Higher Cell Volume Research Articles

High Cell Volume Zn-Metal–organic Framework: Unveiling the Pseudo-capacitor Properties and Selective Colorimetric Recognition of Oxyanions

High Cell Volume Zn-Metal–organic Framework: Unveiling the Pseudo-capacitor Properties and Selective Colorimetric Recognition of Oxyanions

Application-Oriented Bulk Cryopreservation of Human iPSCs in Cryo Bags Followed by Direct Inoculation in Scalable Suspension Bioreactors for Expansion and Neural Differentiation.

Stem cell-based therapies are promising tools for regenerative medicine and require bulk numbers of high-quality cells. Currently, cells are produced on demand and have a limited shelf-life as conventional cryopreservation is primarily designed for stock keeping. We present a study on bulk cryopreservation of the human iPSC lines UKKi011-A and BIONi010-C-41. By increasing cell concentration and volume, compared to conventional cryopreservation routines in cryo vials, one billion cells were frozen in 50 mL cryo bags. Upon thawing, the cells were immediately seeded in scalable suspension-based bioreactors for expansion to assess the stemness maintenance and for neural differentiation to assess their differentiation potential on the gene and protein levels. Both the conventional and bulk cryo approach show comparative results regarding viability and aggregation upon thawing and bioreactor inoculation. Reduced performance compared to the non-frozen control was compensated within 3 days regarding biomass yield. Stemness was maintained upon thawing in expansion. In neural differentiation, a delay of the neural marker expression on day 4 was compensated at day 9. We conclude that cryopreservation in cryo bags, using high cell concentrations and volumes, does not alter the cells' fate and is a suitable technology to avoid pre-cultivation and enable time- and cost-efficient therapeutic approaches with bulk cell numbers.

Blastospores from Metarhizium anisopliae and Metarhizium rileyi Are Not Always as Virulent as Conidia Are towards Spodoptera frugiperda Caterpillars and Use Different Infection Mechanisms.

Infective conidia from entomopathogenic fungi are widely used to control insect pests. Many entomopathogenic fungi also produce yeast-like cells called blastospores under specific liquid culture conditions that can directly infect insects. However, little is known about the biological and genetic factors that allow blastospores to infect insects and make them potentially effective for biological control in the field. Here, we show that while the generalist Metarhizium anisopliae produces a higher number of and smaller blastospores, the Lepidoptera specialist M. rileyi produces fewer propagules with a higher cell volume under high-osmolarity conditions. We compared the virulence of blastospores and conidia of these two Metarhizium species towards the economically important caterpillar pest Spodoptera frugiperda. Conidia and blastospores from M. anisopliae were equally infectious, but acted slower, and killed fewer insects than M. rileyi conidia and blastospores did, where M. rielyi conidia had the highest virulence. Using comparative transcriptomics during propagule penetration of insect cuticles, we show that M. rileyi blastospores express more virulence-related genes towards S. frugiperda than do M. anisopliae blastospores. In contrast, conidia of both fungi express more virulence-related oxidative stress factors than blastospores. Our results highlight that blastospores use a different virulence mechanism than conidia use, which may be explored in new biological control strategies.

Asymptotic behavior of two competing cancer cell migration mechanisms driven by interstitial fluid flow

In this paper, we will investigate two competing mechanisms of cancer cell migration, which are driven by interstitial fluid flow and satisfy the suitable boundary conditions in two different radial symmetry domains, respectively. For both mechanisms, we will study their asymptotic behavior in different environments with low and high initial cell volume fraction separately. More precisely, our first result is to analyze the convergence of the solution to a downstream-dominated steady state over time in BR(0)﹨Bρ(0)⊂Rd(d≥2) with 0<ρ<R when the initial unit volume fraction is sufficiently low, while our second result is to investigate the convergence of the solution to an upstream-dominated steady state in BR(0)⊂Rd(d≥2) when the initial cell volume fraction is high enough.

Effect of cells on spatial quantification of proteoglycans in articular cartilage of small animals

ABSTRACT Objective Histochemical characterization of proteoglycan content in articular cartilage is important for the understanding of osteoarthritis pathogenesis. However, cartilage cells may interfere with the measurement of matrix proteoglycan content in small animal models (e.g. mice and rats) due to the high cell volume fraction (38%) in mice compared to human tissue (~1%). We investigated whether excluding the cells from image analysis affects the histochemically measured proteoglycan content of rat knee joint cartilage and assessed the effectiveness of a deep learning algorithm-based tool named U-Net in cell segmentation. Design Histological sections were stained with Safranin-O, after which optical densities were measured using digital densitometry to estimate proteoglycan content. U-Net was trained with 600 annotated Safranin-O cartilage images for exclusion of cells from the cartilage extracellular matrix. Optical densities of the ECM with and without cells were compared as a function of normalized tissue depth. Results U-Net cell segmentation was accurate, with the measured cell area fraction following largely that of ground-truth images (average difference: 4.3%). Cell area fraction varied as a function of tissue depth and took up 8–21% of the tissue area. The exclusion of cells from the analysis led to an increase in the analyzed depth-dependent optical density of cartilage by approximately 0.6–1.8% (p < 0.01). Conclusions Although the effect of cells on the analyzed proteoglycan content is small, it should be considered for improved sensitivity, especially at the onset of the disease during which cells may proliferate in small animals.

Formation mechanism of bi-modal cell structure polystyrene foams by synergistic effect of CO2-philic additive and co-blowing agent

A new method was developed for the formation of bi-modal cell structure polystyrene (PS) foams by synergistic effect of CO2-philic additive (PDMS) and co-blowing agent (alcohol). The results showed that PDMS and alcohol synergistically increased the solubility and diffusion coefficient of CO2 into PS and reduced the desorption rate from PS. The blowing agent formed a high-concentration area in the PDMS (6000 g/mol) phase to promote bubble nucleation and growth in advance. The moderate size and number of cells were pre-formed, slowing the CO2 desorption and increasing the duration of bubble growth, so that bi-modal cell structure PS foams with high cell density and volume expansion ratio were obtained. Moreover, the solution and diffusion behavior of CO2 and the cell morphology could be easily tuned by changing the composition of complex additives, which might be a promising strategy for preparing bi-modal cell structure in different foaming processes.

Correlation between anomalous thermal expansion coefficient and barocaloric effect: Application to spin crossover systems

We report the strong correlation between anomalous thermal expansion in solid-solid volumetric phase transition and the barocaloric effect. This investigation was carried out in {Fe[H2B(pz)2]2(bipy)} (pz = pyrazolyl; bipy = bipyridine) spin crossover system, in which the spin of Fe2+ changes from sLS = 0 to sHS = 2 coupled with high unit cell volume changes from VLS = 2356 to VHS = 2462 Å3, at T1/2–160 K. Giant barocaloric effect, through the isothermal entropy change (ΔST = 60 J kg−1 K−1) upon moderated hydrostatic pressure variation (ΔP = 1 kbar), was indirectly obtained from reported pressure dependence of magnetic susceptibility. Theoretical results, obtained from our microscopic model, updated with the reported metal-ligand asymmetrical stretching z-vibration modes, from DFT calculation, showed a remarkable agreement with the experimental data.

Autofluorescence-based sorting removes senescent cells from mesenchymal stromal cell cultures

Mesenchymal stromal cells (MSC) are used in cell therapy, but results depend on the unknown quality of cell populations. Extended culture time of MSC increases their senescent levels, leading to a critical loss of cell fitness. Here, we tested the suitability of MSC-sorting based on their FACS autofluorescence profile, for a rapid and non-invasive method of senescent cell elimination. Cells were classified in low- (LA) and high- (HA) autofluorescence groups, and results compared to the original MSC population (control). Three days after sorting, cells were screened by replicative senescence markers (cell volume, SA-β-Gal assay and gene/protein expression) and MSC differentiation assays. The transcriptional profiles of sorted MSC were also analyzed by RNA‐Seq. Compared to control, LA cells had 10% lower cell volume and autofluorescence, and 50% less SA-β-Gal + cells. Instead, HA cells had 20% higher cell volume and autofluorescence, and 120% more SA-β-Gal + cells. No changes in replicative senescence and differentiation potentials were observed between all groups. However, 68 genes (16 related to senescence) were significantly differentially expressed (DEG) between LA and other groups. Biological network of DEG identified CXCL12 as topological bottleneck. In summary, MSC sorting may have practical clinical implications to enhance the results of MSC-based therapies.

Hydrogel Micropost Arrays with Single Post Tunability to Study Cell Volume and Mechanotransduction.

The extracellular matrix varies considerably in mechanical properties at the microscale. It remains unclear how cells respond to these properties, in part, due to lack of tools to create precisely defined microenvironments in a discrete manner. Here, freeform stereolithography is leveraged to control the placement and elastic modulus of individual hydrogel microposts that serve as discrete matrix signals to interface with cells. Mesenchymal stromal cells (MSCs) located in the interstitial spaces between microposts above a base layer are analyzed. Cell volume is higher when MSCs interact with more microposts. MSCs show higher strain energy when they interact simultaneously with 4-kPa and 20-kPa microposts than with mechanically homogeneous micropost arrays. MSCs are sensitive to pharmacological inhibition of Rho-associated protein kinase in 4-kPa arrays, but resistant when presented together with 20-kPa arrays. Yes-associated protein (YAP) activity increases with higher cell volume and elastic modulus of microposts. Surprisingly, YAP activity becomes less variable with higher cell volume and decreases with higher average force and strain energy per post when MSCs interact with both 4-kPa and 20-kPa microposts simultaneously. Together, these results describe a material system for systematically investigating how the placement and intrinsic properties of discrete matrix signals impact cell volume and mechanotransduction.

Densification-Induced Structure Changes in Basolite MOFs: Effect on Low-Pressure CH4 Adsorption.

Metal-organic frameworks’ (MOFs) adsorption potential is significantly reduced by turning the original powder into pellets or granules, a mandatory step for their use at industrial scale. Pelletization is commonly performed by mechanical compression, which often induces the amorphization or pressure-induced phase transformations. The objective of this work is the rigorous study of the impact of mechanical pressure (55.9, 111.8 and 186.3 MPa) onto three commercial materials (Basolite C300, F300 and A100). Phase transformations were determined by powder X-ray diffraction analysis, whereas morphological changes were followed by nitrogen physisorption. Methane adsorption was studied in an atmospheric fixed bed. Significant crystallinity losses were observed, even at low applied pressures (up to 69.9% for Basolite C300), whereas a structural change occurred to Basolite A100 from orthorhombic to monoclinic phases, with a high cell volume reduction (13.7%). Consequently, adsorption capacities for both methane and nitrogen were largely reduced (up to 53.6% for Basolite C300), being related to morphological changes (surface area losses). Likewise, the high concentration of metallic active centers (Basolite C300), the structural breathing (Basolite A100) and the mesopore-induced formation (Basolite F300) smooth the dramatic loss of capacity of these materials.

Mathematical Analysis of Two Competing Cancer Cell Migration Mechanisms Driven by Interstitial Fluid Flow

Recent experimental work has revealed that interstitial fluid flow can mobilize two types of tumor cell migration mechanisms. One is a chemotactic-driven mechanism where chemokine (chemical component) bounded to the extracellular matrix (ECM) is released and skewed in the flow direction. This leads to higher chemical concentrations downstream which the tumor cells can sense and migrate toward. The other is a mechanism where the flowing fluid imposes a stress on the tumor cells which triggers them to go in the upstream direction. Researchers have suggested that these two migration modes possibly can play a role in metastatic behavior, i.e., the process where tumor cells are able to break loose from the primary tumor and move to nearby lymphatic vessels. In Waldeland and Evje (J Biomech 81:22–35, 2018), a mathematical cell–fluid model was put forward based on a mixture theory formulation. It was demonstrated that the model was able to capture the main characteristics of the two competing migration mechanisms. The objective of the current work is to seek deeper insight into certain qualitative aspects of these competing mechanisms by means of mathematical methods. For that purpose, we propose a simpler version of the cell–fluid model mentioned above but such that the two competing migration mechanisms are retained. An initial cell distribution in a one-dimensional slab is exposed to a constant fluid flow from one end to the other, consistent with the experimental setup. Then, we explore by means of analytical estimates the long-time behavior of the two competing migration mechanisms for two different scenarios: (i) when the initial cell volume fraction is low and (ii) when the initial cell volume fraction is high. In particular, it is demonstrated in a strict mathematical sense that for a sufficiently low initial cell volume fraction, the downstream migration dominates in the sense that the solution converges to a downstream-dominated steady state as time elapses. On the other hand, with a sufficiently high initial cell volume fraction, the upstream migration mechanism is the stronger in the sense that the solution converges to an upstream-dominated steady state.

Crystal structure, phonon modes, and bond characteristics of AgPb2B2V3O12 (B = Mg, Zn) microwave ceramics

AbstractAgPb2B2V3O12 (B = Mg, Zn) ceramics with low sintering temperature were synthesized via the conventional solid‐state reaction route. Rietveld refinements of the X‐ray diffraction patterns confirm cubic symmetry with space group . The number of observed vibrational modes and those predicted by group theoretical calculations also confirm the space group. At the optimum sintering temperature of 750°C/4 hours, AgPb2Mg2V3O12 has a relative permittivity of 23.3 ± 0.2, unloaded quality factor () of 26 900 ± 500 GHz (), and temperature coefficient of resonant frequency of 19.3 ± 1 ppm/°C, while AgPb2Zn2V3O12 has the corresponding values of 26.4 ± 0.2, 28 400 ± 500 GHz () and –18.4 ± 1 ppm/°C at 590°C/4 hours. Microwave dielectric properties of a few reported garnets and Pb2AgB2V3O12 (B = Mg, Zn) ceramics were correlated with their intrinsic characteristics such as the Raman shifts as well as width of A1g Raman bands. Higher quality factor was obtained for lower full width at half‐maxima (FWHMs) values of A1g modes. The increase in B‐site bond valence contributes to high and low |τf| with the substitution of Zn2+ by Mg2+. Furthermore, the high ionic polarizability and unit cell volume with Zn2+substitution contribute to increased relative permittivity.

Photoautotrophic production of succinate via the oxidative branch of the tricarboxylic acid cycle influences glycogen accumulation in Synechocystis sp. PCC 6803

Cyanobacteria are interesting biocatalysts for the sustainable production of value-added compounds, but a functional link of biocatalytic efficiency and cell physiology is missing. Especially the role and structure of the tricarboxylic acid (TCA) cycle in cyanobacteria is not fully understood. Succinate dehydrogenase (SDH) is an essential enzyme linking the TCA cycle to the quinone pool and its substrate succinate is an important platform chemical. We utilized Synechocystis sp. PCC 6803 as a host organism for the photoautotrophic production of succinate via the oxidative branch of the TCA cycle. In Synechocystis sp. PCC 6803 SDH is described to be encoded by three open reading frames (ORF). Four SDH deletion mutants were created and characterized regarding the formation of succinate. Synechocystis_∆sll1625 was the best performing strain accumulating 420 mg L−1 succinate during cultivation in a 1.8 L-photobioreactor under constant illumination. Furthermore, deletion of the SDH resulted in a non-bleaching phenotype and concomitant accumulation of glycogen and succinate during growth at 5% CO2. This substantially influenced cell physiology of the mutant, reflected in decreased cell numbers and higher cell volumes. Thus, the TCA cycle seems to have a key role in controlling carbon fluxes from CO2 into biomass, storage compounds like glycogen, and other carbon sinks.

Bone marrow morphologic features, MyPRS, and gene mutation correlations in plasma cell myeloma.

Genetics has played an important role in risk stratification for plasma cell myeloma patients, providing therapeutic guidance. In this study, we investigated the correlation of bone marrow morphologic features and genetic aberrations, including gene expression profiles, translocations, and gene mutations. For the first time we show that high plasma cell volume, diffuse sheet growth pattern, immature cell morphology, high mitotic index, and increased reticulin fibrosis, significantly correlates with high risk disease determined by MyPRS gene expression profiles. Furthermore, we show the association between MyPRS risk stratification and chromosomal alterations and specific gene mutations. We also demonstrate the combinational effect of TP53 mutation and 17p loss on the histological changes in bone marrow.

Evaluation of various culture techniques for identification of hookworm species from stool samples of children.

Background & objectives:Different coproculture techniques have been developed for culturing the hookworm (HW) larvae for morphological identification in the resource-limited settings. The objective of this study was to compare the performances of Harada-Mori culture (HMC), agar plate culture (APC) and modified APC (MAPC) of HW positive stool specimens for identification of HW species in East Sikkim.Methods:This prospective study was done in East Sikkim from May 2015 to May 2016. Stool and blood samples were collected from paediatric patients with gastrointestinal symptoms. The HW positive stool specimens by microscopy were subjected to HMC, APC and MAPC techniques to harvest HW larvae. Stoll's dilution egg count for determining egg intensity and blood parameters were performed in all the 12 HW-positive patients.Results:Twelve of the 180 samples were found positive for HW by microscopy and predominance of Necator americanus (75%) over Ancylostoma duodenale (25%) was observed. Blood parameters results showed high pack cell volume (PCV) values in 78.6 per cent, anaemia in 75 per cent and high eosinophil count in most patients. Stoll's dilution egg count showed moderate infection in 66.6 per cent, light and heavy infections in 16.7 per cent each.Interpretation & conclusions:Our results showed that APC yielded 100 per cent results and was easier to perform in the laboratory compared to MAPC and HMC techniques.

Acquired resistance to BRAFi reverses senescence-like phenotype in mutant BRAF melanoma.

Targeting MAPK pathway in mutant BRAF melanoma with the specific BRAF inhibitor vemurafenib showed robust initial responses in the majority of patients followed by relapses due to acquired resistance to the drug. In V600EBRAF melanoma cell lines, senescence-associated β-galactosidase activity is often encountered in a constitutive manner or induced after MAPK inhibition. However, the link between the senescence-like phenotype and the resistance to BRAF inhibition is not fully understood yet. Our data validate a senescence-like phenotype (low cell proliferation, high cell volume, and high β-Gal activity) in mutant BRAF cells. Vemurafenib increased β-Gal activity in 4 out of 5 sensitive lines and in 2 out of 5 lines with intrinsic resistance to the drug. Interestingly, the 3 lines with acquired resistance to vemurafenib became depending on the drug for proliferation. In absence of drug, these lines showed a lower cell proliferation rate together with a substantial increase of β-Gal activity both in vitro and in vivo. In all settings, the senescence-like phenotype was significantly associated with an inhibition of pRB and cyclin D1, explaining the inhibition of cell proliferation. In conclusion, β-Gal activity is increased by V600EBRAF inhibition in the majority of sensitive and intrinsically resistant melanoma cells. Acquired resistance to vemurafenib is associated with a dependence to the drug for cell proliferation and tumor growth, and, in this case, drug removal stimulate β-Gal activity suggesting that the senescence-like phenotype could contribute to the acquired resistance to BRAF inhibition.

Ex vivo biomechanical characterization of syringe-needle ejections for intracerebral cell delivery

Intracerebral implantation of cell suspensions is finding its clinical translation with encouraging results in patients with stroke. However, the survival of cells in the brain remains poor. Although the biological potential of neural stem cells (NSCs) is widely documented, the biomechanical effects of delivering cells through a syringe-needle remain poorly understood. We here detailed the biomechanical forces (pressure, shear stress) that cells are exposed to during ejection through different sized needles (20G, 26G, 32G) and syringes (10, 50, 250 µL) at relevant flow rates (1, 5, 10 µL/min). A comparison of 3 vehicles, Phosphate Buffered Saline (PBS), Hypothermosol (HTS), and Pluronic, indicated that less viscous vehicles are favorable for suspension with a high cell volume fraction to minimize sedimentation. Higher suspension viscosity was associated with greater shear stress. Higher flow rates with viscous vehicle, such as HTS reduced viability by ~10% and also produced more apoptotic cells (28%). At 5 µL/min ejection using a 26G needle increased neuronal differentiation for PBS and HTS suspensions. These results reveal the biological impact of biomechanical forces in the cell delivery process. Appropriate engineering strategies can be considered to mitigate these effects to ensure the efficacious translation of this promising therapy.

Effect of different CO2 concentrations on biomass, pigment content, and lipid production of the marine diatom Thalassiosira pseudonana.

The marine diatom Thalassiosira pseudonana grown under air (0.04% CO2) and 1 and 5% CO2 concentrations was evaluated to determine its potential for CO2 mitigation coupled with biodiesel production. Results indicated that the diatom cultures grown at 1 and 5% CO2 showed higher growth rates (1.14 and 1.29 div day-1, respectively) and biomass productivities (44 and 48mgAFDWL-1day-1) than air grown cultures (with 1.13 div day-1 and 26mgAFDWL-1day-1). The increase of CO2 resulted in higher cell volume and pigment content per cell of T. pseudonana. Interestingly, lipid content doubled when air was enriched with 1-5% CO2. Moreover, the analysis of the fatty acid composition of T. pseudonana revealed the predominance of monounsaturated acids (palmitoleic-16:1 and oleic-18:1) and a decrease of the saturated myristic acid-14:0 and polyunsaturated fatty acids under high CO2 levels. These results suggested that T. pseudonana seems to be an ideal candidate for biodiesel production using flue gases.

Targeting miRNA for Therapeutics Using a Micronome Based Method for Identification of miRNA-mRNA Pairs and Validation of Key Regulator miRNA.

MicroRNAs are 18-22bp long non-coding sequences and play a critical role in diverse biological processes, through modulation of gene expression at the post-transcriptional level by binding at the 3'-untranslated region of target mRNA. Consequent upon the discovery of structural and functional features of miRNA targeting, several molecular methods have been developed to identify miRNA targets. However, these methods suffer several drawbacks, including technical challenges, requirement of high cell volumes, inability to differentiate between direct and indirect targets, cell/tissue as well as experimental-specificity and imprecise binding site information. Alternatively in silico approach enables the exploration of the potential miRNA-mRNA pairs to investigate signature miRNA and proteins involved in the signaling of various diseases. Here, we describe micronome-based standard method for identification of miRNA-mRNA pairs as well as validation of key regulator miRNA.

Budd-Chiari syndrome secondary to polycythemia vera with inferior vena cava thrombosis

A middle aged male presented with abdominal distension since one month. Further workup showed plasma hemoglobin of 18.1 g/dL with a high pack cell volume (PCV), raised urea, creatinine and disturbed liver function tests. Abdominal ultrasonography showed an enlarged caudate lobe with thrombi in the inferior vena cava while CT scan of abdomen confirmed the same findings and was suggestive of Budd-Chiari syndrome. Further workup was conducted to rule out other causes and to find out the possible cause of Budd-Chiari syndrome. A peripheral film was requested, which showed hyper-segmented neutrophils. Later on JAK2 mutation and thrombophilia profile was ordered, which was positive for JAK2 mutation. Even though the patient was started on low molecular weight heparin but he eventually passed away. Key point Budd-Chiari syndrome is due to the obstruction of the hepatic efferent outflow tract from either the hepatic veins to where the inferior vena cava meets the right atrium. Its pathogenesis is due to the decreased outflow of the liver, leading to increase in pressure, stasis and later on damage induced by hypoxia and thrombus in the portal veins. Citation: Majid Z, Tasneem AA, Luck NH, Ul Haque MM, Mandhwani RK, Laeeq SM, et al. Budd-Chiari syndrome secondary to polycythemia vera with inferior vena cava thrombosis Immunopathol Persa. 2018;4(1):e04. DOI: 10.15171/ipp.2018.04.