Tissue Culture Research Articles

The interplay between homeostatic synaptic scaling and homeostatic structural plasticity maintains the robust firing rate of neural networks.

Critical network states and neural plasticity enable adaptive behavior in dynamic environments, supporting efficient information processing and experience-dependent learning. Synaptic-weight-based Hebbian plasticity and homeostatic synaptic scaling are key mechanisms that enable memory while stabilizing network dynamics. However, the role of structural plasticity as a homeostatic mechanism remains less consistently reported, particularly under activity inhibition, leading to an incomplete understanding of its functional impact. In this study, we combined live-cell microscopy of eGFP-labeled neurons in mouse organotypic entorhinal-hippocampal tissue cultures (Thy1-eGFP mice of both sexes) with computational modeling to investigate how synapse-number-based structural plasticity responds to activity perturbations and interacts with homeostatic synaptic scaling. Tracking individual dendritic segments, we found that inhibiting excitatory neurotransmission does not monotonically regulate dendritic spine density. Specifically, inhibition of AMPA receptors with 200 nM 2,3-dioxo-6-nitro-7-sulfamoyl-benzo[f]quinoxaline (NBQX) increased spine density, whereas complete AMPA receptor blockade with 50 μM NBQX reduced it. Motivated by these findings, we developed network simulations incorporating a biphasic structural plasticity rule governing activity-dependent synapse formation. These simulations showed that the biphasic rule maintains neural activity homeostasis under stimulation and permits either synapse formation or synapse loss depending on the degree of activity deprivation. Homeostatic synaptic scaling further modulated recurrent connectivity, network activity, and structural plasticity outcomes. It reduced stimulation-triggered synapse loss by downscaling synaptic weights and rescued silencing-induced synapse loss by upscaling recurrent input, thus reactivating silent neurons. The interaction between these mechanisms provides a mechanistic explanation for divergent findings in the literature. In summary, homeostatic synaptic scaling and homeostatic structural plasticity dynamically compete and compensate for each other, ensuring efficient and robust control of firing rate homeostasis.

Ascorbate Regulates Root Development Through Auxin Response in Arabidopsis thaliana

Ascorbic acid (ASC) is a furan-based lactone derived from 2-ketogluconic acid that functions as a major antioxidant and redox buffer in mature plant tissues, although its content is lower in meristematic cells. ASC is commonly considered a reactive oxygen species (ROS) scavenger; however, its role in the regulation of plant development remains unclear. Additionally, the chemical behavior of ascorbate warrants special attention during ASC supplementation in in vitro plant culture. In this study, I investigated in detail the behavior of ascorbate in plant tissue culture medium and its uptake by plants. As a secondary objective, the role of ascorbate in root growth regulation was evaluated. The effects of low ASC levels on root architecture and its interaction with auxin signaling were studied using the vtc1 and vtc2 mutants of Arabidopsis thaliana, as well as through external ascorbate supplementation. Several marker lines for auxin response/distribution were used, along with direct ascorbate measurement via HPLC. Reducing ascorbate content through mutations had no significant effect on root development or auxin signaling, whereas high-concentration ASC supplementation inhibited both auxin signaling and root development, as demonstrated using auxin response and transport markers. At the organ level, ASC supplementation significantly downregulated auxin response-mediated cell cycle activation during lateral root induction. Based on the data presented, exogenous ascorbate may regulate root development through its interaction with auxin signaling pathways.

Use of phenotypic markers to determine the genetic variability of some local and tissue cultured date palm Phoenix dactylifera L. cultivars

This study was conducted during the growing season (2024) in one of the private orchards in Nasiriyah district, the center of Dhi Qar Governorate, at longitude (46.06) and latitude (31.90) on some date palm cultivars produced by tissue culture and their counterparts grown by the offshoots method (the traditional method). The aim of studying the phenotypic description (vegetative, floral and fruit) of the studied cultivars, which are (local Barhi and produced by tissue culture, local Balka and produced by tissue culture, local Sukkari and produced by tissue culture), in addition to the cultivars produced by tissue culture only (Khalas, Saqai and Majhool). Results of the phenotypic indicators (vegetative, floral and fruit) and the total chlorophyll content of the leaves showed that the local Barhi cultivar was superior in the average traits (the length of the leaves, the number of fronds, the number of thorn , the length of the thorn , the length of the thorns area, the length of the fronds area, the width of the leaf base, the diameter of the trunk, the number of spadix, Length of the spadix, the width of the spadix, panicle weight, number of flowers, fruit stalk length, fruit panicle length and fruit diameter), while the Barhi produced by tissue culture cultivar recorded the highest rate for the two traits (percentage of frond area and the length of fruit panicle), the Balka produced by tissue culture cultivar recorded the highest rate for the trait of percentage of frond, The local Balka cultivar recorded the highest rates for the following traits (frond length, thorn length, total chlorophyll, fruit length, and fruit cap height), while the local Sukkari cultivar recorded the highest rates for the traits (frond width, total chlorophyll, and number of spadix), the Khalas produced by tissue culture cultivar recorded the highest rate for the trait (thorn length), the Majhool produced by tissue culture cultivar recorded the highest rates for the following traits (thorn length, thorns area percentage, total chlorophyll, number of spadix, fruit diameter, fruit and seed weights, fleshy layer, and fruit size), while the Saqai produced by tissue culture cultivar recorded the highest rates for e traits (frond length, frond area percentage, number of panicles, fruit diameter, fruit weight, fleshy layer weight, and fruit size). The results of tree cluster analysis of phenotypic traits among studied cultivars showed that cultivars were distributed into two groups, the first group included cultivars (Saqai produced by tissue culture, local Balka, local Sukkari, local Barhi and produced by tissue culture), while the second group included cultivars (Sukkari, Majhool, Balka and Khalas produced by tissue culture). The Sukkari produced by tissue culture cultivar recorded furthest genetic distance of (14.2%), thus it is furthest genetically from the rest of studied cultivar. The percentage of genetic similarity between local Barhi and produced by tissue culture cultivar reached (100%), and between the local Balka and produced by tissue culture cultivars reached (99.9%). The results showed by analysis the main ingredients of variation of the appearance qualities that there are a range of appearance traits (Vegetable, floral and fruity) can be used to distinguish between the different cultivars of dates palm most important (The length of the fruity panicle , number of panicles, fruiting stalk length , panicle weight, number of flowers, spadix width, fruit diameter, number of spadix , leaf base width, spadix length, thorns area length, seed and fruit weights, the weight of the flesh layer, the length of the fronds area, the diameter of the trunk, number of thorns , the size of the fruit, fruit cap height, number of fronds, fruit length, the ratio of the fronds area, the ratio of the thorns area, the length of the frond and the width of the frond), which represents these traits (85.13%) of the total variation.

Maintenance of pluripotency and osteogenic differentiation of human mesenchymal stem cells on acrylate-based substrates exhibiting gelatin or heparin grafting

This study investigated the in vitro biological behaviour of human mesenchymal stem cells grown on acrylate-based substrates functionalized with biomolecules that play a significant role in the extracellular matrix, specifically gelatin (derived from collagen) and heparin. Synthetic supports were synthesised by free radical polymerization in the form of flat films, prepared by solvent casting, and microspheres, produced via an oil-in-water emulsion, with gelatin or heparin grafted onto theirs surface. The viability and proliferation of primary cells obtained from bone marrow aspirates were analysed in 2D monolayer environments, and in 3D environments using microspheres. In addition, the gene expression of osteogenic and chondrogenic markers was studied in cells expanded in basal medium before reaching confluence in both 2D and 3D environments, to assess the influence of the grafted biomolecules and dimensionality on the early commitment of MSCs. Significant changes were induced by the interaction of cells with the grafted biomolecules, which suggested the commitment of the human bone marrow mesenchymal stem cells (hBMMSCs) to specific lineages and loss of multipotency. The culture was continued in basal or osteogenic medium for up to 21 days in 2D environments after confluence, revealing significant differences between the functionalised supports and either the non-functionalised substrates and standard polystyrene culture wells (tissue culture plastic, TCP). These findings highlight the potential of this approach to advance bone tissue engineering applications and contribute to the development of innovative and effective bone regeneration strategies.

Optimization of Adventitious Shoot Regeneration Protocols for Six Vaccinium corymbosum Cultivars

Limitations in tissue culture and regeneration techniques present significant barriers to the development of improved blueberry (Vaccinium spp.) cultivars, restricting the adoption of tissue culture in commercial production. This challenge emphasizes the urgent need for a standardized regeneration protocol. Addressing this, we developed enhanced in vitro regeneration protocols for six highbush blueberry (Vaccinium corymbosum L.) cultivars—Emerald, NC5288, Rowan (NC3104), Legacy, Pinnacle, and Jewel—each with varying levels of tissue culture recalcitrance. We evaluated the effects of auxins, specifically α-Naphthaleneacetic Acid, cytokinins [including trans-Zeatin (ZT), 1-(2-Chloro-4-pyridyl)-3-phenylurea 98.0+%, TCI America™ (CPPU), and 6-γ-γ-[Dimethylallylamino]-purine (2iP)], and Thidiazuron (TDZ), which exhibits both auxin- and cytokinin-like activities, in various plant species cultured in vitro, on callus induction and regeneration from leaf explants. Media supplemented with ZT + 2iP (PZ9, PZ10) consistently outperformed all other treatments, enhancing regeneration significantly across all cultivars with high genotype-independent reproducibility, particularly in treatment PZ10. In contrast, TDZ-based treatments yielded highly variable results and were ineffective in recalcitrant cultivars such as Jewel and Pinnacle. ZT + CPPU treatments performed moderately across all metrics and may offer utility for cultivars with intermediate responsiveness. These findings provide critical insights into refining tissue culture techniques, offering a foundation for improving blueberry cultivar development and efficient genome editing. This work represents the first published report of in vitro regeneration for the highbush blueberry cultivars Rowan (NC3104), NC5288, and Pinnacle, expanding the scope of possibilities for advancing blueberry crop improvement.

Melatonin supplementation does not improve ovine pre- or post-hatching development in vitro.

Melatonin supplementation does not improve ovine pre- or post-hatching development in vitro.

Peptidomic and proteomic analysis of precision-cut lung slice supernatants.

Peptidomic and proteomic analysis of precision-cut lung slice supernatants.

High-dimensional droplet digital PCR of multiple hepatitis B splice variants in serum, liver, and tissue culture.

High-dimensional droplet digital PCR of multiple hepatitis B splice variants in serum, liver, and tissue culture.

Metagenomic next-generation sequencing (mNGS) versus tissue culture technique (TCT) in diagnosis of spinal infection: a systematic review and meta-analysis

Spinal infections pose a significant clinical challenge due to the difficulty of early diagnosis. Traditional Tissue Culture Technique (TCT) has limitations, while Metagenomic Next-Generation Sequencing (mNGS) has emerged as a promising diagnostic tool. However, reports on the diagnostic performance of mNGS and TCT for spinal infections are inconsistent, and there has been a lack of systematic analysis of the evidence. This systematic review and meta-analysis included 10 studies involving a total of 770 patients to compare the diagnostic accuracy of mNGS and TCT. A comprehensive literature search was conducted using PubMed, Embase, Web of Science, Cochrane Library, and SinoMed to identify studies evaluating the diagnostic accuracy of mNGS and TCT for spinal infections. Data were analyzed using Review Manager 5.3 and Stata 16.0 to compute the sensitivity, specificity, and Area Under the Summary Receiver Operating Characteristic Curve (AUC). The meta-analysis revealed pooled estimates for mNGS with a sensitivity of 0.81 (95% CI, 0.74–0.87), specificity of 0.75 (95% CI, 0.48–0.91), and an AUC of 0.85 (95% CI, 0.82–0.88). In contrast, pooled estimates for TCT showed a sensitivity of 0.34 (95% CI, 0.27–0.43), specificity of 0.93 (95% CI, 0.79–0.98), and an AUC of 0.59 (95% CI, 0.55–0.63). While mNGS demonstrates higher sensitivity and overall diagnostic accuracy than TCT, its lower specificity suggests that it may be most effective when used alongside conventional methods to enhance diagnostic reliability in spinal infections.

Surface-elastic hydrogels delay senescence via the modulation of redox homeostasis and cytoskeletal tension

The Bone marrow-derived mesenchymal stem cells (MSCs) are widely used in clinical applications owing to their therapeutic properties. However, in vitro expansion of MSCs in tissue culture dishes induces aging, which reduces their quality through an undefined mechanism. This study delineates the role of substrate stiffness as a potential modulator to delay MSC aging by elucidating the senescence progression of preconditioned and serially passaged MSCs on engineered stiffness-tunable gelatinous hydrogels. We demonstrated that mechanoactivation of MSCs increased their radical-scavenging capacity, maintained redox homeostasis, restored actin dynamics, and maintained their therapeutic properties. The hydrogels alleviated hydrogen peroxide-induced oxidative stress, linking mechanical signaling to redox balance and senescence. These hydrogels restored actin remodeling, highlighting the importance of cytoskeletal tension and dynamics in cellular senescence. We established a new culture method to maintain the stemness, proliferation, motility, and osteogenic differentiation potential of MSCs by serially passaging the cells on stepwise surface-elastic gels. Evidence points toward the complex interplay between mechanical memories and actin dynamics and their implications for autophagic activity in the delaying of senescent MSCs via hydrogels. Our findings suggest that mechanoregulation of culture substrates finely tunes the balance between cellular stress, redox homeostasis, and cytoskeletal dynamics to delay the progression of MSC senescence.

Effects of serial passaging of field isolates of Bangladeshi PPR virus in Vero cells on the fusion protein

ObjectivesFusion (F) protein is crucial for facilitating viral entry into host cells and contributes to the virulence of Morbilliviruses. Serial passaging of the Peste Des Petits Ruminants virus (PPRV) in nonnative hosts can lead to mutations that potentially reduce pathogenicity. Hence, this study aimed to investigate the effects of serial passaging of a Bangladeshi strain of PPR virus in Vero cells on the Fusion protein and pathogenicityMaterials and methodsPPR viruses were initially isolated from natural PPR outbreaks, confirmed through reverse transcriptase polymerase chain reaction (RT‒PCR), passaged to the 9th passage in Vero cells, sequenced, and preserved in a previous study. The 9th passage virus from the repository was utilized as the viral inoculant for further passaging in Vero cells, and the 60th passage was completed. The presence of PPR viral RNA was confirmed in tissue culture fluid (TCF) by RT‒PCR at different passage numbers. TCF at the 60th passage was sequenced and used for immunogenicity studies via live animal experiments, and subsequent immunity was measured via cELISA.ResultsComparative analysis of the sequences from the 9th and 60th passages, along with other sequences, revealed substitutions of 14 nucleotides (nts) and 4 amino acids (aa) within the leucine zipper structure of the fusion protein. Notably, live animal experiments demonstrated the occurrence of protective immunity.ConclusionThis study suggests that amino acid substitution and genetic divergence may positively affect viral virulence, highlighting their importance in the development of a potent vaccine.

Advances in gene editing-led route for hybrid breeding in crops.

Advances in gene editing-led route for hybrid breeding in crops.

Substrate stiffness modifies gene expression and transcriptional response of equine endometrial fibroblasts to TGF-β1.

Substrate stiffness modifies gene expression and transcriptional response of equine endometrial fibroblasts to TGF-β1.

The Impact of Cytopathogenic and Non-cytopathogenic Biotypes of Bovine Viral Diarrhea Virus on Total Antioxidant Capacity of Bovine Oocytes In-vitro

Background: Bovine viral diarrhea virus (BVDV) is the most common pathogen in dairy cattle herds. Objectives: This study aims to examine the impact of cytopathic (CP) and non-cytopathic (NCP) BVDV biotypes on the total antioxidant capacity (TAC) of bovine oocytes in vitro. Methods: Oocytes were obtained from slaughtered bovine ovaries, washed, and matured in a maturation medium. Oocytes were divided into five groups, each consisting of at least 60. The control group was not exposed to BVDV biotypes. Oocytes were challenged with CP and NCP, BVDV at two concentrations of 104 and 105 tissue culture infectious doses (TCID)50/mL. To determine the antioxidant capacity of oocytes, a TAC assay was conducted after two hours of incubation. Graph Pad Prism software, version 8.4.3 was utilized to analyze the data. A one-way analysis of variance (ANOVA) was performed, and a post-hoc Tukey’s honestly significant difference (HSD) test was accompanied. Results: The results indicated that only the CP biotype of BVDV significantly decreased the TAC of infected oocytes compared to the control group, whereas the NCP biotype did not significantly alter the TAC of the infected groups. Conclusion: Our study showed that only CP BVDV in 104 and 105 TCID50/mL doses affected the infected oocytes and significantly decreased the oocyte’s TAC. At the same time, the NCP biotype did not significantly alter the infected oocyte’s TAC.

Current Progress on Passiflora caerulea L. In Vitro Culturing

Passiflora caerulea L., commonly known as the blue passionflower, is traditionally grown as an ornamental plant, but has a diverse chemical composition resulting in a wide range of biological activities that determine its pharmacological properties and use in medicine. Traditional propagation methods, including seed germination and vegetative cuttings, are often inefficient due to low germination rates, susceptibility to pathogens, and slow growth. In particular, P. caerulea presents significant challenges in germination due to its slow development. In this context, in vitro cultivation is used to enable rapid, large-scale plant production while maintaining genetic fidelity. The study of Passiflora tissue cultures began in 1966 and has since attracted increasing attention from researchers around the world. However, despite growing interest, studies specifically focused on the in vitro propagation of P. caerulea remain limited. This review aims to summarize existing knowledge on the main techniques used for in vitro culturing and propagation of P. caerulea, including organogenesis, somatic embryogenesis, and callogenesis. Particular attention is paid to the key factors that influence the initiation, growth, and regeneration of cultures, including the type of explant, the composition of the media, and the environmental conditions. Advances in the in vitro cultivation of P. caerulea have greatly improved the understanding and propagation of this species. Although in vitro cultivation offers several advantages, it is crucial to conduct thorough research on the selection of explants, their age, and the appropriate culture media to ensure optimal growth and development.

Optimization of micropropagation protocols and assessment of epigenetic changes in tissue cultures of diverse medical cannabis (Cannabis sativa L.) genotypes

Optimization of micropropagation protocols and assessment of epigenetic changes in tissue cultures of diverse medical cannabis (Cannabis sativa L.) genotypes

Black swimming dots in cell culture: first report of Bradyrhizobium as a cause.

Black swimming dots in cell culture: first report of Bradyrhizobium as a cause.

Occurrence of molds (fungi) in indoor tissue culture laboratories

Laboratory indoor environments play important roles in human health and laboratory experiments. Contamination wastes time and resources as well as posing serious risks to human health due to toxicity. Fungi (molds), like other microorganisms are ubiquitous in distribution and in complexity towards causing human diseases. However, the awareness of the number and nature of molds in any giving laboratory per time will help to better strategize their control mechanisms. The aim of this study was to evaluate some tissue culture laboratory surfaces and indoor air for fungal contamination in some selected Laboratories in the Federal Capital territory (FCT), Abuja, Nigeria. Using swab and air sampling procedures, fungi were isolated from plantlets, tables, wall and the air of the laboratory rooms of fifteen tissue culture laboratories and cultured on Streptomycin-supplemented Potato Dextrose Agar (PDA). The discrete colonies that developed were identified using morphological and microscopical characterization. The fungal species isolated from these laboratories include; Aspergillus niger, Alterneria sp., Aspergillus terreus, Aspergillus flavus, Cladosporium sp., Rhizopus sp., Penicillium sp., Fusarium sp. and Geotrichum sp. The results shown that most of the laboratories were highly contaminated and that Aspergillus was the predominant fungal genera and A. niger recorded the highest overal occurence. Since using plant tissue culture techniques holds promise in substantially augmenting the number of novel cultivars and genotypes of many fruit crops, adhering to standard operating procedures is highly recommended in the studied areas and Nigeria as a whole.

Unraveling the Photoregulatory Mechanisms of Capsaicinoids Biosynthesis and Accumulation of Capsaicinoids in Capsicum annuum InVitro Cultures.

Light is one of the essential environmental factors that significantly influences the biosynthesis of secondary metabolites in plants. To explicate their effect, exposure of monochromatic LED light spectra (yellow, blue, green, red, and white) on the accumulation of capsaicinoids, phenolic compounds, antioxidant capacity, and regulation of transcriptional changes in capsaicinoids biosynthesis genes at 24, 48, and 72 h in chili callus cultures was investigated. Blue light significantly increased capsaicin and dihydrocapsaicin contents by 41.43-fold and 4.36-fold, respectively, compared to white light. Pearson correlation analysis confirmed a strong positive correlation between total phenolics, total flavonoids, and antioxidant capacity (r > 0.85). Gene expression analysis revealed that the biosynthetic genes (pAMT, KAS, ACS, ACL, CL, C4H, and CS) were upregulated, and their associated transcription factors-MYB, ERF, and JERF-showed varied expression, indicating their putative regulatory role in capsaicinoid biosynthesis under different monochromatic LED conditions. Our findings demonstrate that the targeted monochromatic LED lights can enhance bioactive compound production in C. annuum callus cultures, providing a favorable strategy for feasible production of the industrially important compound "capsaicin" on a large scale for pharmaceutical and food applications.

Antimicrobial, Antibiofilm, and Antioxidant Activity of Bacterial Cellulose and Bacterial Nanocellulose from Vinegar’s Mother Pellicles

Recent research indicates that bacterial cellulose (BC) and bacterial nanocellulose (BNC) exhibit promising adsorptive properties for antimicrobial agents, owing to their complex network structure, high surface area, and substantial porosity. The aim of this study was to analyze the impact of the antimicrobial, antibiofilm, activity of BC and BNC on different bacterial isolates, were collected from wounds and burn wound patients. Also, a study examined the antioxidant activity. In this study, (BC) and (BNC) was produced from vingar mothers. The Pathogenic bacterial isolates were collected from wounds and burn wound patients of different age groups. The antibiotic susceptibility testing by disk diffusion method and Vitek 2 compact system method. The antimicrobial activity of BC and BNC on different bacterial isolates by the Resazurin-based 96-well plate microdilution method. Detection of biofilm formation and Inhibition of Biofilm Formation was quantified using the 96-well tissue culture plate method (TCP). Finally, the DPPH radical scavenging assay was used to investigate the ability of (BC) and (BNC) to scavenge free radicals. Seventy bacterial isolates were collected from wounds and burn wound patients of different age groups. It belongs to five species: Citrobacter freundii, ten isolates; Escherichia coli, 17 isolates; Klebsiella pneumonia, 16 isolates; Pseudomonas aeruginosa, 15 isolates and Staphylococcus aureus, 20 isolates. The Antibiotic susceptibility testing by disk diffusion method and Vitek 2 compact system method showed that most bacterial isolates were resistant to the antibiotics used. The antimicrobial activity of BC and BNC on different bacterial isolates by the Resazurin-based 96-well plate microdilution method showed different results in tests bacteria using the minimum inhibitory concentration (MIC). The study found that 60 out of the 70 bacterial isolates were able to produce biofilm, accounting for 85.71% of isolates, while 10 isolates (14.28%) did not produce biofilm. The effect of the BC and BNC on the biofilm formation using SUB-MIC 1 shows that BC and BNC prevent the biofilm formation in 17 (78%) and 25 (100%) isolates. Meanwhile, SUB-MIC 2 of BC and BNC prevent biofilm formation in 10 (42%) and 23 (80%) isolates. The results show that the % PDDH Radical Scavenging Activity for BC was (10.316 and 75.122) when the concentrations were (1.95 and 1000) µg/ml, respectively. The % PDDH Radical Scavenging Activity for BNC was (14.806 and 98.837) when the concentrations were (1.95 and 1000) µg/ml respectively. The % PDDH Radical Scavenging Activity for AA was (16.213 and 98.940) when the concentrations were (1.95 and 1000) µg/ml respectively. The bacterial cellulose and bacterial nanocellulose are characterized by their antimicrobial, antibiofilm, and antioxidant properties, which allow for a wide range of medical, industrial, and environmental applications.