Chemical Inducers Research Articles

Creatinine combined with light increases the contents of lutein and β-carotene, the main carotenoids of Dunaliella bardawil

Dunaliella bardawil, a unicellular green alga, can accumulate a large amount of lutein and β-carotene under stresses. Using chemical inducers combined with abiotic stress to promote the accumulation of high value-added products such as lipids and carotenoids in microalgae has attracted more and more attention. In this study, creatinine was added into autotrophic medium to investigate its effects on the growth, chlorophyll content, and the ingredients and content of carotenoids in D. bardawil. The results showed that creatinine alone could significantly increase the biomass, chlorophyll and carotenoid contents of D. bardawil, among which the contents of lutein and β-carotene were further increased, while the content of zeaxanthin was decreased. In order to further improve the content of the two carotenoids, different light intensities combined with creatinine have been adopted. Under 6.589 W/m2 light intensity, creatinine could effectively increase the production of lutein, zeaxanthin, α-carotene and β-carotene. Compared with the control, the content of lutein increased by 46 % and the content of β-carotene increased by 77 % when the concentration of creatinine was 500 μg/mL. In conclusion, creatinine can effectively improve the production lutein and β-carotene in D. bardawil, which is more conducive under lower light intensity.

Use of capsaicin as a model in the study of migraine: a literature review

Capsaicin is able to induce mast cell degranulation, an event probably related to the pathophysiologyof a migraine attack. The present review study aimed to address the mechanisms of action of capsaicin and other chemical inducers in mast cell degranulation and an interaction of nerves and events that happen in the dura mater with the activation of mast cells. A survey was carried out in the literature, from 1980 to 2019, in different databases, using the following terms: capsaicin, mast cell and dura mater. 36 articles were selected for this review. Studies indicate that the main mechanisms of action of capsaicin are chemical induction through the activation of TRPV1 channels,allowing calcium influx into neurons in the trigeminal ganglion of the dura mater, activating mast cell degranulation, releasing pro-inflammatory (e.g., histamine, oxide nitric) and vasoactive (e.g., CGRP and substance P) substances. Therefore, the use of capsaicin may be a tool to be used in an animal model to better understand the pathophysiology of migraine.

The role of biological and chemical inducers in activating bean defense responses against Rhizoctonia solani

Rhizoctonia root rot is an important bean disease that causes significant yield losses. Increasing the levels of plant resistance using biological and chemical inducers is a sustainable strategy for plant disease management. In this study, the efficiency of thiamine, riboflavin, pyridoxine, folic acid, menadione, quercetin, naringenin, humic acid, fulvic acid, boric acid, zinc sulfate, manganese sulfate, Piriformospora indica and binucleate Rhizoctonia were evaluated to improve bean resistance to root rot pathogen Rhizoctonia solani AG-4 HGII. Under laboratory conditions, effect of various biological agents and chemical compounds on mycelial growth of the pathogen was evaluated. Results showed that applying boric acid, zinc sulfate and manganese sulfate at 1000 and 2000 mg L−1 concentrations and menadione at 2000, 5000, 10000 and 20000 μM completely inhibited the pathogen growth and 10000 and 20000 μM concentrations of thiamine reduced linear growth of R. solani. Under greenhouse conditions, application of the inducers significantly reduced the disease progress on treated plants compared to the non-treated plants. The highest level of disease reduction was observed using zinc sulfate and thiamine applied at 3 days before the pathogen inoculation, and also by using P. indica 14 days before inoculation with the pathogen. Thiamine, zinc sulfate and P. indica significantly reduced the disease index and improved plant growth parameters compared to the controls. Induction of resistance against R. solani by individual and combined treatments using thiamine, zinc sulfate and P. indica was associated with increased activity of various antioxidants and increased levels of membrane stability index (MSI), relative water content (RWC), lignin accumulation and total soluble phenolics in bean seedlings. Our findings revealed a practical possibility of inducing resistance and protection of bean plants against Rhizoctonia root rot with the help of biological and chemical resistance inducers.

Osteoinductive and antimicrobial mechanisms of graphene-based materials for enhancing bone tissue engineering.

Graphene-based materials (GMs) have great application prospects in bone tissue engineering due to their osteoinductive ability and antimicrobial activity. GMs induce osteogenic differentiation through several mechanisms and pathways in bone tissue engineering. First of all, the surface and high hardness of the porous folds of graphene or graphene oxide (GO) can generate mechanical stimulation to initiate a cascade of reactions that promote osteogenic differentiation without any chemical inducers. In addition, change of the extracellular matrix (ECM), regulation of macrophage polarization, the oncostatin M (OSM) signaling pathway, the MAPK signaling pathway, the BMP signaling pathway, the Wnt/β-catenin signaling pathway, and other pathways are involved in GMs' regulation of osteogenesis. In bone tissue engineering, GMs prevent the formation of microbial biofilms mainly through preventing microbial adhesion and killing them. The former is mainly achieved by reducing surface free energy (SFE) and increasing hydrophobicity. The latter mainly includes oxidative stress and photothermal/photodynamic effects. Graphene and its derivatives (GDs) are mainly combined with bioactive ceramic materials, metal materials and macromolecular polymers to play an antimicrobial effect in bone tissue engineering. Concentration, number of layers, and type of GDs often affect the antimicrobial activity of GMs. In this paper, we reviewed relevant osteoinductive and antimicrobial mechanisms of GMs and their applications in bone tissue engineering.

In vitro differentiation of human bone marrow stromal cells into neural precursor cells using small molecules

BackgroundNeurogenic differentiation of human marrow stromal stem cells (hMSCs) into neural precursor cells (NPCs) offers new hope in many neurological diseases. Stromal cells can be differentiated into NPCs using small molecules acting as chemical inducers. The aim of this study is to formulate an efficient, direct, fast and safe protocol to differentiate hMSCs into NPCs using different inducers: b-mercaptoethanol (BME), triiodothyronine (T3), and curcumin (CUR). New methodhMSCs were subjected to either 1 mM BME, 0.5 µM T3, or 5 µM CUR. Neurogenic differentiation was determined by assessing the protein expression of PAX6, SOX2, DLX2, and GAP-43 with flow cytometry and immunofluorescence, along with Nissl staining of differentiated cells. Results and Comparison with Existing MethodIt was revealed that T3 and CUR are 70–80% better than BME in terms of efficiency and safety, and surprisingly BME was a good promoting factor for cell preconditioning with limited effects on neural trans-differentiation related to its toxic effects on cell viability. ConclusionReprogramming of bone marrow stromal cells into neural cells gives hope for treating different neurological disorders. Our study shows that T3 and CUR were effective in generation of NPCs from hMSCs with preservation of cell viability. BME was a good promoting factor for cell preconditioning with limited effects on neural transdifferentiation related to its toxic effects on cell viability.

The multifunctionality of expression systems in Bacillus subtilis: Emerging devices for the production of recombinant proteins.

Bacillus subtilis is a successful host for producing recombinant proteins. Its GRAS (generally recognized as safe) status and its remarkable innate ability to absorb and incorporate exogenous DNA into its genome make this organism an ideal platform for the heterologous expression of bioactive substances. The factors that corroborate its value can be attributed to the scientific knowledge obtained from decades of study regarding its biology that has fostered the development of several genetic engineering strategies, such as the use of different plasmids, engineering of constitutive or double promoters, chemical inducers, systems of self-inducing expression with or without a secretion system that uses a signal peptide, and so on. Tools that enrich the technological arsenal of this expression platform improve the efficiency and reduce the costs of production of proteins of biotechnological importance. Therefore, this review aims to highlight the major advances involving recombinant expression systems developed in B. subtilis, thus sustaining the generation of knowledge and its application in future research. It was verified that this bacterium is a model in constant demand and studies of the expression of recombinant proteins on a large scale are increasing in number. As such, it represents a powerful bacterial host for academic research and industrial purposes.

Biomineralization potential and biological properties of a new tantalum oxide (Ta2O5)\u2013containing calcium silicate cement

ObjectiveThe present study evaluated the biological effects and biomineralization potential of a new tantalum oxide (Ta2O5)–containing material designed for vital pulp therapy or perforation repair (NeoMTA 2), compared to NeoMTA Plus and Bio-C Repair.Material and methodsHuman dental pulp stem cells (hDPSCs) were exposed to different eluates from NeoMTA Plus, NeoMTA 2, and Bio-C Repair. Ion release from each material was determined using inductively coupled plasma-optical emission spectrometry (ICP-MS). The biological experiments performed were MTT assays, apoptosis/necrosis assays, adhesion assays, migration assays, morphology evaluation, and reactive oxygen species (ROS) production analysis. Biomineralization was assessed by Alizarin red S staining. Finally, osteo/odontogenic gene expression was determined by real-time quantitative reverse-transcriptase polymerase chain reaction (RT-qPCR). Data were analyzed using one-way ANOVA followed by Tukey’s multiple comparison test.ResultsNeoMTA 2 displayed a significantly higher calcium release compared to the other materials (p < 0.05). When hDPSCs were cultured in presence of the different material eluates, all groups exhibited similar hDPSC viability and migration rates when compared to untreated cells. Substantial cell attachment and spreading were observed in all materials’ surfaces, without significant differences. hDPSCs treated with NeoMTA 2 displayed an upregulation of ALP, Col1A1, RUNX2 (p < 0.001), ON, and DSPP genes (p < 0.05), and showed the highest mineralization potential compared to other groups (p < 0.001). Finally, the more concentrated eluates from these materials, specially NeoMTA Plus and NeoMTA 2, promoted higher ROS production in hDPSCs compared to Bio-C Repair and control cells (p < 0.001), although these ROS levels did not result in increased cell death.ConclusionsThe new tantalum oxide (Ta2O5)–containing material shows an adequate cytocompatibility and the ability to promote biomineralization without using chemical osteogenic inducers, showing great potential as a new material for vital pulp therapy.Clinical relevanceNeoMTA 2 seems to be a promising material for vital pulp therapy. Further studies considering its biocompatibility and biomineralization potential are necessary.

A precisely adjustable, variation-suppressed eukaryotic transcriptional controller to enable genetic discovery.

Conditional expression of genes and observation of phenotype remain central to biological discovery. Current methods enable either on/off or imprecisely controlled graded gene expression. We developed a 'well-tempered' controller, WTC846, for precisely adjustable, graded, growth condition independent expression of genes in Saccharomyces cerevisiae. Controlled genes are expressed from a strong semisynthetic promoter repressed by the prokaryotic TetR, which also represses its own synthesis; with basal expression abolished by a second, 'zeroing' repressor. The autorepression loop lowers cell-to-cell variation while enabling precise adjustment of protein expression by a chemical inducer. WTC846 allelic strains in which the controller replaced the native promoters recapitulated known null phenotypes (CDC42, TPI1), exhibited novel overexpression phenotypes (IPL1), showed protein dosage-dependent growth rates and morphological phenotypes (CDC28, TOR2, PMA1 and the hitherto uncharacterized PBR1), and enabled cell cycle synchronization (CDC20). WTC846 defines an 'expression clamp' allowing protein dosage to be adjusted by the experimenter across the range of cellular protein abundances, with limited variation around the setpoint.

Research progress in establishment of N-methyl-N'-nitro-N-nitroso-guanidine-induced rat model of Precancerous lesion of gastric cancer

Gastric cancer(GC), one of the most common malignancies worldwide, seriously threatens human health due to its high morbidity and mortality. Precancerous lesion of gastric cancer(PLGC) is a critical stage for preventing the occurrence of gastric cancer, and PLGC therapy has frequently been investigated in clinical research. Exploring the proper animal modeling methods is necessary since animal experiment acts as the main avenue of the research on GC treatment. At present, N-methyl-N'-nitro-N-nitroso-guanidine(MNNG) serves as a common chemical inducer for the rat model of GC and PLGC. In this study, MNNG-based methods for modeling PLGC rats in related papers were summarized, and the applications and effects of these methods were demonstrated by examples. Additionally, the advantages, disadvantages, and precautions of various modeling methods were briefly reviewed, and the experience of this research group in exploring modeling methods was shared. This study is expected to provide a reference for the establishment of MNNG-induced PLGC animal model, and a model support for the following studies on PLGC.

Shear type and magnitude affect aortic valve endothelial cell morphology, orientation, and differentiation.

Valvular endothelial cells line the outer layer of heart valves and can withstand shear forces caused by blood flow. In contrast to vascular endothelial cells, there is limited amount of research over valvular endothelial cells. For this reason, the exact physiologic behavior of valvular endothelial cells is unclear. Prior studies have concluded that valvular endothelial cells align perpendicularly to the direction of blood flow, while vascular endothelial cells align parallel to blood flow. Other studies have suggested that different ranges of shear stress uniquely impact the behavior of valvular endothelial cells. The goal of this study was to characterize the response of valvular endothelial cell under different types, magnitudes, and durations of shear stress. In this work, the results demonstrated that with increased shear rate and duration of exposure, valvular endothelial cells no longer possessed the traditional cuboidal morphology. Instead through the change in cell circularity and aspect ratio, valvular endothelial cells aligned in an organized manner. In addition, different forms of shear exposure caused the area and circularity of valvular endothelial cells to decrease while inducing mesenchymal transformation validated through αSMA and TGFβ1 expression. This is the first investigation showing that valvular endothelial cells alignment is not as straightforward as once thought (perpendicular to flow). Different types and magnitudes of shear induce different local behaviors. This is also the first demonstration of valvular endothelial cells undergoing EndMT without chemical inducers on a soft surface in vitro. Findings from this study provide insights to understanding the pathophysiology of valvular endothelial cells which can potentially propel future artificial engineered heart valves.

Chemical Induction of Trophoblast Hypoxia by Cobalt Chloride Leads to Increased Expression of DDIT3

Choriocarcinoma cells BeWo b30 are used to model human placental trophoblast hypoxia using cobalt (II) chloride and hydroxyquinoline derivative (HD) as chemical inducers of hypoxia-inducible factor (HIF). In this study, it was shown that both substances activate the hypoxic pathway and the epithelial–mesenchymal transition and inhibit the pathways of cell proliferation. However, CoCl2 caused activation of the apoptosis pathway, increased the activity of effector caspases 3 and 7, and increased the expression of the unfolded protein response target DDIT3. The mTORC1 pathway was activated upon exposition to CoCl2, while HD suppressed this pathway, as it happens during real trophoblast hypoxia. Thus, effect of CoCl2 on BeWo cells can be a model of severe hypoxia with activation of apoptosis, while HD mimics moderate hypoxia.

Abstract 2685: Tumor hypoxia drives immunosuppression via upregulation of galectin-1 in recurrent Glioblastoma

Abstract Background: Glioblastoma (GB) recurrence is associated with a highly immunosuppressive tumor microenvironment. Previous research in our lab has illustrated that the carbohydrate-binding protein Galectin-1 (Gal-1) has high expression in GB patient samples and plays vital pro-tumorigenic roles within the tumor microenvironment, regulated by hypoxia-inducible factor-1 (HIF-1). However, since Gal-1 displays many different activities by binding to a number of diverse N- or O-glycan modified target proteins, it has been difficult to fully understand how Gal–1 supports tumor growth and promotes resistance to anti-VEGF mAb. Thus, our aim is to elucidate the role of tumor hypoxia which may in turn increase tumor aggressiveness by up-regulating the expression of Gal-1 in relevant GB in vitro and in vivo model. Methods and Results: GB cells were cultured in a hypoxic environment simulated by cobalt chloride (CoCl2), which is a chemical inducer of hypoxia-inducible factor-1α (HIF-1α). HIF-1α is a transcription factor that has an important role in tumor cell adaptation to hypoxia and controls the expression of several genes. Various CoCl2 concentrations were used to simulate degrees of hypoxia in GB cells. Following treatment with 250 µM CoCl2, an MTT assay was conducted to determine the growth and anti-VEGF mAb (+/- Radiation + Temozolomide) sensitivity of GB cells. We observed that CoCl2(chemical activator of HIF-1α)-treatment simulated hypoxia and induced resistance to anti-VEGF mAb treatment via upregulation of HIF-1α, MDR1/P-gp and MRP protein levels. In addition, proapoptotic members of the Bcl-2 protein family, Bax and Bad, were downregulated. The anti-apoptotic member Bcl-2 exhibited no significant change in expression, whereas the ratio of Bcl-2/Bax was increased. We observed that hypoxia induced Gal-1 through mechanisms that are independent of HIF 1α and HIF-2α but involved reactive oxygen species-dependent activation of the transcription factor nuclear factor κB. Since upregulation of hypoxia inducible transcription factors (HIFs) via Gal-1 is the primary adaptive cellular response to a hypoxic environment, we analyzed the effects of Gal-1 inhibition treatment in vivo. We observed significant alteration of hypoxic tumor microenvironment (TME) in recurrent GB by alteration of tumor associate microglia phenotypes from M2 to. M1, suppression of CD4+CD25+Foxp3+ regulatory T cells and activation of CD 8+ T cells in in vivo GB models. Conclusion: The present study revealed that a CoCl2-simulated hypoxic microenvironment was able to effectively induce resistance to anti-VEGF mAb treatment in GB cells. The results demonstrate the inhibitory effect of Gal-1 in the tumor hypoxia microenvironment and inhibition may be a viable therapeutic target for recurrent GB treatment. Citation Format: Wayne Glore, Arabinda Das, Scott M. Lindhorst, Abhay K. Varma, William A. Vandergrift, Sunil J. Patel, David Cachia. Tumor hypoxia drives immunosuppression via upregulation of galectin-1 in recurrent Glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2685.

LED control of gene expression in a nanobiosystem composed of metallic nanoparticles and a genetically modified E. coli strain

BackgroundWithin the last decade, genetic engineering and synthetic biology have revolutionized society´s ability to mass-produce complex biological products within genetically-modified microorganisms containing elegantly designed genetic circuitry. However, many challenges still exist in developing bioproduction processes involving genetically modified microorganisms with complex or multiple gene circuits. These challenges include the development of external gene expression regulation methods with the following characteristics: spatial–temporal control and scalability, while inducing minimal permanent or irreversible system-wide conditions. Different stimuli have been used to control gene expression and mitigate these challenges, and they can be characterized by the effect they produce in the culture media conditions. Invasive stimuli that cause permanent, irreversible changes (pH and chemical inducers), non-invasive stimuli that cause partially reversible changes (temperature), and non-invasive stimuli that cause reversible changes in the media conditions (ultrasound, magnetic fields, and light).MethodsOpto-control of gene expression is a non-invasive external trigger that complies with most of the desired characteristics of an external control system. However, the disadvantage relies on the design of the biological photoreceptors and the necessity to design them to respond to a different wavelength for every bioprocess needed to be controlled or regulated in the microorganism. Therefore, this work proposes using biocompatible metallic nanoparticles as external controllers of gene expression, based on their ability to convert light into heat and the capacity of nanotechnology to easily design a wide array of nanostructures capable of absorbing light at different wavelengths and inducing plasmonic photothermal heating.ResultsHere, we designed a nanobiosystem that can be opto-thermally triggered using LED light. The nanobiosystem is composed of biocompatible gold nanoparticles and a genetically modified E. coli with a plasmid that allows mCherry fluorescent protein production at 37 °C in response to an RNA thermometer.ConclusionsThe LED-triggered photothermal protein production system here designed offers a new, cheaper, scalable switchable method, non-destructive for living organisms, and contribute toward the evolution of bioprocess production systems.

Substrate-activated expression of a biosynthetic pathway in Escherichia coli.

Microbes can facilitate production of valuable chemicals more sustainably than traditional chemical processes in many cases: they utilize renewable feedstocks, require less energy intensive process conditions, and perform a variety of chemical reactions using endogenous or heterologous enzymes. In response to the metabolic burden imposed by production pathways, chemical inducers are frequently used to initiate gene expression after the cells have reached sufficient density. While chemically inducible promoters are a common research tool used for pathway expression, they introduce a compound extrinsic to the process along with the associated costs. We developed an expression control system for a biosynthetic pathway for the production of d-glyceric acid that utilizes galacturonate as both the inducer and the substrate, thereby eliminating the need for an extrinsic chemical inducer. Activation of expression in response to the feed is actuated by a galacturonate-responsive transcription factor biosensor. We constructed variants of the galacturonate biosensor with a heterologous transcription factor and cognate hybrid promoter, and selected for the best performer through fluorescence characterization. We showed that native E. coli regulatory systems do not interact with our biosensor and favorable biosensor response exists in the presence and absence of galacturonate consumption. We then employed the control circuit to regulate the expression of the heterologous genes of a biosynthetic pathway for the production d-glyceric acid that was previously developed in our lab. Productivity via substrate-induction with our control circuit was comparable to IPTG-controlled induction and significantly outperformed a constitutive expression control, producing 2.13±0.03g L-1 d-glyceric acid within 6h of galacturonate substrate addition. This work demonstrated feed-activated pathway expression to be an attractive control strategy for more readily scalable microbial biosynthesis.

Designing an irreversible metabolic switch for scalable induction of microbial chemical production

Bacteria can be harnessed to synthesise high-value chemicals. A promising strategy for increasing productivity uses inducible control systems to switch metabolism from growth to chemical synthesis once a large population of cell factories are generated. However, use of expensive chemical inducers limits scalability of this approach for biotechnological applications. Switching using cheap nutrients is an appealing alternative, but their tightly regulated uptake and consumption again limits scalability. Here, using mathematical models of fatty acid uptake in E. coli as an exemplary case study, we unravel how the cell’s native regulation and program of induction can be engineered to minimise inducer usage. We show that integrating positive feedback loops into the circuitry creates an irreversible metabolic switch, which, requiring only temporary induction, drastically reduces inducer usage. Our proposed switch should be widely applicable, irrespective of the product of interest, and brings closer the realization of scalable and sustainable microbial chemical production.

Chemical tools for dissecting cell division.

Components of the cell division machinery typically function at varying cell cycle stages and intracellular locations. To dissect cellular mechanisms during the rapid division process, small-molecule probes act as complementary approaches to genetic manipulations, with advantages of temporal and in some cases spatial control and applicability to multiple model systems. This Review focuses on recent advances in chemical probes and applications to address select questions in cell division. We discuss uses of both enzyme inhibitors and chemical inducers of dimerization, as well as emerging techniques to promote future investigations. Overall, these concepts may open new research directions for applying chemical probes to advance cell biology.

Synthetic neural-like computing in microbial consortia for pattern recognition

Complex biological systems in nature comprise cells that act collectively to solve sophisticated tasks. Synthetic biological systems, in contrast, are designed for specific tasks, following computational principles including logic gates and analog design. Yet such approaches cannot be easily adapted for multiple tasks in biological contexts. Alternatively, artificial neural networks, comprised of flexible interactions for computation, support adaptive designs and are adopted for diverse applications. Here, motivated by the structural similarity between artificial neural networks and cellular networks, we implement neural-like computing in bacteria consortia for recognizing patterns. Specifically, receiver bacteria collectively interact with sender bacteria for decision-making through quorum sensing. Input patterns formed by chemical inducers activate senders to produce signaling molecules at varying levels. These levels, which act as weights, are programmed by tuning the sender promoter strength Furthermore, a gradient descent based algorithm that enables weights optimization was developed. Weights were experimentally examined for recognizing 3 × 3-bit pattern.

Effectiveness of some biotic and abiotic agents to control tomato early blight disease caused by Alternaria solani

Bacillus megaterium, Bacillus subtilis, Pseudomonas fluorescens, Trichoderma harzianum, Trichoderma album and Trichoderma viride isolates, as well as different concentrations of chemical inducers, such as chitosan and salicylic acid were evaluated against Tomato early blight, under laboratory and greenhouse conditions. Under laboratory conditions, all tested bio agents, as well as the different concentrations of the tested chemical inducers, decreased the linear growth of A. solani, as the inhibition of fungal growth was increased by increasing the concentration of both chemical inducers in the growing medium. The chemical inducer chitosan at a concentration of 4.0 mg/mL PDA medium, caused the highest inhibition of linear growth of the fungus, followed by anti-fungus T. harzianum. When, different concentrations of chitosan and salicylic acid were tested on spore germination of A. solani, a concentration of 4.0 (mg/mL), for chitosan and a concentration of 25.0 (mM), for salicylic acid were the best concentrations used to reduce the spore germination of A. solani. Under greenhouse conditions, spraying tomato plants (cv. Super Strain B hybrid), with any of the tested biological agents as well as different concentrations of both the chemical inducers before infection of tomato plants by A. solani reduced the severity of early blight, in addition, to increase the fresh and dry weight of tomato plants. The chemical inducer chitosan at a concentration of 4.0 (mg/mL), caused the highest decrease of the disease severity, and the highest increase in fresh and dry weight of tomato plants, followed by the bio agent T. harzianum. The activities of defense-related enzymes i.e., polyphenoloxidase, peroxidase and chitinase were significantly increased in all treated plants with the tested biotic and abiotic agents. Chitosan at a concentration of 4.0 (mg/mL) resulted in the highest activity of oxidative enzymes, followed by T. harzianum. Meanwhile, the total content of phenols was higher in treated plants than in the untreated ones. The tested bio agents and chemical inducers might be playing an important role in management of tomato early blight through induction of induced systemic resistance.

Osteogenic Regulation by Graphene Nanoparticles in Human Mesenchymal Stem Cells

Background Degenerative bone diseases caused by trauma, cancer, or aging imposes a global health care burden every year. Treatment strategies include directing stem cell differentiation through osteogenesis, thereby creating new bone material for repairment. We previously identified that adult mesenchymal stem cells (MSCs) cultured on graphene nanoparticles spontaneously supports osteoblast development in vitro. However, the osteo-genetic signaling of MSCs in the presence of graphene is largely unknown. Therefore, this study sought to measure genetic expressions of MSCs cultured on graphene using focused PCR arrays. Methods MSCs were extracted from human adipose tissue (hAD-MSCs) and human bone marrow (hBM-MSCs) and expanded in growth media. For experimental conditions, hAD-MSCs and hBM-MSCs were seeded on a graphene surface and cultured for either 7 or 21 days. At each time point, cells were extracted for total RNA and reverse transcribed to cDNA before loading onto RT2 Profiler PCR Human Osteogenesis Array (Qiagen, Hilden, Germany). Gene expressions of both cell lines were compared either over time (day 7 as the control to day 21 as the treated group) or between both cell lines at each time point (hAD-MSCs to hBM-MSCs at day 7 or day 21). Results were analyzed using Qiagen Gene Globe software to determine the relative fold change in comparison to the control. Fold changes were considered statistically significant at p < 0.05. Results We examined expression of four transcriptional genes known to control stem cell fate: DLX5, RUNX2, SOX9, and SP7. In hAD-MSCs, RUNX2 was up-regulated while SOX9 and SP7 were down-regulated, suggesting RUNX2 to be a master regulator in hAD-MSCs. In contrast, all genes were down-regulated in hBM-MSCs. However, when hBM-MSCs were compared to hAD-MSCs, all genes were up-regulated at both time points. Interestingly, the fold changes at day 7 were more robust compared to that observed at day 21, suggesting transcriptional regulation of osteogenesis in hBM-MSCs occurs at an earlier time point. Secondly, we examined common markers of osteogenesis including ALPL, BGLAP, PHEX, and SPP1. In hAD-MSCs, ALPL, BGLAP, and PHEX were upregulated, while SPP1 was down-regulated.In hBM-MSCs, ALP, PHEX, and SPP1 were up-regulated, overall suggesting osteo-genetic signaling by graphene occurs in both cell types. Conclusion hAD-MSCs and hBM-MSCs cultured on graphene showed significant changes in osteogenic gene expressions. These cells were cultured in absence of any chemical inducers, suggesting graphene nanoparticles alone commits osteo-genetic signaling of MSCs. We also observed that osteogenesis of hBM-MSCs may occur at an earlier time point than hAD-MSCs. Lastly, we identified DLX5 as a novel osteogenic regulator supported by graphene. Overall, we show that MSCs isolated from two independent tissue sources spontaneously express osteogenic genes in the presence of graphene.

Thermoinducible expression system for producing recombinant proteins in Escherichia coli: advances and insights

Recombinant protein (RP) production from Escherichia coli has been extensively studied to find strategies for increasing product yields. The thermoinducible expression system is commonly employed at the industrial level to produce various RPs which avoids the addition of chemical inducers, thus minimizing contamination risks. Multiple aspects of the molecular origin and biotechnological uses of its regulatory elements (pL/pR promoters and cI857 thermolabile repressor) derived from bacteriophage λ provide knowledge to improve the bioprocesses using this system. Here, we discuss the main aspects of the potential use of the λpL/pR-cI857 thermoinducible system for RP production in E. coli, focusing on the approaches of investigations that have contributed to the advancement of this expression system. Metabolic and physiological changes that occur in the host cells caused by heat stress and by RP overproduction are also described. Therefore, the current scenario and the future applications of systems that use heat to induce RP production is discussed to understand the relationship between the activation of the bacterial heat shock response, RP accumulation, and its possible aggregation to form inclusion bodies.