Theory and Calculation of Nontraditional Stable Isotope Fractionation

This paper conducts processing on isotope anharmonic effect with molecular dynamics method and Monte Carlo method based on path integration. It introduces the theoretical calculation method of pressure effect, and finally the nuclear volume effect and its theoretical calculation method, stressing that the nuclear volume effect is an important part of isotopic studies of heavy metals in the future. This paper makes an analysis on the equilibrium fractionation theory based on simple harmonic approximation.

Theory and simulations of condensin mediated loop extrusion in DNA

Condensation of hundreds of mega-base-pair-long human chromosomes in a small nuclear volume is a spectacular biological phenomenon. This process is driven by the formation of chromosome loops. The ATP consuming motor, condensin, interacts with chromatin segments to actively extrude loops. Motivated by real-time imaging of loop extrusion (LE), we created an analytically solvable model, predicting the LE velocity and step size distribution as a function of external load. The theory fits the available experimental data quantitatively, and suggests that condensin must undergo a large conformational change, induced by ATP binding, bringing distant parts of the motor to proximity. Simulations using a simple model confirm that the motor transitions between an open and a closed state in order to extrude loops by a scrunching mechanism, similar to that proposed in DNA bubble formation during bacterial transcription. Changes in the orientation of the motor domains are transmitted over ~50 nm, connecting the motor head and the hinge, thus providing an allosteric basis for LE.

Balance of osmotic pressures determines the volume of the cell nucleus

The volume of the cell nucleus varies across cell-types and species, and is commonly thought to be determined by the size of the genome and degree of chromatin compaction. However, this notion has been challenged over the years by multiple experimental evidence. Here, we consider the physical condition of mechanical force balance as a determining condition of the nuclear volume and use quantitative, order-of-magnitude analysis to estimate the forces from different sources of nuclear and cellular pressure. Our estimates suggest that the dominant pressure within the nucleus and cytoplasm originates from the osmotic pressure of proteins and RNA molecules that are localized to the nucleus or cytoplasm by out-of-equilibrium, active nucleocytoplasmic transport rather than from chromatin or its associated ions. This motivates us to formulate a physical model for the ratio of the cell and nuclear volumes in which osmotic pressures of localized proteins determine the relative volumes. In accordance with unexplained observations that are century-old, our model predicts that the ratio of the cell and nuclear volumes is a constant, robust to a wide variety of biochemical and biophysical manipulations, and is changed only if gene expression or nucleocytoplasmic transport are modulated.

The Correlation Between Cell and Nucleus Size is Explained by an Eukaryotic Cell Growth Model

In eukaryotes, the cell volume is observed to be strongly correlated with the nuclear volume. The slope of this correlation depends on the cell type, growth condition, and the physical environment of the cell. We develop a computational model of cell growth and proteome increase, incorporating the kinetics of amino acid import, protein/ribosome synthesis and degradation, and active transport of proteins between the cytoplasm and the nucleoplasm. We also include a simple model of ribosome biogenesis and assembly. Results show that the cell volume is tightly correlated with the nuclear volume, and the cytoplasm-nucleoplasm transport rates strongly influences the cell growth rate as well as the cytoplasm/nucleoplasm ratio. Ribosome assembly and the ratio of ribosomal proteins to mature ribosomes also influence the cell volume and the cell growth rate. We find that in order to regulate the cell growth rate and the cytoplasm/nucleoplasm ratio, the cell must optimally control groups of kinetic parameters together, which could explain the quantitative roles of canonical growth pathways. Finally, using an extension of our model and single cell RNAseq data, it is possible to construct a detailed proteome distribution, provided that a cell division mechanism is known.

miRNA-489-3p Improves Sepsis-Induced Lung Injury by Regulating Toll-Like Receptor 4 (TLR4) Signaling Pathway

Aim: To discuss miRNA-489-3p in sepsis-induced lung injury. Materials and methods: Using NR-8383 as research cell in our study, and using LPS stimulation to sepsis induced lung injury vitro model. Measuring cell proliferation by MTS assay; using Elisa assay to measure IL-1β, IL-6 and TNF-α concentration; apoptosis cell number and apoptosis rate were evaluated using TUNEL and flow cytometry; gene and protein were measured by RT-PCR and WB assay; measuring p-NF-κB(p65) nuclear volume by Immunofluorescence (IF); using Luciferase reporter assay to analysis miRNA-489-3p and TLR4 correlation. Results: Cell proliferation rate significantly down-regulated (P < 0.001), apoptosis cell number and apoptosis rate significantly increased (P < 0.001); IL-1β, IL-6 and TNF-α concentrations significantly up-regulated (P < 0.001); TLR4 and MyD88 gene and protein significantly increased (P < 0.001), NF-κB(p65) mRNA and p-NF-κB(p65) protein and nuclear volume significantly increased (P < 0.001). However, with miRNA-489-3p supplement, the cell proliferation rate, apoptosis cell number and apoptosis rate were significantly improved (P < 0.001, respectively) via TLR4, MyD88 and NF-κB(p65) mRNA and protein significantly depressing (P < 0.001, respectively). By LUC assay, miRNA-489-3p could target to TLR4 in NR-8383 cell. Conclusion: miRNA-489-3p overexpression had effect to improve sepsis induced lung injury via regulation TLR4/MyD88/NF-κB(p65).

Loops, TADs, Compartments, and Territories are Elastic and Robust to Dramatic Nuclear Volume Swelling

Layers of genome organization are becoming increasingly better characterized, but less is known about how these structures respond to perturbation or shape changes. Low-salt swelling of isolated chromatin fibers or nuclei has been used for decades to investigate the structural properties of chromatin. But, visible changes in chromatin appearance have not been linked to known building blocks of genome structure or features along the genome sequence. We combine low-salt swelling of isolated nuclei with genome-wide chromosome conformation capture (Hi-C) and imaging approaches to probe the effects of chromatin extension genome-wide. Photoconverted patterns on nuclei during expansion and contraction indicate that global genome structure is preserved after dramatic nuclear volume swelling, suggesting a highly elastic chromosome topology. Hi-C experiments before, during, and after nuclear swelling show changes in average contact probabilities at short length scales, reflecting the extension of the local chromatin fiber. But, surprisingly, during this large increase in nuclear volume, there is a striking maintenance of loops, TADs, active and inactive compartments, and chromosome territories. Subtle differences after expansion are observed, suggesting that the local chromatin state, protein interactions, and location in the nucleus can affect how strongly a given structure is maintained under stress. From these observations, we propose that genome topology is robust to extension of the chromatin fiber and isotropic shape change, and that this elasticity may be beneficial in physiological circumstances of changes in nuclear size and volume.

Interphase Nuclear Structure in Ten Species of Pteridophytes

In thelypterids, interphase chromosomes of Ampelopteris prolifera were accompanied by two or three nucleoli. One nucleolus was seen floating outside the nucleus while other remained attached with the nucleus in annular cells of Christella arida. The mean of chromocentres determined in Adiantum species ranged from 49.8 to 53.6, whereas, it ranged from 46.6 to 49.5 in Pteris species. On the other hand, the value for chromocentres ranged from 62.5 to 123.5 in thelypterids. Nuclear organization was observed to be chromocentric type. In case of interphase nuclear volume it was observed that in Adiantum it was highest in A. capillus-veneris followed by A. caudatum and A. lunulatum, in Pteris it was highest in P. biaurita followed by P. vittata and P. griffithi, and in case of thelypterids, it was highest in C. dantata followed by C. Cylindrothix, A. Prolifera and C. arida. J. Bio-Sci. 29(2): 93-98, 2021 (December)

Live imaging of chromatin distribution reveals novel principles of nuclear architecture and chromatin compartmentalization

The three-dimensional organization of chromatin contributes to transcriptional control, but information about native chromatin distribution is limited. Imaging chromatin in live Drosophila larvae, with preserved nuclear volume, revealed that active and repressed chromatin separates from the nuclear interior and forms a peripheral layer underneath the nuclear lamina. This is in contrast to the current view that chromatin distributes throughout the nucleus. Furthermore, peripheral chromatin organization was observed in distinct Drosophila tissues, as well as in live human effector T lymphocytes and neutrophils. Lamin A/C up-regulation resulted in chromatin collapse toward the nuclear center and correlated with a significant reduction in the levels of active chromatin. Physical modeling suggests that binding of lamina-associated domains combined with chromatin self-attractive interactions recapitulate the experimental chromatin distribution profiles. Together, our findings reveal a novel mode of mesoscale organization of peripheral chromatin sensitive to lamina composition, which is evolutionary conserved.

Prolactin induced cyto-architectural changes in the corpuscles of Stannius of stinging catfish, Heteropneustes fossilis acclimated to different calcium media

Adult fish Heteropneustes fossilis were divided into 4 groups –(i) Group A: kept in artificial freshwater and daily injected intraperitoneally with vehicle; (ii) Group B: kept in artificial freshwater and were daily injected intraperitoneally with 0.1 mg/100 g body wt of oProlactin; (iii) Group C: maintained in calcium-deficient freshwater and daily injected intraperitoneally with vehicle; (iv) Group D: kept in calcium-deficient freshwater and daily injected intraperitoneally with 0.1 mg/100 g body wt of oProlactin. Blood samples were taken 2 h after the last injection on 1, 3, 5, 10 and 15 days of the treatment. Plasma calcium levels were analyzed. The corpuscles of Stannius (CS) were fixed for histological studies. Artificial freshwater: The plasma calcium levels of vehicle-injected specimens (group A) remained unaltered throughout the experiment. Following prolactin treatment (group B) the plasma calcium levels progressively increased from day 3 to day 5. The values became normocalcemic at day 10 and day 15. After day 5 following prolactin administration (group B), the nuclear volume of AF-positive cells increased and the cells were seen degranulated. After day 10, there was an increased dilatation of sinusoids and the nuclear volume of AF-positive cells showed further increase. On day 15, these changes were exaggerated. The AFnegative cells of the corpuscles of Stannius of prolactin-treated fish (group B) showed no change in their histological structure and nuclear volume. Calcium-deficient freshwater: The plasma calcium level decreased in vehicle-injected fish (group C) from day 1 to day 3 (as compared to level of the fish kept in artificial freshwater). Thereafter, the level increased from day 5 resulting in hypercalcemia at day 10 and day 15. In prolactin treated fish (group D) the plasma calcium level indicated progressive increase from day 5 to day 15. In the vehicle-injected fish (group C) the AF-positive cells of corpuscles of Stannius showed accumulati

Histomorphometric and immunohistochemical changes in interstitial cells and ovarian follicles of rats treated with metformin during and after induction of permanent estrus

Objective: To evaluate the histomorphometric and immunohistochemical changes in interstitial cells and ovarian follicles of rats treated with metformin during and after induction of permanent estrus. Methods: Thirty-two adult-female rats with regular estrous cycle were equally divided into four groups: 1) GCtrl - at estrous phase. 2) GPCOS - at permanent-estrous phase. 3) GMet1 - rats and daily treated with metformin (12.5 mg/Kg) during 60 consecutive days, as preventive form and 4) GMet2 - PCOS rats, which remained exposed to 60 days of continuous illumination and treated with metformin. After that, the animals were euthanized, and the ovaries were removed and processed for paraffin embedding. Sections were stained with H.E. for histomorphometry or subjected to immunohistochemistry for Ki-67 and cleaved caspase-3 (Casp-3) detections. Results: The GPCOS showed lack of corpus luteum and several ovarian cysts, as well as interstitial-like cells. The presence of corpus luteum and a significant increase in primary and antral follicles were observed in Mel-treated groups, which also showed a decrease in the number of ovarian cysts and in the area occupied by interstitial-like cells. The presence of corpus luteum along with an increase in the number of primary follicles in the Met2 group were noticed (p<0,01). A significant reduction in number of cysts and in the area occupied by interstitial cells, as well as a decrease in nuclear volume of interstitial cells, were noticed in the Met-treated groups, mainly in the Met2 group. The percentage of cell proliferation was significantly higher in granulosa cells of the Met-treated groups than PCOS group, mainly in the GMet2 (p<0,01), which was similar to the GCtrl group. On the other hand, the percentage of apoptosis (cleaved-caspase-3- positive cells) was significantly higher in the granulosa cells of GPCOS and Met-treated groups than the GCtrl group, but without significant difference, which showed weak cleaved caspase-3 immunoreactivity in those cells. Conclusion: The ovaries of rats treated with metformin showed a decrease in nuclear volume and in the area occupied by interstitial cells, presence of corpus luteum, in addition to a decrease in the number of cysts.