Cell Growth Kinetics Research Articles

Deciphering the Role of Inorganic Nanoparticles' Surface Functionalization on Biohybrid Microbial Photoelectrodes.

Shedding light on the interaction between inorganic nanoparticles (NPs) and living microorganisms is at the basis of the development of biohybrid technologies with improved performance. Au NPs have been shown to be able to improve the extracellular electron transfer (EET) in intact bacterial cells interfaced with an electrode; however, detailed information on the role of NP-surface properties in their interaction with bacterial membranes is still lacking. Herein, we unveil how the surface functionalization of Au NPs influences their interaction with photosynthetic bacteria, focusing on cell morphology, growth kinetics, NPs localization, and electrocatalytic performance. We show that functionalization of Au NPs with cysteine in the zwitterionic form results in a uniform NPs distribution in purple bacteria, specifically locating the NPs within the outer-membrane/periplasmic space of bacterial cells. These biohybrid cells, when coupled with an electrode, exhibit enhanced EET and increased (photo)current generation, paving the way for the future development of rationally designed biohybrid electrochemical systems.

Tip extension and simultaneous multiple fission in a filamentous bacterium

Organisms display an immense variety of shapes, sizes, and reproductive strategies. At microscopic scales, bacterial cell morphology and growth dynamics are adaptive traits that influence the spatial organization of microbial communities. In one such community-the human dental plaque biofilm-a network of filamentous Corynebacterium matruchotii cells forms the core of bacterial consortia known as hedgehogs, but the processes that generate these structures are unclear. Here, using live-cell time-lapse microscopy and fluorescent D-amino acids to track peptidoglycan biosynthesis, we report an extraordinary example of simultaneous multiple division within the domain Bacteria. We show that C. matruchotii cells elongate at one pole through tip extension, similar to the growth strategy of soil-dwelling Streptomyces bacteria. Filaments elongate rapidly, at rates more than five times greater than other closely related bacterial species. Following elongation, many septa form simultaneously, and each cell divides into 3 to 14 daughter cells, depending on the length of the mother filament. The daughter cells then nucleate outgrowth of new thinner vegetative filaments, generating the classic "whip handle" morphology of this taxon. Our results expand the known diversity of bacterial cell cycles and help explain how this filamentous bacterium can compete for space, access nutrients, and form important interspecies interactions within dental plaque.

Isolation and Growth Kinetics of Bogoria Virus from Phlebotomine Sandflies Sampled in Baringo, Kenya.

Phleboviruses are an emerging threat to public health. Recent surveillance efforts in Kenya have unveiled novel phleboviruses. Despite these efforts, there remain knowledge gaps. This study tested female sandflies from diverse ecological settings in Kenya for arboviruses. Sandfly pools were cultured in Vero-CCL cells. Pools showing reproducible cytopathic effects were subjected to next-generation sequencing, followed by phylogenetic analysis. In vitro, cell kinetics analysis was performed using both Vero-E6 cells and C6/36 mosquito cells. One pool from Baringo, Kenya, tested positive for Bogoria virus (BOGV). The BOGV genome clustered in a single clade with previously obtained BOGV genomes. No significant differences were observed between Vero and C6/36 cell growth kinetics. This study has confirmed the presence of BOGV among sandflies in Baringo Kenya and demonstrated growth in mosquito cells.

Extrusion-Based 3D Bioprinting of Bioactive and Piezoelectric Scaffolds as Potential Therapy for Treating Critical Soft Tissue Wounds.

Objective: This study focuses on developing bioactive piezoelectric scaffolds that could deliver bioelectrical cues to potentially treat injuries to soft tissues such as skeletal muscles and promote active regeneration. Approach: To address the underexplored aspect of bioelectrical cues in skeletal muscle tissue engineering (SMTE), we developed piezoelectric bioink based on natural bioactive materials such as sodium alginate, gelatin, and chitosan. Extrusion-based 3D bioprinting was utilized to develop scaffolds that mimic muscle stiffness and generate electrical stimulation (E-stim) when subjected to forces. The biocompatibility of these scaffolds was tested with the C2C12 muscle cell line. Results: The bioink demonstrated suitable rheological properties for 3D bioprinting, resulting in high-resolution composite sodium alginate-gelatin-chitosan scaffolds with good structural fidelity. The scaffolds exhibited a 42-60 kPa stiffness, similar to muscle. When a controlled force of 5N was applied to the scaffolds at a constant frequency of 4 Hz, they generated electrical fields and impulses (charge), indicating their suitability as a stand-alone scaffold to generate E-stim and instill bioelectrical cues in the wound region. The cell viability and proliferation test results confirm the scaffold's biocompatibility with C2C12s and the benefit of piezoelectricity in promoting muscle cell growth kinetics. Our study indicates that our piezoelectric bioink and scaffolds offer promise as autonomous E-stim-generating regenerative therapy for SMTE. Innovation: A novel approach for treating skeletal muscle wounds was introduced by developing a bioactive electroactive scaffold capable of autonomously generating E-stim without stimulators and electrodes. This scaffold offers a unique approach to enhancing skeletal muscle regeneration through bioelectric cues, addressing a major gap in the SMTE, that is, fibrotic tissue formation due to delayed muscle regeneration. Conclusion: A piezoelectric scaffold was developed, providing a promising solution for promoting skeletal muscle regeneration. This development can potentially address skeletal muscle injuries and offers a unique approach to facilitating skeletal muscle wound healing.

High cell density culture of microalgae in horizontal thin-layer algal reactor: Modeling of light attenuation and cell growth kinetics

This study delved into the light intensity effects and light attenuation modeling in high cell density culture (HCDC) of green alga Neochloris oleoabundans. The research primarily focused on how different light intensities influence cell growth in terms of cell division and cell mass, and the biochemical composition within the cells. Recognizing the need to avoid overestimating light path by excluding data point in the data zone where the light intensity was zero, we proposed and verified a novel modified Beer-Lambert model, which was superb in fitting experimental data and predicting light attenuation in both low and high cell density cultures. Taking advantage of the prediction power of the modified Beer-Lambert model, we devised an approach to maintain constant mean light intensity (Imean) and by adjusting the incident light intensity (I0). The results indicate that at an Imean of approximately 66.67 μmol/m2/s and 6 mm culture thickness, maximum volumetric and areal biomass productivities of 2.72 g/L/day and 16.32 g/m2/day, respectively, were achieved. Whereas the highest biomass concentration of 28.20 g/L was produced in 20 days at Imean 50 μmol/m2/s. Photoinhibition to cell division become evident at I0 750 μmol/m2/s and above. Using the data generated at constant Imean in the constant specific growth rate range, superb fitting to the Monod model with a R2 of 0.9910 was demonstrated, highlighting the importance of generating reliable data for the modelling of the kinetics of photoautotrophic growth of microalgae, which had considered to be challenging.

Bioactive peptides (cryptides) obtained by Bothrops jararaca serine peptidases action on myoglobin

Serine peptidases and metallopeptidases are the primary toxins found in Bothrops snakes venoms, which act on proteins in the tissues of victims or prey, and release of peptides formed through proteolytic activity. Various studies have indicated that these peptides, released by the proteolytic activity of heterologous enzymes, generate molecules with unidentified functions, referred to as cryptids. To address this, we purified serine peptidases from Bothrops jararaca venom using molecular exclusion chromatography and then incubated them with the endogenous substrate myoglobin. As a control, we also incubated the substrate with trypsin. The resulting proteolytic fragments were analyzed, separated, and collected via HPLC. These fractions were then tested on cell cultures, the active fractions were sequenced (ALELFR and TGHPETLEK) and synthesized. After confirming their activity, the peptides underwent sequencing and synthesis for additional cell tests, including the increase of cell viability, cycle phases, proliferation, signaling, growth kinetics, angiogenesis, and migration. The results revealed that the synthesized peptides exhibited cellular repair properties, suggesting a potential role in tissue repair in the range of 0.05–5 μ M. Additionally, the effects of fragments resulting from myoglobin degradation isolated (ALELFR and TGHPETLEK) revealed a regenerative action on tissue.

The Impact of Amotosalen Photochemical Pathogen Inactivation on Human Platelet Lysate

Background: Human Platelet Lysate (hPL) is a platelet-derived and growth factor-rich supplement for cell culture. It can be prepared from surplus platelet concentrates initially intended for transfusion. Amotosalen-based photochemical pathogen inactivation of platelet concentrates is used in a number of blood establishments worldwide to minimize the risk of pathogen transmission from donor to patient. Method: This pathogen inactivation method has not been formally validated for direct use on hPL. Here, we have studied the impact of pathogen inactivation on hPL and compared it to untreated hPL prepared from pathogen-inactivated platelet concentrates or control hPL. We used mass spectrometry, ELISA, and in vitro mesenchymal stem cell culture for determining residual amotosalen, final growth factor content, and cell doubling, respectively. Result: The data have shown amotosalen concentrations to be reduced a thousand-fold following pathogen inactivation, leaving trace quantities of photosensitizer molecules in the final hPL product. Some growth factors have been reported to be significantly more impacted in hPL that is directly pathogen-inactivated compared to both control conditions. This has no significant effect on the growth kinetics of adipose-derived mesenchymal stem cells. Conclusion: We have concluded direct amotosalen-based pathogen inactivation to have a measurable impact on certain growth factors in hPL, but this does not outweigh the likely benefits of reducing the odds of donor-to-patient pathogen transmission.

Syzygium aromaticum essential oil and its major constituents: Assessment of activity against Candida spp. and toxicity.

Syzigium aromaticum essential oil (EO), eugenol, and β-caryophyllene were evaluated regarding antifungal, antibiofilm, and in vitro toxicity. Additionally, in vivo toxicity of EO was observed. Anti-Candida activity was assessed through broth microdilution assay for all compounds. Time-kill assay (0, 1, 10, 30 min, 1, 2, and 4 h) was used to determine the influence of EO and eugenol on Candida Growth kinetics. Thereafter, both compounds were evaluated regarding their capacity to act on a biofilm formation and on mature biofilm, based on CFU/ml/g of dry weight. Cell Titer Blue Viability Assay was used for in vitro cytotoxicity, using oral epithelial cells (TR146) and human monocytes (THP-1). Lastly, Galleria mellonella model defined the EO in vivo acute toxicity. All compounds, except β-cariofilene (MIC > 8000 μg/ml), presented antifungal activity against Candida strains (MIC 500-1000 μg/ml). The growth kinetics of Candida was affected by the EO (5xMIC 30 min onward; 10xMIC 10 min onward) and eugenol (5xMIC 10 min onward; 10xMIC 1 min onward). Fungal viability was also affected by 5xMIC and 10xMIC of both compounds during biofilm formation and upon mature biofilms. LD50 was defined for TR146 and THP1 cells at, respectively, 59.37 and 79.54 μg/ml for the EO and 55.35 and 84.16 μg/ml for eugenol. No sign of toxicity was seen in vivo up to 10mg/ml (20 x MIC) for the EO. S. aromaticum and eugenol presented antifungal and antibiofilm activity, with action on cell growth kinetics. In vivo acute toxicity showed a safe parameter for the EO up to 10 mg/ml.

IQGAP3 Is an Important Mediator of Skin Inflammatory Diseases.

IQGAP3 (IQ Motif Containing GTPase Activating Protein 3) is member of the IQGAP family of scaffold proteins, which are essential for assembling multiprotein complexes that coordinate various intracellular signaling pathways. Previous research has shown that IQGAP3 is overexpressed in psoriatic skin lesions. Given its involvement in processes like cell proliferation and chemokine signaling, we sought to explore its molecular role in driving the psoriatic phenotype of keratinocytes. By conducting transcriptome profiling of HaCaT keratinocytes, we identified numerous psoriasis-associated pathways that were affected when IQGAP3 was knocked down. These included alterations in NFkB signaling, EGFR signaling, activation of p38/MAPK and ERK1/ERK2, lipid metabolism, cytokine production, and the response to inflammatory cytokine stimulation. Real-time analysis further revealed changes in cell growth dynamics, including proliferation and wound healing. The balance between cell proliferation and apoptosis was altered, as were skin barrier functions and the production of IL-6 and IFNγ. Despite these significant findings, the diversity of the alterations observed in the knockdown cells led us to conclude that IQGAP3 may not be the best target for the therapeutic inhibition to normalize the phenotype of keratinocytes in psoriasis.

Exploring microalgal nutrient-light synergy to enhance CO2 utilization and lipid productivity in sustainable long-term water recycling cultivation

In this work the effects of nutrient availability and light conditions on CO2 utilization and lipid production in Micractinium pusillum KMC8 is reported. The study investigated the ideal nitrogen concentrations for growth and nitrogen utilization in a 15% CO2 environment. Logistic and Gompertz models were employed to analyze the kinetics of KMC8 cell growth. Compared to 17.6 mmol L−1 control nitrogen, which generated 1.6 g L−1 growth, doubling and quadrupling nitrogen concentrations boosted biomass growth by 12.5% and 28.78%. At 8.6 mmol L−1 nitrogen, the growth decreased but lipid productivity increased to 18.62 mg L−1 day−1. At 70.6 mmol L−1 nitrogen, elevated nitrogen levels maintained an alkaline pH above 7 and enhanced CO2 mitigation, achieving 2.27% CO2 utilization efficiency. Nitrogen shows a positive correlation with higher rates of carbon and nitrogen fixation. The investigation extends to find out the influence of phosphorus and light conditions on microalgae. Increasing light intensity incrementally from 150 to 1200 μmol m−2 s−1 with more phosphorus increased biomass productivity by 85% (255 mg L-1 day−1) and lipid productivity by 2.5-fold (84.76 mg L-1 day−1), with 3.3% CO2 utilization efficiency compared to directly using 1200 μmol m−2 s−1. This study suggests a water recycling-fed batch cycle with gradual light feeding, which results in high CO2 fixation (1.1 g L−1 day−1), 7% CO2 utilization, and significant biomass and lipid productivity (577.23 and 150 mg L−1 day−1). This approach promotes lipid synthesis, maintains carbon fixation, and minimizes biomass loss, thus supporting sustainable bioenergy development in a circular bio-economy framework.

A mathematical model for predicting the spatiotemporal response of breast cancer cells treated with doxorubicin

ABSTRACT Tumor heterogeneity contributes significantly to chemoresistance, a leading cause of treatment failure. To better personalize therapies, it is essential to develop tools capable of identifying and predicting intra- and inter-tumor heterogeneities. Biology-inspired mathematical models are capable of attacking this problem, but tumor heterogeneity is often overlooked in in-vivo modeling studies, while phenotypic considerations capturing spatial dynamics are not typically included in in-vitro modeling studies. We present a data assimilation-prediction pipeline with a two-phenotype model that includes a spatiotemporal component to characterize and predict the evolution of in-vitro breast cancer cells and their heterogeneous response to chemotherapy. Our model assumes that the cells can be divided into two subpopulations: surviving cells unaffected by the treatment, and irreversibly damaged cells undergoing treatment-induced death. MCF7 breast cancer cells were previously cultivated in wells for up to 1000 hours, treated with various concentrations of doxorubicin and imaged with time-resolved microscopy to record spatiotemporally-resolved cell count data. Images were used to generate cell density maps. Treatment response predictions were initialized by a training set and updated by weekly measurements. Our mathematical model successfully calibrated the spatiotemporal cell growth dynamics, achieving median [range] concordance correlation coefficients of > .99 [.88, >.99] and .73 [.58, .85] across the whole well and individual pixels, respectively. Our proposed data assimilation-prediction approach achieved values of .97 [.44, >.99] and .69 [.35, .79] for the whole well and individual pixels, respectively. Thus, our model can capture and predict the spatiotemporal dynamics of MCF7 cells treated with doxorubicin in an in-vitro setting.

Kinetics of microbial cell growth, utilization of organic substrates and methane production in upflow anaerobic filters in two and three separated stages treating sanitary landfill leachates

AbstractThis paper deals with the kinetics of microbial cell growth, utilization of organic substrates and methane production in upflow anaerobic filters in two and three separated stages (UAF‐2SS and UAF‐3SS) treating sanitary landfill leachates, under three temperatures (20, 27, and 34°C), fed with volumetric organic loads (VOLs) for UAF‐2SS (1.8, 2.25, 2.76, 3.45, 3.71, 4.64 kgCOD (chemical oxygen demand)/m3/d) and UAF‐3SS (1.7, 2.25, 2.44, 2.6, 3.15, 3.45, 3.5, 4.57, 4.64, 6.31, 11.61, 14.18, 17.48 kgCOD/m3/d). Kinetic parameters were obtained at temperature (20–34°C), pH (8.8–9.1), each one of three stages within the UAF‐2SS and UAF‐3SS reactors containing methanogenic bacteria (Methanococcus sp., and Methanobacterium sp.) and Clostridium sp.: Maximum substrate utilization rate (rm,s) resulted in 102 mg COD/L/h, maximum microbial cell growth rate (μm) varied 102–103 CFU (colony forming units)/mL/h, methane (CH4) production rate (rm,CH4) tended to vary 100–101 mg CH4/d and the maximum production coefficient (Y) varied 0–102 CFU/mL/h/mgCOD/L/h.

Kinetic studies and dynamic modeling of sophorolipids production by Candida catenulata using different carbon sources

Abstract The kinetic study on sophorolipids (SLs) production by Candida catenulata from glucose, raw sunflower soapstock was investigated at different initial concentrations ranging from 0 to 100 g L−1. The Monod model with a maximum specific growth rate (μ max) of 0.0167 h−1 and half-saturation coefficient (K S ) of 6.91 g L−1 best described the cell growth kinetics of C. catenulata on glucose. The best-fitted constants of the Monod model for raw sunflower soapstock were μ max = 0.0157 h−1 and K S = 16.01 g L−1. Determination of Luedeking-Piret constants indicated SLs mainly produced as an associated growth product in the systems. Dynamic features of the fermentation were modeled using the obtained constants and results showed the prediction power of the developed model in describing the behavior of the process. Also, a modified kinetic model was developed for the dynamic modeling of the dual carbon sources system.

Engineering mammalian cell growth dynamics for biomanufacturing

Precise control over mammalian cell growth dynamics poses a major challenge in biopharmaceutical manufacturing. Here, we present a multi-level cell engineering strategy for the tunable regulation of growth phases in mammalian cells. Initially, we engineered mammalian death phase by employing CRISPR/Cas9 to knockout pro-apoptotic proteins Bax and Bak, resulting in a substantial attenuation of apoptosis by improving cell viability and extending culture lifespan. The second phase introduced a growth acceleration system, akin to a “gas pedal”, based on an abscidic acid inducible system regulating cMYC gene expression, enabling rapid cell density increase and cell cycle control. The third phase focused on a stationary phase inducing system, comparable to a “brake pedal”. A tetracycline inducible genetic circuit based on BLIMP1 gene led to cell growth cessation and arrested cell cycle upon activation. Finally, we developed a dual controllable system, combining the “gas and brake pedals”, enabling for dynamic and precise orchestration of mammalian cell growth dynamics. This work exemplifies the application of synthetic biology tools and combinatorial cell engineering, offering a sophisticated framework for manipulating mammalian cell growth and providing a unique paradigm for reprogramming cell behaviour for enhancing biopharmaceutical manufacturing and further biomedical applications.

To click or not to click for short pulse-labeling of the bacterial cell wall.

A method of choice to study the spatio-temporal dynamics of bacterial cell growth and division is to analyze the localization of cell wall synthesis regions by fluorescence microscopy. For this, nascent cell wall biopolymers need to be labeled with fluorescent reporters, like fluorescent d-alanines (FDAs) that can be incorporated into the peptidoglycan. To achieve high spatial and temporal resolution, dense, high-intensity fluorescence labeling must be obtained in the shortest possible time. However, modifications carried by d-Ala can hinder their uptake by the enzymes that incorporate them into the peptidoglycan, such as the d,d-transpeptidases. Conversely, these modifications can impede the elimination of the incorporated d-Ala derivatives by d,d-carboxypeptidases, making the labeling more persistent. In this context, we synthesized clickable d-Alas and tested their incorporation into the peptidoglycan using different labeling approaches, prior or after their conjugation to clickable fluorescent dyes through SPAAC reaction. Our data allow ranking of the d-Ala derivatives in terms of their ease of incorporation and resistance to trimming during one-step, "one-pot" two-step or sequential two-step labeling strategies. We further show that a hybrid "one-step" approach, in which a FDA is used in combination with clickable choline and fluorescent dye, enables two-color co-labeling of peptidoglycan and teichoic acids. Finally, we identify a strategy compatible with the cell fixation required for super-resolution microscopy, by combining one-step labeling with FDA and sequential two-step labeling with clickable choline and fluorescent dye, allowing to obtain two-color high-resolution images of peptidoglycan and teichoic acid synthesis regions.

Nasoseptal chondroprogenitors isolated through fibronectin-adherence confer no biological advantage for cartilage tissue engineering compared to nasoseptal chondrocytes.

The ability to bioprint facial cartilages could revolutionise reconstructive surgery, but identifying the optimum cell source remains one of the great challenges of tissue engineering. Tissue specific stem cells: chondroprogenitors, have been extracted previously using preferential adhesion to fibronectin based on the expression of CD49e: a perceived chondroprogenitor stem cell marker present on <1% of cartilage cells. This study sought to determine whether these fibronectin-adherent chondroprogenitor cells could be exploited for cartilage tissue engineering applications in isolation, or combined with differentiated chondrocytes. Nasoseptal cartilage samples from 20 patients (10 male, 10 female) were digested to liberate cartilage-derived cells (CDCs) from extracellular matrix. Total cell number was counted using the Trypan Blue exclusion assay and added to fibronectin coated plates for 20min, to determine the proportion of fibronectin-adherent (FAC) and non-adherent cells (NFACs). All populations underwent flow cytometry to detect mesenchymal stem/progenitor cell markers and were cultured in osteogenic, chondrogenic and adipogenic media to determine trilineage differentiation potential. Cell adherence and growth kinetics of the different populations were compared using iCELLigence growth assays. Chondrogenic gene expression was assessed using RT-qPCR for Type 2 collagen, aggrecan and SOX9 genes. Varying proportions of NFAC and FACs were cultured in alginate beads to assess tissue engineering potential. 52.6% of cells were fibronectin adherent in males and 57.7% in females, yet on flow cytometrical analysis, only 0.19% of cells expressed CD49e. Moreover, all cells (CDC, FAC and NFACs) demonstrated an affinity for trilineage differentiation by first passage and the expression of stem/progenitor cell markers increased significantly from digest to first passage (CD29, 44, 49e, 73 and 90, p < 0.0001). No significant differences were seen in adhesion or growth rates. Collagen and aggrecan gene expression was higher in FACs than CDCs (2-fold higher, p = 0.008 and 0.012 respectively), but no differences in chondrogenic potential were seen in any cell mixtures in 3D culture models. The fibronectin adhesion assay does not appear to reliably isolate a chondroprogenitor cell population from nasoseptal cartilage, and these cells confer no advantageous properties for cartilage tissue engineering. Refinement of cell isolation methods and chondroprogenitor markers is warranted for future nasoseptal cartilage tissue engineering efforts.

The impact of acemannan, an extracted product from Aloe vera, on proliferation of dental pulp stem cells and healing of mandibular defects in rabbits.

Dental pulp stem cells (DPSCs) were shown to play an important role in regenerative medicine including reconstruction of various bone lesions. This study determined the impact of acemannan, an extracted product from Aloe vera, on in vitro proliferation of DPSCs and in vivo healing of mandibular defects in rabbits. DPSCs were isolated and characterized. The growth kinetics of cells exposed to acemannan (8 mg/mL) and Hank's balanced salt solution (HBSS) were compared in vitro. Fifteen male rabbits were divided into 3 groups. Five animals were left as control group without any therapeutic intervention. Five rabbits were considered as experimental group 1 and received 20 µL of a cell suspension containing 106 DPSCs in the bone defect. Another 5 rabbits were regarded as experimental group 2 and were injected in the bone defect with 20 µL of a cell suspension containing 106 DPSCs treated with acemannan for 24 h. After 60 days, the animals were assessed by radiography and histologically. The mesenchymal properties of DPSCs were confirmed. Population doubling time (PDT) of DPSCs treated with acemannan (29.8 h) was significantly shorter than cells were just exposed to HBSS (45.9 h). DPSCs together with acemannan could significantly accelerate the healing process and osteogenesis in mandibular defects. As DPSCS showed an increased proliferation when treated with acemannan and accelerated the healing process in mandibular defects, these findings can open a new avenue in dentistry regenerative medicine when remedies of bone defects are targeted.

Chlorophyllin attenuates the effects of benzo[a]pyrene in human hepatoma HepG2/C3A cells

Chlorophyllin, a semisynthetic compound derived from chlorophyll, has been a focus in cancer prevention because it exerts important biological activities, such as antigenotoxic, antioxidative and anticarcinogenic activities. The aim of this study was to evaluate the effect of chlorophyllin against benzo[a]pyrene toxicity in HepG2/C3A cells. Cells were co-treated (chlorophyllin plus benzo[a]pyrene) and the cell viability, cell growth kinetics, cell cycle, and apoptosis were evaluated. The mRNA levels of cell cycle and apoptotic genes were analyzed. Our results showed that chlorophyllin reduce the antiproliferative effects of benzo[a]pyrene in a multi-specific manner, restoring the normal distribution of the cell cycle and inhibiting the cell death induced by the xenobiotic. Gene expression analysis showed that benzo[a]pyrene decreased the mRNA levels of genes involved in cell cycle control (cyclins and cyclin-dependent kinases) and apoptosis. Cells co-treated with 200 µM of chlorophyllin and benzo[a]pyrene also showed a downregulation of mRNA levels of the genes, but the expression levels of the gene that encodes to tumor protein p53 were maintained near to control. Thus, chlorophyllin is a good candidate as a chemoprotective agent that mitigates the cytotoxic effects benzo[a]pyrene and, thus, might be a promising tool to prevent liver cancer.

Evaluation of a Peptide Hydrogel as a Chondro-Instructive Three-Dimensional Microenvironment.

Articular cartilage injuries are inherently irreversible, even with the advancement in current therapeutic options. Alternative approaches, such as the use of mesenchymal stem/stromal cells (MSCs) and tissue engineering techniques, have gained prominence. MSCs represent an ideal source of cells due to their low immunogenicity, paracrine activity, and ability to differentiate. Among biomaterials, self-assembling peptide hydrogels (SAPH) are interesting given their characteristics such as good biocompatibility and tunable properties. Herein we associate human adipose-derived stem cells (hASCs) with a commercial SAPH, Puramatrix™, to evaluate how this three-dimensional microenvironment affects cell behavior and its ability to undergo chondrogenic differentiation. We demonstrate that the Puramatrix™ hydrogel comprises a highly porous matrix permissible for hASC adhesion and in vitro expansion. The morphology and cell growth dynamics of hASCs were affected when cultured on the hydrogel but had minimal alteration in their immunophenotype. Interestingly, hASCs spontaneously formed cell aggregates throughout culturing. Analysis of glycosaminoglycan production and gene expression revealed a noteworthy and donor-dependent trend suggesting that Puramatrix™ hydrogel may have a natural capacity to support the chondrogenic differentiation of hASCs. Altogether, the results provide a more comprehensive understanding of the potential applications and limitations of the Puramatrix™ hydrogel in developing functional cartilage tissue constructs.

Chaos in a bacterial stress response

Cellular responses to environmental changes are often highly heterogeneous and exhibit seemingly random dynamics. The astonishing insight of chaos theory is that such unpredictable patterns can, in principle, arise without the need for any random processes, i.e., purely deterministically without noise. However, while chaos is well understood in mathematics and physics, its role in cell biology remains unclear because the complexity and noisiness of biological systems make testing difficult. Here, we show that chaos explains the heterogeneous response of Escherichia coli cells to oxidative stress. We developed a theoretical model of the gene expression dynamics and demonstrate that chaotic behavior arises from rapid molecular feedbacks that are coupled with cell growth dynamics and cell-cell interactions. Based on theoretical predictions, we then designed single-cell experiments to show we can shift gene expression from periodic oscillations to chaos on demand. Our work suggests that chaotic gene regulation can be employed by cell populations to generate strong and variable responses to changing environments.