Changes In Cell Size Research Articles

Empagliflozin Reduces High Glucose-Induced Cardiomyopathy in hiPSC-Derived Cardiomyocytes : Glucose-induced Lipotoxicity in hiPSC-Derived Cardiomyocytes.

Human-induced pluripotent stem cell (hiPSC) technology has been applied in pathogenesis studies, drug screening, tissue engineering, and stem cell therapy, and patient-specific hiPSC-derived cardiomyocytes (hiPSC-CMs) have shown promise in disease modeling, including diabetic cardiomyopathy. High glucose (HG) treatment induces lipotoxicity in hiPSC-CMs, as evidenced by changes in cell size, beating rate, calcium handling, and lipid accumulation. Empagliflozin, an SGLT2 inhibitor, effectively mitigates the hypertrophic changes, abnormal calcium handling, and contractility impairment induced by HG. Glucose concentration influences SGLT2 expression in cardiomyocytes, highlighting its potential role in diabetic cardiomyopathy. These findings support the potential utility of hiPSC-CMs in studying diabetic cardiomyopathy and the efficacy of empagliflozin in ameliorating HG-induced cardiomyocyte dysfunction. Such research may advance developments in precision medicine and therapeutic interventions for patients with diabetic cardiomyopathy.

High-Quality Factor Ultrathin Polarization-Insensitive Wide Angular Stable Elliptical-Shaped Microwave Absorber with Reconfigurability Features

This paper introduces an ultrathin, polarization-insensitive, wide angular stable microwave absorber characterized by a high-Quality (Q) factor. The absorber consists of elliptical metallic rings with a thickness of 0.035 mm on the top of Roger’s RT 5880 substrate with a thickness of 0.51 mm and a metallic ground plane with a thickness of 0.035 mm at the bottom. The proposed structure is fourfold symmetric which makes it polarization insensitive. The structure offers angular stability till 60° for the angle of incidence for both Transverse Electric (TE) and Transverse Magnetic (TM) modes under oblique incidence. The structure provides a high-quality factor of 155 at an operating frequency of 7.75 GHz. Also, the structure provides reconfigurability with a change in the axis ratio of the elliptical structure from 7.30 to 9.48 GHz when the axis ratio changes from 0.26 to 0.36 without any change in the unit cell size. The unit cell dimension is 1.07 λg x 1.07 λg x 0.022 λg. With its high Q-factor, thin size, structural simplicity, insensitivity to polarization, and incident angle stability suggested prototype is apt for microwave sensors and antennas to reduce Radar cross-section and X-band satellite communication.

MRI-based analysis of the microstructure of the thalamus and hypothalamus and functional connectivity between cortical networks in episodic cluster headache

BackgroundNeuroimaging studies have shown that hypothalamic/thalamic nuclei and other distant brain regions belonging to complex cerebral networks are involved in cluster headache (CH). However, the exact relationship between these areas, which may be dependent or independent, remains to be understood. We investigated differences in resting-state functional connectivity (FC) between brain networks and its relationship with the microstructure of the hypothalamus and thalamus in patients with episodic CH outside attacks and healthy controls (HCs).MethodsWe collected 3T MRI data from 26 patients with CH during the in-bout period outside the attacks and compared them with data from 20 HCs. From resting-state data we derived independent component (IC) networks. We calculated the fractional anisotropy (FA) and mean (MD), axial (AD), and radial (RD) diffusivity values of the hypothalamus and bilateral thalami and correlated them with resting-state IC Z-scores and CH clinical features.ResultsPatients with CH had less FC between the salience network (SN) and left executive control network (ECN) than HCs, but more FC between the default mode network and right ECN. Patients with CH showed lower FA and higher MD microstructural hypothalamic metrics than HCs. Patients with CH had a higher bilateral FA metric in the thalamus than HCs. The AD and RD diffusivity metrics of the hypothalamus were positively correlated with the disease history duration. We found no correlations between the hypothalamic and thalamic diffusivity metrics and the FC of the cortical networks.ConclusionOur findings presented the possibility of a correlation between the FC of the SN and the inability to switch between internalizing and externalizing brain activity during demanding cognitive tasks, such as recurring headaches. Moreover, we found differences in the thalamic and hypothalamic microstructures that may independently contribute to the pathophysiology of CH. These differences may reflect changes in directional organization, cell size, and density.

Sensitivity of bulk electrical impedance spectroscopy (bio-capacitance) probes to cell and culture properties: Study on CHO cell cultures.

Bulk electrical impedance spectroscopy (bio-capacitance) probes, hold significant promise for real-time cell monitoring in bioprocesses. Focusing on Chinese hamster ovary (CHO) cells, we present a sensitivity analysis framework to assess the impact of cell and culture properties on the complex permittivity spectrum, εmix, and its associated parameters, permittivity increment, Δε, critical frequency, fc, and Cole-Cole parameter, α, measured by bio-capacitance probes. Our sensitivity analysis showed that Δε is highly sensitive to cell size and concentration, making it suitable for estimating biovolume during the exponential growth phase, whereas fc provides information about cumulative changes in cell size, membrane permittivity, and cytoplasm conductivity during the transition to death phase. The analysis indicated that specific information about cell membrane permittivity or internal conductivity cannot be extracted from εmix spectrum. Based on the sensitivity analysis, we proposed two alternative parameters for monitoring cells in bioprocesses: Δε1 MHz and Δε1 MHz/Δε0.3 MHz, using measurements at 300 kHz, 1 MHz, and 10 MHz. Δε1 MHz is suitable for estimating viable cell density during the exponential growth phase due to its lower sensitivity to cell size. Δε1 MHz/Δε0.3 MHz can replace fc due to similar sensitivities to cell size and dielectric properties. These frequencies are within most bio-capacitance probes' optimal operation range, eliminating the need for low-frequency electrode polarization and high-frequency stray capacitances corrections. Experimental measurements on CHO cells confirmed the results of sensitivity analysis.

Bwa, an ortholog of alkaline ceramidase-ACER2, promotes intestinal stem cell proliferation through pro-inflammatory cytokine signaling in Drosophila melanogaster.

Sphingolipids, including ceramides, are an important component of high-fat diets. These molecules can regulate fatty acid oxidation and intestinal stem cell proliferation, predisposing the gut to tumorigenesis. However, the molecular mechanisms involved in ceramide metabolism-mediated intestinal stem cell (ISC) proliferation and tumorigenesis are poorly understood. To understand how changes in sphingolipid metabolite flux affect intestinal stem cells, we manipulated the activities of each of the enzymes of the ceramide synthetic pathway using cell type-specific over-expression or depletion of the corresponding mRNAs in each intestinal cell type of the Drosophila midgut. We documented cell-autonomous and non-cell-autonomous effects, including alterations in cell size, number, differentiation, and proliferation. In our screen, the altered expression of several ceramide metabolism enzymes led to changes in ISC proliferation, cell sizes, and overall cellularity. Among other genes, over-expression of ceramidase homolog, Brain washing (bwa) in gut enteroblasts (EB) increased EB cell size and caused a non-cell-autonomous, 7-8-fold increase in ISC proliferation. Our analysis confirmed previous reports that bwa does not have ceramidase activity, and lipidomic studies indicated that bwa increases the saturation status of sphingolipids, free fatty acids, and other lipids. The pro-proliferative effects of bwa could be counter-acted by depleting a serine palmitoyltransferase, Lace , or a sphingosine acyltransferase, Schlank , which are needed for ceramide synthesis, or by co-expressing a ceramide desaturase enzyme, ifc , indicating that increased saturated ceramides were causal for ISC proliferation and the disruption of gut homeostasis. Accumulating saturated sphingolipids and fatty acids induced inflammatory signaling in the gut, and activated ISC proliferation through the pro-inflammatory cytokines, Upd3 and Upd2. We propose that saturated sphingolipids promote ISC proliferation through pro-inflammatory pathways.

Detonation effect of hydrogen-oxygen mixtures at various initial pressures and hydrogen concentrations in obstructed channels

Numerical investigations have been conducted for the effects of initial ambient pressure, hydrogen concentration, and obstacle distribution on the chemical kinetic characteristics of stable detonation propagation, as well as the changes in the shape and size of detonation cells. The configuration studied is a rectangular channel with regularly spaced obstacles containing the mixture of hydrogen and oxygen. The results indicate that under the same blockage ratio conditions, the position where detonation is generated in the channel with obstacles distributed on one side is farther than in the channel with obstacles distributed on both sides. However, there is no significant change in flame speed or detonation cell size after reaching stable detonation in both cases. The study reveals that under fuel-rich conditions, an increase in hydrogen concentration leads to an increasing duration for the flame to propagate to the end of the channel, accompanied by the distance from the ignition point to the occurrence of detonation also increases. Additionally, under fuel-lean conditions, as the hydrogen concentration decreases, the distance from the ignition point to the occurrence of detonation also increases. The study identifies the limiting concentration for hydrogen detonation is in the range of 18.3 %–60 %. As the initial pressure within the channel decreases, the location of detonation occurrence is farther away from the ignition point. And higher initial ambient pressures make detonation more likely to occur within the channel and the size of stable detonation cells decreases as the initial ambient pressure increases.

Effect of temperature, nutrients and growth rate on picophytoplankton cell size across the Atlantic Ocean

The cell size of picophytoplankton populations affects their ecology and biogeochemical role, but how different environmental drivers control its variability is still not well understood. To gain insight into the role of temperature and nutrient availability as determinants of picophytoplankton population mean cell size, we carried out five microcosm experiments across the Atlantic Ocean (45°N-27°S) in which surface plankton assemblages were incubated under all combinations of three temperatures (in situ, 3 °C cooling and 3 °C warming) and two nutrient levels (unamended and addition of nitrogen and phosphorus). The overall range of variability in cell volume was 5-fold for Prochlorococcus, 8-fold for Synechococcus and 6-fold for the picoeukaryotes. We observed, in all the treatments and in the control, a consistent trend toward larger mean cell sizes over time for both Prochlorococcus and Synechococcus, which was likely the result of sample confinement. Changes in temperature and nutrient status alone did not cause clear changes in cell size, relative to the control, but the combination of warming and nutrient addition resulted in an increase in Prochlorococcus and Synechococcus cell size. The largest increases in cell volume were associated with slow or negative population net growth rates. Our results emphasize the importance of considering changes in biovolume to obtain accurate estimates of picophytoplankton biomass and suggest that the inverse relationship between growth rate and population mean cell size may be a general pattern in marine phytoplankton.

Structural changes in Nelumbo flower petals during opening and closing.

Nelumbo nucifera is one of several plant species with flowers that typically open in the early morning and close by noon. This movement normally repeats for 3 days, with all petals falling off on day 4. However, detailed observations of flower movement in Nelumbo species are limited. Movement of flowers on plants in growth chambers were observed using time-lapse photography. Petals were examined with scanning electron microscopy or fixed and sectioned for light microscopy to determine whether changes in cell size contribute to petal elongation during flowering. This study is the first to microscopically observe petal cells of a Nelumbo species during flowering in controlled conditions. Petals elongated during the 4-day flowering period. Inner petals were more elongated than the outer petals. Among the basal, central, and tip regions of a single petal, the cells in the basal region enlarged the most. Outer and inner epidermal cells in the basal region of the inner petals, on both adaxial and abaxial sides, gradually enlarged during the flowering period through cycles of repeated increases as the petals opened and decreases as they closed. Petals opened and closed repeatedly as they elongated, primarily in their basal region. Cells in the basal region of the petal on the adaxial and abaxial sides periodically fluctuated in size, increased during flower opening and decreased during closing, but differences in these patterns were observed between the two sides, suggesting that one of the driving forces behind the opening and closing of flowers of Nelumbo species is the increase and decrease in the size of the petal cells.

OrgaMapper: a robust and easy-to-use workflow for analyzing organelle positioning

BackgroundEukaryotic cells are highly compartmentalized by a variety of organelles that carry out specific cellular processes. The position of these organelles within the cell is elaborately regulated and vital for their function. For instance, the position of lysosomes relative to the nucleus controls their degradative capacity and is altered in pathophysiological conditions. The molecular components orchestrating the precise localization of organelles remain incompletely understood. A confounding factor in these studies is the fact that organelle positioning is surprisingly non-trivial to address e.g., perturbations that affect the localization of organelles often lead to secondary phenotypes such as changes in cell or organelle size. These phenotypes could potentially mask effects or lead to the identification of false positive hits. To uncover and test potential molecular components at scale, accurate and easy-to-use analysis tools are required that allow robust measurements of organelle positioning.ResultsHere, we present an analysis workflow for the faithful, robust, and quantitative analysis of organelle positioning phenotypes. Our workflow consists of an easy-to-use Fiji plugin and an R Shiny App. These tools enable users without background in image or data analysis to (1) segment single cells and nuclei and to detect organelles, (2) to measure cell size and the distance between detected organelles and the nucleus, (3) to measure intensities in the organelle channel plus one additional channel, (4) to measure radial intensity profiles of organellar markers, and (5) to plot the results in informative graphs. Using simulated data and immunofluorescent images of cells in which the function of known factors for lysosome positioning has been perturbed, we show that the workflow is robust against common problems for the accurate assessment of organelle positioning such as changes of cell shape and size, organelle size and background.ConclusionsOrgaMapper is a versatile, robust, and easy-to-use automated image analysis workflow that can be utilized in microscopy-based hypothesis testing and screens. It effectively allows for the mapping of the intracellular space and enables the discovery of novel regulators of organelle positioning.

A Sensitivity Test on the Modifiable Areal Unit Problem in the Spatial Aggregation of Fossil Data

In paleobiology and macroevolution research, the spatial aggregation of fossil data can be influenced by the modifiable areal unit problem (MAUP), wherein the selection of different grid-cell sizes for data aggregation can lead to variations in statistical results. This study presents a case analysis focused on the spatial extent of marine bivalves and brachiopods over time across three Areas of Interest (AOIs) to evaluate the potential impact of the MAUP in grid-based fossil data processing. By employing rectangular grid matrices with cell sizes of 50, 100, 200, and 400 km, this research assesses the MAUP-related sensitivity of two commonly used grid-based proxies for species’ spatial distribution. The results reveal that the proxy based on the number of occupied grid cells (OGCs) is particularly sensitive to changes in cell size, whereas the proxy based on minimum-spanning-tree distance (MST distance) demonstrates greater robustness across varying grid scales. This study underscores that when constructing proxies for species’ spatial distribution ranges using grid matrices, the OGC method is more susceptible to MAUP effects than the MST distance method, warranting increased caution in studies employing the OGC approach.

Quantitative label-free digital holographic imaging of cardiomyocyte optical volume, nucleation, and cell division

Cardiac regeneration in newborn rodents depends on the ability of pre-existing cardiomyocytes to proliferate and divide. This capacity is lost within the first week of postnatal development when these cells rapidly switch from hyperplasia to hypertrophy, withdraw from the cell cycle, become binucleated, and increase in size. How these dynamic changes in cell size and nucleation impact cardiomyocyte proliferative potential is not well understood. In this study, we innovate the application of a commercially available digital holographic imaging microscope, the Holomonitor M4, to evaluate the proliferative responses of mononucleated and binucleated cardiomyocytes after CHIR99021 treatment, a model proliferative stimulus. This system enables long-term label-free quantitative tracking of primary cardiomyocyte dynamics in real-time with single-cell resolution. Our results confirm that chemical inhibition of glycogen synthase kinase 3 with CHIR99021 promotes complete cell division of both mononucleated and binucleated cardiomyocytes with high frequency. Quantitative tracking of cardiomyocyte volume dynamics during these proliferative events revealed that both mononucleated and binucleated cardiomyocytes reach a similar size-increase threshold prior to attempted cell division. Binucleated cardiomyocytes attempt to divide with lower frequency than mononucleated cardiomyocytes, which may be associated with inadequate increases in cell size. By defining the interrelationship between cardiomyocyte size, nucleation, and cell cycle control, we may better understand the cellular mechanisms that drive the loss of mammalian cardiac regenerative capacity after birth.

Temperature-dependent akinete formation strategies of the harmful cyanobacterium Dolichospermum circinale

Cyanobacteria from the orders Nostocales and certain Stigonematales form akinetes, spore-like dormant cells that allow them to survive adverse environmental conditions. Temperature is known to be one of the key factors affecting akinete formation, but there is currently little known about akinete formation during cell growth over a wide range of temperature conditions and its relation to the overall survival strategy of cyanobacteria. Therefore, in the current study, we conducted a temperature-controlled experiment to analyze the akinete formation of a harmful cyanobacterium Dolichospurmum circinale using a growth chamber. We measured the concentration and size of both vegetative cells and different types of akinetes (free, attached, and empty type) under varying temperatures (5–25 °C). We also analyzed the buoyant ability and vertical migration velocity of trichomes along with changes in the volume of vegetative cells and akinetes. The total akinete concentration and ratio (number of akinetes to total number of cells) were both found to be higher at high temperatures (20–25°C) than they were at low temperatures (5–10°C) (p<0.05). Meanwhile, the rate of formation of akinetes (both free and attached akinetes) was highest at low temperature (10 °C) and decreased with increasing temperature. The rate of empty akinete formation increased with increasing temperature and was highest at 25°C, indicating that most of the akinetes produced under high temperature conditions germinated. The change in vegetative cell size was proportional to the increase in the growth rate in response to increasing temperature (p<0.05). At high temperature, vegetative cells exhibited positive buoyancy and higher vertical migration velocity, while at low temperature, they exhibited negative buoyancy and relatively low migration velocity. Akinete size was larger at low temperature than it was at high temperature. These findings suggest that akinetes play an important role in maintaining populations in the water column, with a link between akinete formation and germination during summer cyanobacteria blooms. This information is expected to contribute to a deeper understanding of the D. circinale life cycle.

Cell size regulates human endoderm specification through actomyosin-dependent AMOT-YAP signaling

Cell size is a crucial physical property that significantly impacts cellular physiology and function. However, the influence of cell size on stem cell specification remains largely unknown. Here, we investigated the dynamic changes in cell size during the differentiation of human pluripotent stem cells into definitive endoderm (DE). Interestingly, cell size exhibited a gradual decrease as DE differentiation progressed with higher stiffness. Furthermore, the application of hypertonic pressure or chemical to accelerate the reduction in cell size significantly and specifically enhanced DE differentiation. By functionally intervening in mechanosensitive elements, we have identified actomyosin activity as a crucial mediator of both DE differentiation and cell size reduction. Mechanistically, the reduction in cell size induces actomyosin-dependent angiomotin (AMOT) nuclear translocation, which suppresses Yes-associated protein (YAP) activity and thus facilitates DE differentiation. Together, our study has established a novel connection between cell size diminution and DE differentiation, which is mediated by AMOT nuclear translocation. Additionally, our findings suggest that the application of osmotic pressure can effectively promote human endodermal lineage differentiation.

Cell-to-cell signaling in cell populations with large cell size variability.

Sizes of multiple cells vary when they communicate with each other. Differences in cell size result in variations in the cell surface area and volume, as well as the number of enzymes and receptors involved in signal transduction. Although heterogeneity in cell size may inhibit uniformity in signaling, cell-to-cell signaling is still possible. The outcome when cell size changes to an extreme degree remains unclear. Hence, we inhibited cell division in Dictyostelium cells, a model organism for signal transduction, to gain insights into the consequences of extreme cell size variations. Measurements of cell signals in this population using fluorescence microscopy indicated that the giant cells can communicate with normal-sized cells by suppressing the signal level. Simulations of signal transduction based on the FitzHugh-Nagumo model also suggested similar results.Our findings suggest that signaling mechanism homogenizes cell-to-cell signaling in response to cell size.

Direct and indirect cumulative effects of temperature, nutrients, and light on phytoplankton growth.

Temperature and resource availability are pivotal factors influencing phytoplankton community structures. Numerous prior studies demonstrated their significant influence on phytoplankton stoichiometry, cell size, and growth rates. The growth rate, serving as a reflection of an organism's success within its environment, is linked to stoichiometry and cell size. Consequently, alterations in abiotic conditions affecting cell size or stoichiometry also exert indirect effects on growth. However, such results have their limitations, as most studies used a limited number of factors and factor levels which gives us limited insights into how phytoplankton respond to environmental conditions, directly and indirectly. Here, we tested for the generality of patterns found in other studies, using a combined multiple-factor gradient design and two single species with different size characteristics. We used a structural equation model (SEM) that allowed us to investigate the direct cumulative effects of temperature and resource availability (i.e., light, N and P) on phytoplankton growth, as well as their indirect effects on growth through changes in cell size and cell stoichiometry. Our results mostly support the results reported in previous research thus some effects can be identified as dominant effects. We identified rising temperature as the dominant driver for cell size reduction and increase in growth, and nutrient availability (i.e., N and P) as dominant factor for changes in cellular stoichiometry. However, indirect effects of temperature and resources (i.e., light and nutrients) on species' growth rates through cell size and cell stoichiometry differed across the two species suggesting different strategies to acclimate to its environment.

Sucrose promotes cone enlargement via the TgNGA1-TgWRKY47-TgEXPA2 module in Torreya grandis.

Cone enlargement is a crucial process for seed production and reproduction in gymnosperms. Most of our knowledge of cone development is derived from observing anatomical structure during gametophyte development. Therefore, the exact molecular mechanism underlying cone enlargement after fertilization is poorly understood. Here, we demonstrate that sucrose promotes cone enlargement in Torreya grandis, a gymnosperm species with relatively low rates of cone enlargement, via the TgNGA1-TgWRKY47-TgEXPA2 pathway. Cell expansion plays a significant role in cone enlargement in T. grandis. 13C labeling and sucrose feeding experiments indicated that sucrose-induced changes in cell size and number contribute to cone enlargement in this species. RNA-sequencing analysis, transient overexpression in T. grandis cones, and stable overexpression in tomato (Solanum lycopersicum) suggested that the expansin gene TgEXPA2 positively regulates cell expansion in T. grandis cones. The WRKY transcription factor TgWRKY47 directly enhances TgEXPA2 expression by binding to its promoter. Additionally, the NGATHA transcription factor TgNGA1 directly interacts with TgWRKY47. This interaction suppresses the DNA-binding ability of TgWRKY47, thereby reducing its transcriptional activation on TgEXPA2 without affecting the transactivation ability of TgWRKY47. Our findings establish a link between sucrose and cone enlargement in T. grandis and elucidate the potential underlying molecular mechanism.

Overexpression of oHIOMT results in various morphological, anatomical, physiological and molecular changes in switchgrass.

Melatonin (N-acetyl-5-methoxytryptamine) is a molecule implicated in multiple biological functions, but exerts contrasting effects on plants owing to concentration differences. Hydroxyindole O-methyltransferase (HIOMT), which catalyzes the last step of melatonin synthesis, plays a crucial role in this context. Transgenic switchgrass overexpressing oHIOMT with different melatonin levels displayed distinct morphological changes in a concentration-dependent manner. In this study, we divided the transgenic switchgrass into two groups: melatonin-moderate transgenic (MMT) plants and melatonin-rich transgenic (MRT) plants. To determine the concentration-dependent effect of melatonin on switchgrass growth and stress resistance, we conducted comparative morphological, physiological, omics and molecular analyses between MMT, MRT and wild-type (WT) plants. We found that oHIOMT overexpression, with moderate melatonin levels, was crucial in regulating switchgrass growth through changes in cell size rather than cell number. Moderate levels of melatonin were vital in regulating carbon fixation, stomatal development and chlorophyll metabolism. Regarding salt tolerance, melatonin with moderate levels activated numerous defense (e.g. morphological characteristics, anatomical structure, antioxidant enzymatic properties, non-enzymatic capacity and Na+/K+ homeostasis). Additionally, moderate levels of oHIOMT overexpression were sufficient to increase lignin content and alter monolignol compositions with an increase in the S/G lignin ratio. Taken together, oHIOMT overexpression in switchgrass with different melatonin levels resulted in morphological, anatomical, physiological and molecular changes in a concentration-dependent manner, which characterized by stimulation at low doses and inhibition at high doses. Our study presents new ideas and clues for further research on the mechanisms of the concentration-dependent effect of melatonin.

A Timeline of Cardiac Maturation

Background: Much is known about the differences between adult and immature fetal cardiac myocytes. For example, the switch from glycolysis to oxidative phosphorylation, the increase in sarcomere distance and cell size, along with changes in mitochondrial structure and organization are all known to occur at some point between late embryonic development and adulthood. However, the precise timing of when these changes take place is ill-defined. Here, we report in high temporal resolution when hallmarks of maturation develop to provide a timeline and the order of events that are necessary for structural and physiological cardiomyocyte maturation. Methods and Results: Using murine samples from embryonic day 15 all the way to adulthood, we performed characterization of basic organelles, contractile machinery, and extracellular matrix composition, all shown to be necessary for cardiac maturation. We observed clear differences in when maturation hallmarks became established. Features that increased continuously include growth of the size of the heart and cardiomyocytes. Accompanying cell size changes, an increase in endothelial cell count was observed, although cardiomyocyte growth continued after the endothelial cell population reached a plateau. Seahorse assays revealed that mitochondrial respiration increased continuously over time for Complex 1 and 2 until about postnatal day 15, with complex 4 being relatively stable throughout maturation. Lipid droplets appeared at a higher frequency in ventricular tissue starting at postnatal day 5, coinciding with an increase in sarcomere length. T-tubules began to appear at postnatal day 7 alongside changes in calcium handling genes. Mitochondrial size was found to increase rapidly at postnatal day 19. Discussion: Overall, we found that multiple stages of development seem important to target for enhancing maturation as not all changes occur in a linear fashion. Ultimately, knowledge of when maturational hallmarks develop might inform strategies to enhance maturation of human induced pluripotent stem cell derived cardiomyocytes. T32 Minnesota Muscle Training Grant; Summer’s Wish; Regenerative Medicine Minnesota; Stem Cell Institute Predoctoral Infuse Award. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Determination of Ploidy Levels and Nuclear DNA Content in Cryptococcus neoformans by Flow Cytometry: Drawbacks with Variability.

Flow cytometry is commonly employed for ploidy determination and cell cycle analysis in cryptococci. The cells are subjected to fixation and staining with DNA-binding fluorescent dyes, most commonly with propidium iodide (PI), before undergoing flow cytometric analysis. In ploidy determination, cell populations are classified according to variations in DNA content, as evidenced by the fluorescence intensity of stained cells. As reported in Saccharomyces cerevisiae, we found drawbacks with PI staining that confounded the accurate analysis of ploidy by flow cytometry when the size of the cryptococci changed significantly. However, the shift in the fluorescence intensity, unrelated to ploidy changes in cells with increased size, could be accurately interpreted by applying the ImageStream system. SYTOX Green or SYBR Green I, reported to enable DNA analysis with a higher accuracy than PI in S. cerevisiae, were nonspecific for nuclear DNA staining in cryptococci. Until dyes or methods capable of reducing the variability inherent in the drastic changes in cell size or shape become available, PI appears to remain the most reliable method for cell cycle or ploidy analysis in Cryptococcus.

Exercise training decreases lactylation and prevents myocardial ischemia-reperfusion injury by inhibiting YTHDF2.

Exercise improves cardiac function and metabolism. Although long-term exercise leads to circulating and micro-environmental metabolic changes, the effect of exercise on protein post-translational lactylation modifications as well as its functional relevance is unclear. Here, we report that lactate can regulate cardiomyocyte changes by improving protein lactylation levels and elevating intracellular N6-methyladenosine RNA-binding protein YTHDF2. The intrinsic disorder region of YTHDF2 but not the RNA m6A-binding activity is indispensable for its regulatory function in influencing cardiomyocyte cell size changes and oxygen glucose deprivation/re-oxygenation (OGD/R)-stimulated apoptosis via upregulating Ras GTPase-activating protein-binding protein 1 (G3BP1). Downregulation of YTHDF2 is required for exercise-induced physiological cardiac hypertrophy. Moreover, myocardial YTHDF2 inhibition alleviated ischemia/reperfusion-induced acute injury and pathological remodeling. Our results here link lactate and lactylation modifications with RNA m6A reader YTHDF2 and highlight the physiological importance of this innovative post-transcriptional intrinsic regulation mechanism of cardiomyocyte responses to exercise. Decreasing lactylation or inhibiting YTHDF2/G3BP1 might represent a promising therapeutic strategy for cardiac diseases.