Improved Cell Proliferation Research Articles

Exploring the physicomechanical properties and biocompatibility traits of CuO Substituted 45S5 bioactive glass through in-vitro analysis

While clinically approved, 45S5 bioactive glass possesses limitations in terms of mechanical and biological properties for broader biomedical applications. To address this, we have synthesized 45S5 bioactive glass with partial substitution of CuO for CaO using the melt-quench technique. We conducted extensive characterization using XRD (X-Ray Diffraction), FTIR (Fourier Transform Infrared Spectroscopy), SEM (Scanning Electron Microscopy), and UTM (Universal Testing Machine), in-vitro simulated body fluid (SBF) studies, MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide), antibacterial and hemolysis assays. Our results demonstrate that CuO-substituted glasses exhibit enhanced bioactivity, as evidenced by the formation of a hydroxyapatite layer on their surface after immersion in SBF, confirmed from XRD, FTIR and SEM. Additionally, MTT assay with MG-63 cells reveals improved cell proliferation, particularly with 1.5 mol% of CuO addition. Furthermore, Cu_2 to Cu_4 compositions demonstrate effective inhibition of E. coli (Escherichia coli) colonization. The substitution of CuO also leads to significant improvements in the mechanical properties of these bioactive glasses. The density of 45S5 glass increases from 2.72 to 2.86 g/cc, compressive strength from 53 to 64 MPa and young's modulus from 76.74 to 82.21 GPa after the substitution of copper from 0.0 to 2.5 mol%, respectively. Partial substitution of CuO for CaO in 45S5 bioactive glass significantly enhances its physicomechanical properties and biocompatibility compared to the original 45S5 bioactive glass that can be used as implant materials for various biomedical applications.

Gouqi-derived nanovesicles (GqDNVs) inhibited dexamethasone-induced muscle atrophy associating with AMPK/SIRT1/PGC1α signaling pathway

With the increasing trend of global aging, sarcopenia has become a significant public health issue. Goji berry, also known as “Gou qi zi” in China, is a traditional Chinese herb that can enhance the structure and function of muscles and bones. Otherwise, previous excellent publications illustrated that plant-derived exosome-like nanoparticles can exert good bioactive functions in different aging or disease models. Thus, we issued the hypothesis that Gouqi-derived nanovesicles (GqDNVs) may also have the ability to improve skeletal muscle health, though the effect and its mechanism need to be explored. Hence, we have extracted GqDNVs from fresh berries of Lycium barbarum L. (goji) and found that the contents of GqDNVs are rich in saccharides and lipids. Based on the pathway annotations and predictions in non-targeted metabolome analysis, GqDNVs are tightly associated with the pathways in metabolism. In muscle atrophy model mice, intramuscular injection of GqDNVs improves the cross-sectional area of the quadriceps muscle, grip strength and the AMPK/SIRT1/PGC1α pathway expression. After separately inhibiting AMPK or PGC1α in C2C12 cells with dexamethasone administration, we have found that the activated AMPK plays the chief role in improving cell proliferation induced by GqDNVs. Furthermore, the energy-targeted metabolome analysis in the quadriceps muscle demonstrates that the GqDNVs up-regulate the metabolism of amino sugar and nucleotide sugar, autophagy and oxidative phosphorylation process, which indicates the activation of muscle regeneration. Besides, the Spearman rank analysis shows close associations between the quality and function of skeletal muscle, metabolites and expression levels of AMPK and SIRT1. In this study, we provide a new founding that GqDNVs can improve the quality and function of skeletal muscle accompanying the activated AMPK/SIRT1/PGC1α signaling pathway. Therefore, GqDNVs have the effect of anti-aging skeletal muscle as a potential adjuvant or complementary method or idea in future therapy and research.Graphical

Sanshentongmai Mixture Improves Oxidative Damage in Rat Cardiomyocytes H9C2 via Upregulation of microRNA-146a

To investigate the effect of Sanshentongmai (SSTM) mixture on the regulation of oxidative damage to rat cardiomyocytes (H9C2) through microRNA-146a and its mechanism. H9C2 were cultured in vitro, H2O2 was used as an oxidant to create an oxidative damage model in H9C2 cells. SSTM intervention was administered to the H9C2 cells. Then, the changes in H2O2-induced oxidative damage in H9C2 cells and the expression of microRNA-146a were observed to explore the protective effect of SSTM on H9C2 and its mechanism. H9C2 cells cultured i n vitro were divided into 3 groups, including a control group, a model group of H2O2-induced oxidative damage (referred to hereafter as the model group), and a group given H2O2 modeling plus SSTM intervention at 500 μg/L for 72 h (referred to hereafter as the treatment group). The cell viability was measured by CCK8 assay. In addition, the levels of N-terminal pro-brain natriuretic peptide (Nt-proBNP), nitric oxide (NO), high-sensitivity C-reactive protein (Hs-CRP), and angiotensin were determined by enzyme-linked immunosorbent assay (ELISA). The expression level of microRNA-146a was determined by real-time PCR (RT-PCR). H9C2 cells were pretreated with SSTM at mass concentrations ranging from 200 to 1500 μg/L. Then, CCK8 assay was performed to measure cell viability and the findings showed that the improvement in cell proliferation reached its peak when the mass concentration of SSTM was 500 μg/L, which was subsequently used as the intervention concentration. ELISA was performed to measure the indicators related to heart failure, including Nt-proBNP, NO, Hs-CRP, and angiotensin Ⅱ. Compared with those of the control group, the expressions of Nt-proBNP and angiotensin Ⅱ in the treatment group were up-regulated (P<0.05), while the expression of NO was down-regulated (P<0.05). There was no significant difference in the expression of Hs-CRP between the treatment group and the control group. These findings indicate that SSTM could effectively ameliorate oxidative damage in H9C2 rat cardiomyocytes. Finally, according to the RT-PCR findings for the expression of microRNA-146a in each group, H2O2 treatment at 15 μmol/L could significantly reduce the expression of microRNA-146a, and the expression of microRNA-146a in the treatment group was nearly doubled compared with that in the model group. There was no significant difference between the treatment group and the control group. SSTM can significantly resist the H2O2-induced oxidative damage of H9C2 cells and may play a myocardial protective role by upregulating microRNA-146a.

Hypoxic adipose stem cell-derived exosomes carrying high-abundant USP22 facilitate cutaneous wound healing through stabilizing HIF-1α and upregulating lncRNA H19.

Hypoxic preconditioning has been recognized as a promotive factor for accelerating cutaneous wound healing. Our previous study uncovered that exosomal lncRNA H19, derived from adipose-derived stem cells (ADSCs), plays a crucial role in orchestrating cutaneous wound healing. Herein, we aimed to explore whether there is a connection between hypoxia and ADSC-derived exosomes (ADSCs-exos) in cutaneous wound healing. Exosomes extracted from ADSCs under normoxic and hypoxic conditions were identified using transmission electron microscope (TEM) and particle size analysis. The effects of ADSCs-exos on the proliferation, migration, and angiogenesis of human umbilical vein endothelial cells (HUVECs) were evaluated by CCK-8, EdU, wound healing, and tube formation assays. Expression patterns of H19, HIF-1α, and USP22 were measured. Co-immunoprecipitation, chromatin immunoprecipitation, ubiquitination, and luciferase reporter assays were conducted to confirm the USP22/HIF-1α/H19 axis, which was further validated in a mice model of skin wound. Exosomes extracted from hypoxia-treated ADSCs (termed as H-ADSCs-exos) significantly increased cell proliferation, migration, and angiogenesis in H2O2-exposed HUVECs, and promoted cutaneous wound healing invivo. Moreover, H-ADSCs and H-ADSCs-exos, which exhibited higher levels of H19, were found to be transcriptionally activated by HIF-1α. Mechanically, H-ADSCs carrying USP22 accounted for deubiquitinating and stabilizing HIF-1α. Additionally, H-ADSCs-exos improved cell proliferation, migration, and angiogenesis in H2O2-triggered HUVECs by activating USP22/HIF-1α axis and promoting H19 expression, which may provide a new clue for the clinical treatment of cutaneous wound healing.

Hydrogen Sulfide Delivery to Enhance Bone Tissue Engineering Cell Survival.

Though crucial for natural bone healing, local calcium ion (Ca2+) and phosphate ion (Pi) concentrations can exceed the cytotoxic limit leading to mitochondrial overload, oxidative stress, and cell death. For bone tissue engineering applications, H2S can be employed as a cytoprotective molecule to enhance mesenchymal stem cell (MSC) tolerance to cytotoxic Ca2+/Pi concentrations. Varied concentrations of sodium hydrogen sulfide (NaSH), a fast-releasing H2S donor, were applied to assess the influence of H2S on MSC proliferation. The results suggested a toxicity limit of 4 mM for NaSH and that 1 mM of NaSH could improve cell proliferation and differentiation in the presence of cytotoxic levels of Ca2+ (32 mM) and/or Pi (16 mM). To controllably deliver H2S over time, a novel donor molecule (thioglutamic acid-GluSH) was synthesized and evaluated for its H2S release profile. Excitingly, GluSH successfully maintained cytoprotective level of H2S over 7 days. Furthermore, MSCs exposed to cytotoxic Ca2+/Pi concentrations in the presence of GluSH were able to thrive and differentiate into osteoblasts. These findings suggest that the incorporation of a sustained H2S donor such as GluSH into CaP-based bone graft substitutes can facilitate considerable cytoprotection, making it an attractive option for complex bone regenerative engineering applications.

Prophylactic zinc and therapeutic selenium administration in adult rats prevents long-term cognitive and behavioral sequelae by a transient ischemic attack

The transient hypoxic-ischemic attack, also known as a minor stroke, can result in long-term neurological issues such as memory loss, depression, and anxiety due to an increase in nitrosative stress. The individual or combined administration of chronic prophylactic zinc and therapeutic selenium is known to reduce nitrosative stress in the first seven days post-reperfusion and, due to an antioxidant effect, prevent cell death. Besides, zinc or selenium, individually administered, also causes antidepressant and anxiolytic effects. Therefore, this work evaluated whether combining zinc and selenium could prevent stroke-elicited cognition and behavior deficits after 30 days post-reperfusion. Accordingly, we assessed the expression of growth factors at 7 days post-reperfusion, a four-time course of memory (from 7 to 28 days post-learning test), and cell proliferation, depression, and anxiety-like behavior at 30 days post-reperfusion. Male Wistar rats with a weight between 190 and 240 g) were treated with chronic prophylactic zinc administration with a concentration of 0.2 mg/kg for 15 days before common carotid artery occlusion (10 min) and then with therapeutic selenium (6 μg/kg) for 7 days post-reperfusion. Compared with individual administrations, the administration combined of prophylactic zinc and therapeutic selenium decreased astrogliosis, increased growth factor expression, and improved cell proliferation and survival in two regions, the hippocampus, and cerebral cortex. These effects prevented memory loss, depression, and anxiety-like behaviors. In conclusion, these results demonstrate that the prophylactic zinc administration combined with therapeutic selenium can reduce the long-term sequelae caused by the transient ischemic attack.Significance statement.A minor stroke caused by a transient ischemic attack can result in psychomotor sequelae that affect not only the living conditions of patients and their families but also the economy. The incidence of these micro-events among young people has increased in the world. Nonetheless, there is no deep understanding of how this population group responds to regular treatments (Ekker and et al., 2018) [1]. On the basis that zinc and selenium have antioxidant, anti-inflammatory, and regenerative properties in stroke animal models, our work explored whether the chronic combined administration of prophylactic zinc and therapeutic selenium could prevent neurological sequelae in the long term in a stroke rat model of unilateral common carotid artery occlusion (CCAO) by 10-min. Our results showed that this combined treatment provided a long-term neuroprotective effect by decreasing astrogliosis, memory loss, anxiety, and depression-like behavior.

A Bioinspired Dopamine‐Amorphous Calcium Phosphate Coating on Titanium Implant for Osteogenesis and Angiogenesis

AbstractTitanium (Ti) is widely used in dental implantation. However, their lack of biological activity causes significant clinical risks associated with immediate loading and early osteointegration failure. Amorphous calcium phosphate (ACP) is reported to be a precursor of bone mineralization. Dopamine (dopa) is well known to possess strong adhesive force in natural mussels. Combining the bio‐related advantages of these materials, a dopa‐polyacrylic acid (PAA)‐ACP hybrid material on the surface of the Ti plate is judiciously fabricated. The biological adhesiveness of dopa to graft it together with PAA onto the surface of Ti is utilized. Then, the dopa‐PAA layer serves as a matrix to grow ACP species. The modified interface allows for regulations of bioactivities. MC3T3‐E1 osteoblast (MC3T3) cells indeed show improved cell proliferation and adhesion abilities on the Ti with dopa‐PAA‐ACP coatings compared to those on bare Ti or Ti with only dopa‐PAA coating. The osteogenic potential of MC3T3 cells is also considerably upregulated on dopa‐PAA‐ACP coatings. Moreover, human umbilical vein endothelial cells (HUVECs) on dopa‐PAA‐ACP coatings show higher levels of angiogenic markers. This report proves that dopa‐PAA‐ACP composite is a new and simple bioactive titanium coating to facilitate osteogenesis and angiogenesis for dental applications.

ZNF554 Inhibits Endometrial Cancer Progression via Regulating RBM5 and Inactivating WNT/β-Catenin Signaling Pathway.

Uterine corpus endometrial carcinoma (UCEC), a kind of gynecologic malignancy, poses a significant risk to women's health. The precise mechanism underlying the development of UCEC remains elusive. Zinc finger protein 554 (ZNF554), a member of the Krüppel-associated box domain zinc finger protein superfamily, was reported to be dysregulated in various illnesses, including malignant tumors. This study aimed to examine the involvement of ZNF554 in the development of UCEC. The expression of ZNF554 in UCEC tissues and cell lines were examined by qRT-PCR and Western blot assay. Cells with stably overexpressed or knocked-down ZNF554 were established through lentivirus infection. CCK-8, wound healing, and Transwell invasion assays were employed to assess cell proliferation, migration, and invasion. Propidium iodide (PI) staining combined with fluorescence-activated cell sorting (FACS) flow cytometer was utilized to detect cell cycle distribution. qRT-PCR and Western blotting were conducted to examine relative mRNA and protein levels. Chromatin immunoprecipitation assay and luciferase reporter assay were used to explore the regulatory role of ZNF554 in RNA binding motif 5 (RBM5). The expression of ZNF554 was found to be reduced in both UCEC samples and cell lines. Decreased expression of ZNF554 was associated with higher tumor stage, decreased overall survival, and reduced disease-free survival in UCEC. ZNF554 overexpression suppressed cell proliferation, migration, and invasion, while also inducing cell cycle arrest. In contrast, a decrease in ZNF554 expression resulted in the opposite effect. Mechanistically, ZNF554 transcriptionally regulated RBM5, leading to the deactivation of the Wingless (WNT)/β-catenin signaling pathway. Moreover, the findings from rescue studies demonstrated that the inhibition of RBM5 negated the impact of ZNF554 overexpression on β-catenin and p-glycogen synthase kinase-3β (p-GSK-3β). Similarly, the deliberate activation of RBM5 reduced the increase in β-catenin and p-GSK-3β caused by the suppression of ZNF554. In vitro experiments showed that ZNF554 overexpression-induced decreases in cell proliferation and migration were counteracted by RBM5 knockdown. Additionally, when RBM5 was overexpressed, it hindered the improvements in cell proliferation and migration caused by reducing the ZNF554 levels. ZNF554 functions as a tumor suppressor in UCEC. Furthermore, ZNF554 regulates UCEC progression through the RBM5/WNT/β-catenin signaling pathway. ZNF554 shows a promise as both a prognostic biomarker and a therapeutic target for UCEC.

Hesperidin ameliorates H2O2-induced bovine mammary epithelial cell oxidative stress via the Nrf2 signaling pathway

BackgroundHesperidin is a citrus flavonoid with anti-inflammatory and antioxidant potential. However, its protective effects on bovine mammary epithelial cells (bMECs) exposed to oxidative stress have not been elucidated.ResultsIn this study, we investigated the effects of hesperidin on H2O2-induced oxidative stress in bMECs and the underlying molecular mechanism. We found that hesperidin attenuated H2O2-induced cell damage by reducing reactive oxygen species (ROS) and malondialdehyde (MDA) levels, increasing catalase (CAT) activity, and improving cell proliferation and mitochondrial membrane potential. Moreover, hesperidin activated the Keap1/Nrf2/ARE signaling pathway by inducing the nuclear translocation of Nrf2 and the expression of its downstream genes NQO1 and HO-1, which are antioxidant enzymes involved in ROS scavenging and cellular redox balance. The protective effects of hesperidin were blocked by the Nrf2 inhibitor ML385, indicating that they were Nrf2 dependent.ConclusionsOur results suggest that hesperidin could protect bMECs from oxidative stress injury by activating the Nrf2 signaling pathway, suggesting that hesperidin as a natural antioxidant has positive potential as a feed additive or plant drug to promote the health benefits of bovine mammary.Graphical

On 3D printed thermoresponsive PCL-PLA nanofibers based architected smart nanoporous scaffolds for tissue reconstruction

Previous studies have reported that polycaprolactone (PCL)-polylactic acid (PLA) based scaffolds are one of the most acceptable composite materials in tissue engineering/reconstruction applications. However, little has been reported on the fabrication of nanoporous smart scaffolds (having sensing capability, programmable cell growth, and programmable shape memory capabilities, etc.). In this study, nanoporous smart scaffolds have been fabricated using the fused filament fabrication (FFF) process by architecting the PLA nanofibers in between the PCL polymer matrix to introduce the smart properties. The results have revealed an acceptable modulus of toughness (7.304 MPa) (as crash resistance property for scaffolds) when fabricated using thermal stress relieved/pre-heat treated (PHT), PCL granules at 64 °C-barrel processing temperature (optimized setting) of a screw extruder. The recrystallization has ensured the tuning of crystallinity (increased from 6.06 % to 9.53) as confirmed by differential scanning calorimetry (DSC) analysis. On the optimized extrusion setting, the dielectric constant (ϵr) (∼12) has shown acceptable sensing capabilities in scaffolds. The results of the crash resistance, thermal, and sensing properties have been supported by X-ray diffraction (XRD) and Fourier transform infrared (FTIR) spectroscopic analysis. Deposition of the PLA nanofibers mat in the PCL matrix ensured improved cell proliferation and it was 180 % when 1 mat was deposited. Also, PCL + 1 PLA nanofiber mat scaffolds recovered their original shape in less time (by 25 %) as compared to PCL scaffolds under the thermal stimulus.

Abstract LB341: Clinical validation and mechanistic insights of ThisCART: A novel allogeneic platform enhancing CAR-T signaling via intracellular TCRαβ/CD3 retention

Abstract Background: Current allogeneic CAR-T cell therapies face challenges with post-infusion expansion and efficacy. In response, we developed ThisCART, a non-genetic editing, allogeneic platform that significantly enhances CAR-T cell expansion and efficacy. This is achieved through a novel approach of intracellular retention of membrane proteins, reducing surface expression of TCRαβ/CD3 complexes and HLA-I molecules. Our lead product, ThisCART19, has demonstrated superior efficacy compared to conventional AutoCART19 in preclinical and early clinical studies for B-ALL. Methods: In the clinical trial NCT04384393, we assessed the safety and efficacy of ThisCART19 in patients with B-cell malignancies. Post-lymphodepletion, participants received infusions of ThisCART19 and AutoCART19, both personalized from the patient's own T cells, in equal proportions. Safety and efficacy were the primary endpoints, evaluated on Day 28 post-infusion and throughout extended follow-up. Mechanistic insights were gained through comprehensive multi-omics studies, including RNAseq, whole-cell proteomics, and phosphoproteomics. Results: ThisCART19 showed enhanced expansion and an improved safety profile in vivo. As of 01 January 2024, Two patients diagnosed as relapse/refractory acute B cell leukemia (R/R B-ALL) were enrolled in the clinical trial. Pt1 is a 55yo male with Ph-positive B-ALL (36% blasts) who relapsed after multiagent chemotherapy and TKIs. Pt2 is a 16yo male with CNS leukemia (78.5% blasts) after multiagent chemotherapy. No DLTs or ICANS were observed. Pt1 experienced Grade 1 CRS that resolved without treatment, and Pt2 experienced Grade 2 CRS that resolved after tocilizumab and corticosteroids. Complete remission with MRD-negative status was achieved by Day 28. Pharmacodynamic analysis indicated a significant expansion of CAR-T cells, predominantly in the CAR+TCR- subset, highlighting autonomous CAR signaling enhancement. Multi-omics analysis identified upregulation in JAK-STAT, PI3K-Akt, and NFKB pathways, suggesting improved cell proliferation, survival, anti-apoptotic activity, and effector function. This molecular profiling provides a mechanistic understanding of ThisCART19A's therapeutic efficacy. Conclusion: The ThisCART platform marks a transformative advancement in allogeneic CAR-T cell therapy. By enhancing CAR signaling autonomously through a unique intracellular retention of TCRαβ/CD3 complexes, ThisCART has shown promising safety and efficacy in early clinical trials. Its potential in reshaping the treatment landscape for hematological and autoimmune disorders positions ThisCART as a crucial development in cellular therapeutics. Citation Format: Jun Li, Lei Xue, Sujun Li, Ling He, Kaichun Liu, Rong Jin, Tao Wang, Yinhang Zhang, Shanshan Chen, Yulian Gao, Dan Liu, Liyun Yang, Xingbing Wang. Clinical validation and mechanistic insights of ThisCART: A novel allogeneic platform enhancing CAR-T signaling via intracellular TCRαβ/CD3 retention [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB341.

Black soldier fly bioconversion to cultivated meat media components using blue catfish gut microbiome

This study aimed to develop low-cost media for cell-cultivated seafood using gut-microbial community-assisted fermentation. Black soldier fly larvae (Hermetia illucens) were used as the substrate and exposed to gut microbial communities isolated from blue catfish (Ictalurus furcatus). Fermentation led to a decrease in species richness, with the loss of important chitinase and protease-producing genera such as Pseudomonas and Clostridiaceae. However, there was an increase in Paraclostridium and members of the Enterobacteriaceae. In addition, the effect of fermented hydrolysates from BSFL on the proliferation of zebrafish embryo fibroblasts was tested in comparison to fetal bovine serum (FBS) in in vitro cell cultivation. Lower concentrations of FBS resulted in decreased cell density and altered cell morphology. The supplementation of hydrolysate B at high peptide concentrations had cytotoxic effects on the cells, while at lower peptide concentrations, it improved cell proliferation only in cultures with 2.5 % FBS.

Current Research, Industrialization Status, and Future Perspective of Cultured Meat.

Expectations for the industrialization of cultured meat are growing due to the increasing support from various sectors, such as the food industry, animal welfare organizations, and consumers, particularly vegetarians, but the progress of industrialization is slower than initially reported. This review analyzes the main issues concerning the industrialization of cultured meat, examines research and media reports on the development of cultured meat to date, and presents the current technology, industrialization level, and prospects for cultured meat. Currently, over 30 countries have companies industrializing cultured meat, and around 200 companies that are developing or industrializing cultured meat have been surveyed globally. By country, the United States has over 50 companies, accounting for more than 20% of the total. Acquiring animal cells, developing cell lines, improving cell proliferation, improving the efficiency of cell differentiation and muscle production, or developing cell culture media, including serum-free media, are the major research themes related to the development of cultured meat. In contrast, the development of devices, such as bioreactors, which are crucial in enabling large-scale production, is relatively understudied, and few of the many companies invested in the development of cultured meat have presented products for sale other than prototypes. In addition, because most information on key technologies is not publicly available, it is not possible to determine the level of technology in the companies, and it is surmised that the technology of cultured meat-related startups is not high. Therefore, further research and development are needed to promote the full-scale industrialization of cultured meat.

Optimization of Zwitterionic Polymers for Cell Cryopreservation.

Cryopreservation techniques are valuable for the preservation of genetic properties in cells, and the development of this technology contributes to various fields. In a previous study, an isotonic freezing medium composed of poly(zwitterion) (polyZI) has been reported, which alleviates osmotic shock, unlike typical hypertonic freezing media. In this study, the primitive freezing medium composed of emerging polyZI is optimized. Imidazolium/carboxylate-type polyZI (VimC3C) is the optimal chemical structure. The molecular weight and degree of ion substitution (DSion) are not significant factors. There is an impediment with the primitive polyZI freezing media. While the polyZI forms a matrix around the cell membrane to protect cells, the matrix is difficult to remove after thawing, resulting in low cell proliferation. Unexpectedly, increasing the poly(VimC3C) concentration from 10% to 20% (w/v) improves cell proliferation. The optimized freezing medium, 20% (w/v) poly(VimC3C)_DSion(100%)/1% (w/v) NaCl aqueous solution, exhibited a better cryoprotective effect.

Reduced miR-99a-3p levels in systemic lupus erythematosus may promote B cell proliferation via NCAPG and the PI3K/AKT signaling pathway.

Systemic lupus erythematosus is an immunologically dysregulated disease characterized by the presence of multiple autoantibodies. In SLE, B lymphocytes contribute to the dysregulated production of autoantibodies and cytokines. Recently, we discovered that miR-99a-3p binds to both EIF4EBP1 and NCAPG mRNA and that lowering miR-99a-3p can promote B cell autophagy in SLE by increasing EIF4EBP1 expression. However, the functions of miR-99a-3p and NCAPG in SLE have not been extensively investigated. This work aims to evaluate the levels of miR-99a-3p and NCAPG expression in SLE B cells and to determine whether the aberrant expression of miR-99a-3p and NCAPG contributes to the pathological mechanisms in SLE. B lymphocytes were obtained through immunomagnetic negative selection. Using RT-qPCR, miR-99a-3p and NCAPG mRNA expressions in B lymphocytes and in the BALL-1 cell line were measured. To determine the relative abundance of NCAPG, PI3K, p-PI3K, AKT, and p-AKT, we normalize them to the level of β-actin using Western blotting. Evaluation of miR-99a-3p and NCAPG's impact on cell proliferation was done utilizing CCK-8 assay. Using flow cytometry, the cell cycle and apoptosis were both measured. Comparing SLE B cells to healthy controls, miR-99a-3p expression was significantly downregulated. Additionally, it was observed that SLE B cells had significantly higher NCAPG mRNA expression. Blocking miR-99a-3p expression in BALL-1 cells with an antagomir elevated NCAPG expression, facilitated PI3K/AKT pathway activation, improved cell proliferation, raised the fraction of S-phase cells, and prevented cell apoptosis. The opposite effects of upregulated miR-99a-3p levels on BALL-1 cells were observed by using an agomir. Furthermore, the effect of decreased miR-99a-3p expression on cell proliferation was partially mediated by elevating NCAPG levels and activating the PI3K/AKT pathway. Our research indicates that lower miR-99a-3p expression in SLE B cells appears to boost B cell number via the NCAPG and PI3K/AKT pathways.

Polyelectrolyte Multilayer Coating on 3D Printed PEGDA/TCP Scaffold with Improved Cell Proliferation

Bioactive coating of ceramic scaffolds is an effective way to ameliorate osseointegration and attenuate implant-induced inflammatory responses, which should be biocompatible and possess suitable mechanical properties to regulate cell adhesion and migration. In this study, a poly (ethylene glycol) diacrylate/tricalcium phosphate (PEGDA/TCP) ceramic scaffold was prepared using SLA-3D printing, and its compressive strength was 8.9 ± 1.0 MPa. Chitosan (Chi) and chondroitin sulfate (CS) were assembled on the surface of the PEGDA/TCP scaffolds and crosslinked with 1-ethyl-3-(3-dimethylaminopropyl) -carbodiimide/ N-hydroxysuccinimide (EDC/NHS). Scanning electron microscope (SEM), Fourier transform infrared (FTIR), and laser scanning microscope were used to evaluate the surface modification of the PEGDA/TCP scaffolds. Cellular tests showed that polyelectrolyte multilayers (PEMs) promoted cell adhesion and proliferation of osteoblasts relative to unmodified scaffolds. Furthermore, it can be demonstrated that the SLA-3D printed TCP scaffolds could meet the compressive requirements of trabecular bones, and the bioactivity of the bone scaffolds could be effectively improved by combining them with Chi/CS PEM.

Chondrogenesis of mesenchymal stromal cells on the 3D printed polycaprolactone/fibrin/decellular cartilage matrix hybrid scaffolds in the presence of piascledine

Nowadays, cartilage tissue engineering (CTE) is considered important due to lack of repair of cartilaginous lesions and the absence of appropriate methods for treatment. In this study, polycaprolactone (PCL) scaffolds were fabricated by three-dimensional (3D) printing and were then coated with fibrin (F) and acellular solubilized extracellular matrix (ECM). After extracting adipose-derived stem cells (ADSCs), 3D-printed scaffolds were characterized and compared to hydrogel groups. After inducing the chondrogenic differentiation in the presence of Piascledine and comparing it with TGF-β3 for 28 days, the expression of genes involved in chondrogenesis (AGG, COLII) and the expression of the hypertrophic gene (COLX) were examined by real-time PCR. The expression of proteins COLII and COLX was also determined by immunohistochemistry. Glycosaminoglycan was measured by toluidine blue staining. 3D-printed scaffolds clearly improved cell proliferation, viability, water absorption and compressive strength compared to the hydrogel groups. Moreover, the use of compounds such as ECM and Piascledine in the process of ADSCs chondrogenesis induction increased cartilage-specific markers and decreased the hypertrophic marker compared to TGF-β3. In Piascledine groups, the expression of COLL II protein, COLL II and Aggrecan genes, and the amount of glycosaminoglycan showed a significant increase in the PCL/F/ECM compared to the PCL and PCL/F groups.

Design-rules for stapled peptides with in vivo activity and their application to Mdm2/X antagonists

Although stapled α-helical peptides can address challenging targets, their advancement is impeded by poor understandings for making them cell permeable while avoiding off-target toxicities. By synthesizing >350 molecules, we present workflows for identifying stapled peptides against Mdm2(X) with in vivo activity and no off-target effects. Key insights include a clear correlation between lipophilicity and permeability, removal of positive charge to avoid off-target toxicities, judicious anionic residue placement to enhance solubility/behavior, optimization of C-terminal length/helicity to enhance potency, and optimization of staple type/number to avoid polypharmacology. Workflow application gives peptides with >292x improved cell proliferation potencies and no off-target cell proliferation effects ( > 3800x on-target index). Application of these ‘design rules’ to a distinct Mdm2(X) peptide series improves ( > 150x) cellular potencies and removes off-target toxicities. The outlined workflow should facilitate therapeutic impacts, especially for those targets such as Mdm2(X) that have hydrophobic interfaces and are targetable with a helical motif.

Melatonin alleviates ischemic stroke by inhibiting ferroptosis through the CYP1B1/ACSL4 pathway.

This study utilized middle cerebral artery occlusion (MCAO) mouse models and HT-22 cell oxygen and glucose deprivation/reoxygenation (OGD/R) models to investigate the therapeutic effects of melatonin on ischemic brain injury. In the experiments, MCAO mice were treated with 5 and 10 mg/kg doses of melatonin, and H-T22 cells underwent OGD/R treatment and were administered different concentrations of melatonin. The results showed that melatonin significantly reduced ischemic brain area, neural damage, cerebral edema, and neuronal apoptosis in MCAO mice. In the HT-22 cell model, melatonin also improved cell proliferation ability, reduced apoptosis, and ROS production. Further mechanistic studies found that melatonin exerts protective effects by inhibiting ferroptosis, an iron-dependent form of regulated cell death, through regulation of the ACSL4/CYP1B1 pathway. In MCAO mice, melatonin decreased lipid peroxidation, ROS production, and ACSL4 protein expression. Overexpression of CYP1B1 increased ACSL4 ubiquitination and degradation, thereby increasing cell tolerance to ferroptosis, reducing ACSL4 protein levels, and decreasing ROS production. CYP1B1 knockdown obtained opposite results. The CYP1B1 metabolite 20-HETE induces expression of the E3 ubiquitin ligase FBXO10 by activating PKC signaling, which promotes ACSL4 degradation. In the OGD/R cell model, inhibition of CYP1B1 expression reversed the therapeutic effects of melatonin. In summary, this study demonstrates that melatonin protects the brain from ischemic injury by inhibiting ferroptosis through regulation of the ACSL4/CYP1B1 pathway, providing evidence for new therapeutic targets for ischemic brain injury.

Electrospun nylon 6/hyaluronic acid/chitosan bioactive nanofibrous composite as a potential antibacterial wound dressing.

Hyaluronic acid (HA) and chitosan (CS), as natural biomaterials, display excellent biocompatibility and stimulate the growth and proliferation of fibroblasts. Furthermore, nylon 6 (N6) is a low-cost polymer with good compatibility with human tissues and high mechanical stability. In this study, HA and CS were applied to modify N6 nanofibrous mat (N6/HA/CS) for potential wound dressing. N6/HA/CS nanofibrous composite mats were developed using a simple one-step electrospinning technique at different CS concentrations of 1, 2, and 3 wt%. The results demonstrated that incorporating HA and CS into N6 resulted in increased hydrophilicity, as well as favorable physical and mechanical properties. In addition, the minimum inhibitory concentration and (MIC) optical density techniques were used to determine the antibacterial properties of N6/HA/CS nanofibrous composite mats, and the results demonstrated that the composites could markedly inhibit the growth of Gram-positive bacteria Staphylococcus aureus and Gram-negative bacteria Escherichia coli. Because of its superior mechanical properties, substantial antimicrobial effects, and hydrophilic surface, N6/HA/CS at 2 wt% of CS (N6/HA/CS2) was chosen as the most suitable nanofibrous mat. The swelling, porosity, gel content, and in vitro degradation studies imply that N6/HA/CS2 nanofibrous composite mat has proper moisture retention and biodegradability. Furthermore, the N6/HA/CS2 nanofibrous composite mat was discovered to be nontoxic to L929 fibroblast cells and to even improve cell proliferation. Based on the findings, this research offers a simple and rapid method for creating material that could be utilized as prospective wound dressings in clinical environments.