Suspension Growth Research Articles

CRISPR-Cas9 screening identifies the role of FER as a tumor suppressor.

It is important to systematically identify tumor suppressor genes (TSGs) to improve our understanding of tumorigenesis and develop strategies for early diagnosis and mitigating disease progression. In the present study, we used an in vivo genome-wide clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein 9 (Cas9) screen and identified FPS/FES-related (FER) as a TSG. Single-cell RNA sequencing (scRNA-seq) revealed that normal cells with low FER expression exhibited elevated malignant transformation potential and stemness properties. FER knockout promoted the tumorigenic transformation, characterized by high colony-forming efficiency and suspension growth ability, acquired tumorigenicity in vivo, increased metabolic activity, dedifferentiation properties, and immune evasion. Moreover, analysis revealed that low FER expression tumors share molecular phenotypes with FER knockout cells, suggesting the consistent role of FER in tumor initiation and progression. Taken together, our findings not only provide insights into the essential role of FER as a tumor suppressor in tumor initiation and progression but also highlight its potential as a target for future clinical diagnosis. © 2024 The Pathological Society of Great Britain and Ireland.

Preparation and Biological Activity of Lignin-Silver Hybrid Nanoparticles.

Silver nanoparticles (AgNPs) are excellent antimicrobial agents and promising candidates for preventing or treating bacterial infections caused by antibiotic resistant strains. However, their increasing use in commercial products raises concerns about their environmental impact. In addition, traditional physicochemical approaches often involve harmful agents and excessive energy consumption, resulting in AgNPs with short-term colloidal stability and silver ion leaching. To address these issues, we designed stable hybrid lignin-silver nanoparticles (AgLigNPs) intended to effectively hit bacterial envelopes as a main antimicrobial target. The lignin nanoparticles (LigNPs), serving as a reducing and stabilizing agent for AgNPs, have a median size of 256 nm and a circularity of 0.985. These LigNPs were prepared using the dialysis solvent exchange method, producing spherical particles stable under alkaline conditions and featuring reducing groups oriented toward a wrinkled surface, facilitating AgNPs synthesis and attachment. Maximum accumulation of silver on the LigNP surface was observed at a mass reaction ratio mAg:mLig of 0.25, at pH 11. The AgLigNPs completely inhibited suspension growth and reduced biofilm development by 50% in three tested strains of Pseudomonas aeruginosa at a concentration of 80/9.5 (lignin/silver) mg L-1. Compared to unattached AgNPs, AgLigNPs required two to eight times lower silver concentrations to achieve complete inhibition. Additionally, our silver-containing nanosystems were effective against bacteria at safe concentrations in HEK-293 and HaCaT tissue cultures. Stability experiments revealed that the nanosystems tend to aggregate in media used for bacterial cell cultures but remain stable in media used for tissue cultures. In all tested media, the nanoparticles retained their integrity, and the presence of lignin facilitated the prevention of silver ions from leaching. Overall, our data demonstrate the suitability of AgLigNPs for further valorization in the biomedical sector.

Growing a single suspended perfect protein crystal in a fully noncontact manner

Nucleation is a fundamental process that determines the structure, morphology, and properties of crystalline materials, and is difficult to control because it is unpredictable. Here, we demonstrate a new method to control the protein crystal nucleation using a magnetic force, where we manipulate the movement and coalescence of nucleation precursors by adding paramagnetic salt into the crystallization solution to constrain the number and position of nucleation. We found that protein nucleation could be significantly affected by the magnetic force that the gradient magnetic fields generate. When the magnetization force is sufficiently enough, nucleation can be confined to the crystallization solution with no interface contact; therefore, only one crystal nucleus appears, which results in noncontact suspension growth of a single crystal in the crystallization solution system. Under these situations, the nucleation rate significantly decreases due to the coalescence of the dense liquid phase, and the crystal growth rate also decreases due to the suppression of convection, which increases the crystal quality. Our findings provide a new method for the noncontact control of crystal nucleation and demonstrate that externally applied physical environments can be used to affect the liquid-liquid phase separation process.

Analysis of the Plasticity of Circulating Tumor Cells Reveals Differentially Regulated Kinases During the Suspension-to-Adherent Transition.

Research on circulating tumor cells (CTCs) offers the opportunity to better understand the initial steps of blood-borne metastasis as main cause of cancer-related deaths. Here, we have used the colon cancer CTC-MCC-41 and breast cancer CTC-ITB-01 lines, which were both established from human CTCs as permanent cell lines as models to further study CTC biology with special emphasis on anchorage-independent survival and growth. Both cell lines showed a marked intrinsic plasticity to switch between suspension and adherent invitro growth, in 2D adherent culture conditions, and established an equilibrium of both growth patterns with predominant adherent cells in the CTC-MCC-41 line (77%) and suspension cells in the CTC-ITB-01 line (85%). Western blot analysis revealed a higher expression of pERK1/2 in CTC-ITB-01 adherent cells compared to the suspension counterpart that suggested the involvement of kinases in this process. Subsequent functional kinome profiling identified several serine/threonine as well as tyrosine kinases that were differentially regulated in adherent and suspension CTCs. In the adherent cells of the breast cancer line CTC-ITB-01 the activity of MSK1, Src family kinases and the PKG family was increased compared to the suspension counterpart. In adherent cells of the colorectal CTC-MCC-41 line, an increased activity of TYRO3 and JAK2 was detected, whereas p38 MAPK was strongly impaired in the suspension CTC-MCC-41 cells. Some of the regulated kinases, which include the Src family, TYRO3, MSK1, JAK2 and p38 MAPK, have been associated with crucial cellular processes including proliferation, migration and dormancy in the past. The investigated CTC lines exhibit a high plasticity, similar to the concept of 'adherent-to-suspension transition (AST)' that was recently suggested as a new hallmark of tumor biology by Huh etal. Moreover, we identified differentially regulated kinome profiles that may represent potential targets for future studies on therapeutic interventions.

High‐temperature oxidation and TGO growth behavior of Sr.9(Zr.9Yb.05Y.05)O2.85 thermal barrier coatings

AbstractHigh‐temperature oxidation (1050°C) of Sr.9(Zr.9Yb.05Y.05)O2.85 (SZYY) thermal barrier coatings (TBCs) by suspension plasma spraying (SPS) and growth behavior of thermally grown oxide (TGO) were investigated. When the TBCs were exposed to high temperature for a period of time (∼5 h), the BC oxidized and TGO inevitably formed between the bond coating (BC) and the ceramic top coating (TC). The high‐temperature oxidation behavior of the BC is generally manifested as the growth of TGO, which has four specific stages as follows: (1) formative oxidation stage (0‒10 h), (2) rapid oxidation stage (10‒50 h), (3) stable oxidation stage (50‒100 h), and (4) complex oxidation stage (100‒200 h). The main component of early TGO is α‐Al2O3. It has a very low oxygen ion diffusivity and provides an excellent diffusion barrier, which has a positive effect on preventing further BC oxidation. However, as the heat treatment time increased, the Al consumption and the formation of a CNS layer (NiO, Co3O4, and spinel) in the BC eventually led to coating failure. The working life of TBCs can be improved by improving the ceramic TC structure and the Al content of BC. SZYY‐TBCs have certain potential application value.

Differential Competitive Growth of Transgenic Subclones of Neuroblastoma Cells Expressing Different Levels of Cathepsin D Co-Cultured in 2D and 3D in Response to EGF: Implications in Tumor Heterogeneity and Metastasis.

Neuroblastoma (NB) is an embryonal tumor arising from the sympathetic central nervous system. The epidermal growth factor (EGF) plays a role in NB growth and metastatic behavior. Recently, we have demonstrated that cathepsin D (CD) contrasts EGF-induced NB cell growth in 2D by downregulating EGFR/MAPK signaling. Aggressive NB is highly metastatic to the bone and the brain. In the metastatic process, adherent cells detach to form clusters of suspended cells that adhere once they reach the metastatic site and form secondary colonies. Whether CD is involved in the survival of metastatic NB clones is not known. Therefore, in this study, we addressed how CD differentially affects cell growth in suspension versus the adherent condition. To mimic tumor heterogeneity, we co-cultured transgenic clones silenced for or overexpressing CD. We compared the growth kinetics of such mixed clones in 2D and 3D models in response to EGF, and we found that the Over CD clone had an advantage for growth in suspension, while the CD knocked-down clone was favored for the adherent growth in 2D. Interestingly, on switching from 3D to 2D culture conditions, the expression of E-cadherin and of N-cadherin increased in the KD-CD and Over CD clones, respectively. The fact that CD plays a dual role in cancer cell growth in 2D and 3D conditions indicates that during clonal evolution, subclones expressing different level of CD may arise, which confers survival and growth advantages depending on the metastatic step. By searching the TCGA database, we found up to 38 miRNAs capable of downregulating CD. Interestingly, these miRNAs are associated with biological processes controlling cell adhesion and cell migration. The present findings support the view that during NB growth on a substrate or when spreading as floating neurospheres, CD expression is epigenetically modulated to confer survival advantage. Thus, epigenetic targeting of CD could represent an additional strategy to prevent NB metastases.

Synthetic auxin herbicide 2,4-D and its influence on a model BY-2 suspension.

2,4-D is a broadly used auxin herbicide. The presence of the 2,4-D synthetic auxin in the medium is imperative for long-term BY-2 tobacco suspension viability. The precise mechanism of this symbiosis of the suspension and the synthetic auxin remains unclear. Our goal was to study the hormonal regulation of the growth of the cell suspension; and to describe the experiments clarifying the interaction between the chosen growth regulators and phytohormones on the cellular level, specifically between the 2,4-D synthetic auxin and the native stress phytohormone - ethylene. This study examined the influence of low 2,4-D concentrations stimulating cell growth in vitro as well as the influence of high herbicide concentrations on the model tobacco BY-2 suspension. The culture took 6 days. Different parameters were evaluated, including the influence of different 2,4-D concentrations on the production of the phytohormone ethylene and its precursor 1-Aminocyclopropane-1-carboxylic acid (ACC) in the tobacco cells. The content of 2,4-D in the cells and the medium was established. The observations of the morphological changes showed that a heavy impregnation of the cell walls taking place depending on the concentration of 2,4-D. A dramatic increase in protective polysaccharides and a remodulation of the cell walls by the formation of a pectin shield in artificial conditions were expected and observed. At the same time, massive production of the stress phytohormone ethylene took place, and, because of that, plant mutagenicity, anomalous tumour-type proliferation growth, and the production of supercells were observed. The hypothesis of the protective shield is discussed.

Biofilm and suspension-based cultivation of microalgae to treat anaerobic digestate food effluent (ADFE)

Anaerobic digestion of organic waste produces effluent (ADE) that requires further treatment. Biofilm-based microalgal cultivation is a favoured approach to ADE treatment. This study compared Chlorella sp. MUR 268 and Scenedesmus sp. MUR 269 in biofilm and suspension cultures to treat anaerobic digestate food effluent (ADFE). Chlorella sp. MUR 268 biofilm had significantly higher biomass (50.38 g m−2) than Scenedesmus sp. biofilm (9.39 g m−2). Conversely, Scenedesmus sp. yielded 1.5 times more biomass (1.2 g L−1) than Chlorella sp. in suspension. Chlorella sp. biofilm had 49.3 % higher areal productivity than suspension, while Scenedesmus sp. showed 87.3 % higher areal growth in suspension. Chlorella sp. MUR 268 and Scenedesmus sp. MUR 269 significantly removed nutrients in ADFE. In suspension, COD, ammoniacal nitrogen, and phosphate were reduced to 94.9, 5.2, and 5.98 mg L−1 for Chlorella sp. MUR 268, and 245, 2.89, and 3.22 mg L−1 for Scenedesmus sp. MUR 269, respectively. In biofilm, Chlorella sp. MUR 268 achieved reductions to 149.9, 1.16, and 3.57 mg L−1, while Scenedesmus sp. MUR 269 achieved 100.2, 6.9 and 2.07 mg L−1. Most of these values are below the recommended effluent discharge standard, highlighting the efficacy of this system in ADFE treatment. Biofilm cultures fixed 68–81 % of removed nitrogen in biomass, while in suspension, only 55–71 % ended in the biomass. Chlorella sp. MUR 268 biofilm fixed 88 % of removed phosphorus, while Scenedesmus sp. MUR 269 suspension fixed more phosphorus (55 %) than the biofilm counterpart (34 %). This biofilm design offers advantages like simplified, cost-effective operation, easy biomass recovery, and reduced water usage.

Effects of destruction of Euglena gracilis by ultrasonic cavitation

Euglena gracilis has attracted attention because it contains the polysaccharide paramylon. In this study, we aimed to destruct E. gracilis by applying ultrasonic cavitation and to elucidate the mechanism. We also examined the breakdown of paramylon particles and attempted to extract paramylon nanofibers. It was suggested that the damage caused by ultrasonic waves was frequency dependent and influenced by the size ratio of the cell to cavitation bubbles, yield strength, and inhibition of cavitation bubble growth in suspension. It is also assumed that the cell destruction rate decreased because it was also dependent on the initial cell density, and an increase in the initial cell density resulted in a decrease in acoustic pressure. The fracture strength of the paramylon particles was much greater than the microjet stress at the acoustic power used in this study, and the paramylon particles did not fracture.

Establishment of feijoa (Acca sellowiana) callus and cell suspension cultures and identification of arctigenin - a high value bioactive compound.

Feijoa (Acca sellowiana (O. Berg.) Burret), also known as pineapple guava, is a member of the Myrtaceae family and is well known for its fruit. Chemical profiling of the different tissues of the feijoa plant has shown that they generate an array of useful bioactive compounds which have health benefits such as significant antioxidant activities. In this study, an in vitro culture system has been developed, which could be explored to extract high-value bioactive compounds from feijoa. Feijoa tissue culture was initiated by the induction of callus from floral buds. Sections of floral buds were plated on MS medium supplemented with 2,4-D and BAP at 2.0mg/L and 0.2mg/L concentrations, respectively. Cell suspension cultures of feijoa were established using a liquid MS medium with different concentrations of 2,4-D and BAP and cultured on a rotary shaker. The growth of the cell suspension was evaluated with different parameters such as different carbohydrate sources, concentration of MS media, and inoculum density. When the cell suspensions were treated with different concentrations of MeJA at different time points, phytochemicals UPLC - QTOF MS analysis identified extractables of interest. The main compounds identified were secondary metabolites (flavonoids and flavonoid-glucosides) and plant hormones. These compounds are of interest for their potential use in therapeutics or skin and personal care products. This report investigates essential methodology parameters for establishing cell suspension cultures from feijoa floral buds, which could be used to generate in vitro biomass to produce high-value bioactive compounds. This is the first study reporting the identification of arctigenin from feijoa, a high-value compound whose pharmaceutical properties, including anti-tumour, anti-inflammatory and anti-colitis effects, have been widely reported. The ability of feijoa cell cultures to produce such high-value bioactive compounds is extremely promising for its use in pharmaceuticals, cosmeceuticals and nutraceuticals applications.

Effect of Hyaluronic Acid on the Self-Assembly of a Dipeptide-Based Supramolecular Gel.

The combination of polymers and low molecular weight (LMW) compounds is a powerful approach to prepare new supramolecular materials. Here we prepare two-component hydrogels made by a well-known and biologically active polymer, hyaluronic acid (HA), and a dipeptide-based supramolecular gelator. We undertake a detailed study of materials with different compositions including macroscopic (hydrogel formation, rheology) and micro/nanoscopic characterization (electron microscopy, X-ray powder diffraction). We observe that the two components mutually benefit in the new materials: a minimum amount of HA helps to reduce the polymorphism of the LMW network leading to reproducible hydrogels with improved mechanical properties; the LMW component network holds HA without the need for an irreversible covalent crosslinking. These materials have a great potential for biomedical application as, for instance, extracellular matrix mimetics for cell growth. As a proof of concept, we have observed that this material is effective for cell growth in suspension and avoids cell sedimentation even in the presence of competing cell-adhesive surfaces. This may be of interest to advanced cell delivery techniques.

Seawater biodesalination treatment using Phormidium keutzingianum in attached growth-packed bed continuous flow stirred tank reactor

Water scarcity is increasing worldwide due to rising population which is creating opportunities to unlock alternative green desalination techniques for seawater, such as biodesalination. Therefore, this study presents the utilization of the Phormidium keutzingianum strain in an attached growth-packed bed reactor to treat seawater in real-time in a continuous-flow stirred tank reactor for biodesalination. Two reactors were designed and developed, in which zeolites were used as the support media for the attached growth. The experiment was conducted in an open outdoor environment with a continuous air flow rate of 3 mL/min and two hydraulic retention times (HRT) of 7 and 15 d. Parameters such as the pH, chloride ion concentration, total organic carbon (TOC), and optical density were monitored regularly. The pH change was not significant in either reactor and remained within the range of 7.25–8.0. Chloride ion removal was the most crucial component of biodesalination efficiency, with d 7 removal efficiencies of approximately 40% and 32% for reactors 1 and 2, respectively. Reactor 1 exhibited a TOC reduction of 36% within the first 10 d at a HRT of 7, and when the HRT was set to 15 d, a TOC removal efficiency of 89% was achieved on d 53. For reactor 2, a TOC removal efficiency of approximately 81% was achieved on d 11 at HRT 7, and it reduced to less than 50% at an HRT of 15. The chloride ion and TOC removal phenomena were similar in both reactors. The optical density (OD) showed low measurement recordings, ranging from 0.005 to 0.01, indicating low cell detachment in the seawater effluent. Therefore, using the attached growth method for the biodesalination of seawater is feasible. Furthermore, biomass harvesting in attached growth systems is easier than that in suspension growth systems.

Interferon alpha and beta receptor 1 knockout in human embryonic kidney 293 cells enhances the production efficiency of proteins or adenoviral vectors related to type I interferons.

Human embryonic kidney (HEK) 293 cells are widely used in protein and viral vector production owing to their high transfection efficiency, rapid growth, and suspension growth capability. Given their antiviral, anticancer, and immune-enhancing effects, type I interferons (IFNs) have been used to prevent and treat human and animal diseases. However, the binding of type I IFNs to the IFN-α and-β receptor (IFNAR) stimulates the expression of IFN-stimulated genes (ISGs). This phenomenon induces an antiviral state and promotes apoptosis in cells, thereby impeding protein or viral vector production. In this study, we generated an IFNAR subtype 1 knockout (KO) HEK 293 suspension (IFNAR-KO) cell line by using clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein-9 nuclease (Cas9) technology. Upon treatment with human IFN-α, the IFNAR-KO cells showed a constant expression of ISGs, including 2'-5'-oligoadenylate synthetase 1 (OAS1), myxovirus resistance 1 (Mx1), protein kinase RNA-activated (PKR), and IFN-induced transmembrane protein 1 (IFITM1), when compared with the wild-type HEK 293 (WT) cells, wherein the ISGs were significantly upregulated. As a result, the titer of recombinant adenovirus expressing porcine IFN-α was significantly higher in the IFNAR-KO cells than in the WT cells. Furthermore, the IFNAR-KO cells continuously produced higher amounts of IFN-α protein than the WT cells. Thus, the CRISPR-Cas9-mediated IFNAR1 KO cell line can improve the production efficiency of proteins or viral vectors related to IFNs. The novel cell line may be used for producing vaccines and elucidating the type I IFN signaling pathway in cells.

Titanium biogenic nanoparticles to help the growth of Trichoderma harzianum to be used in biological control

BackgroundThe biogenic synthesis of metallic nanoparticles is a green alternative that reduces the toxicity of this nanomaterials and may enable a synergy between the metallic core and the biomolecules employed in the process enhancing biological activity. The aim of this study was to synthesize biogenic titanium nanoparticles using the filtrate of the fungus Trichoderma harzianum as a stabilizing agent, to obtain a potential biological activity against phytopathogens and mainly stimulate the growth of T. harzianum, enhancing its efficacy for biological control.ResultsThe synthesis was successful and reproductive structures remained in the suspension, showing faster and larger mycelial growth compared to commercial T. harzianum and filtrate. The nanoparticles with residual T. harzianum growth showed inhibitory potential against Sclerotinia sclerotiorum mycelial growth and the formation of new resistant structures. A great chitinolytic activity of the nanoparticles was observed in comparison with T. harzianum. In regard to toxicity evaluation, an absence of cytotoxicity and a protective effect of the nanoparticles was observed through MTT and Trypan blue assay. No genotoxicity was observed on V79-4 and 3T3 cell lines while HaCat showed higher sensitivity. Microorganisms of agricultural importance were not affected by the exposure to the nanoparticles, however a decrease in the number of nitrogen cycling bacteria was observed. In regard to phytotoxicity, the nanoparticles did not cause morphological and biochemical changes on soybean plants.ConclusionThe production of biogenic nanoparticles was an essential factor in stimulating or maintaining structures that are important for biological control, showing that this may be an essential strategy to stimulate the growth of biocontrol organisms to promote more sustainable agriculture.Graphical

Improving biomass and carbohydrate production of microalgae in the rotating cultivation system on natural carriers

Biofilm-based algal technologies have gained popularity due to higher biomass productivity, efficient harvesting, and water-saving over suspended growth systems. A rotating attached system was designed to assess the biofilm-forming capacity of different isolated microalgal strains from the Persian Gulf. Four microalgal strains, including two Chlorella sp., one Picochlorum sp. and one filamentous cyanobacterium Desmonostoc sp. were cultivated on four carriers: jute, cotton, yarn and nylon. The carriers’ physicochemical surface characteristics and attachment effects, like contact angle, were investigated. The incorporated biomass and exopolysaccharides (EPS) content in the suspended and biofilm system was calculated and compared. The results showed that the cyanobacterium strain had the biofilm formation capability on both jute and cotton in the attached cultivation system. Under the same culture conditions, the biomass productivity on jute and cotton carriers was significantly higher (4.76 and 3.61 g m− 2 respectively) than the growth in aqueous suspension (1.19 g m− 2 d− 1). The greatest incorporated exopolysaccharides amount was observed on jute (43.62 ± 4.47%) and the lowest amount was obtained from the growth on positive charge yarn (18.62 ± 1.88%). This study showed that in comparison with planktonic growth, the colonization of cyanobacterial cells and subsequent production of extracellular matrix and biofilm formation can lead to increased biomass production.

Abstract 6149: STAT3 activation promotes invasiveness and cancer stemness in lung cancer

Abstract Lung cancer is the leading cause of cancer-related death worldwide, featuring a high incidence and low 5-year survival rate. Nearly 40% of patients are diagnosed with metastatic disease when they are first presented to the clinic. However, drug intervention targeting established metastatic tumors remains challenging, a better understanding of therapeutically exploitable mechanisms is urgently needed. This present study aims to explore novel therapeutic intervention against metastatic lung cancer. To understand the tumor-cell-intrinsic pathway that drives metastatic progression, we generated a highly invasive subline of lung cancer cell - A549 (ATCC, USA) by serial selections for Matrigel-coated transwells. The invasive subline A549-i3 shows about 5 times higher invasion potential than the parental cell line A549-Par (p<0.05, Student t-test). Colony-formation Assay and suspension growth in ultra-low attachment plates were used to measure the 2D proliferation ability and anoikis resistance, albeit with no difference. The differential efficacies between A549-Par and A549-i3 against multiple pathway inhibitors were screened using MTT assay in vitro. Notably, A549-i3 is more sensitive towards napabucasin (STAT3 inhibitor) and tipifarnib (FTase inhibitor) when compared with A549-Par, suggesting their potency in targeting cancer stemness and addicted oncogenic driver. Furthermore, napabucasin reduced the migration and invasion abilities of A549-i3 to similar extent as their parent counterpart. Overall, our preliminary data identified STAT3 as a potential therapeutic vulnerability in highly invasive cancer cells. Future work will be focused on identifying pathways responsible to sensitize highly invasive NSCLC cells toward STAT3 inhibition using multiomics approach. This project was supported by the Research Impact Fund (Ref. No.: R4015-19) from the Research Grants Council in Hong Kong. Citation Format: Sai Fung Yeung, Sze Man Chan, Chun Ho Law, Hoi Yin Chan, Kwok Wing Tsui. STAT3 activation promotes invasiveness and cancer stemness in lung cancer. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6149.

Novel active edible food packaging films based entirely on citrus peel wastes

This is the first report on mobilizing three value-added compounds from citrus peel wastes to fabricate novel/cost-effective active edible films with multiple functionalities. Pectin extracted from grapefruit peels (GFP) with improved recovery yields (48.35%), displayed better thermal stability and superior physico-chemical properties to commercial citrus pectin ( CPec ) with regard to methoxylation degree, anhydrouronic acid and moisture contents. Active films plasticized with PEG 400 (25% w/w of polymer) were prepared by solution casting upon incorporating 0.04% (w/v) of grapefruit peel methanolic extract (GFPE) and maltodextrin-encapsulated lemon peel extract (MD-LPE) into grapefruit pectin ( GFPec ; 3% w/v) matrix. The bioactive ingredients and the optimally active ( GFPec -GFPE/MD-LPE) film-forming solutions exhibited strong radical scavenging and antimicrobial activities against foodborne pathogens. SEM micrographs of the optimal active film revealed good dispersion and compatibility of ingredients in agreement with the FTIR data that confirmed no modifications in the film's functional groups. GFPE and MD-LPE capsules synergistically enhanced the tensile strength, thermal, water vapor/UV barrier properties and soil biodegradability of the elaborated transparent flexible films compared to CPec -based films. Wrapping of artificially contaminated cherry tomatoes with optimal active films inhibited the growth of a four-strain cocktail suspension of E. coli O157:H7 by ∼1.6 log units after 6 days of chilled storage. Overall, the study highlights the benefits of citrus wastes valorization, provides evidence of potential applicability of the fabricated novel edible GFPec -based films as candidates for antimicrobial food packaging, shelf life extension of fresh-cut produce, and supports their consideration as promising eco-friendly alternatives to synthetic food packaging materials. • Three value-added compounds were mobilized from citrus peels for film fabrication. • High methoxyl grapefruit pectin showed superiority to commercial citrus pectin. • Free and encapsulated extracts had remarkable antioxidant and cytotoxic activities. • Incorporating both ingredients enhanced thermal and barrier properties of films. • The novel active film inhibited E. coli O157:H7 growth on wrapped cherry tomatoes.

INFLUENCE OF DIFFERENT PREPARATIONS ON SOWING QUALITY OF SPRING WHEAT SEEDS

The article is devoted to the study of the effect of pre-sowing treatment with various preparations on germination, growth and development of spring wheat. Spring wheat seeds were pre-soaked for 24 hours in Biohumus liquid organomineral fertilizer and Chlorella Suspension growth stimulator, presowing treatment was also carried out with agromineral fertilizers - zeolite from the Tatarsko-Shatrashanskiy deposit and tripoli from Kaluga region. The experiments were conducted in laboratory conditions in vessels and rolls. An increase of germination energy and seed germination was noted in rolls, when seeds were soaked in biohumus and treated with zeolite - 45 and 22% higher than in the control. The maximum values of plant height were observed in the variant with zeolite - 9% higher than the control. The number of roots increased by 22%, their length in the variant with biohumus was, on average, 20% more than the control values. The coleoptile length was at the same level in all variants. The maximum germination in vessels was obtained in the variant with soaking seeds in chlorella suspension - 92%, which is 9.52% more than the control variant. The maximum plant height was observed in the variant with biohumus - 7% more than the control, the largest root length - in the variant with seed treatment with tripoli - 6% more than the control, and the largest number of roots in the variant with chlorella suspension and zeolite - 9% more than in the control variant.

Novel transfer learning schemes based on Siamese networks and synthetic data

Transfer learning schemes based on deep networks which have been trained on huge image corpora offer state-of-the-art technologies in computer vision. Here, supervised and semi-supervised approaches constitute efficient technologies which work well with comparably small data sets. Yet, such applications are currently restricted to application domains where suitable deep network models are readily available. In this contribution, we address an important application area in the domain of biotechnology, the automatic analysis of CHO-K1 suspension growth in microfluidic single-cell cultivation, where data characteristics are very dissimilar to existing domains and trained deep networks cannot easily be adapted by classical transfer learning. We propose a novel transfer learning scheme which expands a recently introduced Twin-VAE architecture, which is trained on realistic and synthetic data, and we modify its specialized training procedure to the transfer learning domain. In the specific domain, often only few to no labels exist and annotations are costly. We investigate a novel transfer learning strategy, which incorporates a simultaneous retraining on natural and synthetic data using an invariant shared representation as well as suitable target variables, while it learns to handle unseen data from a different microscopy technology. We show the superiority of the variation of our Twin-VAE architecture over the state-of-the-art transfer learning methodology in image processing as well as classical image processing technologies, which persists, even with strongly shortened training times and leads to satisfactory results in this domain. The source code is available at https://github.com/dstallmann/transfer_learning_twinvae, works cross-platform, is open-source and free (MIT licensed) software. We make the data sets available at https://pub.uni-bielefeld.de/record/2960030.

Breast Cancer MCF-7 Cells Acquire Heterogeneity during Successive Co-Culture with Hematopoietic and Bone Marrow-Derived Mesenchymal Stem/Stromal Cells.

During disease progression and bone metastasis, breast tumor cells interact with various types of bystander cells residing in the tumor microenvironment. Such interactions prompt tumor cell heterogeneity. We used successive co-culture as an experimental model to examine cancer-bystander cell interaction. RMCF7-2, a clone of the human breast cancer MCF-7 cells tagged with a red fluorescent protein, was tracked for morphologic, behavioral, and gene expression changes. Co-cultured with various types of hematopoietic cells, RMCF7-2 adopted stable changes to a rounded shape in suspension growth of red fluorescent cells, from which derivative clones displayed marked expressional changes of marker proteins, including reduced E-cadherin and estrogen receptor α, and loss of progesterone receptor. In a successive co-culture with bone marrow-derived mesenchymal stem/stromal cells, the red fluorescent clones in suspension growth changed once more, adopting an attachment growth, but in diversified shapes. Red fluorescent clones recovered from the second-round co-culture were heterogeneous in morphology, but retained the altered marker protein expression while displaying increased proliferation, migration, and xenograft tumor formation. Interaction with bystander cells caused permanent morphologic, growth behavioral, and gene expressional changes under successive co-culture, which is a powerful model for studying cancer cell heterogeneity during breast cancer progression and metastasis.