Synergistic strategy for multicolor two-photon microscopy: application to the analysis of germinal center reactions in vivo

Simultaneous detection of multiple cellular and molecular players in their native environment, one of the keys to a full understanding of immune processes, remains challenging for in vivo microscopy. Here, we present a synergistic strategy for spectrally multiplexed in vivo imaging composed of (i) triple two-photon excitation using spatiotemporal synchronization of two femtosecond lasers, (ii) a broad set of fluorophores with emission ranging from blue to near infrared, (iii) an effective spectral unmixing algorithm. Using our approach, we simultaneously excite and detect seven fluorophores expressed in distinct cellular and tissue compartments, plus second harmonics generation from collagen fibers in lymph nodes. This enables us to visualize the dynamic interplay of all the central cellular players during germinal center reactions. While current in vivo imaging typically enables recording the dynamics of 4 tissue components at a time, our strategy allows a more comprehensive analysis of cellular dynamics involving 8 single-labeled compartments. It enables to investigate the orchestration of multiple cellular subsets determining tissue function, thus, opening the way for a mechanistic understanding of complex pathophysiologic processes in vivo. In the future, the design of transgenic mice combining a larger spectrum of fluorescent proteins will reveal the full potential of our method

Optimal Resting-Growth Strategies of Microbial Populations in Fluctuating Environments

Bacteria spend most of their lifetime in non-growing states which allow them to survive extended periods of stress and starvation. When environments improve, they must quickly resume growth to maximize their share of limited nutrients. Cells with higher stress resistance often survive longer stress durations at the cost of needing more time to resume growth, a strong disadvantage in competitive environments. Here we analyze the basis of optimal strategies that microorganisms can use to cope with this tradeoff. We explicitly show that the prototypical inverse relation between stress resistance and growth rate can explain much of the different types of behavior observed in stressed microbial populations. Using analytical mathematical methods, we determine the environmental parameters that decide whether cells should remain vegetative upon stress exposure, downregulate their metabolism to an intermediate optimum level, or become dormant. We find that cell-cell variability, or intercellular noise, is consistently beneficial in the presence of extreme environmental fluctuations, and that it provides an efficient population-level mechanism for adaption in a deteriorating environment. Our results reveal key novel aspects of responsive phenotype switching and its role as an adaptive strategy in changing environments

“I should have …”:A Photovoice Study With Women Who Have Lost a Man to Suicide

While the gendered nature of suicide has received increased research attention, the experiences of women who have lost a man to suicide are poorly understood. Drawing on qualitative photovoice interviews with 29 women who lost a man to suicide, we completed a narrative analysis, focused on describing the ways that women constructed and accounted for their experiences. We found that women’s narratives drew upon feminine ideals of caring for men’s health, which in turn gave rise to feelings of guilt over the man’s suicide. The women resisted holding men responsible for the suicide and tended to blame themselves, especially when they perceived their efforts to support the man as inadequate. Even when women acknowledged their guilt as illogical, they were seemingly unable to entirely escape regret and self-blame. In order to reformulate and avoid reifying feminine ideals synonymous with selflessly caring for others regardless of the costs to their own well-being, women’s postsuicide bereavement support programs  hould integrate a critical gender approach

Toll-like receptor 4 signaling in liver injury and hepatic fibrogenesis

Toll-like receptors (TLRs) are a family of transmembrane pattern recognition receptors (PRR) that play a key role in innate and adaptive immunity by recognizing structural components unique to bacteria, fungi and viruses. TLR4 is the most studied of the TLRs, and its primary exogenous ligand is lipopolysaccharide, a component of Gram-negative bacterial walls. In the absence of exogenous microbes, endogenous ligands including damage-associated molecular pattern molecules from damaged matrix and injured cells can also activate TLR4 signaling. In humans, single nucleotide polymorphisms of the TLR4 gene have an effect on its signal transduction and on associated risks of specific diseases, including cirrhosis. In liver, TLR4 is expressed by all parenchymal and non-parenchymal cell types, and contributes to tissue damage caused by a variety of etiologies. Intact TLR4 signaling was identified in hepatic stellate cells (HSCs), the major fibrogenic cell type in injured liver, and mediates key responses including an inflammatory phenotype, fibrogenesis and anti-apoptotic properties. Further clarification of the function and endogenous ligands of TLR4 signaling in HSCs and other liver cells could uncover novel mechanisms of fibrogenesis and facilitate the development of therapeutic strategies

Undiagnosed Phenylketonuria Can Exist Everywhere: Results From an International Survey

peer reviewedMany countries do not have a newborn screening (NBS) program, and immigrants from such countries are at risk for late diagnosis of phenylketonuria (PKU). In this international survey, 52 of 259 patients (20%) with late diagnosed PKU were immigrants, and 145 of the 259 (55%) were born before NBS or in a location without NBS. © 2021 The Author