Search

Extended abstract of a paper presented at Microscopy and Microanalysis 2010 in Portland, Oregon, USA, August 1 – August 5, 2010. [ABSTRACT FROM PUBLISHER]

The aim of this study is to determine the effect of repeated vaccinations on neutralizing SARS-CoV-2 IgG antibody titers, evaluate risk factors for immunological non-response, and to report breakthrough infections in chronic hemodialysis patients., Methods: A prospective, multi-center cohort study in 163 chronic hemodialysis patients was conducted. Antibody titers were measured three months after second, third, and fourth (10 pts) booster vaccinations. SARS-CoV-2 neutralizing antibody titers in BAU/mL and % inhibition were divided into three categories (<216, 216-433, >433 and <33, 33-66, and >66%). Somers's test, paired t-test, and univariable and multivariable logistic regression analysis were applied to evaluate differences in antibody levels and search for risk factors for vaccination failure defined as neutralizing titers <50% and/or need for repeated booster vaccinations. Furthermore, we report on a case series to describe characteristics of patients after four vaccinations ( n = 10) and breakthrough infections ( n = 20)., Results: Third dose boosters resulted in higher proportions of patients with neutralizing antibody levels >66% as compared to after the second dose (64.7% after second dose vs. 88.9% after third dose, p = 0.003), as well as in a respective increase in neutralizing titer levels in % from 68 ± 33% to 89 ± 24 ( p <0.001). The proportion of patients with IgG-titers below 216 BAU/mL decreased from 38.6 to 10.5% ( p ≤ 0.001). Age ( p = 0.004, OR 1.066, 95% CI 1.020-1.114) and presence of immunosuppressive medications ( p = 0.002, OR 8.267, 95% CI 2.206-30.975) were identified as major risk factors for vaccination failure. Repeated booster vaccinations ≥4 times were effective in 8 out of 10 former low-responders (80%) without any side effects or safety concerns. Breakthrough infections showed a clinically mild course but were associated with prolonged viral shedding on PCR-testing ranging 7-29 (mean 13) days., Conclusions: Third and fourth mRNA-based booster vaccinations resulted in higher and longer lasting SARS-CoV-2 antibody levels as compared to after two dosages. The presence of immunosuppressive medication and repeat vaccinations are major potentially modifiable measures to increase antibody levels in non-or low-responders. Breakthrough infections with SARS-CoV-2 Omicron were associated with prolonged viral shedding but clinically mild disease courses.

The aim of this investigation was to determine the effect of SARS-Cov-2 vaccination in hemodialysis patients, search for risk factors for non- or low-response, and to measure the effect of a third booster vaccination in non- or low-responders. Methods SARS-CoV-2 IgG antibodies and the virus-neutralizing capacity were measured 4-5 weeks after a full standard vaccination in 95 chronic hemodialysis patients and 60 controls. IgG titers > 30 AU/mL served to classify participants as responders. Multivariable binary logistic regression analysis was used to search for risk factors of reduced vaccination success. Patients with vaccination failure were offered a third booster dosage. Results 82.1% of the patient cohort as compared to 98.3% of the healthy control group were able to mount SARS-CoV-2 titers above 30 AU/mL after two standard vaccine doses. Mean IgG antibody titers were lower in hemodialysis patients than controls (78 ± 35 vs. 90 ± 20 AU/mL, p = 0.002). Multivariable binary logistic regression analysis showed age and immunosuppressive medication as major risk factors for vaccination failure with a decreased probability of successful vaccination of -6.1% (95% CI -1.2 to -10.9) per increase in age of one year and -87.4% (95% CI -98.0 to -21.5) in patients on immunosuppressive therapy (crude odds ratio for vaccination failure for immunosuppressive therapy 6.4). Ten out of 17 patients with non-response to vaccination were offered a third dose. Booster vaccination after the second dose induced an increase in effective antibody titers of >30 AU/mL in seven out of ten patients 4-5 weeks later (70%). Conclusion Standard SARS-CoV-2 vaccination schemes are highly effective in mounting protective neutralizing IgG antibodies in chronic hemodialysis patients. Nevertheless, response to vaccination is diminished as compared to a healthy control group. Major risk factors for vaccination failure are older age and immunosuppressive therapy. In non- or low-responders to standard vaccination a third booster vaccination was able to induce effective antibody titers in about 70% of patients, indicating that a third booster vaccination might be preferable to decreasing immunosuppressive therapy.

Spatiotemporal control of integrin-mediated cell adhesions to extracellular matrix regulates cell behavior with has numerous implications for biotechnological applications. In this work, two approaches for regulating cell adhesions in space and time with high precision are reported, both of which utilize green light. In the first design, CarH, which is a tetramer in the dark, is used to mask cRGD adhesion-peptides on a surface. Upon green light illumination, the CarH tetramer dissociates into its monomers, revealing the adhesion peptide so that cells can adhere. In the second design, the RGD motif is incorporated into the CarH protein tetramer such that cells can adhere to surfaces functionalized with this protein. The cell adhesions can be disrupted with green light, due to the disassembly of the CarH-RGD protein. Both designs allow for photoregulation with noninvasive visible light and open new possibilities to investigate the dynamical regulation of cell adhesions in cell biology., (© 2020 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA.)

Cell adhesions to the extracellular matrix and to neighboring cells are fundamental to cell behavior and have also been implemented into minimal synthetic cells, which are assembled from molecular building blocks from the bottom-up. Investigating adhesion in cell mimetic models with reduced complexity provides a better understanding of biochemical and biophysical concepts underlying the cell adhesion machinery. In return, implementing cell-matrix and cell-cell adhesions into minimal synthetic cells allows reconstructing cell functions associated with cell adhesions including cell motility, multicellular prototissues, fusion of vesicles, and the self-sorting of different cell types. Cell adhesions have been mimicked using both the native cell receptors and reductionist mimetics providing a variety of specific, reversible, dynamic, and spatiotemporally controlled interactions. This review gives an overview of different minimal adhesion modules integrated into different minimal synthetic cells drawing inspiration from cell and colloidal science., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The dynamic and spatiotemporal control of integrin-mediated cell adhesion to RGD motifs in its extracellular matrix (ECM) is important for understating cell biology and biomedical applications because cell adhesion fundamentally regulates cellular behavior. Herein, the first photoswitchable synthetic ECM protein, Photo-ECM, based on the blue light switchable protein LOV2 is engineered. The Photo-ECM protein includes a RGD sequence, which is hidden in the folded LOV2 protein structure in the dark and is exposed under blue light so that integrins can bind and cells can adhere. The switchable presentation of the RGD motif allows to reversibly mediate and modulate integrin-based cell adhesions using noninvasive blue light. With this protein cell adhesions in live cells could be reversed and the dynamics at the cellular level is observed. Hence, the Photo-ECM opens a new possibility to investigate the spatiotemporal regulation of cell adhesions in cell biology and is the first step toward a genetically encoded and light-responsive ECM., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Biofilms are not only a leading cause of chronic infections and biofouling, but they also have a tremendous positive potential in biotechnology for biocatalysis and waste treatment. Biofilms are spatially structured communities of microbes, which exchange chemicals and communicate with each other. By spatially controlling bacterial adhesion to surfaces, and therefore the microstructure of biofilms, a promising method of understanding social interactions between bacteria and designed biofilms is developed. The bacterial photolithography approach described here allows to photopattern specific bacteria adhesion molecules, to control surface adhesion, and to guide the formation of biofilms. To do this, α-D-mannoside, which is recognized by the Escherichia coli FimH receptor, is linked to a nonadhesive polyethylene glycol surface through a photocleavable 2-nitrobenzyl linker. When a pattern of UV light in a specific shape is projected onto these surfaces, the light-exposed areas become nonadhesive and bacteria only adhere to the dark, unexposed areas in the photopattern. Bacterial photolithography enables bacterial patterning with high spatial resolution down to 10 µm without mechanical interference. Additionally, patterning biofilms with complicated geometries allows studying the importance of microscale spatial organization on the collective behavior of bacteria such as quorum sensing., (© 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

We use plasmon rulers to follow the conformational dynamics of a single protein for up to 24 h at a video rate. The plasmon ruler consists of two gold nanospheres connected by a single protein linker. In our experiment, we follow the dynamics of the molecular chaperone heat shock protein 90 (Hsp90), which is known to show "open" and "closed" conformations. Our measurements confirm the previously known conformational dynamics with transition times in the second to minute time scale and reveals new dynamics on the time scale of minutes to hours. Plasmon rulers thus extend the observation bandwidth 3-4 orders of magnitude with respect to single-molecule fluorescence resonance energy transfer and enable the study of molecular dynamics with unprecedented precision.

Independent control over multiple cell-material interactions with high spatiotemporal resolution is a key for many biomedical applications and understanding cell biology, as different cell types can perform different tasks in a multicellular context. In this study, the binding of two different cell types to materials is orthogonally controlled with blue and red light providing independent regulation in space and time. Cells expressing the photoswitchable protein cryptochrome 2 (CRY2) on cell surface bind to N-truncated CRY-interacting basic helix-loop-helix protein 1 (CIBN)-immobilized substrates under blue light and cells expressing the photoswitchable protein phytochrome B (PhyB ) on cell surface bind to phytochrome interaction factor 6 (PIF6)-immobilized substrates under red light, respectively. These light-switchable cell interactions provide orthogonal and noninvasive control using two wavelengths of visible light. Moreover, both cell-material interactions are dynamically switched on under light and reversible in the dark. The specificity of the CRY2/CIBN and PhyB/PIF6 interactions and their response to different wavelengths of light allow selectively activating the binding of one cell type with blue and the other cell type with red light in the presence of the other cell type.

Neurospora crassa has been systematically investigated for circadian entrainment behavior. Many aspects of synchronization can be investigated in this simple, cellular system, ranging from systematic entrainment and drivenness to masking. Clock gene expression during entrainment and entrainment without clock genes suggest that the known transcription/translation feedback loop is not alone responsible for entrainment in Neurospora.

This review is intended as a summary of our work carried out as part of the German Research Association (DFG) Center Program on Circadian Rhythms. Over the last six years, our approach to understanding circadian systems combined theoretical and experimental tools, and Gonyaulax and Neurospora have proven ideal for these efforts. Both of these model organisms demonstrate that even simple circadian systems can have multiple light input pathways and more than one rhythm generator. They have both been used to elaborate basic circadian features in conjunction with formal models. The models introduce the "zeitnehmer," i.e., a clock-regulated input pathway, to the conceptual framework of circadian systems, and proposes networks of individual feedbacks as the basis for circadian rhythmicity.

Objective: To examine the effects of peptidyl membrane interactive molecule D4B in a murine model of lethal burn wound infection., Experimental Design: Four experiments were performed: (1) growth inhibition assays of Pseudomonas aeruginosa treated with D4B, 0 to 100 micromol/L; (2) in vitro coculture of bone marrow cells with D4B, 0 to 100 micromol/L; (3) D4B treatment survival studies after burn injury only or burn wound infection in mice; and (4) peripheral white blood cell count, burn wound tissue bacterial culture, and burn wound morphological analysis at days 1, 2, and 3 after injury., Setting: University medical center laboratory., Subjects: Groups of B6D2F1 male mice (20 each) were studied., Interventions: Full-thickness scald burn, 15% of total body surface area, with P aeruginosa topical infection, and subeschar injections of D4B at 200 microg or 0.25 mL of placebo per mouse at 2 and 24 hours after injury., Main Outcome Measures: Animal survival after thermal burn wound bacterial infection, circulating leukocyte numbers, in vitro clonal cell culture of granulocyte-macrophage progenitor cells, and wound histopathological analysis., Results: The survival rate in the D4B-treated group was nearly 2-fold greater than that in controls (P<.01) during 14 days of study. Bacterial quantitative wound cultures disclosed significant reductions in bacterial numbers at days 1, 2, and 3 in D4B-treated animals as compared with controls (P<.05 to <.01). D4B induced a dose-dependent inhibition of bacterial cell growth when added to in vitro P aeruginosa cultures (P<.01). Granulocyte-macrophage progenitor cell growth in culture was not altered by D4B treatment. D4B-treated animals displayed no signs of toxic effects or impairment in wound healing., Conclusions: The peptidyl membrane interactive molecule D4B had the ability to improve survival after gram-negative burn wound sepsis via direct antimicrobial effects. Peptidyl membrane interactive molecules may offer the potential of alternative treatments to standard topical agents or in patients with drug-resistant microbes.

Suppressed granulocyte and macrophage growth after burn infection or endotoxicosis appears to be mediated by macrophage-derived products. In this study, we found that after burn, burn plus infection, or endotoxicosis, peritoneal-elicited macrophages or bone marrow cells released increased amounts of prostaglandin E2 (PGE2) and inhibited growth of granulocyte-macrophage progenitor cells (GM-CFC). PGE2, when added in culture, inhibited in vitro GM-CFC growth in a dose-dependent manner. Pretreatment of bone marrow cells with either dibutyryl cyclic adenosine monophosphate or Forskolin in vitro mimicked the PGE2 inhibition, further aggravated the inhibition induced by burn, burn plus infection, or endotoxicosis, and was not blocked by co-culture with indomethacin. Pretreatment of bone marrow cells with SQ22536, an adenylate cyclase inhibitor, significantly restored the suppressed GM-CFC growth found after burn, burn plus infection, or endotoxicosis. Alterations in myeloid production after burn infection appear to be related in part to the level of intracellular cyclic adenosine monophosphate for the GM-CFC and are responsive to PGE2.

The serotonergic system in brain is adversely affected by both aging and chronic ethanol consumption. The present study examined the combined effects of aging and chronic ethanol consumption on two components of the serotonergic system. Serotonin (5-HT) reuptake sites and 5-HT2A receptors were quantitated in brain areas of 5-, 14-, and 24-month-old male Fischer 344 rats that were pair-fed a control or 6.6% (v/v) ethanol-containing liquid diet on a chronic basis. The regions examined include those containing the cell bodies and projections of serotonergic neurons. These experiments demonstrated the sensitivity of the serotonergic system of male Fischer 344 rats to both aging and chronic ethanol consumption. In control rats, aging was associated with a decline in the concentration of 5-HT2A receptors in the nucleus accumbens and four cortical regions: frontal, parietal, piriform, and cingulate cortex. 5-HT2A receptors were also reduced in the frontal, parietal, and cingulate cortex of aged ethanol-fed rats. In contrast, 5-HT reuptake sites were increased in older rats in the frontal cortex, nucleus accumbens, amygdala, and CA3 region of the hippocampus. If comparable changes in 5-HT2A receptors and 5-HT reuptake sites occur in elderly humans, they may contribute to ethanol consumption, and lead to cognitive and other age-related problems. These changes may also alter the effectiveness of serotonergic drugs used in the treatment of alcoholism and mental disorders. The effects of chronic ethanol consumption were more limited. The only significant ethanol effect was an increase of 5-HT2A receptors in the nucleus accumbens of 5-month-old ethanol-fed rats.

The present study examined the hypothesis that chronic alcoholism augments the age-related loss of dopamine D1 receptors. This hypothesis was investigated because previous studies reported that both aging and chronic alcoholism produce significant changes in dopaminergic systems, and because chronic alcoholism potentiates some age-related CNS losses. In addition, this study investigated the effects of aging on D1 receptors in animals 1 and 7 days after withdrawal from chronic ethanol. Quantitative autoradiography was used to measure [3H]SCH 23390 binding to D1 receptors in brain areas associated with both the nigrostriatal and mesocorticolimbic dopamine systems. Receptors were assessed in 5-, 14-, and 24-month-old male Fischer 344 rats that were pair-fed a control or 6.6% (v/v) ethanol-containing liquid diet for 6 weeks. The results of these studies demonstrated that aging is associated with a significant decline in D1 receptors in the rostral and caudal striatum, and substantia nigra of both control and ethanol-fed rats. These receptor changes in the nigrostriatal system may be associated with motor abnormalities. In addition, there was an age-related decline in D1 receptors in two brain areas of the mesocorticolimbic system: the nucleus accumbens and frontal cortex. The latter findings may be important because of the involvement of this system with the rewarding properties of ethanol and other drugs of abuse. There were no age-related differences in the response of D1 receptors to ethanol withdrawal in the caudal and rostral striatum, substantia nigra, and nucleus accumbens.(ABSTRACT TRUNCATED AT 250 WORDS)