Overexpression of the limk1 Gene in Drosophila melanogaster Can Lead to Suppression of Courtship Memory in Males.

Courtship suppression is a behavioral adaptation of the fruit fly. When majority of the females in a fly population are fertilized and non-receptive for mating, a male, after a series of failed attempts, decreases its courtship activity towards all females, saving its energy and reproductive resources. The time of courtship decrease depends on both duration of unsuccessful courtship and genetically determined features of the male nervous system. Thereby, courtship suppression paradigm can be used for studying molecular mechanisms of learning and memory. p-Cofilin, a component of the actin remodeling signaling cascade and product of LIM-kinase 1 (LIMK1), regulates Drosophila melanogaster forgetting in olfactory learning paradigm. Previously, we have shown that limk1 suppression in the specific types of nervous cells differently affects fly courtship memory. Here, we used Gal4 > UAS system to induce limk1 overexpression in the same types of neurons. limk1 activation in the mushroom body, glia, and fruitless neurons decreased learning index compared to the control strain or the strain with limk1 knockdown. In cholinergic and dopaminergic/serotoninergic neurons, both overexpression and knockdown of limk1 impaired Drosophila short-term memory. Thus, proper balance of the limk1 activity is crucial for normal cognitive activity of the fruit fly.

Karyopherin α2 is a maternal effect gene required for early embryonic development and female fertility in mice.

The nuclear transport of proteins plays an important role in mediating the transition from egg to embryo and distinct karyopherins have been implicated in this process. Here, we studied the impact of KPNA2 deficiency on preimplantation embryo development in mice. Loss of KPNA2 results in complete arrest at the 2cell stage and embryos exhibit the inability to activate their embryonic genome as well as a severely disturbed nuclear translocation of Nucleoplasmin 2. Our findings define KPNA2 as a new maternal effect gene.

Suspension cell cultures of Panax vietnamensis as a biotechnological source of ginsenosides: growth, cytology, and ginsenoside profile assessment.

Introduction: Panax vietnamensis is a valuable medicinal plant and a source of a broad spectrum of biologically active ginsenosides of different structural groups. Overexploitation and low adaptability to planation cultivation have made this species vulnerable to human pressure and prompted the development of cell cultivation in vitro as a sustainable alternative to harvesting wild plants for their bioactive components. Despite high interest in biotechnological production, little is known about the main factors affecting cell growth and ginsenoside biosynthesis of this species under in vitro conditions. In this study, the potential of cell cultures of P. vietnamensis as a biotechnological source of ginsenosides was was assessed. Methods: Six suspension cell lines that were developed from different sections of a single rhizome through a multi-step culture optimization process and maintained for over 3 years on media with different mineral salt base and varying contents of auxins and cytokinins. These cell lines were evaluated for productivity parameters and cytological characteristics. Ginsenoside profiles were assessed using a combination of the reversed-phase ultra-high-performance liquid chromatography-Orbitrap-tandem mass spectrometry (UHPLC-Orbitrap-MS/MS) and ultra-performance liquid chromatography-time of flight-mass spectrometry (UPLC-TOF-MS). Results: All lines demonstrated good growth with a specific growth rate of 0.1-0.2 day-1, economic coefficient of 0.31-0.70, productivity on dry weight (DW) of 0.30-0.83 gDW (L·day)-1, and maximum biomass accumulation varying from 10 to 22 gDW L-1. Ginsenosides of the protopanaxadiol (Rb1, Rb2/Rb3, malonyl-Rb1, and malonyl-Rb2/Rb3), oleanolic acid (R0 and chikusetsusaponin IV), and ocotillol (vinaginsenoside R1) groups and their isomers were identified in cell biomass extracts. Chikusetsusaponin IV was identified in P. vietnamensis cell culture for the first time. Discussion: These results suggest that suspension cell cultures of Vietnamese ginseng have a high potential for the biotechnological production of biomass containing ginsenosides, particularly of the oleanolic acid and ocotillol groups.

Bioreactor Systems for Plant Cell Cultivation at the Institute of Plant Physiology of the Russian Academy of Sciences: 50 Years of Technology Evolution from Laboratory to Industrial Implications.

The cultivation of plant cells in large-scale bioreactor systems has long been considered a promising alternative for the overexploitation of wild plants as a source of bioactive phytochemicals. This idea, however, faced multiple constraints upon realization, resulting in very few examples of technologically feasible and economically effective biotechnological companies. The bioreactor cultivation of plant cells is challenging. Even well-growing and highly biosynthetically potent cell lines require a thorough optimization of cultivation parameters when upscaling the cultivation process from laboratory to industrial volumes. The optimization includes, but is not limited to, the bioreactor's shape and design, cultivation regime (batch, fed-batch, continuous, semi-continuous), aeration, homogenization, anti-foaming measures, etc., while maintaining a high biomass and metabolite production. Based on the literature data and our experience, the cell cultures often demonstrate cell line- or species-specific responses to parameter changes, with the dissolved oxygen concentration (pO2) and shear stress caused by stirring being frequent growth-limiting factors. The mass transfer coefficient also plays a vital role in upscaling the cultivation process from smaller to larger volumes. The Experimental Biotechnological Facility at the K.A. Timiryazev Institute of Plant Physiology has operated since the 1970s and currently hosts a cascade of bioreactors from the laboratory (20 L) to the pilot (75 L) and a semi-industrial volume (630 L) adapted for the cultivation of plant cells. In this review, we discuss the most appealing cases of the cell cultivation process's adaptation to bioreactor conditions featuring the cell cultures of medicinal plants Dioscorea deltoidea Wall. ex Griseb., Taxus wallichiana Zucc., Stephania glabra (Roxb.) Miers, Panax japonicus (T. Nees) C.A.Mey., Polyscias filicifolia (C. Moore ex E. Fourn.) L.H. Bailey, and P. fruticosa L. Harms. The results of cell cultivation in bioreactors of different types and designs using various cultivation regimes are covered and compared with the literature data. We also discuss the role of the critical factors affecting cell behavior in bioreactors with large volumes.

Plant Cryopreservation: Principles, Applications, and Challenges of Banking Plant Diversity At Ultralow Temperatures.

Progressive loss of plant diversity requires the protection of wild and agri-/horticultural species. For species whose seeds are extremely short-lived, or rarely or never produce seeds, or whose genetic makeup must be preserved, cryopreservation offers the only possibility for long-term conservation. At temperatures below freezing, most vegetative plant tissues suffer severe damage from ice crystal formation and require protection. In this review, we describe how increasing the concentration of cellular solutes by air drying or adding cryoprotectants, together with rapid cooling, results in a vitrified, highly viscous state in which cells can remain viable and be stored. On this basis, a range of dormant bud-freezing, slow-cooling, and (droplet-)vitrification protocols have been developed, but few are used to cryobank important agricultural/horticultural/timber and threatened species. To improve cryopreservation efficiency, the effects of cryoprotectants and molecular processes need to be understood and the costs for cryobanking reduced. However, overall, the long-term costs of cryopreservation are low, while the benefits are huge. Expected final online publication date for the Annual Review of Plant Biology, Volume 75 is May 2024. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

Effect of Phytic Acid Addition on the Structure of Collagen-Hyaluronic Acid Composite Gel.

Composite collagen gels with hyaluronic acid are developed tissue-engineered structures for filling and regeneration of defects in various organs and tissues. For the first time, phytic acid was used to increase the stability and improve the mechanical properties of collagen gels with hyaluronic acid. Phytic acid is a promising cross-linker for collagen hydrogels and is a plant-derived antioxidant found in rich sources of beans, grains, and oilseeds. Phytic acid has several benefits due to its antioxidant, anticancer, and antitumor properties. In this work, studies were carried out on the kinetics of the self-assembly of collagen molecules in the presence of phytic and hyaluronic acids. It was shown that both of these acids do not lead to collagen self-assembly. Scanning electron microscopy showed that in the presence of phytic and hyaluronic acids, the collagen fibrils had a native structure, and the FTIR method confirmed the chemical cross-links between the collagen fibrils. DSC and rheological studies demonstrated that adding the phytic acid improved the stability and modulus of elasticity of the collagen gel. The presence of hyaluronic acid in the collagen gel slightly reduced the effect of phytic acid. The presence of phytic acid in the collagen gel improved the stability of the scaffold, but, after 1 week of cultivation, slightly reduced the viability of mesenchymal stromal cells cultured in the gel. The collagen type I gel with hyaluronic and phytic acids can be used to replace tissue defects, especially after the removal of cancerous tumors.

Dynamics of Organic Acids during the Droplet-Vitrification Cryopreservation Procedure Can Be a Signature of Oxidative Stress in Pogostemon yatabeanus.

Cryopreservation in liquid nitrogen (LN, -196 °C) is a unique option for the long-term conservation of threatened plant species with non-orthodox or limitedly available seeds. In previous studies, a systematic approach was used to develop a droplet-vitrification (DV) cryopreservation protocol for Postemon yatabeanus shoot tips that includes preculture with 10% sucrose, osmoprotection with C4-35%, cryoprotection with A3-80% vitrification solution, and a three-step regrowth starting with the ammonium-free medium. The tricarboxylic acid (TCA) cycle is a crucial component of plant cell metabolism as it is involved in redox state regulation and energy provision. We hypothesized that organic acids (OAs) associated with the TCA and its side reactions indirectly indicate metabolism intensity and oxidative stress development in shoot tips under the cryopreservation procedure. In this study, the contents of 14 OAs were analyzed using gas chromatography-tandem mass spectrometry (GC-MS/MS) in P. yatabeanus shoot tips in a series of treatments including individual steps of the DV procedure, additional stress imposed by non-optimum protocol conditions (no preculture, no osmoprotection, various vitrification solution composition, using vials instead of aluminum foils, etc.) and regrowth on different media with or without ammonium or growth regulators. The possible relation of OA content with the total cryoprotectant (CPA) concentration and shoot tips regeneration percentage was also explored. Regeneration of cryopreserved shoot tips reduced in descending order as follows: standard protocol condition (91%) > non-optimum vitrification solution (ca. 68%) > non-optimum preculture (60-62%) > regrowth medium (40-64%) > no osmoprotection, cryopreservation in vials (28-30%). Five OAs (glycolic, malic, citric, malonic, and lactic) were the most abundant in the fresh (control) shoot tips. The dynamic pattern of OAs during the DV procedure highly correlated (r = 0.951) with the total CPA concentration employed: it gradually increased through the preculture, osmoprotection, and cryoprotection, peaked at cooling/rewarming (6.38-fold above control level), and returned to the fresh control level after 5 days of regrowth (0.89-fold). The contents of four OAs (2-hydroxybutyric, 3-hydroxypropionic, lactic, and glycolic) showed the most significant (10-209-fold) increase at the cooling/rewarming step. Lactic and glycolic acids were the major OAs at cooling/rewarming, accounting for 81% of the total OAs content. The OAs were categorized into three groups based on their dynamics during the cryopreservation protocol, and these groups were differently affected by protocol step modifications. However, there was no straightforward relationship between the dynamics of OAs and shoot tip regeneration. The results suggest that active modulation of OAs metabolism may help shoot tips to cope with osmotic stress and the chemical cytotoxicity\ of CPAs. Further intensive studies are needed to investigate the effect of cryopreservation on cell primarily metabolism and identify oxidative stress-related biomarkers in plant materials.

Suspension Cell Culture of Polyscias fruticosa (L.) Harms in Bubble-Type Bioreactors-Growth Characteristics, Triterpene Glycosides Accumulation and Biological Activity.

Polyscias fruticosa (L.) Harms, or Ming aralia, is a medicinal plant of the Araliaceae family, which is highly valued for its antitoxic, anti-inflammatory, analgesic, antibacterial, anti-asthmatic, adaptogenic, and other properties. The plant can be potentially used to treat diabetes and its complications, ischemic brain damage, and Parkinson's disease. Triterpene glycosides of the oleanane type, such as 3-O-[ß-D-glucopyranosyl-(1→4)-ß-D-glucuronopyranosyl] oleanolic acid 28-O-ß-D-glucopyranosyl ester (PFS), ladyginoside A, and polysciosides A-H, are mainly responsible for biological activities of this species. In this study, cultivation of the cell suspension of P. fruticosa in 20 L bubble-type bioreactors was attempted as a sustainable method for cell biomass production of this valuable species and an alternative to overexploitation of wild plant resources. Cell suspension cultivated in bioreactors under a semi-continuous regime demonstrated satisfactory growth with a specific growth rate of 0.11 day-1, productivity of 0.32 g (L · day)-1, and an economic coefficient of 0.16 but slightly lower maximum biomass accumulation (~6.8 g L-1) compared to flask culture (~8.2 g L-1). Triterpene glycosides PFS (0.91 mg gDW-1) and ladyginoside A (0.77 mg gDW-1) were detected in bioreactor-produced cell biomass in higher concentrations compared to cells grown in flasks (0.50 and 0.22 mg gDW-1, respectively). In antibacterial tests, the minimum inhibitory concentrations (MICs) of cell biomass extracts against the most common pathogens Staphylococcus aureus, methicillin-resistant strain MRSA, Pseudomonas aeruginosa, and Escherichia coli varied within 250-2000 µg mL-1 which was higher compared to extracts of greenhouse plant leaves (MIC = 4000 µg mL-1). Cell biomass extracts also exhibited antioxidant activity, as confirmed by DPPH and TEAC assays. Our results suggest that bioreactor cultivation of P. fruticosa suspension cell culture may be a perspective method for the sustainable biomass production of this species.

Effect of Hydroxyl-Containing Fragments on the Structure and Properties of Membrane-Forming Polyamide-Imides.

The structural features and thermophysical and transport properties of dense nonporous membranes of the casting type from (co)polyamide-imides synthesized by the polycondensation of the diacid chloride of 2-(4-carboxyphenyl)-1,3-dioxoisoindoline-5-carboxylic acid and diamines 5,5'-methylene-bis (2-aminophenol) (DADHyDPhM) and 4,4'-methylenebis(benzeneamine) (DADPhM), taken in molar ratios of 7:3, 1:1, and 3:7, have been studied. The effect of hydroxyl-containing modifying fragments of dihydroxy diphenylmethane introduced in various amounts into the main polymer chain on the pervaporation properties of the formed films is discussed. It has been shown that the presence of the residual solvent N-methyl-2-pyrrolidone in the films not only has a plasticizing effect on the characteristics of film membranes but also promotes the preferential transmembrane transport of polar liquids, primarily methanol (permeation rate over 2 kg for a copolymer with a ratio of DADHyDPhM:DADPhM = 7:3). The removal of the residual solvent from the polymer film, both thermally (heating to 200 °C) and by displacement with another solvent as a result of sequential pervaporation, led to a significant decrease in the rate of transfer of polar liquids and a decrease in the selectivity of the membrane. However, the dehydrocyclization reaction resulted in more brittle films with low permeability to penetrants of different polarities. The results of our comprehensive study made it possible to assume the decisive influence of structural changes in membranes occurring in connection with the competitive formation of intra- and intermolecular hydrogen bonds.

Characteristic Length for Pinning Force Density in Nb3Sn.

The pinning force density, Fp, is one of the main parameters that characterize the resilience of a superconductor to carrying a dissipative-free transport current in an applied magnetic field. Kramer (1973) and Dew-Hughes (1974) proposed a widely used scaling law for this quantity, where one of the parameters is the pinning force density maximum, Fp,max, which represents the maximal performance of a given superconductor in an applied magnetic field at a given temperature. Since the late 1970s to the present, several research groups have reported experimental data on the dependence of Fp,max on the average grain size, d, in Nb3Sn-based conductors. Fp,maxd datasets were analyzed and a scaling law for the dependence Fp,maxd=A×ln1/d+B was proposed. Despite the fact that this scaling law is widely accepted, it has several problems; for instance, according to this law, at T=4.2 K and d≥650 nm, Nb3Sn should lose its superconductivity, which is in striking contrast to experiments. Here, we reanalyzed the full inventory of publicly available Fp,maxd data for Nb3Sn conductors and found that the dependence can be described by the exponential law, in which the characteristic length, δ, varies within a remarkably narrow range of δ=175±13 nm for samples fabricated using different technologies. The interpretation of this result is based on the idea that the in-field supercurrent flows within a thin surface layer (thickness of δ) near grain boundary surfaces (similar to London's law, where the self-field supercurrent flows within a thin surface layer with a thickness of the London penetration depth, λ, and the surface is a superconductor-vacuum surface). An alternative interpretation is that δ represents the characteristic length of the exponential decay flux pinning potential from the dominant defects in Nb3Sn superconductors, which are grain boundaries.

3'UTR of mRNA Encoding CPEB Protein Orb2 Plays an Essential Role in Intracellular Transport in Neurons.

Intracellular trafficking plays a critical role in the functioning of highly polarized cells, such as neurons. Transport of mRNAs, proteins, and other molecules to synaptic terminals maintains contact between neurons and ensures the transmission of nerve impulses. Cytoplasmic polyadenylation element binding (CPEB) proteins play an essential role in long-term memory (LTM) formation by regulating local translation in synapses. Here, we show that the 3'UTR of the Drosophila CPEB gene orb2 is required for targeting the orb2 mRNA and protein to synapses and that this localization is important for LTM formation. When the orb2 3'UTR is deleted, the orb2 mRNAs and proteins fail to localize in synaptic fractions, and pronounced LTM deficits arise. We found that the phenotypic effects of the orb2 3'UTR deletion were rescued by introducing the 3'UTR from the orb, another Drosophila CPEB gene. In contrast, the phenotypic effects of the 3'UTR deletion were not rescued by the 3'UTR from one of the Drosophila α-tubulin genes. Our results show that the orb2 mRNAs must be targeted to the correct locations in neurons and that proper targeting depends upon sequences in the 3'UTR.

Influence of Molecular Weight on Thermal and Mechanical Properties of Carbon-Fiber-Reinforced Plastics Based on Thermoplastic Partially Crystalline Polyimide.

For the first time, a study of the influence of the molecular weight of the thermoplastic partially crystalline polyimide R-BAPB on the thermophysical and mechanical properties of carbon plastics was presented. The molecular weight of polyimide was determined using the method of light scattering and the study of the intrinsic viscosity of polyamic acid solutions. To obtain CFRPs, the uniform distribution of polyimide powder on continuous carbon fibers via electrostatic spraying and further hot calendering and pressing were applied. The study of the structure of the obtained carbon plastics via scanning electron microscopy has shown that the growth of the molecular weight of polyimide prevents the impregnation of carbon fiber with the introduced polyimide. Moreover, an increase in the molecular weight of polyimide leads to a rise in glass transition and thermal decomposition temperatures up to 590 °C, while the degree of crystallinity of CFRP falls. Nonetheless, raising the molecular weight from 22,000 to 70,000 g/mol of a binder polymer improves the interlayer fracture toughness G1C by more than five times.

Plants, Cells, Algae, and Cyanobacteria In Vitro and Cryobank Collections at the Institute of Plant Physiology, Russian Academy of Sciences-A Platform for Research and Production Center.

Ex situ collections of algae, cyanobacteria, and plant materials (cell cultures, hairy and adventitious root cultures, shoots, etc.) maintained in vitro or in liquid nitrogen (-196 °C, LN) are valuable sources of strains with unique ecological and biotechnological traits. Such collections play a vital role in bioresource conservation, science, and industry development but are rarely covered in publications. Here, we provide an overview of five genetic collections maintained at the Institute of Plant Physiology of the Russian Academy of Sciences (IPPRAS) since the 1950-1970s using in vitro and cryopreservation approaches. These collections represent different levels of plant organization, from individual cells (cell culture collection) to organs (hairy and adventitious root cultures, shoot apices) to in vitro plants. The total collection holdings comprise more than 430 strains of algae and cyanobacteria, over 200 potato clones, 117 cell cultures, and 50 strains of hairy and adventitious root cultures of medicinal and model plant species. The IPPRAS plant cryobank preserves in LN over 1000 specimens of in vitro cultures and seeds of wild and cultivated plants belonging to 457 species and 74 families. Several algae and plant cell culture strains have been adapted for cultivation in bioreactors from laboratory (5-20-L) to pilot (75-L) to semi-industrial (150-630-L) scale for the production of biomass with high nutritive or pharmacological value. Some of the strains with proven biological activities are currently used to produce cosmetics and food supplements. Here, we provide an overview of the current collections' composition and major activities, their use in research, biotechnology, and commercial application. We also highlight the most interesting studies performed with collection strains and discuss strategies for the collections' future development and exploitation in view of current trends in biotechnology and genetic resources conservation.

Effect of Salinity Stress on Phenolic Compounds and Antioxidant Activity in Halophytes Spergularia marina (L.) Griseb. and Glaux maritima L. Cultured In Vitro.

The study of halophytes as sources of phenolic compounds, as well as conditions that further enhance the accumulation of biologically active compounds in them, is of particular interest. In this paper, the effect of different salinity levels (25-500 mM in the form of NaCl) on the content of phenolic compounds and the antioxidant activity of two rare halophyte species Spergularia marina (L.) Griseb. and Glaux maritima L. cultured in vitro was investigated. A species-specific reaction of plants to salinization was established. In G. maritima, the maximum total content of phenolic compounds was observed at 50-100 mM, flavonoids 75-400 mM, and hydroxycinnamic acids 200-300 mM, as well as individual phenolics (protocatechuic acid, catechin, astragalin, hyperoside, rutin, isoquercitrin, and apigenin derivative) at 100-300 mM NaCl. For S. marina, on the contrary, there was a slight decrease in the content of phenolic compounds when NaCl was added to the nutrient medium compared to the control. The content of protocatechuic acid, rosmarinic acid, and apigenin derivative significantly decreased with increased salt stress. The change in antioxidant activity at different salinity levels was also species specific. The maximum values of different groups of phenolic compounds in G. maritima were observed at 50-300 mM NaCl. The cultivation of S. marina without the addition of NaCl and at 500 mM NaCl allowed the production of plants with the highest content of phenolic compounds. The obtained results can be further used in the development of protocols for the cultivation of these plants in vitro in order to induce the biosynthesis of phenolic compounds in them.

Transgenic angiotensin-converting enzyme 2 overexpression in the rat vasculature protects kidneys from ageing-induced injury.

Chronic kidney disease is one of the leading causes of morbidity and mortality especially among the aged population. A decline in kidney function with ageing comparable to ageing-related processes in human kidneys has also been described in Sprague-Dawley (SD) rats. The renin-angiotensin-system (RAS) plays a pivotal role in the pathophysiology of cardiovascular and kidney disease and is a successful therapeutic target. The discovery of angiotensin-(1-7) (Ang(1-7)), mainly produced by angiotensin-converting enzyme 2 (ACE2), and its receptor MAS offered a new view on the RAS. This ACE2/Ang(1-7)/MAS axis counteracts most deleterious actions of the RAS in the kidney. In order to evaluate if activation of this axis has a protective effect in ageing-induced kidney disease we generated a transgenic rat model (TGR(SM22hACE2)) overexpressing human ACE2 in vascular smooth muscle cells. These animals showed a specific transgene expression pattern and increased ACE2 activity in the kidney. Telemetric recording of cardiovascular parameters and evaluation of kidney function by histology and urine analysis revealed no alterations in blood pressure regulation and basal kidney function in young transgenic rats when compared to young SD rats. However, with ageing, SD rats developed a decline in kidney function characterized by severe albuminuria which was significantly less pronounced in TGR(SM22hACE2) rats. Concomitantly, we detected lower mRNA expression levels of kidney damage markers in aged transgenic animals. Thus, our results indicate that vascular ACE2-overexpression protects the kidney against ageing-induced decline in kidney function, supporting the kidney-protective role of the ACE2/Ang(1-7)/MAS axis.

Critical Role of Regrowth Conditions in Post-Cryopreservation of In Vitro Plant Germplasm.

Cryopreservation is an effective option for the long-term conservation of plant genetic resources, including vegetatively propagated crops and ornamental plants, elite tree genotypes, threatened plant species with non-orthodox seeds or limited seed availability, as well as cell and root cultures useful for biotechnology. With increasing success, an arsenal of cryopreservation methods has been developed and applied to many species and material types. However, severe damage to plant material accumulating during the multi-step cryopreservation procedure often causes reduced survival and low regrowth, even when the optimized protocol is applied. The conditions at the recovery stage play a vital role in supporting material regrowth after cryopreservation and, when optimized, may shift the life-and-death balance toward a positive outcome. In this contribution, we provide an overview of the five main strategies available at the recovery stage to improve post-cryopreservation survival of in vitro plant materials and their further proliferation and development. In particular, we discuss the modification of the recovery medium composition (iron- and ammonium-free), exogenous additives to cope with oxidative stress and absorb toxic chemicals, and the modulation of medium osmotic potential. Special attention is paid to plant growth regulators used at various steps of the recovery process to induce the desired morphological response in cryopreserved tissues. Given studies on electron transport and energy provision in rewarmed materials, we discuss the effects of light-and-dark conditions and light quality. We hope that this summary provides a helpful guideline and a set of references for choosing the recovery conditions for plant species that have not been cryopreserved. We also propose that step-wise recovery may be most effective for materials sensitive to cryopreservation-induced osmotic and chemical stresses.

Sustainable Production of Ajuga Bioactive Metabolites Using Cell Culture Technologies: A Review.

The genus Ajuga (Lamiaceae) is rich in medicinally important species with biological activities ranging from anti-inflammatory, antitumor, neuroprotective, and antidiabetic to antibacterial, antiviral, cytotoxic, and insecticidal effects. Every species contains a unique and complex mixture of bioactive metabolites-phytoecdysteroids (PEs), iridoid glycosides, withanolides, neo-clerodane terpenoids, flavonoids, phenolics, and other chemicals with high therapeutic potential. Phytoecdysteroids, the main compounds of interest, are natural anabolic and adaptogenic agents that are widely used as components of dietary supplements. Wild plants remain the main source of Ajuga bioactive metabolites, particularly PEs, which leads to frequent overexploitation of their natural resources. Cell culture biotechnologies offer a sustainable approach to the production of vegetative biomass and individual phytochemicals specific for Ajuga genus. Cell cultures developed from eight Ajuga taxa were capable of producing PEs, a variety of phenolics and flavonoids, anthocyanins, volatile compounds, phenyletanoid glycosides, iridoids, and fatty acids, and demonstrated antioxidant, antimicrobial, and anti-inflammatory activities. The most abundant PEs in the cell cultures was 20-hydroxyecdysone, followed by turkesterone and cyasterone. The PE content in the cell cultures was comparable or higher than in wild or greenhouse plants, in vitro-grown shoots, and root cultures. Elicitation with methyl jasmonate (50-125 µM) or mevalonate and induced mutagenesis were the most effective strategies that stimulated cell culture biosynthetic capacity. This review summarizes the current progress in cell culture application for the production of pharmacologically important Ajuga metabolites, discusses various approaches to improve the compound yield, and highlights the potential directions for future interventions.

The Hypoglycemic and Hypocholesterolemic Activity of Dioscorea deltoidea, Tribulus terrestris and Panax japonicus Cell Culture Biomass in Rats with High-Fat Diet-Induced Obesity.

Obesity, and its consequences for human health, is a huge and complicated problem that has no simple solution. The constant search for natural and safe compounds with systemic action that can be used for obesity prophylactics and treatment is hampered by the limited availability and variable quality of biomass of wild medicinal plants. Plant cell biotechnology is an alternative approach for the sustainable production of vegetative biomass or individual phytochemicals with high therapeutic potential. In this study, the suspension cell biomass of the medicinal plants, Dioscorea deltoidea Wall., Tribulus terrestris L., and Panax japonicus (T. Nees) C.A. Mey, produced in 20 L and 630 L bioreactors, were tested for therapeutic effects in rat models with alimentary-induced obesity. Three-month intake of water infusions of dry cell biomass (100 mg/g body weight) against the background of a hypercaloric diet reduced weight gain and the proportion of fat mass in the obese animals. In addition, cell biomass preparation reduced the intracellular dehydration and balanced the amounts of intra- and extracellular fluids in the body as determined by bioimpedance spectroscopy. A significant decrease in the glucose and cholesterol levels in the blood was also observed as a result of cell biomass administration for all species. Hypocholesterolemic activity reduced in the line P. japonicus > D. deltoidea > T. terrestris/liraglutide > intact group > control group. By the sum of parameters tested, the cell culture of D. deltoidea was considered the most effective in mitigating diet-induced obesity, with positive effects sometimes exceeding those of the reference drug liraglutide. A safety assessment of D. deltoidea cell phytopreparation showed no toxic effect on the reproductive function of the animals and their offspring. These results support the potential application of the biotechnologically produced cell biomass of medicinal plant species as safe and effective natural remedies for the treatment of obesity and related complications, particularly for the long-term treatment and during pregnancy and lactation periods when conventional treatment is often contraindicated.

Long-Term Memory Formation in Drosophila Depends on the 3'UTR of CPEB Gene orb2.

Activation of local translation in neurites in response to stimulation is an important step in the formation of long-term memory (LTM). CPEB proteins are a family of translation factors involved in LTM formation. The Drosophila CPEB protein Orb2 plays an important role in the development and function of the nervous system. Mutations of the coding region of the orb2 gene have previously been shown to impair LTM formation. We found that a deletion of the 3'UTR of the orb2 gene similarly results in loss of LTM in Drosophila. As a result of the deletion, the content of the Orb2 protein remained the same in the neuron soma, but significantly decreased in synapses. Using RNA immunoprecipitation followed by high-throughput sequencing, we detected more than 6000 potential Orb2 mRNA targets expressed in the Drosophila brain. Importantly, deletion of the 3'UTR of orb2 mRNA also affected the localization of the Csp, Pyd, and Eya proteins, which are encoded by putative mRNA targets of Orb2. Therefore, the 3'UTR of the orb2 mRNA is important for the proper localization of Orb2 and other proteins in synapses of neurons and the brain as a whole, providing a molecular basis for LTM formation.

Effect of Phase Heterogeneity on the Properties of Poly(vinyl alcohol)-Based Composite Pervaporation Membranes.

The structure, thermophysical characteristics, and pervaporation properties of composite membranes based on poly(vinyl alcohol) (PVA) are studied in dependence of the film preparation conditions. It is shown that the nature of the supramolecular organization of the composite polymer film determines which of the components of the separated mixtures of toluene and heptane predominantly penetrate through the corresponding pervaporation membrane. The observed structural effects can become more pronounced if the second component of a polymer mixture is purposefully selected (in this case, poly(N,N-dimethylaminoethyl methacrylate) instead of poly(acrylic acid)) or a nano-sized filler that can be well dispersed in the polymer matrix is introduced. Multi-wall carbon nanotubes are introduced into binary PVA-containing polymer blends. The influence of these fillers on the structure and transport properties of the obtained membranes is studied.