each segment was randomly instrumented with pedicle screws (6/45 mm) with either the standard technique of sagittal parallel or non-parallel screw placement, respectively, under fluoroscopic control. CT was used to verify the intrapedicular positioning of all screws. The maximum pull-out forces and type of failure were registered and compared between the groups.

Results

The pull-out forces were 5,394 N (range 4,221 N to 8,342 N) for the sagittal non-parallel screws and 5,263 N (range 3,589 N to 7,554 N) for the sagittal-parallel screws (p?=?0.838). Interlevel comparisons also showed no statistically significant differences between the groups with no relevant difference in failure mode.

Conclusion

Non-parallel pedicle screws in the sagittal plane have at least equal initial fixation strength compared to parallel pedicle screws in the setting of the here performed cadaveric calf spine experiments.     

T cell responses are governed by avidity and co-stimulatory thresholds

We analyzed the avidity and CD28-mediated co-stimulatory requirements for the activation of T cells in vivo and in vitro. The strength of the T cell/antigen-presenting cell interaction was varied by using altered peptide ligands for stimulation. Co-stimulatory requirements were studied using T cells from CD28-deficient mice. The results indicate that T cell activation is not an all-or-nothing event, but occurs in distinct steps. For each step, a certain avidity, co-stimulatory threshold or both, must be met. Depending upon the strength of the interaction between the T cell receptor and the major histocompatibility complex/peptide and the presence of CD28 co-stimulatory signals, T cells may undergo blast formation alone or proliferate or eventually both proliferate and differentiate to effector cells. Thus, T cell activation is governed by both avidity and co-stimulatory thresholds.     

Complex decay dynamics of HIV virions,intact and defective proviruses,and 2LTR circles following initiation of antiretroviral therapy

In persons living with HIV-1 (PLWH) who start antiretroviral therapy (ART), plasma virus decays in a biphasic fashion to below the detection limit. The first phase reflects the short half-life (<1 d) of cells that produce most of the plasma virus. The second phase represents the slower turnover (t_1/2 = 14 d) of another infected cell population, whose identity is unclear. Using the intact proviral DNA assay (IPDA) to distinguish intact and defective proviruses, we analyzed viral decay in 17 PLWH initiating ART. Circulating CD4⁺ T cells with intact proviruses include few of the rapidly decaying first-phase cells. Instead, this population initially decays more slowly (t_1/2 = 12.9 d) in a process that largely represents death or exit from the circulation rather than transition to latency. This more protracted decay potentially allows for immune selection. After ∼3 mo, the decay slope changes, and CD4⁺ T cells with intact proviruses decay with a half-life of 19 mo, which is still shorter than that of the latently infected cells that persist on long-term ART. Two-long-terminal repeat (2LTR) circles decay with fast and slow phases paralleling intact proviruses, a finding that precludes their use as a simple marker of ongoing viral replication. Proviruses with defects at the 5′ or 3′ end of the genome show equivalent monophasic decay at rates that vary among individuals. Understanding these complex early decay processes is important for correct use of reservoir assays and may provide insights into properties of surviving cells that can constitute the stable latent reservoir.

For persons living with HIV-1 (PLWH), lifelong adherence to antiretroviral therapy (ART) is critical for maintaining suppression of viral replication and forestalling the development of fatal immunodeficiency. Following initiation of ART, plasma virus levels decay rapidly to below the limit of detection of clinical assays (1–6). Because antiretroviral drugs block new infection of susceptible cells, but not virus production by cells that have an integrated viral genome, this decay must reflect the loss of productively infected cells, cells that were infected prior to the initiation of ART. Productively infected cells could die from viral cytopathic effects, cytolytic host effector mechanisms, or virus-independent T cell turnover. In principle, the decay of plasma virus could also be explained by transition to a nonproductive or latent state of infection. Importantly, the decay is biphasic, indicating the presence of two populations of productively infected cells with different half-lives. Most of the plasma virus is produced by cells that decay very rapidly, with a half-life of less than 1 d. Perelson et al. (4) showed that after most of these cells have decayed, the slope changes, reflecting the slower elimination of a second population of productively infected cells. This population decays with a variable half-life (mean ∼ 2 wk). Although this biphasic decay is a consistent feature of the response to ART, there is still uncertainty about the nature, anatomic location, and fate of the cells responsible for virus production during the first and second phases of decay (referred to here as first- and second-phase cells, respectively). The differences between these two populations have never been elucidated.The first and second phases of decay bring viremia down to below the limit of detection of clinical assays (typically 20 to 50 copies of HIV-1 RNA per mL of plasma) within months of ART initiation, initially raising hope for eradication. However, a latent form of the virus persists in resting memory CD4⁺ T cells (7–14). Initial studies used a quantitative viral outgrowth assay (QVOA) to demonstrate that latently infected resting CD4⁺ T cells purified from PLWH on long-term suppressive ART could be induced to produce replication-competent virus by global T cell activation (8, 9). Longitudinal studies using the QVOA demonstrated that the half-life of the latent reservoir in resting CD4⁺ T cells is 44 mo in PLWH who are adherent to ART. This half-life is long enough to guarantee lifetime persistence of HIV-1 despite ART (12–14). Strategies targeting the latent reservoir in resting CD4⁺ T cells are a major focus of HIV cure research (15–17). In addition to resting CD4⁺ T cells, other cell types may contribute to HIV-1 persistence (18–20).Prior to and immediately following initiation of ART, the frequency of latently infected cells detected by QVOA is substantially higher than frequencies observed in PLWH on long-term ART (21). In principle, several different types of decay processes occurring over the first 6 to 12 mo of treatment could reduce the frequency of latently infected cells to the more stable frequencies observed in PLWH on long-term ART. Early studies by Jerome Zack and Mario Stevenson demonstrated that infected resting CD4⁺ T cells could harbor linear, unintegrated HIV-1 DNA in a state of preintegration latency (22, 23). Following cellular activation, linear unintegrated HIV-1 DNA can be integrated and transcribed, allowing production of virus (22, 23). The half-life of linear, unintegrated forms of the viral genome is not clear, with some studies suggesting that these forms are labile (22, 24–26). Some reverse-transcribed viral genomes can undergo homology-dependent or end-to-end ligation, generating one-long-terminal repeat or two-long-terminal repeat (2LTR) circles, respectively (reviewed in ref. 27). The stability of these forms is also controversial, but they are clearly replication-defective (27–31). Following integration of linear viral cDNA, decay dynamics depend on dynamics of the infected host cells, which can be eliminated by viral cytopathic effects, immune cytolytic effector mechanisms, and normal contraction-phase death of previously activated CD4⁺ T cells (32, 33).While the QVOA provides a definitive minimal estimate of the frequency of latently infected cells, it underestimates reservoir size because not all proviruses in resting CD4⁺ T cells are induced upon one round of maximum T cell activation (34–36). Many replication-competent proviruses require multiple rounds of stimulation for induction. As an alternative to the QVOA, many studies use PCR-based assays to measure proviral DNA. However, the vast majority of HIV-1 proviruses are defective due to apolipoprotein B messenger RNA editing enzyme, catalytic polypeptide-like (APOBEC)-mediated hypermutation or large internal deletions (34, 37–39). PCR-based assays do not distinguish between defective and intact proviruses (40, 41). Although infected cell dynamics have been explored using PCR-based assays (42), the results likely reflect the dynamics of defective proviruses (41). The recently developed intact proviral DNA assay (IPDA) uses two carefully chosen amplicons to probe informative regions of individual proviruses to provide better discrimination between intact and defective proviruses (41, 43). This assay has proven useful in evaluating the long-term dynamics of cells with intact and defective proviruses, demonstrating differences in decay rates that may reflect some vulnerability of cells with intact proviruses to immune effector mechanisms (41, 44, 45).In this study, we use the IPDA to explore the decay of intact and defective proviruses at early time points following initiation of ART. We identify decay processes occurring over intermediate time scales, but with pronounced differences between intact and defective proviruses. Of particular importance is the second-phase decay because infected cells that survive second-phase decay may down-regulate HIV-1 gene expression and enter the stable latent reservoir. Our findings also provide insight into mechanisms for the elimination of the cells with intact viral genomes and into the proper use of assays for the latent reservoir.     

Free Zinc as a Predictive Marker for COVID-19 Mortality Risk

Free zinc is considered to be the exchangeable and biological active form of zinc in serum, and is discussed to be a suitable biomarker for alterations in body zinc homeostasis and related diseases. Given that coronavirus disease 2019 (COVID-19) is characterized by a marked decrease in total serum zinc, and clinical data indicate that zinc status impacts the susceptibility and severity of the infection, we hypothesized that free zinc in serum might be altered in response to SARS-CoV-2 infection and may reflect disease severity. To test this hypothesis, free zinc concentrations in serum samples of survivors and nonsurvivors of COVID-19 were analyzed by fluorometric microassay. Similar to the reported total serum zinc deficit measured by total reflection X-ray fluorescence, free serum zinc in COVID-19 patients was considerably lower than that in control subjects, and surviving patients displayed significantly higher levels of free zinc than those of nonsurvivors (mean ± SD; 0.4 ± 0.2 nM vs. 0.2 ± 0.1 nM; p = 0.0004). In contrast to recovering total zinc concentrations (r = 0.706, p < 0.001) or the declining copper–zinc ratio (r = −0.646; p < 0.001), free zinc concentrations remained unaltered with time in COVID-19 nonsurvivors. Free serum zinc concentrations were particularly low in male as compared to female patients (mean ± SD; 0.4 ± 0.2 nM vs. 0.2 ± 0.1 nM; p = 0.0003). This is of particular interest, as the male sex is described as a risk factor for severe COVID-19. Overall, results indicate that depressed free serum zinc levels are associated with increased risk of death in COVID-19, suggesting that free zinc may serve as a novel prognostic marker for the severity and course of COVID-19.