-
Materials (Basel, Switzerland) Dec 2023In recent decades, there has been increased attention to the role of layer-by-layer assembled bio-polymer 3D structures (capsules, beads, and microgels) for biomedical...
In recent decades, there has been increased attention to the role of layer-by-layer assembled bio-polymer 3D structures (capsules, beads, and microgels) for biomedical applications. Such free-standing multilayer structures are formed via hard templating onto sacrificial cores such as vaterite CaCO crystals. Immobilization of these structures onto solid surfaces (e.g., implants and catheters) opens the way for the formulation of advanced bio-coating with a patterned surface. However, the immobilization step is challenging. Multiple approaches based mainly on covalent binding have been developed to localize these multilayer 3D structures at the surface. This work reports a novel strategy to formulate multilayer surface-supported microgels (ss-MG) directly on the surface via hard templating onto ss-CaCO pre-grown onto the surface via the direct mixing of NaCO and CaCl precursor solutions. ss-MGs were fabricated using biopolymers: polylysine (PLL) as polycation and three polyanions-hyaluronic acid (HA), heparin sulfate (HS), and alginate (ALG). ss-MG biodegradation was examined by employing the enzyme trypsin. Our studies indicate that the adhesion of the ss-MG to the surface and its formation yield directly correlate with the mobility of biopolymers in the ss-MG, which decreases in the sequence of ALG > HA > HS-based ss-MGs. The adhesion of HS-based ss-MGs is only possible via heating during their formation. Dextran-loading increases ss-MG formation yield while reducing ss-MG shrinking. ss-MGs with higher polymer mobility possess slower biodegradation rates, which is likely due to diffusion limitations for the enzyme in more compact annealed ss-MGs. These findings provide valuable insights into the mechanisms underlying the formation and biodegradation of surface-supported biopolymer structures.
PubMed: 38203957
DOI: 10.3390/ma17010103 -
International Journal of Molecular... Dec 2023Undifferentiated germ cells, including the spermatogonial stem cell subpopulation required for fertility restoration using human immature testicular tissue (ITT), are...
Optimized Recovery of Immature Germ Cells after Prepubertal Testicular Tissue Digestion and Multi-Step Differential Plating: A Step towards Fertility Restoration with Cancer-Cell-Contaminated Tissue.
Undifferentiated germ cells, including the spermatogonial stem cell subpopulation required for fertility restoration using human immature testicular tissue (ITT), are difficult to recover as they do not easily adhere to plastics. Due to the scarcity of human ITT for research, we used neonatal porcine ITT. Strategies for maximizing germ cell recovery, including a comparison of two enzymatic digestion protocols (P1 and P2) of ITT fragment sizes (4 mm and 8 mm and multi-step differential plating were explored. Cellular viability and yield, as well as numbers and proportions of DDX4+ germ cells, were assessed before incubating the cell suspensions overnight on uncoated plastics. Adherent cells were processed for immunocytochemistry (ICC) and floating cells were further incubated for three days on Poly-D-Lysine-coated plastics. Germ cell yield and cell types using ICC for SOX9, DDX4, ACTA2 and CYP19A1 were assessed at each step of the multi-step differential plating. Directly after digestion, cell suspensions contained >92% viable cells and 4.51% DDX4+ germ cells. Pooled results for fragment sizes revealed that the majority of DDX4+ cells adhere to uncoated plastics (P1; 82.36% vs. P2; 58.24%). Further incubation on Poly-D-Lysine-coated plastics increased germ cell recovery (4.80 ± 11.32 vs. 1.90 ± 2.07 DDX4+ germ cells/mm, respectively for P1 and P2). The total proportion of DDX4+ germ cells after the complete multi-step differential plating was 3.12%. These results highlight a reduced proportion and number of germ cells lost when compared to data reported with other methods, suggesting that multi-step differential plating should be considered for optimization of immature germ cell recovery. While Poly-D-Lysine-coating increased the proportions of recovered germ cells by 16.18% (P1) and 28.98% (P2), future studies should now focus on less cell stress-inducing enzymatic digestion protocols to maximize the chances of fertility restoration with low amounts of cryo-banked human ITT.
Topics: Humans; Animals; Swine; Neoplasms; Germ Cells; Fertility; Lysine; Plastics; Poly A; Digestion; Polylysine
PubMed: 38203691
DOI: 10.3390/ijms25010521 -
Theranostics 2024Intravesical chemotherapy is highly recommended after transurethral resection of bladder tumor for patients with bladder cancer (BCa). However, this localized adjuvant...
Intravesical chemotherapy is highly recommended after transurethral resection of bladder tumor for patients with bladder cancer (BCa). However, this localized adjuvant therapy has drawbacks of causing indiscriminate damage and inability to penetrate bladder mucosal. Fluorinated polylysine micelles (PLLF) were synthesized by reacting polylysine (PLL) with heptafluorobutyrate anhydride. Anti-apoptotic gene defender against cell death 1 (DAD1) was selected by different gene expression analysis between BCa patients and healthy individuals and identified by several biological function assays. The gene transfection ability of PLLF was verified by multiple and assays. The therapeutic efficiency of PLLF nanoparticles (NPs) targeting DAD1 were confirmed by intravesical administration using an orthotopic BCa mouse model. Decorated with fluorinated chains, PLL can self-assemble to form NPs and condense plasmids with excellent gene transfection efficiency . Loading with the CRISPR-Cas9 system designed to target DAD1 (Cas9-sgDAD1), PLLF/Cas9-sgDAD1 NPs strongly inhibited the expression of DAD1 in BCa cells and induced BCa cell apoptosis through the MAPK signaling pathway. Furthermore, intravesical administration of PLLF/Cas9-sgDAD1 NPs resulted in significant therapeutic outcomes without systemic toxicity . The synthetized PLLF can transmucosally deliver the CRISPR-Cas9 system into orthotopic BCa tissues to improve intravesical instillation therapy for BCa. This work presents a new strategy for targeting DAD1 gene in the intravesical therapy for BCa with high potential for clinical applications.
Topics: Mice; Animals; Humans; Urinary Bladder; Polylysine; CRISPR-Cas Systems; Urinary Bladder Neoplasms; Nanoparticles; Genetic Therapy
PubMed: 38164146
DOI: 10.7150/thno.88550 -
Journal of the American Chemical Society Jan 2024Gd-L1 is a macrocyclic Gd-HPDO3A derivative functionalized with a short spacer to a trisulfonated pyrene. When compared to Gd-HPDO3A, the increased relaxivity appears to...
Seeking for Innovation with Magnetic Resonance Imaging Paramagnetic Contrast Agents: Relaxation Enhancement via Weak and Dynamic Electrostatic Interactions with Positively Charged Groups on Endogenous Macromolecules.
Gd-L1 is a macrocyclic Gd-HPDO3A derivative functionalized with a short spacer to a trisulfonated pyrene. When compared to Gd-HPDO3A, the increased relaxivity appears to be determined by both the higher molecular weight and the occurrence of an intramolecularly catalyzed prototropic exchange of the coordinated OH moiety. In water, Gd-L1 displayed a relaxivity of 7.1 mM s (at 298 K and 0.5 T), slightly increasing with the concentration likely due to the onset of intermolecular aggregation. A remarkably high and concentration-dependent relaxivity was measured in human serum (up to 26.5 mM s at the lowest tested concentration of 0.005 mM). The acquisition of H-nuclear magnetic relaxation dispersion (NMRD) and O- vs profiles allowed to get an in-depth characterization of the system. experiments in the presence of human serum albumin, γ-globulins, and polylysine, as well as using media mimicking the extracellular matrix, provided strong support to the view that the trisulfonated pyrene fosters binding interactions with the exposed positive groups on the surface of proteins, responsible for a remarkable hyperintensity in MR images. The MR images of the liver, kidneys, and spleen showed a marked contrast enhancement in the first 10 min after the i.v. injection of Gd-L1, which was 2-6-fold higher than that for Gd-HPDO3A, while maintaining a very similar excretion behavior. These findings may pave the way to an improved design of MRI GBCAs, for the first time, based on the setup of weak and dynamic interactions with abundant positive groups on serum and ECM proteins.
Topics: Humans; Contrast Media; Static Electricity; Magnetic Resonance Imaging; Organometallic Compounds; Pyrenes; Gadolinium; Heterocyclic Compounds
PubMed: 38152996
DOI: 10.1021/jacs.3c06275 -
Pharmaceutics Dec 2023Antiangiogenic therapy with sorafenib (SF) alone is ineffective in eradicating tumors, and its long-term application can exacerbate tumor hypoxia, which in turn...
Antiangiogenic therapy with sorafenib (SF) alone is ineffective in eradicating tumors, and its long-term application can exacerbate tumor hypoxia, which in turn restricts SF's therapeutic efficacy. Here, a redox-responsive fluorinated peptide (DEN-TAT-PFC) consisting of dendritic poly-lysine, cell-penetrating peptide TAT, and perfluorocarbon was designed and synthesized to co-load siRNA-targeting hypoxia-inducible factors (siHIF-1α) and SF. The unique architecture of the peptide and fluorinated modifications enhanced the siRNA delivery efficiency, including increased siRNA binding, GSH-responsive release, cellular uptake, endosomal escape, and serum resistance. Simultaneously, the DEN-TAT-PFC/SF/siHIF-1α co-delivery system achieved efficient knockdown of HIF-1α at mRNA and protein levels, thus alleviating hypoxia and further substantially reducing VEGF expression. Additionally, the excellent oxygen-carrying ability of DEN-TAT-PFC may facilitate relief of the hypoxic microenvironment. As a result of these synergistic effects, DEN-TAT-PFC/SF/siHIF-1α exhibited considerable anti-tumor cell proliferation and anti-angiogenesis effects. Therefore, DEN-TAT-PFC can be a versatile platform for fabricating fluorine-containing drugs/siRNA complex nano-systems.
PubMed: 38140129
DOI: 10.3390/pharmaceutics15122789 -
Cells Nov 2023Sperm DNA fragmentation (SDF) that occurs during the freezing-thawing of sperm may negatively impact the treatment outcomes of assisted reproductive technologies (ART)....
Sperm DNA fragmentation (SDF) that occurs during the freezing-thawing of sperm may negatively impact the treatment outcomes of assisted reproductive technologies (ART). In a previous study, we developed a human sperm cryopreservation reagent containing carboxylated poly-L-lysine (CPLL) that reduced SDF after freeze-thawing compared with clinically popular cryopreservation reagents containing human serum albumin. However, it is unclear whether CPLL reduces SDF, as it differed from the constituents of the commercial cryopreservation reagents used for comparison. Therefore, here, we examined whether CPLL reduces the SDF of human sperm and evaluated reactive oxygen species (ROS) levels and lipid peroxidation (LPO), which are the causes of SDF; mitochondrial injury, ROS production; and impaired sperm motility. Furthermore, optimal antioxidants and their concentrations that could further enhance the reduction in SDF were determined for future clinical application in ART and underwent the same functional evaluations. CPLL can reduce SDF via inhibition of intracytoplasmic ROS and LPO. Furthermore, the addition of 0.1 mM resveratrol avoided the enhancement of SDF, which potentially affects mitochondrial and cytoplasmic ROS and LPO. This novel human sperm cryopreservation reagent containing CPLL and resveratrol has the potential to improve treatment outcomes in ART using frozen sperm.
Topics: Humans; Male; Freezing; Resveratrol; Polylysine; Reactive Oxygen Species; DNA Fragmentation; Cryoprotective Agents; Sperm Motility; Semen; Semen Preservation; Spermatozoa; Cryopreservation
PubMed: 37998320
DOI: 10.3390/cells12222585 -
Journal of Controlled Release :... Dec 2023Adipose-derived mesenchymal stem cell-derived small extracellular vesicles (Ad-MSC-sEVs/AMEs) combined with scaffold materials are used in tissue-engineered bladders;...
Negatively charged bladder acellular matrix loaded with positively charged adipose-derived mesenchymal stem cell-derived small extracellular vesicles for bladder tissue engineering.
Adipose-derived mesenchymal stem cell-derived small extracellular vesicles (Ad-MSC-sEVs/AMEs) combined with scaffold materials are used in tissue-engineered bladders; however, the lack of retention leads to limited distribution of AMEs in the scaffold areas and low bioavailability of AMEs after bladder reconstruction. To improve retention of AMEs, we developed a novel strategy that modifies the surface charge of the bladder acellular matrix (BAM) via oxidative self-polymerization of dopamine-reducing graphene oxide (GO) and AMEs using ε-polylysine-polyethylene-distearyl phosphatidylethanolamine (PPD). We evaluated two BAM surface modification methods and evaluated the biocompatibility of materials and PPD and electrostatic adherence effects between PPD-modified AMEs and rGO-PDA/BAM in vivo and in vitro. Surface modification increased retention of AMEs, enhanced regeneration of bladder structures, and increased electrical conductivity of rGO-PDA/BAM, thereby improving bladder function recovery. RNA-sequencing revealed 543 miRNAs in human AMEs and 514 miRNAs in rat AMEs. A Venn diagram was used to show target genes of miRNA with the highest proportion predicted by the four databases; related biological processes and pathways were predicted by KEGG and GO analyses. We report a strategy for improving bioavailability of AMEs for bladder reconstruction and reveal that enriched miR-21-5p targets PIK3R1 and activates the PI3K/Akt pathway to promote cell proliferation and migration.
Topics: Rats; Humans; Animals; Tissue Engineering; Urinary Bladder; Phosphatidylinositol 3-Kinases; Extracellular Matrix; Mesenchymal Stem Cells; MicroRNAs; Extracellular Vesicles
PubMed: 37944669
DOI: 10.1016/j.jconrel.2023.10.048 -
Polymers Oct 2023A major goal of regenerative medicine of the central nervous system is to accelerate the regeneration of nerve tissue, where astrocytes, despite their positive and...
A major goal of regenerative medicine of the central nervous system is to accelerate the regeneration of nerve tissue, where astrocytes, despite their positive and negative roles, play a critical role. Thus, scaffolds capable of producing astrocytes from neural precursor cells (NPCs) are most desirable. Our study shows that NPCs are converted into reactive astrocytes upon cultivation on coralline-derived calcium carbonate coated with poly-D-lysine (PDL-CS). As shown via nuclei staining, the adhesion of neurospheres containing hundreds of hippocampal neural cells to PDL-CS resulted in disaggregation of the cell cluster as well as the radial migration of dozens of cells away from the neurosphere core. Migrating cells per neurosphere averaged 100 on PDL-CS, significantly higher than on uncoated CS (28), PDL-coated glass (65), or uncoated glass (20). After 3 days of culture on PDL-CS, cell migration plateaued and remained stable for four more days. In addition, NPCs expressing nestin underwent continuous morphological changes from round to spiky, extending and elongating their processes, resembling activated astrocytes. The extension of the process increased continuously during the maturation of the culture and doubled after 7 days compared to day 1, whereas bifurcation increased by twofold during the first 3 days before plateauing. In addition, nestin positive cells' shape, measured through the opposite circularity level correlation, decreased approximately twofold after three days, indicating spiky transformation. Moreover, nestin-positive cells co-expressing GFAP increased by 2.2 from day 1 to 7, reaching 40% of the NPC population on day 7. In this way, PDL-CS promotes NPC differentiation into reactive astrocytes, which could accelerate the repair of neural tissue.
PubMed: 37896298
DOI: 10.3390/polym15204054 -
Foods (Basel, Switzerland) Oct 2023In this paper, amino-carboxymethyl chitosan (ACC) was prepared through amino carboxymethylation, which introduces -COOH and -NH groups to the chitosan (CS) chains....
In this paper, amino-carboxymethyl chitosan (ACC) was prepared through amino carboxymethylation, which introduces -COOH and -NH groups to the chitosan (CS) chains. Meanwhile, dialdehyde starch (DAS) was produced by oxidizing corn starch using sodium periodate. To attain the optimal loading and long-time release of ε-polylysine (ε-PL), the ACC/DAS hydrogels were synthesized through the Schiff base reaction between the amino group on ACC and the aldehyde group in DAS. The molecular structure, microcosmic properties, loading capacity, and bacteriostatic properties of the four types of hydrogels containing different mass concentrations of ACC were investigated. The results showed that the dynamic imine bond C=N existed in the ACC/DAS hydrogels, which proved that the hydrogels were formed by the cross-linking of the Schiff base reaction. With the increasing mass concentration of the ACC, the cross-sectional morphology of the hydrogel became smoother, the thermal stability increased, and the swelling behavior was gradually enhanced. The tight network structure improved the ε-PL loading efficiency, with the highest value of 99.2%. Moreover, the loading of ε-PL gave the hydrogel good antibacterial properties. These results indicate that ACC/DAS hydrogel is potential in food preservation.
PubMed: 37893700
DOI: 10.3390/foods12203807 -
Foods (Basel, Switzerland) Oct 2023Yan 73 () is a dyed grape variety cultivated in China. Currently, most studies have focused on the mechanism of anthocyanins or the impact of anthocyanins as auxiliary...
Yan 73 () is a dyed grape variety cultivated in China. Currently, most studies have focused on the mechanism of anthocyanins or the impact of anthocyanins as auxiliary color varieties on wine color. There is little research on its direct use or direct processing of products such as juice. In order to investigate the effects of different processing methods on the juice of Yan 73 grapes, the physicochemical and functional properties, as well as the sensory indexes of the juice, were analyzed by using thermal pasteurization (TP), thermosonication (TS), TS combined with nisin (TSN), TS combined with ε-Polylysine (TSε), irradiation (IR), and high hydrostatic pressure (HHP). The physicochemical indexes, functional properties, and sensory indexes of Smoke 73 grape juice were determined and analyzed. The results of the study showed that among the seven sterilization methods, total polyphenol content (TPC) in juice was significantly increased in all treatments except HHP. TPC was the highest in TP (3773.33 mg GAE/L). Total anthocyanin content (TAC) was increased except IR5, and TSN (1202.67 mg/L) had the highest TAC. In terms of color, TP (a* = 36.57, b* = 19.70, L* = 14.81, C* = 41.55, h° = 28.30, ΔE = 5.9) promotes the dissolution of anthocyanins because of high temperatures, which basically improves all the color indicators of grape juice and makes the color of grape juice more vivid. After HHP treatment, the color (ΔE = 1.72) and aroma indicators are closer to the grape juice itself. The Entropy weight-TOPSIS, CRITIC-Topsis, and PCA integrated quality evaluation models showed that all selected TP as the best integrated quality.
PubMed: 37893615
DOI: 10.3390/foods12203722