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Introduction: Microcystis aeruginosa (M. aeruginosa), one of the most prevalent blue-green algae in aquatic environments, produces microcystin by causing harmful algal blooms (HAB). This study investigated the combined effects of nutrients and cyanobacterial subpopulation competition on synthesizing microcystin-LR. Method: In varied nitrogen and phosphorus concentrations, cyanobacterial coculture, and algicidal DCMU presence, the growth was monitored by optical density analysis or microscopic counting, and the microcystin production was analyzed using high-performance liquid chromatography-UV. Furthermore, growth and toxin production were predicted using MATLAB. Results and discussion: First, coculturing with a fast-growing cyanobacterium Synechococcus elongatus UTEX 2973 (S. elongatus) reduced M. aeruginosa biomass and microcystin production at 30oC. Under high nitrogen and low phosphorus conditions, S. elongatus is mostly effective, with up to 94.7% and 92.4% limitation of M. aeruginosa growth and toxin synthesis, respectively. Second, this biological strategy became less effective at 23oC, where S. elongatus grew slower. Third, photosynthesis inhibitor DCMU (3-(3,4-dichlorophenyl)-1,1-dimethylurea) hindered M. aeruginosa growth (at 0.1 mg/L) and microcystin production (at 0.02 mg/L). DCMU was also effective for controlling microcystin production in S. elongatus – M. aeruginosa cocultures. Based on experimental results, a multi-substrate, multi-species kinetic model was built to describe coculture growth and population interactions. Conclusion: Future research should examine more complex models to further develop and refine to facilitate the derivation of more effective recommendations for health prevention programs, particularly for mothers and girls. [ABSTRACT FROM AUTHOR]

This research pivots around screening of idoneous lactic acid bacteria (LAB) from cow milk and subjecting them to adaptive evolution experiments to aid superior growth/robustness necessary for 3-phenyllactic acid (3-PLA) production. Conventional and statistical fermentation studies were conducted at batch scale using a non-axenic coculture of three novel LAB strains: Lactiplantibacillus plantarum str. nov. plantharim, Lactobacillus delbrueckki str. nov. delharim, and Pediococcus pentasaceous str. nov. pentharim. Statistically optimized fermentation using Box Behnken technique resulted in 1225 mg/L 3-PLA production using the growth medium: cheese whey—MRS medium mixture (5:2 ratio), phenylalanine (2.69% w/v), and glucose (9.6% w/v). Statistical optimization of fermentation parameters resulted in a substantial increase (17 times higher) compared to the non-optimized fermentation conditions (72 mg/L). Monad growth kinetics of the cow milk whey (CMW) coculture were calculated and estimated as: μmax = 0.336 h−1, Ks = 11.64 mg/mL, Yx/s = 0.835 mg/g, YP/S = 1.66 mg/g, YX/P = 0.112 mg/mg. The purified 3-PLA (1.93 mg/mL) showed antimicrobial activity with pathogenic bacteria like Pseudomonas aeruginosa, Escherichia coli, and Staphylococcus aureus, with a minimum inhibitory concentration of 12 mg/mL. [ABSTRACT FROM AUTHOR]

The influence of the initial pH on the morphology and secondary metabolite production in cocultures and axenic cultures of Aspergillus terreus and Streptomyces rimosus was investigated. The detected secondary metabolites (6 of bacterial and 4 of fungal origin) were not found in the cultures initiated at pH values less than or equal to 4.0. The highest mean levels of oxytetracycline were recorded in S. rimosus axenic culture at pH 5.0. Initiating the axenic culture at pH 5.9 led to visibly lower product levels, yet the presence of A. terreus reduced the negative effect of non-optimal pH and led to higher oxytetracycline titer than in the corresponding S. rimosus axenic culture. The cocultivation initiated at pH 5.0 or 5.9 triggered the formation of oxidized rimocidin. The products of A. terreus were absent in the cocultures. At pH 4.0, the striking morphological differences between the coculture and the axenic cultures were recorded. [ABSTRACT FROM AUTHOR]

Pararamosis, also known as Pararama-associated phalangeal periarthritis, is a neglected tropical disease primarily affecting rubber tappers in the Amazon region. It is caused by contact with the urticating hairs of the Premolis semirufa moth caterpillar, which resides in rubber plantations. The condition is marked by the thickening of the articular synovial membrane and cartilage impairment, features associated with chronic synovitis. Given the significance of synovial inflammation in osteoarticular diseases, in this study, the role of synoviocytes and their interactions with macrophages and chondrocytes are examined when stimulated by Pararama toxins. Synoviocytes and macrophages treated with Pararama hair extract showed an increased production of cytokines IL-6, IL-1β, and TNF-α, indicating a direct effect on these cells. In cocultures, there was a significant rise in inflammation, with levels of IL-1β, IL-6, and chemokines CCL2, CCL5, and CXCL8 increasing up to seven times compared to monocultures. Additionally, matrix-degrading enzymes MMP-1 and MMP-3 were significantly elevated in cocultures. Chondrocytes exposed to the extract also produced IL-6, CCL2, and CCL5, and in cocultures with synoviocytes, there was a notable increase in IL-6, CCL5, and CXCL8, as well as a doubling of MMP-1 and MMP-3 levels. These findings underscore the critical role of cell crosstalk in the inflammatory and catabolic processes associated with pararamosis and demonstrate how Pararama hair extract can influence factors affecting cartilage health, providing valuable insights into this condition. [ABSTRACT FROM AUTHOR]

The ability of microorganisms to decompose brown algae has attracted attention. This study aims to clarify the characteristics of marine microbial communities in which prokaryotic and eukaryotic microorganisms interact via the metabolism of brown algae carbohydrates. Amplicon-based microbiome analysis revealed the predominance of the genera Marinomonas and Vibrio in seawater and seaweed samples mixed with alginate and mannitol, which are the primary carbohydrates in brown algae. Three Vibrio species and Candida intermedia were isolated via alginate enrichment culture. Although C. intermedia did not utilize alginate as a nutrient source, the yeast grew in the spent alginate medium in which Vibrio algivorus had been cultured. Coculture with C. intermedia and the Vibrio isolates, especially V. algivorus, also enhanced the growth of the yeast on alginate. These results suggested that C. intermedia grew because of the supply of nutrients via alginate metabolism by Vibrio species. In the coculture medium, the amount of phosphatidylserine increased in the early phase but decreased with the growth of C. intermedia, indicating that phosphatidylserine secreted by Vibrio is involved in the putative mutualistic interaction. We examined whether such interaction is applicable to the production of useful substances and succeeded in lipid production by oleaginous marine yeast Yarrowia lipolytica through coculture with V. algivorus. Our study suggested the potential of mutualistic interaction via degradation of alginate by marine Vibrio for production of industrially useful substances in yeast cells. [ABSTRACT FROM AUTHOR]

Background: Typical difficulties encountered during in vitro fertilization (IVF) to produce embryos in pigs include poor pronucleus formation and poor-quality fertilized embryos because of high polysperm invasion. In this study, we evaluated the effects of supplementation with apple seed extract (ASE) and coculture systems on porcine in vitro-fertilized embryo culture. Methods: Slaughterhouse-derived ovaries were used to obtain cumulus-oocyte complexes (COCs). COCs were conventionally used to perform IVF. We examined the differences in apoptosis and metabolism during development following addition of ASE to normal culture and coculture systems. Matrix metalloproteinases (MMPs), cell development-related factors, and apoptotic proteins were compared in porcine embryos produced under different conditions. Results: The expression of genes related to insulin-like growth factor (IGF) signaling was increased in the coculture system. In the ASE group, early apoptosis and necrosis were reduced in fertilized embryos and the late survival rate increased. Supplementation of the coculture system with ASE led to increased expression of BCL-2 and decreased expression of Casp-3 in the cytoplasm, thereby lowering the apoptosis rate and inducing MMP expression. In addition, compared with the extract-supplemented group in normal culture, the activity of MMP-2 decreased in the coculture system supplemented with ASE, activity of MMP-9 increased, and the expression of dynactin p62 and BrdU in the cytoplasm was higher than that in the other groups. Conclusions: The coculture system increased the activity of the embryonic cytoplasm compared with the non-coculture system. Supplementation with ASE may induce cell activity and inhibit the expression of apoptotic factors.

The two-species microbial cocultures are effective in terms of awakening the cryptic biosynthetic pathways. They may also lead to the improved production of previously discovered molecules. Importantly, only a few outcomes of the cocultures may prove desirable, namely those leading to the formation of useful secondary metabolites. To address this issue, a method allowing for the evaluation of the final outcome of the co-culture process and fine-tune the cocultivation strategy was proposed. The systematic approach was supported by the experimental data from the bioreactor runs with the participation of Aspergillus terreus and Penicillium rubens confronted with Streptomyces rimosus and Streptomyces noursei. Kinetic, morphological and metabolic aspects of dominance were analysed via the newly proposed formula describing the dominance pattern. The suggested method involved the determination of the numerical value representing the dominance level. When it was high (value 1) no useful metabolites were formed apart from those originating from the winning counterpart. But either for the partial dominances or when the winning organism changed within the run or when the competition ended in draw, the number of the secondary metabolites of interest in the broth was the highest. Next, the systematic approach illustrated how the delayed inoculation strategy influenced the level of dominance leading to the change of winning counterpart and the set of metabolites produced. The proposed systematic approach allows for the reliable determination of the level of dominance in the two-species cocultures to seek for the potentially useful substances for future applications. [ABSTRACT FROM AUTHOR]

Aims: Tumorigenicity is a significant concern in stem cell‐based therapies. However, traditional tumorigenicity tests using animal models often produce inaccurate results. Consequently, a more sensitive method for assessing tumorigenicity is required. This study aimed to enhance sensitivity by exposing functional progenitors derived from human pluripotent stem cells (hPSCs) to the tumor microenvironment (TME) in vitro before transplantation, potentially making them more prone to abnormal proliferation or tumorigenicity. Methods: Midbrain dopamine (mDA) cells derived from hPSCs were exposed to the TME by coculturing with medulloblastoma. The cellular characteristics of these cocultured mDA cells were evaluated both in vitro and in vivo, and the mechanisms underlying the observed alterations were investigated. Results: Our findings demonstrated increased proliferation of cocultured mDA cells both in vitro and in vivo. Moreover, these proliferating cells showed a higher expression of Ki67 and SOX1, suggesting abnormal proliferation. The observed abnormal proliferation in cocultured mDA cells was attributed to the hyperactivation of proliferation‐related genes, the JAK/STAT3 pathway, and cytokine stimulation. Conclusion: This study indicates that exposing functional progenitors to the TME in vitro before transplantation can induce abnormal proliferation, thereby increasing the sensitivity of tumorigenicity tests. [ABSTRACT FROM AUTHOR]

Fungi generate a diverse array of bioactive compounds with significant pharmaceutical applications. However, the chemical diversity of natural products in fungi remains largely unexplored. Here, we present a paradigm for specifically discovering diverse and bioactive compounds from fungi by integrating genome mining with building block molecular network and coculture analysis. Through pangenome and sequence similarity network analysis, we identified a rare type I polyketide enzyme from Penicillium sp. ZJUT-34. Subsequent building block molecular network and coculture strategy led to the identification and isolation of a pair of novel polyketides, (±)-peniphenone E [(±)−1], three known polyketides (2–4), and three precursor compounds (5–7) from a combined culture of Penicillium sp. ZJUT-34 and Penicillium sp. ZJUT23. Their structures were established through extensive spectroscopic analysis, including NMR and HRESIMS. Chiral HPLC separation of compound 1 yielded a pair of enantiomers (+)−1 and (−)−1, with their absolute configurations determined using calculated ECD methods. Compound (±)−1 is notable for its unprecedented structure, featuring a unique 2-methyl-hexenyl-3-one moiety fused with a polyketide clavatol core. We proposed a hypothetical biosynthetic pathway for (±)−1. Furthermore, compounds 2, 5, and 6 exhibited strong antioxidant activity, whereas (−)−1, (+)−1, 3, and four exhibited moderate antioxidant activity compared to the positive control, ascorbic acid. Our research demonstrates a pioneering strategy for uncovering novel polyketides by merging genome mining, metabolomics, and cocultivation methods. This approach addresses the challenge of discovering natural compounds produced by rare biosynthetic enzymes that are often silent under conventional conditions due to gene regulation. IMPORTANCE Polyketides, particularly those with complex structures, are crucial in drug development and synthesis. This study introduces a novel approach to discover new polyketides by integrating genomics, metabolomics, and cocultivation strategies. By combining genome mining, building block molecular networks, and coculturing techniques, we identified and isolated a unique polyketide, (±)-peniphenone E, along with three known polyketides and three precursor compounds from Penicillium sp. ZJUT-34 and Penicillium sp. ZJUT23. This approach highlights the potential of using combined strategies to explore fungal chemical diversity and discover novel bioactive compounds. The successful identification of (±)-peniphenone E, with its distinctive structure, demonstrates the effectiveness of this integrated method in enhancing natural product discovery and underscores the value of innovative approaches in natural product research. [ABSTRACT FROM AUTHOR]

In this study, the interaction of pure Mg and WE43 alloy under the presence of osteoblast (OB) and osteoclast (OC) cells and their influence on the degradation of materials have been deeply analyzed. Since OB and OC interaction has an important role in bone remodeling, we examined the surface morphology and dynamic changes in the chemical composition and thickness of the corrosion layers formed on pure Mg and WE43 alloy by direct monoculture and coculture of pre-differentiated OB and OC cells in vitro. Electrochemical techniques examined the corrosion performance. The corrosion products were characterized using a combination of the focused ion beam (FIB), scanning electron microscopy (SEM), and energy-dispersive X-ray spectroscopy (EDX). Cell viability and morphology were assessed by fluorescent microscopy and SEM. Our findings demonstrate cell spread and attachment variations, which differ depending on the Mg substrates. It was clearly shown that cell culture groups delayed degradation processes with the lowest corrosion rate observed in the presence of OBOC coculture for the WE43 substrate. Ca-P enrichment was observed in the outer-middle region of the corrosion layer but only after 7 days of OBOC coculture on WE43 and after 14 days on the pure Mg specimens. Magnesium metallic materials that can degrade over time provide distinct opportunities for orthopedic application. However, there is still a lack, especially in elucidating cell-material interface characterization. This study investigated the influence of osteoblast-osteoclast coculture in direct Mg-material contact. Our findings demonstrated that pre-differentiated osteoblasts and osteoclasts cocultured on Mg substrates influenced the chemistry of the corrosion layers. The cell spread and attachment were Mg substrate-dependent. The findings of coculturing bone cells directly on Mg materials within an in vitro model provide an effective approach for studying the dynamic degradation processes of Mg alloys while also elucidating cell behavior and their potential contribution to the degradation of these alloys. [Display omitted] [ABSTRACT FROM AUTHOR]

The efficiency of in vitro embryo production in mammals is influenced by variables associated with culture conditions during maturation, fertilization, and embryonic development. The embryos obtained often exhibit low quality due to suboptimal in vitro culture conditions compared to the in vivo environment. Co-culturing gametes and embryos with somatic cells has been developed to enhance in vitro culture conditions. This study aimed to assess the impact of coculturing in vitro-produced porcine embryos with porcine oviductal epithelial cells (POEC) on embryo development and quality. Firstly, a pure culture of POEC suitable for coculture systems was established. The epithelial origin of the cells was confirmed by the expression of E-cadherin and cytokeratin. The expression pattern of hormone receptors aligned with the diestrous oviduct, and POEC also secreted oviductal glycoprotein type 1 (OVGP-1). Secondly, POEC from passage 1 (POEC-1) were used to coculture with in vitro-produced porcine embryos. A successful coculture system was established without the addition of fetal bovine serum as a supplement. Coculturing POEC-1 in monolayers with in vitro-produced porcine embryos during the initial two days of culture enhanced the percentage of blastocysts and their hatching. Although the coculture did not alter the number of cells in the blastocysts or apoptosis assessed by TUNEL, it significantly reduced reactive oxygen species (ROS) levels in cleaved porcine embryos. This study represents the first report evaluating the quality of porcine embryos produced by IVF in coculture systems and assessing ROS levels in cleaved porcine embryos obtained by IVF. • The presence of oviductal epithelial cells during embryo in vitro culture enhanced porcine blastocyst formation. • The oviductal epithelial cells during embryo culture decrease ROS levels in porcine in vitro produced cleaved embryos. • The porcine oviductal epithelial cells expressed OVGP-1 during in vitro culture. [ABSTRACT FROM AUTHOR]

Background: In vitro single‐cell experiments may yield inconsistent results compared to clinical trials. To enhance the reliability of cosmetic active ingredient screening, a coculture model of B16F10‐HaCaT cells was established in vitro based on the structural characteristics of human skin, thereby improving the credibility of experimental outcomes. Currently, most cosmetic whitening additives primarily target simple efficacy goals such as inhibiting tyrosinase activity or melanin transfer. Therefore, investigating novel and efficient whitening additives has become a prominent research focus. Objectives: The aim is to establish an in vitro cell coculture model for more reliable experimental results and investigate the mechanism by which Paeonia lactiflora Pall seeds oil inhibits melanin production and transfer. Methods: The impact of different concentrations of Paeonia lactiflora Pall seeds oil on cocultured cell proliferation rate was assessed using cck8 assay. Tyrosinase inhibition ability in cocultured cells was tested using levodopa as a substrate. Melanin production inhibition ability in coculture cells was evaluated by lysing cells with sodium hydroxide. The effect of Paeonia lactiflora Pall seeds oil on dendrite‐related gene expression levels was examined through qPCR analysis. Additionally, Western blotting was employed to study the effect of Paeonia lactiflora Pall seeds oil on dendrite‐related protein expression levels. Results: Different concentrations of Paeonia lactiflora Pall seeds oil did not affect the proliferation activity of cocultured cells. A specific concentration of α‐MSH increased cell tyrosinase activity, cellular melanin content, as well as Rac1, Cdc42, and PAR‐2 gene and protein expression related to dendritic formation. Treatment with a certain concentration of Paeonia lactiflora Pall seeds oil resulted in decreased tyrosinase activity and melanin content in cells along with downregulated expression levels of Rac1, Cdc42, and PAR‐2 genes and proteins associated with dendritic formation. Conclusions: Paeonia lactiflora Pall seeds oil at specific concentrations exhibits the ability to inhibit tyrosinase activity, decrease melanin content, and possesses the potential to impede melanin transfer. [ABSTRACT FROM AUTHOR]

Legumes enhance pasture health and soil productivity by fixing atmospheric nitrogen and boosting soil microbiota. We investigated the effects of tropical pasture legumes, including butterfly pea (Clitoria ternatea), seca stylo (Stylosanthes scabra), desmanthus (Desmanthus virgatus), lablab (Lablab purpureus), and Wynn cassia (Chamaecrista rotundifolia), on the soil microbial community and buffel grass (Cenchrus ciliaris) gene expression. Additionally, we explored the impact of a phytogenic bioactive product (PHY) in the coculture system. A pot trial using soil enriched with cow paunch compost included four treatments: monoculture of buffel grass and five legume species with and without PHY supplementation and coculture of buffel grass with each legume species with and without PHY supplementation. Actinobacteriota and Firmicutes were the dominant bacterial phyla. Regardless of PHY application, the coculture of buffel grass with legumes positively influenced microbial composition and diversity. Transcriptomic analysis revealed significant gene expression changes in buffel grass shoots and roots, with each legume uniquely affecting nitrogen metabolism. Lablab and Wynn cassia exhibited similarities in modulating metabolic processes, butterfly pea contributed to mycotoxin detoxification, and desmanthus balanced cell death and growth. Seca stylo enhanced root cell growth and regeneration. These findings offer insights for optimizing legume–grass coculture systems, enhancing soil activity and promoting sustainable agriculture. [ABSTRACT FROM AUTHOR]

The engineering of microbial cell factories for the production of high-value chemicals from renewable resources presents several challenges, including the optimization of key enzymes, pathway fluxes and metabolic networks. Addressing these challenges involves the development of synthetic auxotrophs, a strategy that links cell growth with enzyme properties or biosynthetic pathways. This linkage allows for the improvement of enzyme properties by in vivo directed enzyme evolution, the enhancement of metabolic pathway fluxes under growth pressure, and remodeling of metabolic networks through directed strain evolution. The advantage of employing synthetic auxotrophs lies in the power of growth-coupled selection, which is not only high-throughput but also labor-saving, greatly simplifying the development of both strains and enzymes. Synthetic auxotrophs play a pivotal role in advancing microbial cell factories, offering benefits from enzyme optimization to the manipulation of metabolic networks within single microbes. Furthermore, this strategy extends to coculture systems, enabling collaboration within microbial communities. This review highlights the recently developed applications of synthetic auxotrophs as microbial cell factories, and discusses future perspectives, aiming to provide a practical guide for growth-coupled models to produce value-added chemicals as part of a sustainable biorefinery. [ABSTRACT FROM AUTHOR]

The human microbiota impacts a variety of diseases and responses to therapeutics. Due to a lack of robust in vitro models, detailed mechanistic explanations of host‐microbiota interactions cannot often be recapitulated. We describe the design and development of a novel, versatile and modular in vitro system that enables indirect coculture of human epithelial cells with anaerobic bacteria for the characterization of host‐microbe secreted metabolite interactions. This system was designed to compartmentalize anaerobes and human cells in separate chambers conducive to each organism's requisite cell growth conditions. Using perfusion, fluidic mixing, and automated sample collection, the cells continuously received fresh media, while in contact with their corresponding compartments conditioned supernatant. Supernatants from each chamber were collected in a cell‐free time‐resolved fashion. The system sustained low oxygen conditions in the anaerobic chamber, while also supporting the growth of a representative anaerobe (Bacteroides thetaiotaomicron) and a human colonic epithelial cell line (Caco‐2) in the aerobic chamber. Caco‐2 global gene expression changes in response to coculture with B. thetaiotaomicron was characterized using RNA sequencing. Extensive, targeted metabolomics analysis of over 150 central carbon metabolites was performed on the serially collected supernatants. We observed broad metabolite changes in host‐microbe coculture, compared to respective mono‐culture controls. These effects were dependent both on sampling time and the compartment probed (apical vs. basolateral). Coculturing resulted in the depletion of several important metabolites, including guanine, uridine 5'‐monophosphate, asparagine, and thiamine. Additionally, while Caco‐2 cells cultured alone predominantly affected the basolateral metabolite milieu, increased abundance of 2,3‐dihydroxyisovalerate and thymine on the basolateral side, occurred when the cells were cocultured with B. thetaiotaomicron. Thus, our system can capture the dynamic, competitive and cooperative processes between host cells and gut microbes. [ABSTRACT FROM AUTHOR]

IntroductionMicrocystis aeruginosa (M. aeruginosa), one of the most abundant blue-green algae in aquatic environments, produces microcystin by causing harmful algal blooms (HABs). This study investigated the combined effects of nutrients and competition among cyanobacterial subpopulations on the synthesis of microcystin-LR.MethodsUnder varying nitrogen and phosphorus concentrations, cyanobacterial coculture, and the presence of algicidal DCMU, the growth was monitored by optical density analysis or microscopic counting, and the microcystin production was analyzed using high-performance liquid chromatography-UV. Furthermore, growth and toxin production were predicted using a kinetic model.Results and discussionFirst, coculture with the fast-growing cyanobacterium Synechococcus elongatus UTEX 2973 (S. elongatus) reduced M. aeruginosa biomass and microcystin production at 30°C. Under high nitrogen and low phosphorus conditions, S. elongatus was most effective, limiting M. aeruginosa growth and toxin synthesis by up to 94.7% and 92.4%, respectively. Second, this biological strategy became less effective at 23°C, where S. elongatus grew more slowly. Third, the photosynthesis inhibitor DCMU (3-(3,4-dichlorophenyl)-1,1-dimethylurea) inhibited M. aeruginosa growth (at 0.1 mg/L) and microcystin production (at 0.02 mg/L). DCMU was also effective in controlling microcystin production in S. elongatus–M. aeruginosa cocultures. Based on the experimental results, a multi-substrate, multi-species kinetic model was built to describe coculture growth and population interactions.ConclusionMicrocystin from representative toxin-producing M. aeruginosa can be controlled by coculturing fast-growing benign cyanobacteria, which can be made even more efficient if appropriate algicide is applied. This study improved the understanding of the biological control of microcystin production under complex environmental conditions.

Chronic white adipose tissue (WAT) inflammation has been recognized as a critical early event in the pathogenesis of obesity-related disorders. This process is characterized by the increased residency of proinflammatory M1 macrophages in WAT. However, the lack of an isogenic human macrophage-adipocyte model has limited biological studies and drug discovery efforts, highlighting the need for human stem cell-based approaches. Here, human induced pluripotent stem cell (iPSC) derived macrophages (iMACs) and adipocytes (iADIPOs) are cocultured in a microphysiological system (MPS). iMACs migrate toward and infiltrate into the 3D iADIPOs cluster to form crown-like structures (CLSs)-like morphology around damaged iADIPOs, recreating classic histological features of WAT inflammation seen in obesity. Significantly more CLS-like morphologies formed in aged and palmitic acid-treated iMAC-iADIPO-MPS, showing the ability to mimic inflammatory severity. Importantly, M1 (proinflammatory) but not M2 (tissue repair) iMACs induced insulin resistance and dysregulated lipolysis in iADIPOs. Both RNAseq and cytokines analyses revealed a reciprocal proinflammatory loop in the interactions of M1 iMACs and iADIPOs. This iMAC-iADIPO-MPS thus successfully recreates pathological conditions of chronically inflamed human WAT, opening a door to study the dynamic inflammatory progression and identify clinically relevant therapies.

The dynamic structures of microbial communities emerge from the complex network of interactions between their constituent microorganisms. Quantitative measurements of these interactions are important for understanding and engineering ecosystem structure. Here, we present the development and application of the BioMe plate, a redesigned microplate device in which pairs of wells are separated by porous membranes. BioMe facilitates the measurement of dynamic microbial interactions and integrates easily with standard laboratory equipment. We first applied BioMe to recapitulate recently characterized, natural symbiotic interactions between bacteria isolated from the Drosophila melanogaster gut microbiome. Specifically, the BioMe plate allowed us to observe the benefit provided by two Lactobacillus strains to an Acetobacter strain. We next explored the use of BioMe to gain quantitative insight into the engineered obligate syntrophic interaction between a pair of Escherichia coli amino acid auxotrophs. We integrated experimental observations with a mechanistic computational model to quantify key parameters associated with this syntrophic interaction, including metabolite secretion and diffusion rates. This model also allowed us to explain the slow growth observed for auxotrophs growing in adjacent wells by demonstrating that, under the relevant range of parameters, local exchange between auxotrophs is essential for efficient growth. The BioMe plate provides a scalable and flexible approach for the study of dynamic microbial interactions. IMPORTANCE Microbial communities participate in many essential processes from biogeochemical cycles to the maintenance of human health. The structure and functions of these communities are dynamic properties that depend on poorly understood interactions among different species. Unraveling these interactions is therefore a crucial step toward understanding natural microbiota and engineering artificial ones. Microbial interactions have been difficult to measure directly, largely due to limitations of existing methods to disentangle the contribution of different organisms in mixed cocultures. To overcome these limitations, we developed the BioMe plate, a custom microplate-based device that enables direct measurement of microbial interactions, by detecting the abundance of segregated populations of microbes that can exchange small molecules through a membrane. We demonstrated the possible application of the BioMe plate for studying both natural and artificial consortia. BioMe is a scalable and accessible platform that can be used to broadly characterize microbial interactions mediated by diffusible molecules.

To evaluate the effects of filter-feeding fishes on water quality and bacterial community in the rice–crayfish coculture system, four different stocking densities of bighead carp (0, 500, 1000, 1500 ind./200 m2) were set up in rice–crayfish coculture systems. Water samples in the systems were collected biweekly to detect dissolved oxygen (DO), temperature (T), potential of Hydrogen (pH), ammonia nitrogen (NH4+-N), nitrite nitrogen (NO2−-N), nitrate nitrogen (NO3−-N), total nitrogen (TN), total phosphorus (TP), and Chlorophyll-a (Chl-a); the bacterial community in the water was analyzed simultaneously, then the correlation between water quality and microorganisms were studied. The results showed that concentrations of TN, TP, NO2−-N, and NH4+-N decreased while DO and NO3−-N increased along with the breeding process. NO2−-N, NO3−-N, TN, and NH4+-N were important environmental factors affecting the bacterial community structure in water (p < 0.05). Bighead carp stocking had an impact on the diversity, richness, and evenness of the bacterial communities in the systems. The dominant bacteria in the four different carp density groups were Proteobacteria, Actinomycetes, Bacteroidetes, and Cyanobacteria. Bighead carp increased the abundance of Bacteroidea but reduced that of Actinomycetes, Cyanobacteria, and Proteobacteria. The introduction of bighead carp promoted the conversion of nitrogen and phosphorus, reducing the risk of cyanobacterial blooms. Group 1000 ind./200 m2 exhibited the best effect on the removal of nitrogen and phosphorus from the water body. [ABSTRACT FROM AUTHOR]

There is a need for the assessment of respiratory hazard potential and mode of action of carbon nanotubes (CNTs) before their approval for technological or medical applications. In CNT-exposed lungs, both alveolar macrophages (MФs), which phagocytose CNTs, and alveolar epithelial type II cells (AECII cells), which show tissue injury, are impacted but cell–cell interactions between them and the impacted mechanisms are unclear. To investigate this, we first optimized an air–liquid interface (ALI) transwell coculture of human AECII cell line A549 (upper chamber) and human monocyte cell line THP-1 derived macrophages (lower chamber) in a 12-well culture by exposing macrophages to CNTs at varying doses (5–60 ng/well) for 12–48 h and measuring the epithelial response markers for cell differentiation/maturation (proSP-C), proliferation (Ki-67), and inflammation (IL-1β). In optimal ALI epithelial-macrophage coculture (3:1 ratio), expression of Ki-67 in AECII cells showed dose dependence, peaking at 15 ng/well CNT dose; the Ki-67 and IL-1β responses were detectable within 12 h, peaking at 24–36 h in a time-course. Using the optimized ALI transwell coculture set up with and without macrophages, we demonstrated that direct interaction between CNTs and MФs, but not a physical cell–cell contact between MФ and AECII cells, was essential for inducing immunotoxicity (proliferative and inflammatory responses) in the AECII cells. [ABSTRACT FROM AUTHOR]

The strong appeal to reduce animal testing calls for the development and validation of in vitro, in chemico and in silico models that would replace the need for in vivo testing and ex vivo materials. A category that requires such new approach methods is the assessment of immunosuppression that can be induced by chemicals including environmental pollutants. To assess the immunosuppressive action on monocytes and lymphocytes, we mimicked the whole-blood cytokine-release assay by preparing an in vitro coculture of THP-1 and Jurkat cell lines. We optimised its activation and investigated the effects of known immunosuppressive drugs with different mechanisms of action on the release of proinflammatory cytokines. Decreased secretion of IL-8 was achieved by several immunosuppressive mechanisms and was therefore selected as an appropriate marker of immunosuppression. A set of environmentally occurring bisphenols, BPA, BPAP, BPP, BPZ, BPE, TCBPA and BPS-MAE, were then applied to the model and BPP and BPZ were found to act as potent immunosuppressants at micromolar concentrations. [ABSTRACT FROM AUTHOR]

Cell–cell interactions between fibroblasts and immune cells, like macrophages, are influenced by interaction with the surrounding extracellular matrix during wound healing. In vitro hydrogel models that mimic and modulate these interactions, especially of soluble mediators like cytokines, may allow for a more detailed investigation of immunomodulatory processes. In the present study, a biomimetic extracellular matrix model based on fibrillar 3D collagen I networks with a functionalization with heparin or 6-ON-desulfated heparin, as mimics of naturally occurring heparan sulfate, was developed to modulate cytokine binding effects with the hydrogel matrix. The constitution and microstructure of the collagen I network were found to be stable throughout the 7-day culture period. A coculture study of primary human fibroblasts/myofibroblasts and M-CSF-stimulated macrophages was used to show its applicability to simulate processes of progressed wound healing. The quantification of secreted cytokines (IL-8, IL-10, IL-6, FGF-2) in the cell culture supernatant demonstrated the differential impact of glycosaminoglycan functionalization of the collagen I network. Most prominently, IL-6 and FGF-2 were shown to be regulated by the cell culture condition and network constitution, indicating changes in paracrine and autocrine cell–cell communication of the fibroblast–macrophage coculture. From this perspective, we consider our newly established in vitro hydrogel model suitable for mechanistic coculture analyses of primary human cells to unravel the role of extracellular matrix factors in key events of tissue regeneration and beyond. [ABSTRACT FROM AUTHOR]

ABSTRACT Many microbial biosynthetic gene clusters (BGCs) are inactive under standard laboratory conditions, making characterization of their products difficult. Silent BGCs are likely activated by specific cues in their natural environment, such as the presence of competitors. Growth conditions such as coculture with other microbes, which more closely mimic natural environments, are practical strategies for inducing silent BGCs. Here, we utilize coculture to activate BGCs in nine actinobacteria strains. We observed increased production of the ferrous siderophores siderochelin A and B during coculture of Amycolatopsis strain WAC04611 and Tsukamurella strain WAC06889b. Furthermore, we identified the siderochelin BGC in WAC04611 and discovered that the GntR-family transcription factor sidR3 represses siderochelin production. Deletion of the predicted aminotransferase sidA abolished production of the carboxamides siderochelin A/B and led to the accumulation of the carboxylate siderochelin D. Finally, we deleted the predicted hydroxylase sidB and established that it is essential for siderochelin production. Our findings show that microbial coculture can successfully activate silent BGCs and lead to the discovery and characterization of unknown BGCs for molecules like siderochelin.IMPORTANCESiderophores are vital iron-acquisition elements required by microbes for survival in a variety of environments. Furthermore, many siderophores are essential for the virulence of various human pathogens, making them a possible target for antibacterials. The significance of our work is in the identification and characterization of the previously unknown BGC for the siderophore siderochelin. Our work adds to the growing knowledge of siderophore biosynthesis, which may aid in the future development of siderophore-targeting pharmaceuticals and inform on the ecological roles of these compounds. Furthermore, our work demonstrates that combining microbial coculture with metabolomics is a valuable strategy for identifying upregulated compounds and their BGCs.

ABSTRACT Although most cyanobacteria grow in visible light (VL; λ = 400–700 nm), some cyanobacteria can also use far-red light (FRL; λ = 700–800 nm) for oxygenic photosynthesis by performing far-red light photoacclimation. These two types of cyanobacteria can be found in the same environment. However, how they respond to each other remains unknown. Here, we reveal that coculture stresses FRL-using Chlorogloeopsis fritschii PCC 9212 and VL-using Synechocystis sp. PCC 6803. No significant growth difference was found in Synechocystis sp. PCC 6803 between the coculture and the monoculture. Conversely, the growth of Chlorogloeopsis fritschii PCC 9212 was suppressed in VL under coculture. According to transcriptomic analysis, Chlorogloeopsis fritschii PCC 9212 in coculture shows low transcript levels of metabolic activities and high transcript levels of ion transporters, with the differences being more noticeable in VL than in FRL. The transcript levels of stress responses in coculture were likewise higher than in monoculture in Synechocystis sp. PCC 6803 under FRL. The low transcript level of metabolic activities in coculture or the inhibition of cyanobacterial growth indicates a possible negative interaction between these two cyanobacterial strains.IMPORTANCEThe interaction between two cyanobacterial species is the primary focus of this study. One species harvests visible light, while the other can harvest far-red and visible light. Prior research on cyanobacteria interaction concentrated on its interactions with algal, coral, and fungal species. Interactions between cyanobacterial species were, nevertheless, rarely discussed. Thus, we characterized the interaction between two cyanobacterial species, one capable of photosynthesis using far-red light and the other not. Through experimental and bioinformatic approaches, we demonstrate that when one cyanobacterium thrives under optimal light conditions, it stresses the remaining cyanobacterial species. We contribute to an ecological understanding of these two kinds of cyanobacteria distribution patterns. Cyanobacteria that utilize far-red light probably disperse in environments with limited visible light to avoid competition with other cyanobacteria. From a biotechnological standpoint, this study suggests that the simultaneous cultivation of two cyanobacterial species in large-scale cultivation facilities may reduce the overall biomass yield.

Abstract Multi resistant fungi are on the rise, and our arsenal compounds are limited to few choices in the market such as polyenes, pyrimidine analogs, azoles, allylamines, and echinocandins. Although each of these drugs featured a unique mechanism, antifungal resistant strains did emerge and continued to arise against them worldwide. Moreover, the genetic variation between fungi and their host humans is small, which leads to significant challenges in new antifungal drug discovery. Endophytes are still an underexplored source of bioactive secondary metabolites. Many studies were conducted to isolate and screen endophytic pure compounds with efficacy against resistant yeasts and fungi; especially, Candida albicans, C. auris, Cryptococcus neoformans and Aspergillus fumigatus, which encouraged writing this review to critically analyze the chemical nature, potency, and fungal source of the isolated endophytic compounds as well as their novelty features and SAR when possible. Herein, we report a comprehensive list of around 320 assayed antifungal compounds against Candida albicans, C. auris, Cryptococcus neoformans and Aspergillus fumigatus in the period 1980–2024, the majority of which were isolated from fungi of orders Eurotiales and Hypocreales associated with terrestrial plants, probably due to the ease of laboratory cultivation of these strains. 46% of the reviewed compounds were active against C. albicans, 23% against C. neoformans, 29% against A. fumigatus and only 2% against C. auris. Coculturing was proved to be an effective technique to induce cryptic metabolites absent in other axenic cultures or host extract cultures, with Irperide as the most promising compounds MIC value 1 μg/mL. C. auris was susceptible to only persephacin and rubiginosin C. The latter showed potent inhibition against this recalcitrant strain in a non-fungicide way, which unveils the potential of fungal biofilm inhibition. Further development of culturing techniques and activation of silent metabolic pathways would be favorable to inspire the search for novel bioactive antifungals. Graphic abstract

One strain of Hanseniaspora uvarum YT-35 was screened from fermented sediment of wild cherry. Hanseniaspora uvarum YT-35 and commercial Saccharomyces cerevisiae were used as coculture for manufacture of fermented wild cherry beverage. The dynamics of microbial populations, reducing sugars and ethanol were analyzed at different stages of fermentation using single-strain fermentation with 2 strains of bacteria as a control. Meanwhile, the organic acids and volatile aromatic compounds of the fermented beverages were detected by high-performance liquid chromatography (HPLC) and headspace solid-phase microextraction/gas chromatography-mass spectrometry (HS-SPME/GC-MS). The results showed that H. uvarum YT-35 dominated in the pre-fermentation stage of co-culture. Compared with single fermentation with S. cerevisiae, the coculture fermentation resulted in lower ethanol content (3.51 g/L). Notably, HPLC results revealed that coculture fermented beverage reduced the yield of citric, malic and quinic acids and increased the yield of glacial acetic acid. HS-SPME/GC-MS results revealed that coculture fermented beverage produced more volatile compounds of esters, such as ethyl caproate, methyl benzoate and isoamyl octanoate and showed enhanced contents of ethyl laurate, ethyl octanoate, phenyl ethyl alcohol, benzyl alcohol, octanoic acid and lauric acid. Meanwhile, clustering analysis revealed that coculture fermentation were correlated with the greatest number of volatile aroma compounds in the fermented wild cherry beverage. This study provides scientific basis and theoretical guidance for the research of coculture strains with different metabolic potential in improving the quality of fruit juice fermented beverage.

Multi resistant fungi are on the rise, and our arsenal compounds are limited to few choices in the market such as polyenes, pyrimidine analogs, azoles, allylamines, and echinocandins. Although each of these drugs featured a unique mechanism, antifungal resistant strains did emerge and continued to arise against them worldwide. Moreover, the genetic variation between fungi and their host humans is small, which leads to significant challenges in new antifungal drug discovery. Endophytes are still an underexplored source of bioactive secondary metabolites. Many studies were conducted to isolate and screen endophytic pure compounds with efficacy against resistant yeasts and fungi; especially, Candida albicans, C. auris, Cryptococcus neoformans and Aspergillus fumigatus, which encouraged writing this review to critically analyze the chemical nature, potency, and fungal source of the isolated endophytic compounds as well as their novelty features and SAR when possible. Herein, we report a comprehensive list of around 320 assayed antifungal compounds against Candida albicans, C. auris, Cryptococcus neoformans and Aspergillus fumigatus in the period 1980–2024, the majority of which were isolated from fungi of orders Eurotiales and Hypocreales associated with terrestrial plants, probably due to the ease of laboratory cultivation of these strains. 46% of the reviewed compounds were active against C. albicans, 23% against C. neoformans, 29% against A. fumigatus and only 2% against C. auris. Coculturing was proved to be an effective technique to induce cryptic metabolites absent in other axenic cultures or host extract cultures, with Irperide as the most promising compounds MIC value 1 μg/mL. C. auris was susceptible to only persephacin and rubiginosin C. The latter showed potent inhibition against this recalcitrant strain in a non-fungicide way, which unveils the potential of fungal biofilm inhibition. Further development of culturing techniques and activation of silent metabolic pathways would be favorable to inspire the search for novel bioactive antifungals. [ABSTRACT FROM AUTHOR]

Estimation of the biliary clearance of drugs and their metabolites in humans is crucial for characterizing hepatobiliary disposition and potential drug-drug interactions. Sandwich-cultured hepatocytes, while useful for in vitro bile analysis, require cell destruction for bile recovery, limiting long-term or repeated dose drug effect evaluations. To overcome this limitation, we investigated the feasibility of coculturing a human hepatic carcinoma cell line (HepG2-NIAS cells) and a human cholangiocarcinoma cell line (TFK-1 cells) using the collagen vitrigel membrane in a variety of coculture configurations. The coculture configuration with physiological bile flow increased the permeability of fluorescein-labeled bile acids (CLF) across the HepG2-NIAS cell layer by approximately 1.2-fold compared to the HepG2-NIAS monoculture. This enhancement was caused by paracellular leakage due to the loosened tight junctions of HepG2-NIAS, confirmed by the use of an inhibitor for bile acid transporters, the increase of permeability of dextran, and the decrease of the transepithelial electrical resistance (TEER) value. Based on the results of loosening hepatic tight junctions via coculture with TFK-1 in the CLF permeability assay, we next attempted to collect the CLF accumulated in the bile canaliculi of HepG2-NIAS. The recovery of the CLF accumulated in the bile canaliculi was increased 1.4 times without disrupting hepatic tight junctions by the coculture of HepG2-NIAS cells and TFK-1 cells compared to the monoculture of HepG2-NIAS cells. This non-destructive bile recovery has the potential as a tool for estimating the biliary metabolite and provides valuable insights to improve in vitro bile analysis. [ABSTRACT FROM AUTHOR]

Liver failure represents a critical medical condition with a traditionally grim prognosis, where treatment options have been notably limited. Historically, liver transplantation has stood as the sole definitive cure, yet the stark disparity between the limited availability of liver donations and the high demand for such organs has significantly hampered its feasibility. This discrepancy has necessitated the exploration of hepatocyte transplantation as a temporary, supportive intervention. In light of this, our review delves into the burgeoning field of hepatocyte transplantation, with a focus on the latest advancements in maintaining hepatocyte function, co-microencapsulation techniques, xenogeneic hepatocyte transplantation, and the selection of materials for microencapsulation. Our examination of hepatocyte microencapsulation research highlights that, to date, most studies have been conducted in vitro or using liver failure mouse models, with a notable paucity of experiments on larger mammals. The functionality of microencapsulated hepatocytes is primarily inferred through indirect measures such as urea and albumin production and the rate of ammonia clearance. Furthermore, research on the mechanisms underlying hepatocyte co-microencapsulation remains limited, and the practicality of xenogeneic hepatocyte transplantation requires further validation. The potential of hepatocyte microencapsulation extends beyond the current scope of application, suggesting a promising horizon for liver failure treatment modalities. Innovations in encapsulation materials and techniques aim to enhance cell viability and function, indicating a need for comprehensive studies that bridge the gap between small-scale laboratory success and clinical applicability. Moreover, the integration of bioengineering and regenerative medicine offers novel pathways to refine hepatocyte transplantation, potentially overcoming the challenges of immune rejection and ensuring the long-term functionality of transplanted cells. In conclusion, while hepatocyte microencapsulation and transplantation herald a new era in liver failure therapy, significant strides must be made to translate these experimental approaches into viable clinical solutions. Future research should aim to expand the experimental models to include larger mammals, thereby providing a clearer understanding of the clinical potential of these therapies. Additionally, a deeper exploration into the mechanisms of cell survival and function within microcapsules, alongside the development of innovative encapsulation materials, will be critical in advancing the field and offering new hope to patients with liver failure. [ABSTRACT FROM AUTHOR]

Being the green gold of the future, cyanobacteria have recently attracted considerable interest worldwide. This study investigates the adaptability and biocompatibility of the cyanobacterial strain Synechococcus sp. PCC 7002 with human dermal cells, focusing on its potential application in biomedical contexts. First, we investigated the adaptability of Synechococcus PCC 7002 bacteria to human cell culture conditions. Next, we evaluated the biocompatibility of cyanobacteria with common dermal cells, like 3T3 fibroblasts and HaCaT keratinocytes. Therefore, cells were directly and indirectly cocultured with the corresponding cells, and we measured metabolic activity (AlamarBlue assay) and proliferation (cell count and PicoGreen assay). The lactate dehydrogenase (LDH) assay was performed to determine the cytotoxic effect of cyanobacteria and their nutrition medium on human dermal cells. The cyanobacteria exhibited exponential growth under conventional human cell culture conditions, with the temperature and medium composition not affecting their viability. In addition, the effect of illumination on the proliferation capacity was investigated, showing a significant impact of light exposure on bacterial growth. The measured oxygen production under hypoxic conditions demonstrated a sufficient oxygen supply for further tissue engineering approaches depending on the number of bacteria. There were no significant adverse effects on human cell viability and growth under coculture conditions, whereas the LDH assay assessed signs of cytotoxicity regarding 3T3 fibroblasts after 2 days of coculturing. These negative effects were dismissed after 4 days. The findings highlight the potential of Synechococcus sp. PCC 7002 for integration into biomedical approaches. We found no cytotoxicity of cyanobacteria on 3T3 fibroblasts and HaCaT keratinocytes, thus paving the way for further in vivo studies to assess long-term effects and systemic reactions. [ABSTRACT FROM AUTHOR]

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The lack of human‐derived in vitro models that recapitulate cervical pre‐cancerous lesions has been the bottleneck in researching human papillomavirus (HPV) infection‐associated pre‐cancerous lesions and cancers for a long time. Here, a long‐term 3D organoid culture protocol for high‐grade squamous intraepithelial lesions and cervical squamous cell carcinoma that stably recapitulates the two tissues of origin is described. Originating from human‐derived samples, a small biobank of cervical pre‐tumoroids and tumoroids that faithfully retains genomic and transcriptomic characteristics as well as the causative HPV genome is established. Cervical pre‐tumoroids and tumoroids show differential responses to common chemotherapeutic agents and grow differently as xenografts in mice. By coculture organoid models with peripheral blood immune cells (PBMCs) stimulated by HPV antigenic peptides, it is illustrated that both organoid models respond differently to immunized PBMCs, supporting organoids as reliable and powerful tools for studying virus‐specific T‐cell responses and screening therapeutic HPV vaccines. In this study, a model of cervical pre‐cancerous lesions containing HPV is established for the first time, overcoming the bottleneck of the current model of human cervical pre‐cancerous lesions. This study establishes an experimental platform and biobanks for in vitro mechanistic research, therapeutic vaccine screening, and personalized treatment for HPV‐related cervical diseases. [ABSTRACT FROM AUTHOR]

Introduction: Bioprinting, using “bio-inks” consisting of living cells, supporting structures, and biological motifs to create customized constructs, is an emerging technique that aims to overcome the challenges of cartilaginous reconstruction of head and neck structures. Several living cell lines and culturing methods have been explored as bio-inks with varying efficacy. Coculture of primary chondrocytes and stem cells (SCs) is one technique well established for degenerative joint disease treatment, with potential for use in expanding chondrocyte populations for bio-inks. This study aimed to evaluate the techniques for coculture of primary chondrocytes and SCs for head and neck cartilage regeneration. Methods: A literature review was performed through OVID/Web of Science/MEDLINE/BIOSIS Previews/Embase. Studies reporting on chondrocytes and SCs in conjunction with coculture or cartilage regeneration were included. Studies not reporting on findings from chondrocytes/SCs of the head and neck were excluded. Extracted data included cell sources, coculture ratios, and histological, biochemical, and clinical outcomes. Results: Fifteen studies met inclusion criteria. Auricular cartilage was the most common chondrocyte source (n = 10), then nasal septum (n = 5), articular (n = 1), and tracheal cartilage (n = 1). Bone marrow was the most common SC source (n = 9) then adipose tissue (n = 7). Techniques varied, with coculture ratios ranging from 1:1 to 1:10. All studies reported coculture to be superior to SC monoculture by all outcomes. Most studies reported superiority or equivalence of coculture to chondrocyte monoculture by all outcomes. When comparing clinical outcomes, coculture constructs were equivalent to chondrocyte monoculture in diameter and equivalent or inferior in wet weight and height. Conclusion: Coculture of primary chondrocytes and SCs is a promising technique for expanding chondrocyte populations, with at least equivalence to chondrocyte monoculture and superior to SC monoculture when seeded at the same chondrocyte densities. However, there remains a lack of consensus regarding the optimal cell sources and coculture ratios. [ABSTRACT FROM AUTHOR]

Osteoarthritis (OA) is a whole-joint disorder involving inflammation in cartilage, synovium, and subchondral bone. An ex vivo inflammatory OA model incorporating various intra-articular tissues could facilitate the development of new OA therapeutics targeting whole joint tissue. In this study, an inflammatory ex vivo coculture system with bovine osteochondral explants (OCEs) and synovial explants (SEs) was induced using 1 ng/mL interleukin one beta and tumor necrosis factor alpha. Furthermore, 5-aminosalicylic acid (5-ASA), a promising OA drug reported previously, was assessed in the coculture system to evaluate the potential of the system to be applied in drug screening for OA treatment. Under inflammatory stimulation, cartilage and synovium tissues in monoculture displayed a strong inflammatory response transcriptionally, biochemically, and histologically. Under inflammatory conditions, coculture elevated the gene expression of IL-6 in synovium tissue compared with monocultured synovium. The synovium tissue in the coculture inflammatory group showed the highest synovitis score. The IL-8 release in the coculture medium was significantly higher than the additive amount in the OCE and SE monoculture groups under inflammatory microenvironment. For drug assessment, 5-ASA-treated group could promote the gene expression of CD163/CD86 and inhibit CD86 expression in synovium compared with inflammatory group. Moreover, the cumulative release of IL-6 into the medium was mitigated upon 5-ASA treatment. This study revealed the interaction between OCEs and SEs in an ex vivo coculture system. This preclinical coculture system could be used to evaluate the effects of novel OA drugs on both cartilage and synovium and explore the crosstalk mechanisms between intra-articular tissues.

Abstract Background Pulp inflammation is complex interactions between different types of cells and cytokines. To mimic the interactions of different types of cells in inflamed dental pulp tissues, dental pulp cells (DPCs) were cocultured with different ratios of macrophages (THP-1) or LPS treatment. Methods DPCs were cocultured with various ratios of THP-1, then photographed cell morphology and determined cell viability by MTT assay at preset times. Total RNA was also extracted to measure the inflammation marker-IL-6 and IL-8 expressions by RT-Q-PCR. The DPCs and THP-1 were treated with 0.01 – 1μg/ml lipopolysaccharide (LPS) and extract RNA at preset times, and detected IL-6 and IL-8 expression. DPCs were cocultured with various ratios of THP-1 with 0.1 μg/mL LPS, and detected IL-6 and IL-8 expression after 24 and 48 h. The data were analyzed by unpaired t-test or Mann-Whitney test. Differences were considered statistically significant when p

Recent advances in genetics, molecular biology, and stem cell biology have accelerated our understanding of neuropsychiatric disorders, like autism spectrum disorder (ASD), major depressive disorder (MDD), bipolar disorder (BD), and schizophrenia (SZ). This progress highlights the incredible complexity of both the human brain and mental illnesses from the biochemical to the cellular level. Contributing to the complexity of neuropsychiatric disorders are their polygenic nature, cellular and brain region interconnectivity, and dysregulation of human-specific neurodevelopmental processes. Here, we discuss available tools, including CRISPR-Cas9, and the applications of these tools to develop cell-based two-dimensional (2D) models and 3D brain organoid models that better represent and unravel the intricacies of neuropsychiatric disorder pathophysiology.

Context: In pigs, in vitro fertilisation (IVF) is associated with high polyspermy rates, and for this reason, in vitro embryo production (IVP) is still an inefficient biotechnology. Coculture with somatic cells is an alternative to improve suboptimal in vitro maturation (IVM) conditions. Aim: This study was conducted to test a coculture system of porcine luteal cells (PLC) and cumulus–oocyte complexes (COC) to improve oocyte metabolism. Methods: COC were matured in vitro with PLC. Oocyte lipid content, mitochondrial activity, zona pellucida (ZP) digestibility and pore size, cortical reaction and in vitro embryo development were assessed. Key results: Coculture reduced cytoplasmic lipid content in the oocyte cytoplasm without increasing mitochondrial activity. Although ZP digestibility and ZP pore number were not different between culture systems, ZP pores were smaller in the coculture. Coculture impacted the distribution of cortical granules as they were found immediately under the oolemma, and more of them had released their content in the ZP. Coculture with porcine luteal cells during IVM increased monospermic penetration and embryo development after IVF. Conclusions: The coculture of COC with PLC affects the metabolism of the oocyte and benefits monospermic penetration and embryo development. Implications: The coculture system with PLC could be an alternative for the conventional maturation medium in pigs. In vitro -produced embryos are essential for employing assisted reproductive technologies to establish porcine models. However, polyspermy remains a major obstacle to the production of porcine embryos in vitro. This study was conducted to test a coculture system of porcine luteal cells during in vitro maturation to improve suboptimal conditions. Our model could be an alternative to replace the conventional maturation medium, affecting the metabolism of the oocytes and leading to lower rates of polyspermy and higher rates of embryo development. Image by Gabriela Teplitz and Alexandre Bastien. [ABSTRACT FROM AUTHOR]

The rising number of antibiotic‐resistant bacteria requires alternative antibacterial substances as therapeutics. However, besides a strong effect against bacteria, they should not exhibit cytotoxicity towards human cells. Typically, both properties are tested in separate in vitro experiments that do not take into account naturally occurring interactions. To analyze whether the bacteria‐cell interaction influences the results of antibacterial and cytotoxic testing, surfactant‐free antibacterial silver‐gold alloy nanoparticles produced by laser ablation in liquid were examined both separately and in a straightforward coculture setup. Whereas different commensal bacteria and human skin cells exhibited reduced metabolic activity at comparable concentrations when treated separately, their response completely differed when analyzed in a combined coculture. For the combination of oral keratinocytes and Staphylococcus aureus, reduced cytotoxicity with increasing cell numbers, but a similar antibacterial effect quantified by standard plate counting could be observed. In contrast, coculture of keratinocytes and Porphyromonas gingivalis resulted in a complete absence of cytotoxicity and antibacterial effect, probably due to bacterial invasion of cells. These results clearly showed that the bacteria‐cell interaction greatly influences the results of antibacterial and cytotoxic testing and highlight the importance of more complex in vitro experiments to reliably characterize novel antibacterial substances for efficient clinical translation. [ABSTRACT FROM AUTHOR]

Lactiplantibacillus plantarum OV50 is a novel strain that was isolated from Algerian olives. Prior to its use as a natural biopreservative, OV50 underwent characterization for various functions. OV50 shows no proteolytic, lipolytic, or hemolytic activity. In addition, it is non-cytotoxic to eukaryotic cells and does not exhibit acquired antibiotic resistance. OV50 was tested with Pseudomonas aeruginosa ATCC 27835, Staphylococcus aureus ATCC 6538, Escherichia coli ATCC 8739, and Vibrio cholerae ATCC 14035 in a sardine based-medium at 37 °C and 7 °C. At 37 °C, OV50 completely inhibited the growth of these foodborne pathogens for a maximum of 6 h. At 7 °C, it suppressed their growth for a maximum of 8 days, except for S. aureus ATCC 6538, whose growth was reduced from 4 to 2 log CFU/mL. Microbiological counts, total volatile basic nitrogen (TVB-N), and peroxide values (PV) concentrations were determined in fresh sardines inoculated with OV50 and kept at 7 °C for 12 days. The inoculated sardines showed a significant reduction in TVB-N levels at D8 (34.9 mg/100 g) compared to the control (59.73 mg/100 g) and in PV concentrations at D4 (6.67 meq/kg) compared to the control (11.44 meq/kg), as well as a significant reduction in the numbers of Enterobacterales, Coliforms, Pseudomonas spp., Vibrio spp., and S. aureus At D8 and D12 compared to the control. Taken together, these results indicate that OV50 can improve the microbiological safety, freshness, and quality of sardines. [ABSTRACT FROM AUTHOR]

(–)‐Cannabidiol (CBD), a nonpsychoactive compound isolated from the Cannabis genus, has recently emerged as a promising research subject in neuroscience due to its potential therapeutic benefits against neurological disorders. However, there has been limited fundamental understanding of how CBD affects neuronal development, such as neurite outgrowth and axonal branching, at the cell level. This work investigates the effects of CBD on the early development of primary hippocampal neurons in a noncontact neuron‐astrocyte coculture system. The immunocytochemical and quantitative analyses indicate that CBD of 10 μM promotes neurite growth, including neurite elongation and arborization, and accelerates the process of axon specification. The CBD treatment to the coculture system leads to noticeable increases in the longest‐neurite length, primary‐neurite number, and branch‐point number, as well as the population of axon‐determined neurons. The results provide informative insights into the therapeutic potential of CBD in neurological disorders, such as Alzheimer's disease. [ABSTRACT FROM AUTHOR]

Human adipose tissue-derived stem/stromal cells (hASCs) are adult multipotent mesenchymal stem/stromal cells with immunomodulatory capacities. Here, we present up-to-date knowledge on the impact of different experimental and donor-related factors on hASC immunoregulatory functions in vitro. The experimental determinants include the immunological status of hASCs relative to target immune cells, contact vs. contactless interaction, and oxygen tension. Factors such as the ratio of hASCs to immune cells, the cellular context, the immune cell activation status, and coculture duration are also discussed. Conditioning of hASCs with different approaches before interaction with immune cells, hASC culture in xenogenic or xenofree culture medium, hASC culture in two-dimension vs. three-dimension with biomaterials, and the hASC passage number are among the experimental parameters that greatly may impact the hASC immunosuppressive potential in vitro, thus, they are also considered. Moreover, the influence of donor-related characteristics such as age, sex, and health status on hASC immunomodulation in vitro is reviewed. By analysis of the literature studies, most of the indicated determinants have been investigated in broad non-standardized ranges, so the results are not univocal. Clear conclusions cannot be drawn for the fine-tuned scenarios of many important factors to set a standard hASC immunopotency assay. Such variability needs to be carefully considered in further standardized research. Importantly, field experts' opinions may help to make it clearer. Parameters that promote ASC immunosuppression on immune cells. Activation of immune cells induces their proliferation and differentiation and presence of ASCs modulates/suppresses such consequences. Augmented immunosuppressive effects of ASCs can be introduced in direct contact with the immune cells and via complementing the repeatedly reported experimental settings (texts in grey shapes). Abbreviations: ASCs: adipose tissue-derived stem/stromal cells, IFN-ɤ: Interferon gamma, MLR: Mixed lymphocyte reaction, TNF: Tumor necrosis factor. [ABSTRACT FROM AUTHOR]