3D-bioprinted GelMA/gelatin/amniotic membrane extract (AME) scaffold loaded with keratinocytes, fibroblasts, and endothelial cells for skin tissue engineering.

Gelatin-methacryloyl (GelMA) is a highly adaptable biomaterial extensively utilized in skin regeneration applications. However, it is frequently imperative to enhance its physical and biological qualities by including supplementary substances in its composition. The purpose of this study was to fabricate and characterize a bi-layered GelMA-gelatin scaffold using 3D bioprinting. The upper section of the scaffold was encompassed with keratinocytes to simulate the epidermis, while the lower section included fibroblasts and HUVEC cells to mimic the dermis. A further step involved the addition of amniotic membrane extract (AME) to the scaffold in order to promote angiogenesis. The incorporation of gelatin into GelMA was found to enhance its stability and mechanical qualities. While the Alamar blue test demonstrated that a high concentration of GelMA (20%) resulted in a decrease in cell viability, the live/dead cell staining revealed that incorporation of AME increased the quantity of viable HUVECs. Further, gelatin upregulated the expression of KRT10 in keratinocytes and VIM in fibroblasts. Additionally, the histological staining results demonstrated the formation of well-defined skin layers and the creation of extracellular matrix (ECM) in GelMA/gelatin hydrogels during a 14-day culture period. Our study showed that a 3D-bioprinted composite scaffold comprising GelMA, gelatin, and AME can be used to regenerate skin tissues.

Exploring the various effects of Cu doping in hydroxyapatite nanoparticle.

Adding foreign ions to hydroxyapatite (HAp) is a popular approach for improving its properties. This study focuses on the effects of calcium substitution with copper in HAp. Instead of calcium, copper ions were doped into the structure of hydroxyapatite nanoparticles at 1%, 3%, and 5% concentrations. XRD analysis showed that the amount of substituted copper was less than needed to generate a distinct phase, yet its lattice parameters and crystallinity slightly decreased. Further, the results of degradation tests revealed that copper doping in hydroxyapatite doubled calcium ion release in water. The incorporation of copper into the apatite structure also boosted the HAp zeta potential and FBS protein adsorption onto powders. According to antibacterial investigations, a concentration of 200 mg/ml of hydroxyapatite containing 5% copper was sufficient to effectively eradicate E. coli and S. aureus bacteria. Furthermore, copper improved hydroxyapatite biocompatibility. Alkaline phosphatase activity and alizarin red tests showed that copper in hydroxyapatite did not inhibit stem cell differentiation into osteoblasts. Also, the scratch test demonstrated that copper-containing hydroxyapatite extract increased HUVEC cell migration. Overall, our findings demonstrated the utility of incorporating copper into the structure of hydroxyapatite from several perspectives, including the induction of antibacterial characteristics, biocompatibility, and angiogenesis.

Role of P11 through serotonergic and glutamatergic pathways in LID.

Parkinson's disease is a progressive neurodegenerative disorder caused by the degeneration of dopaminergic neurons. This leads to the pathogenesis of multiple basal ganglia-thalamomotor loops and diverse neurotransmission alterations. Dopamine replacement therapy, and on top of that, levodopa and l-3,4-dihydroxyphenylalanine (L-DOPA), is the gold standard treatment, while it develops numerous complications. Levodopa-induced dyskinesia (LID) is well-known as the most prominent side effect. Several studies have been devoted to tackling this problem. Studies showed that metabotropic glutamate receptor 5 (mGluR5) antagonists and 5-hydroxytryptamine receptor 1B (5HT1B) agonists significantly reduced LID when considering the glutamatergic overactivity and compensatory mechanisms of serotonergic neurons after L-DOPA therapy. Moreover, it is documented that these receptors act through an adaptor protein called P11 (S100A10). This protein has been thought to play a crucial role in LID due to its interactions with numerous ion channels and receptors. Lately, experiments have shown successful evidence of the effects of P11 blockade on alleviating LID greater than 5HT1B and mGluR5 manipulations. In contrast, there is a trace of ambiguity in the exact mechanism of action. P11 has shown the potential to be a promising target to diminish LID and prolong L-DOPA therapy in parkinsonian patients owing to further studies and experiments.

3D printing of complicated GelMA-coated Alginate/Tri-calcium silicate scaffold for accelerated bone regeneration.

Polymer-based composite scaffolds are an attractive class of biomaterials due to their suitable physical and mechanical performance as well as appropriate biological properties. When such composites contain osteoinductive ceramic nanopowders, it is possible, in principle, to stimulate the seeded cells to differentiate into osteoblasts. However, reproducibly fabricating and developing an appropriate niche for cells' activities in three-dimensional (3D) scaffolds remains a challenge using conventional fabrication techniques. Additive manufacturing provides a new strategy for the fabrication of complex 3D structures. Here, an extrusion-based 3D printing method was used to fabricate the Alginate (Alg)/Tri-calcium silicate (C3S) bone scaffolds. To improve physical and biological attributes, scaffolds were coated with gelatin methacryloyl (GelMA), a biocompatible viscose hydrogel. Conducting a combination of experimental techniques and molecular dynamics simulations, it is found that the composition ratio of Alg/C3S governs intermolecular interactions among the polymer and ceramic, affecting the product performance. Investigating the effects of various C3S amounts in the bioinks, the 90/10 composition ratio of Alg/C3S is known as the optimum content in developed bioinks. Accordingly, the printability of high-viscosity inks is boosted by improved hierarchical interactions among assemblies, which in turn leads to better nanoscale alignment in extruded macroscopic filaments. Conducting multiple tests on specimens, the GelMA-coated Alg/C3S scaffolds (with a composition ratio of 90/10) were shown to have improved mechanical qualities and cell adhesion, spreading, proliferation, and osteogenic differentiation, compared to the bare scaffolds, making them better candidates for further future research. Overall, the in-silico and in vitro studies of GelMA-coated 3D-printed Alg/C3S scaffolds open new aspects for biomaterials aimed at the regeneration of large- and complicated-bone defects through modifying the extrusion-based 3D-printed constructs.

Synergy effects of copper and L-arginine on osteogenic, angiogenic, and antibacterial activities.

Copper (Cu) ions have been found to exert antibacterial and angiogenic effects. However, some studies have indicated that it inhibits osteogenesis at high concentrations. On the other hand, L-arginine (Arg) is a semi-essential amino acid required for various biological processes, including osteogenic and angiogenic activities. As a result, we hypothesized that combining Arg with Cu ions would reduce its inhibitory effects on osteogenesis while increasing its angiogenic and antibacterial capabilities. To assess osteogenic and angiogenic activities, we employed rat bone marrow mesenchymal stem cells (MSCs) and human umbilical vein endothelial cells (HUVECs), respectively. The gram-positive bacteria Staphylococcus epidermidis (S. epidermidis), Staphylococcus aureus (S. aureus), and the gram-negative bacterium Escherichia coli (E. coli) were used to investigate bacterial behaviors. According to ALP activity and calcium deposition outcomes, copper ions inhibited osteogenic development of MSCs at 100 µM; however, Arg supplementation somewhat mitigated the inhibitory effects. Furthermore, Copper and Arg synergistically stimulated migration and tube formation of HUVECs. According to our findings, copper ions and Arg in the range of 1-100 µM had no antibacterial effect on any examined bacteria. However, at a dose of 20 mM, copper demonstrated antibacterial activity, which was boosted by Arg. Overall, these findings suggest that a combination of copper and Arg may be more beneficial for bone regeneration than either copper or Arg alone.

Anti-IgE monoclonal antibodies as potential treatment in COVID-19.

Coronavirus disease 2019 (COVID-19) is associated with irreversible effects on vital organs, especially the respiratory and cardiac systems. While the immune system plays a key role in the survival of patients to viral infections, in COVID-19, there is a hyperinflammatory immune response evoked by all the immune cells, such as neutrophils, monocytes, and includes release of various cytokines, resulting in an exaggerated immune response, named cytokine storm. This severe, dysregulated immune response causes multi-organ damage, which eventually leads to high mortality. One of the most important components of hypersensitivity is immunoglobulin E (IgE), which plays a major role in susceptibility to respiratory infections and can lead to the activation of mast cells. There is also a negative association between IgE and IFN-α, which can reduce Toll-like receptor (TLR) nine receptor expression and TLR-7 signaling to disrupt IFN production. Moreover, anti-IgE drugs such as omalizumab reduces the severity and duration of COVID-19. In addition to its anti-IgE effect, omalizumab inhibits inflammatory cells such as neutrophils. Hence, blockade of IgE may have clinical utility as an immunotherapy for COVID-19.

Loss of WD2 subdomain of Apaf-1 forms an apoptosome structure which blocks activation of caspase-3 and caspase-9.

Split luciferase complementary assay has been used to investigate the effect of WD domain deletion on Apaf-1 oligomerization. Apaf-1 is an adaptor molecule in formation of apoptosome that activates caspase-9, an activation that is a key event in the mitochondrial cell death pathway. Structural studies suggest that normally Apaf-1 is held in an inactive conformation by intramolecular interactions between Apaf-1's nucleotide binding domain and one of its WD40 domains (WD1). In the prevailing model of Apaf-1 activation, cytochrome c binds to sites in WD1 and in Apaf-1's second WD40 domain (WD2), moving WD1 and WD2 closer together and rotating WD1 away from the nucleotide binding domain. This allows Apaf-1 to bind dATP or ATP and to form the apoptosome, which activates caspase-9. This model predicts that cytochrome c binding to both WD domains is necessary for apoptosome formation and that an Apaf-1 with only WD1 will be locked in an inactive conformation that cannot be activated by cytochrome c. Here we investigated the effect of removing one WD domain (Apaf-1 1-921) on Apaf-1 interactions and caspase activation. Apaf-1 1-921 could not activate caspase-9, even in the presence of cytochrome c. These data show that a single WD domain is sufficient to lock Apaf-1 in an inactive state and this state cannot be altered by cytochrome c.

A review of fibrin and fibrin composites for bone tissue engineering.

Tissue engineering has emerged as a new treatment approach for bone repair and regeneration seeking to address limitations associated with current therapies, such as autologous bone grafting. While many bone tissue engineering approaches have traditionally focused on synthetic materials (such as polymers or hydrogels), there has been a lot of excitement surrounding the use of natural materials due to their biologically inspired properties. Fibrin is a natural scaffold formed following tissue injury that initiates hemostasis and provides the initial matrix useful for cell adhesion, migration, proliferation, and differentiation. Fibrin has captured the interest of bone tissue engineers due to its excellent biocompatibility, controllable biodegradability, and ability to deliver cells and biomolecules. Fibrin is particularly appealing because its precursors, fibrinogen, and thrombin, which can be derived from the patient's own blood, enable the fabrication of completely autologous scaffolds. In this article, we highlight the unique properties of fibrin as a scaffolding material to treat bone defects. Moreover, we emphasize its role in bone tissue engineering nanocomposites where approaches further emulate the natural nanostructured features of bone when using fibrin and other nanomaterials. We also review the preparation methods of fibrin glue and then discuss a wide range of fibrin applications in bone tissue engineering. These include the delivery of cells and/or biomolecules to a defect site, distributing cells, and/or growth factors throughout other pre-formed scaffolds and enhancing the physical as well as biological properties of other biomaterials. Thoughts on the future direction of fibrin research for bone tissue engineering are also presented. In the future, the development of fibrin precursors as recombinant proteins will solve problems associated with using multiple or single-donor fibrin glue, and the combination of nanomaterials that allow for the incorporation of biomolecules with fibrin will significantly improve the efficacy of fibrin for numerous bone tissue engineering applications.

Mitochondrial complex I deficiency and ATP/ADP ratio in lymphocytes of amyotrophic lateral sclerosis patients.

OBJECTIVES: Several lines of evidence suggest that mitochondrial dysfunction is involved in amyotrophic lateral sclerosis (ALS), but despite the fact that mitochondria play a central role in excitotoxicity, oxidative stress, and apoptosis, the intimate underlying mechanism linking mitochondrial defects to motor neuron degeneration in ALS still remains elusive. This study was performed to assess the mitochondrial respiratory chain dysfunction and cellular energy index (ATP/ADP ratio) in lymphocytes of ALS patients. METHODS: In this study, activity of mitochondrial respiratory chain complex I (measured as NADH-ferricyanide reductase) and both intracellular ATP and ADP measurements were performed on lymphocytes of ALS patients (n = 14) and control subjects (n = 26). Then, ATP/ADP ratio was calculated. RESULTS: Our finding showed that in patients compared with controls, complex I activity and intracellular ATP were significantly reduced (P = 0·001) and intracellular ADP content was increased (P<0·005) and ATP/ADP ratio subsequently was decreased and also we found strong correlation between complex I activity and intracellular ATP content and strong reverse correlation between complex I activity and intracellular ADP content in the patients with ALS (r(2) = 0·90). DISCUSSION: This study suggests that complex I deficiency and both reduction in intracellular ATP and increase in intracellular ADP content may be involved in the progression and pathogenesis of ALS.