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RSC Medicinal Chemistry Sep 2023Cell therapies such as allogenic CAR T-cell therapy, natural killer cell therapy and stem cell transplants must be cryopreserved for transport and storage. This is...
Cell therapies such as allogenic CAR T-cell therapy, natural killer cell therapy and stem cell transplants must be cryopreserved for transport and storage. This is typically achieved by addition of dimethyl sulfoxide (DMSO) but the cryoprotectant does not result in 100% cell recovery. New additives or technologies to improve their cryopreservation could have major impact for these emerging therapies. l-Proline is an amino acid osmolyte produced as a cryoprotectant by several organisms such as the codling moth and the larvae of the fly , and has been found to modulate post-thaw outcomes for several cell lines but has not been studied with Jurkat cells, a T lymphocyte cell line. Here we investigate the effectiveness of l-proline compared to d-proline and l-alanine for the cryopreservation of Jurkat cells. It is shown that 24-hour pre-freezing incubation of Jurkat cells with 200 mM l-proline resulted in a modest increase in cell recovery post-thaw at high cell density, but a larger increase in recovery was observed at the lower cell densities. l-Alanine was as effective as l-proline at lower cell densities, and addition of l-proline to the cryopreservation media (without incubation) had no benefit. The pre-freeze incubation with l-proline led to significant reductions in cell proliferation supporting an intracellular, biochemical, mechanism of action which was shown to be cell-density dependent. Controls with d-proline were found to reduce post-thaw recovery attributed to osmotic stress as d-proline cannot enter the cells. Preliminary analysis of apoptosis/necrosis profiles by flow cytometry indicated that inhibition of apoptosis is not the primary mode of action. Overall, this supports the use of l-proline pre-conditioning to improve T-cell post-thaw recovery without needing any changes to cryopreservation solutions nor methods and hence is simple to implement.
PubMed: 37731697
DOI: 10.1039/d3md00274h -
BBA Advances 2023Antimicrobial peptide buforin II translocates across the cell membrane and binds to DNA. Its sequence is identical to a portion of core histone protein H2A making it a...
Antimicrobial peptide buforin II translocates across the cell membrane and binds to DNA. Its sequence is identical to a portion of core histone protein H2A making it a highly charged peptide. Buforin II has a proline residue in the middle of its sequence that creates a helix-hinge-helix motif which has been found to play a key role in its ability to translocate across the cell membrane. To explore the structure-function relationship of this proline residue this study has replaced P11 with a meta-substituted azobenzene amino acid (Z). The resultant peptide, photobuforin II, retained the secondary structure and membrane activity of the naturally occurring peptide while gaining new spectroscopic properties. Photobuforin II can be isomerized from its to isomer upon irradiation with ultra-violet (UV) light and from its cis to trans isomer upon irradiation with visible (VL). Photobuforin II is also fluorescent with an emission peak at 390 nm. The intrinsic fluorescence of the peptide was used to determine binding to the membrane and to DNA. VL-treated photobuforin II has a 2-fold lower binding constant compared to UV-treated photobuforin and causes 11-fold more membrane leakage in 3:1 POPC:POPG vesicles. Photobuforin II provides insights into the importance of structure function relationships in membrane active peptides while also demonstrating that azobenzene can be used in certain peptide sequences to produce intrinsic fluorescence.
PubMed: 37842183
DOI: 10.1016/j.bbadva.2023.100106 -
International Journal of Molecular... Jul 2023Cold stress is among the most important environmental factors reducing the yield of crops. The present study aimed to investigate the impact of increasing cold stress...
Cold stress is among the most important environmental factors reducing the yield of crops. The present study aimed to investigate the impact of increasing cold stress conditions on winter oilseed rape polyamines, proline, and ethylene metabolism in acclimated and non-acclimated winter oilseed rape. This study was carried out under controlled conditions in the laboratory. The winter oilseed rape hybrid 'Visby' was used in the experiment. Acclimated and non-acclimated plants were subjected to a two-day-long increasing cold (from -1 °C to -3 °C) treatment. HPTLC, RT-qPCR, spectral analysis, and gas chromatography methods were used to analyse the levels of polyamines, gene expression, proline, and ethylene, respectively. This study showed a decrease in putrescine, spermidine, and spermine content during cold acclimation and a decrease in putrescine and spermidine levels at sub-zero temperatures. There were intensive changes in ADC2 gene expression, proline, and ethylene levels in non-acclimated plants: a substantial increase after exposure to -1 °C temperature and a sharp decrease after exposure to -3 °C temperature. The changes in these parameters were lower or absent in acclimated plants. The phenomena observed in this study add new insights to the knowledge about the plant stress response and suggest questions to be answered in the future.
Topics: Polyamines; Spermidine; Putrescine; Proline; Cold Temperature; Brassica napus
PubMed: 37511158
DOI: 10.3390/ijms241411402 -
Plant Signaling & Behavior Dec 2024Tomato ( L.) is one of the most important economic crops in China. However, its quality and yield are susceptible to the adverse effects of low temperatures. In our...
Tomato ( L.) is one of the most important economic crops in China. However, its quality and yield are susceptible to the adverse effects of low temperatures. In our study, two tomato cultivars, showing different tolerance to low temperatures, namely the cold-sensitive tomato cultivar (S708) and cold-tolerant tomato cultivar (T722), were grown at optimal (25/18°C) and sub-optimal (15/10°C) temperature conditions for 5 days. Our study aimed to explore the effect of sub-optimal temperature on fresh weight, chlorophyll content and chlorophyll fluorescence, soluble sugars and proline content of two tomato cultivars. Moreover, we employed RNA-Seq to analyze the transcriptomic response of tomato roots to sub-optimal temperature. The results revealed that S708 showed a more significant reduction in fresh weight, chlorophyll content, photochemical efficiency of PSII (YII), maximum quantum yield of PSII (Fv/Fm), photochemical quenching (qP) and electron transport rate (ETR) compared to T722 under the sub-optimal temperature condition. Notably, T722 maintained higher level of soluble sugars and proline in comparison to S708 uner sub-optimal temperature. RNA-seq data showed that up-regulated DEGs in both tomato cultivars were involved in "plant-pathogen interaction", "MAPK signaling pathway", "plant hormone signal transduction", and "phosphatidylinositol signaling system". Furthermore, "Amino sugar and nucleotide sugar metabolism" pathway was enriched only in T722. Moreover, under sub-optimal temperature, transcription factor genes and osmoregulation genes showed varying degrees of response in both tomato cultivars. Conclusion: In summary, our results offer detailed insights into the response characteristics of tomato to sub-optimal temperature, providing valuable references for the practical management of tomato crops under sub-optimal temperature condition.
Topics: Temperature; Solanum lycopersicum; Chlorophyll; Proline; Gene Expression Profiling; Sugars; Stress, Physiological; Photosynthesis
PubMed: 38511566
DOI: 10.1080/15592324.2024.2332018 -
Journal of Clinical and Translational... Oct 2023Prolyl endopeptidase (PREP) is a serine endopeptidase that participates in many pathological processes including inflammation, oxidative stress, and autophagy. Our...
BACKGROUND AND AIMS
Prolyl endopeptidase (PREP) is a serine endopeptidase that participates in many pathological processes including inflammation, oxidative stress, and autophagy. Our previous studies found that PREP knockout exhibited multiple benefits in high-fat diet (HFD) or methionine choline-deficient diet-induced metabolic dysfunction-associated fatty liver disease (MAFLD). However, cumulative studies have suggested that PREP performs complex functions during disease development. Therefore, further understanding the role of PREP in MAFLD development is the foundation of PREP intervention.
METHODS
In this study, an HFD-induced MAFLD model at different time points (4, 8, 12, and 16 weeks) was used to explore dynamic changes in the PREP proline-glycine-proline (PGP)/N-acetyl-seryl-aspartyl-lysyl-proline (AcSDKP) system. To explore its potential value in MAFLD treatment, saline, or the PREP inhibitor, KYP-2047, was administered to HFD-induced MAFLD mice from the 10th to 16th weeks.
RESULTS
PREP activity and expression were increased in HFD-mice compared with control mice from the 12th week onwards, and increased PREP mainly resulted in the activation of the matrix metalloproteinase 8/9 (MMP8/9)-PREP-PGP axis rather than the thymosin β4-meprin α/PREP-AcSDKP axis. In addition, KYP-2047 reduced HFD-induced liver injury and oxidative stress, improved lipid metabolism through the suppression of lipogenic genes and the induction of β-oxidation-related genes, and attenuated hepatic inflammation by decreasing MMP8/9 and PGP. Moreover, KYP2047 restored HFD-induced impaired autophagy and this was verified in HepG2 cells.
CONCLUSIONS
These findings suggest that increased PREP activity/expression during MAFLD development might be a key factor in the transition from simple steatosis to steatohepatitis, and KYP-2047 might possess therapeutic potential for MAFLD treatment.
PubMed: 37577240
DOI: 10.14218/JCTH.2022.00110 -
Nutrients Dec 2023Alzheimer's disease (AD), is a progressive neurodegenerative disorder that involves the deposition of β-amyloid plaques and the clinical symptoms of confusion, memory...
Alzheimer's disease (AD), is a progressive neurodegenerative disorder that involves the deposition of β-amyloid plaques and the clinical symptoms of confusion, memory loss, and cognitive dysfunction. Despite enormous progress in the field, no curative treatment is available. Therefore, the current study was designed to determine the neuroprotective effects of N-methyl-(2S, 4R)-Trans-4-hydroxy-L-proline (NMP) obtained from Sideroxylon obtusifolium, a Brazilian folk medicine with anti-inflammatory and anti-oxidative properties. Here, for the first time, we explored the neuroprotective role of NMP in the Aβ-injected mouse model of AD. After acclimatization, a single intracerebroventricular injection of Aβ (5 µL/5 min/mouse) in C57BL/6N mice induced significant amyloidogenesis, reactive gliosis, oxidative stress, neuroinflammation, and synaptic and memory deficits. However, an intraperitoneal injection of NMP at a dose of (50 mg/kg/day) for three consecutive weeks remarkably decreased beta secretase1 (BACE-1) and Aβ, activated the astrocyte and microglia expression level as well as downstream inflammatory mediators such as pNF-ĸB, TNF-α, and IL-1β. NPM also strongly attenuated oxidative stress, as evaluated by the expression level of NRF2/HO-1, and synaptic failure, by improving the level of both the presynaptic (SNAP-25 and SYN) and postsynaptic (PSD-95 and SNAP-23) regions of the synapses in the cortexes and hippocampi of the Aβ-injected mice, contributing to cognitive improvement in AD and improving the behavioral deficits displayed in the Morris water maze and Y-maze. Overall, our data suggest that NMP provides potent multifactorial effects, including the inhibition of amyloid plaques, oxidative stress, neuroinflammation, and cognitive deficits.
Topics: Mice; Animals; Alzheimer Disease; Neuroprotective Agents; Neuroinflammatory Diseases; Plaque, Amyloid; Mice, Inbred C57BL; Amyloid beta-Peptides; Memory Disorders; Disease Models, Animal
PubMed: 38068844
DOI: 10.3390/nu15234986 -
International Journal of Molecular... Dec 2023Impaired wound healing is a complication of diabetes, which constitutes a serious problem in clinical practice. Currently, there is a high demand on the market for local... (Review)
Review
Impaired wound healing is a complication of diabetes, which constitutes a serious problem in clinical practice. Currently, there is a high demand on the market for local treatment options for difficult-to-heal wounds caused by diabetes. The development of dressings that accelerate wound healing has recently been the subject of much research. Sheep and camel milk is gaining importance due to the content of many bioactive substances with health-promoting effects, such as insulin, LF, proline, or CLA. Sheep and camel milk proteins are a promising source of insulin, antidiabetic, and antihypertensive peptides. Numerous studies show that local administration of insulin has a significant impact on the healing of diabetic wounds. Sheep and camel milk, due to the highest LF content among ruminants, reduces autoimmune inflammatory processes and protects against bacterial and viral infections in the wound environment. Sheep's milk has the highest content of proline and CLA, and their addition to a hydrogel dressing can help in the development of an effective dressing material. The production of hydrogel dressings containing sheep and camel milk, which are naturally rich in the bioactive substances presented in this review, may be a promising step in the market of specialized dressings for difficult-to-heal diabetic wounds.
Topics: Sheep; Animals; Camelus; Bandages; Diabetic Foot; Insulin; Insulin, Regular, Human; Hydrogels; Proline; Diabetes Mellitus
PubMed: 38139380
DOI: 10.3390/ijms242417551 -
PeerJ 2023Hydrogen sulfide (HS), as an endogenous gas signaling molecule, plays an important role in plant growth regulation and resistance to abiotic stress. This study aims to...
Hydrogen sulfide (HS), as an endogenous gas signaling molecule, plays an important role in plant growth regulation and resistance to abiotic stress. This study aims to investigate the mechanism of exogenous HS on the growth and development of seedlings under salt stress and to determine the optimal concentration for foliar application. To investigate the regulatory effects of exogenous HS (donor sodium hydrosulfide, NaHS) at concentrations ranging from 0 to 1 mM on reactive oxygen species (ROS), antioxidant system, and osmoregulation in seedlings under 300 mM NaCl stress. The growth of seedlings was inhibited by salt stress, which resulted in a decrease in the leaf relative water content (LRWC), specific leaf area (SLA), and soluble sugar content in leaves, elevated activity levels of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT); and accumulated superoxide anion (O), proline, malondialdehyde (MDA), and soluble protein content in leaves; and increased L-cysteine desulfhydrase (LCD) activity and endogenous HS content. This indicated that a high level of ROS was produced in the leaves of seedlings and seriously affected the growth and development of seedlings. The exogenous application of different concentrations of NaHS reduced the content of O , proline and MDA, increased the activity of antioxidant enzymes and the content of osmoregulators (soluble sugars and soluble proteins), while the LCD enzyme activity and the content of endogenous HS were further increased with the continuous application of exogenous HS. The inhibitory effects of salt stress on the growth rate of plant height and ground diameter, the LRWC, biomass, and SLA were effectively alleviated. A comprehensive analysis showed that the LRWC, POD, and proline could be used as the main indicators to evaluate the alleviating effect of exogenous HS on seedlings under salt stress. The optimal concentration of exogenous HS for seedlings under salt stress was 0.025 mM. This study provides an important theoretical foundation for understanding the salt tolerance mechanism of and for cultivating high-quality germplasm resources.
Topics: Salt Tolerance; Hydrogen Sulfide; Reactive Oxygen Species; Seedlings; Antioxidants; Salt Stress; Peroxidase; Peroxidases; Coloring Agents; Deuterium
PubMed: 37641597
DOI: 10.7717/peerj.15881 -
Beilstein Journal of Organic Chemistry 2023Rhodanines and their derivatives are known to have many pharmacological activities that can be modulated through different functionalization sites. One of the most...
Rhodanines and their derivatives are known to have many pharmacological activities that can be modulated through different functionalization sites. One of the most studied modification in those scaffolds is the introduction of a benzylidene moiety on C5 via a Knoevenagel reaction. Here, a facile synthesis of 5-arylidenerhodanines via a Knoevenagel reaction in an ʟ-proline-based deep eutectic solvent (DES) is reported. This method is fast (1 h at 60 °C), easy, catalyst-free and sustainable as no classical organic solvents were used. The expected compounds are recovered by a simple filtration after hydrolysis and no purification is required. Those derivatives were studied for their antioxidant activities and the results are consistent with those reported in the literature indicating that phenolic compounds are the more active ones.
PubMed: 37822921
DOI: 10.3762/bjoc.19.110 -
EMBO Molecular Medicine Jul 2023Prolyl-tRNA synthetase 1 (PARS1) has attracted much interest in controlling pathologic accumulation of collagen containing high amounts of proline in fibrotic diseases....
Prolyl-tRNA synthetase 1 (PARS1) has attracted much interest in controlling pathologic accumulation of collagen containing high amounts of proline in fibrotic diseases. However, there are concerns about its catalytic inhibition for potential adverse effects on global protein synthesis. We developed a novel compound, DWN12088, whose safety was validated by clinical phase 1 studies, and therapeutic efficacy was shown in idiopathic pulmonary fibrosis model. Structural and kinetic analyses revealed that DWN12088 binds to catalytic site of each protomer of PARS1 dimer in an asymmetric mode with different affinity, resulting in decreased responsiveness at higher doses, thereby expanding safety window. The mutations disrupting PARS1 homodimerization restored the sensitivity to DWN12088, validating negative communication between PARS1 promoters for the DWN12088 binding. Thus, this work suggests that DWN12088, an asymmetric catalytic inhibitor of PARS1 as a novel therapeutic agent against fibrosis with enhanced safety.
Topics: Humans; Amino Acyl-tRNA Synthetases; Fibrosis; Proline; Protein Biosynthesis
PubMed: 37212275
DOI: 10.15252/emmm.202216940