Influence of Ultrasound on the Characteristics of CaP Coatings Generated Via the Micro-arc Oxidation Process in Relation to Biomedical Engineering.

Over the past decade, bone tissue engineering has been at the core of attention because of an increasing number of implant surgeries. The purpose of this study was to obtain coatings on titanium (Ti) implants with improved properties in terms of biomedical applications and to investigate the effect of ultrasound (US) on these properties during the micro-arc oxidation (MAO) process. The influence of various process parameters, such as time and current density, as well as US mode, on the properties of such coatings was evaluated. Novel porous calcium-phosphate-based coatings were obtained on commercially pure Ti. Their microstructure, chemical composition, topography, wettability, nanomechanical properties, thickness, adhesion to the substrate, and corrosion resistance were analyzed. In addition, cytocompatibility evaluation was checked with the human osteoblasts. The properties of the coatings varied significantly, depending on applied process parameters. The US application during the MAO process contributes to the increase of coating thickness, porosity, roughness, and skewness, as well as augmented calcium incorporation. The most advantageous coating was obtained at a current of 136 mA, time 450 s, and unipolar rectangular US, as it exhibits high porosity, adequate wettability, and beneficial skewness, which enabled increased adhesion and proliferation of osteoblasts during in vitro studies. Finally, the conducted research demonstrated the influence of various UMAO process parameters, which allowed for the selection of appropriate Ti implant modification for specific biomedical utilization.

PVA-Based Films with Strontium Titanate Nanoparticles Dedicated to Wound Dressing Application.

Bioactive materials may be applied in tissue regeneration, and an example of such materials are wound dressings, which are used to accelerate skin healing, especially after trauma. Here, we proposed a novel dressing enriched by a bioactive component. The aim of our study was to prepare and characterize poly(vinyl alcohol) films modified with strontium titanate nanoparticles. The physicochemical properties of films were studied, such as surface free energy and surface roughness, as well as the mechanical properties of materials. Moreover, different biological studies were carried out, like in vitro hemo- and cyto-compatibility, biocidal activity, and anti-biofilm formation. Also, the degradation of the materials' utilization possibilities and enzymatic activity in compost were checked. The decrease of surface free energy, increase of roughness, and improvement of mechanical strength were found after the addition of nanoparticles. All developed films were cyto-compatible, and did not induce a hemolytic effect on the human erythrocytes. The PVA films containing the highest concentration of STO (20%) reduced the proliferation of Eschericha coli, Pseudomonas aeruginosa, and Staphylococcus aureus significantly. Also, all films were characterized by surface anti-biofilm activity, as they significantly lowered the bacterial biofilm abundance and its dehydrogenase activity. The films were degraded by the compost microorganism. However, PVA with the addition of 20%STO was more difficult to degrade. Based on our results, for wound dressing application, we suggest using bioactive films based on PVA + 20%STO, as they were characterized by high antibacterial properties, favorable physicochemical characteristics, and good biocompatibility with human cells.

Hyaluronic acid/tannic acid films for wound healing application.

In this study, thin films based on hyaluronic acid (HA) with tannic acid (TA) were investigated in three different weight ratios (80HA/20TA, 50HA/50TA, 20HA/80TA) for their application as materials for wound healing. Surface free energy, as well as their roughness, mechanical properties, water vapor permeability rate, and antioxidant activity were determined. Moreover, their compatibility with blood and osteoblast cells was investigated. The irritation effect caused by hyaluronic acid/tannic acid films was also considered with the use of are constructed human epidermis model. The irritation effect for hyaluronic acid/tannic acid films by the in vitro method was also studied. The low surface free energy, surface roughness, and antioxidant activity presented by the obtained films were examined. All the tested compositions of hyaluronic acid/tannic acid films were hemocompatible, but only films based on 50HA/50TA were fully cytocompatible. Regarding the potential implantation, all the films except 80HA/20TA showed appropriate mechanical properties. The specimens did not exert the irritation effect during the studies involving reconstructed human epidermis.

Cytocompatibility, antibacterial, and corrosion properties of chitosan/polymethacrylates and chitosan/poly(4-vinylpyridine) smart coatings, electrophoretically deposited on nanosilver-decorated titania nanotubes.

The development of novel implants subjected to surface modification to achieve high osteointegration properties at simultaneous antimicrobial activity is a highly current problem. This study involved different surface treatments of titanium surface, mainly by electrochemical oxidation to produce a nanotubular oxide layer (TNTs), a subsequent electrochemical reduction of silver nitrate and decoration of a nanotubular surface with silver nanoparticles (AgNPs), and finally electrophoretic deposition (EPD) of a composite of chitosan (CS) and either polymethacrylate-based copolymer Eudragit E 100 (EE100) or poly(4-vinylpyridine) (P4VP) coating. The effects of each stage of this multi-step modification were examined in terms of morphology, roughness, wettability, corrosion resistance, coating-substrate adhesion, antibacterial properties, and osteoblast cell adhesion and proliferation. The results showed that the titanium surface formed nanotubes (inner diameter of 97 ± 12 nm, length of 342 ± 36 nm) subsequently covered with silver nanoparticles (with a diameter of 88 ± 8 nm). Further, the silver-decorated nanotubes were tightly coated with biopolymer films. Most of the applied modifications increased both the roughness and the surface contact angle of the samples. The deposition of biopolymer coatings resulted in reduced burst release of silver. The coated samples revealed potent antimicrobial activity against both Gram-positive and Gram-negative bacteria. Total elimination (99.9%) of E. coli was recorded for a sample with CS/P4VP coating. Cytotoxicity results using hFOB 1.19, a human osteoblast cell line, showed that after 3 days the tested modifications did not affect the cellular growth according to the titanium control. The proposed innovative multilayer antibacterial coatings can be successful for titanium implants as effective postoperative anti-inflammation protection.

Dual-Setting Bone Cement Based On Magnesium Phosphate Modified with Glycol Methacrylate Designed for Biomedical Applications.

Magnesium phosphate cement (MPC) is a suitable alternative for the currently used calcium phosphates, owing to beneficial properties like favorable resorption rate, fast hardening, and higher compressive strength. However, due to insufficient mechanical properties and high brittleness, further improvement is still expected. In this paper, we reported the preparation of a novel type of dual-setting cement based on MPC with poly(2-hydroxyethyl methacrylate) (pHEMA). The aim of our study was to evaluate the effect of HEMA addition, especially its concentration and premix time, on the selected properties of the composite. Several beneficial effects were found: better formability, shortened setting time, and improvement of mechanical strengths. The developed cements were hardening in ∼16-21 min, consisted of well-crystallized phases and polymerized HEMA, had porosity between ∼2-11%, degraded slowly by ∼0.1-4%/18 days, their wettability was ∼20-30°, they showed compressive and bending strength between ∼45-73 and 13-20 MPa, respectively, and, finally, their Young's Modulus was close to ∼2.5-3.0 GPa. The results showed that the optimal cement composition is MPC+15%HEMA and 4 min of polymer premixing time. Overall, our research suggested that this developed cement may be used in various biomedical applications.

Evaluating Gelatin-Based Films with Graphene Nanoparticles for Wound Healing Applications.

In this study, gelatin-based films containing graphene nanoparticles were obtained. Nanoparticles were taken from four chosen commercial graphene nanoplatelets with different surface areas, such as 150 m2/g, 300 m2/g, 500 m2/g, and 750 m2/g, obtained in different conditions. Their morphology was observed using SEM with STEM mode; porosity, Raman spectra and elemental analysis were checked; and biological properties, such as hemolysis and cytotoxicity, were evaluated. Then, the selected biocompatible nanoparticles were used as the gelatin film modification with 10% concentration. As a result of solvent evaporation, homogeneous thin films were obtained. The surface's properties, mechanical strength, antioxidant activity, and water vapor permeation rate were examined to select the appropriate film for biomedical applications. We found that the addition of graphene nanoplatelets had a significant effect on the properties of materials, improving surface roughness, surface free energy, antioxidant activity, tensile strength, and Young's modulus. For the most favorable candidate for wound dressing applications, we chose a gelatin film containing nanoparticles with a surface area of 500 m2/g.

Gelatin and gelatin/starch-based films modified with sorbitol for wound healing.

Gelatin-based films modified with sorbitol were produced from gelatin solution or gelatin/starch blends using a simple and low-cost solvent casting method, and subsequently, their physicochemical, mechanical, and biocompatibility properties were characterized. This work focused on developing and optimizing a biopolymeric blend to improve the pure biopolymers' properties for potential biomedical applications such as wound dressing. The films were characterized in terms of morphology and transparency, mechanical, moisture and swelling properties, thermal stability, and degradation potential. Moreover, hemocompatibility, as well as cytocompatibility of prepared films, were examined. The addition of sorbitol contributed to improving mechanical properties, swelling reduction, and increasing biostability over time. The cytocompatibility of obtained films was confirmed in vitro with two different human cell lines, fibroblastic and osteoblastic, and a more favorable cellular response was received for fibroblasts. Further, in hemocompatibility studies, it was found that all films may be classified as non-hemolytic as they did not have a negative effect on the human erythrocytes. The obtained results indicate the great potential of the gelatin/starch blends modified with sorbitol as regenerative biomaterials intended for wound healing applications.

Chitosan-Based Membranes as Gentamicin Carriers for Biomedical Applications-Influence of Chitosan Molecular Weight.

Over the past decade, much attention has been paid to chitosan as a potential drug carrier because of its non-toxicity, biocompatibility, biodegradability and antibacterial properties. The effect of various chitosan characteristics on its ability to carry different antibiotics is discussed in the literature. In this work, we evaluated the influence of the different molecular weights of this polymer on its potential as an antibacterial membrane after adding gentamicin (1% w/w). Three types of chitosan membranes without and with antibiotic were prepared using a solvent casting process. Their microstructures were analyzed with a 4K digital microscope, and their chemical bonds were studied using FTIR spectroscopy. Furthermore, cytocompatibility on human osteoblasts and fibroblasts as well as antibacterial activity against Staphylococcus aureus (S. aureus.) and Escherichia coli (E. coli) were assessed. We observed that the membrane prepared from medium-molecular-weight chitosan exhibited the highest contact angle (≈85°) and roughness (10.96 ± 0.21 µm) values, and its antibacterial activity was unfavorable. The maximum tensile strength and Young's modulus of membranes improved and elongation decreased with an increase in the molecular weight of chitosan. Membranes prepared with high-molecular-weight chitosan possessed the best antibacterial activity, but mainly against S. aureus. For E. coli, is not advisable to add gentamicin to the chitosan membrane, or it is suggested to deplete its content. None of the fabricated membranes exhibited a full cytotoxic effect on osteoblastic and fibroblast cells. Based on our results, the most favorable membrane as a gentamicin carrier was obtained from high-molecular-weight chitosan.

The characterization of collagen-based scaffolds modified with phenolic acids for tissue engineering application.

The aim of the experiment was to study the morphology of collagen-based scaffolds modified by caffeic acid, ferulic acid, and gallic acid, their swelling, and degradation rate, as well as the biological properties of scaffolds, such as antioxidant activity, hemo- and cytocompatibility, histological observation, and antibacterial properties. Scaffolds based on collagen with phenolic acid showed higher swelling rate and enzymatic stability compared to scaffolds based on pure collagen, and the radical scavenging activity was in the range 85-91%. All scaffolds were non-hemolytic and compatible with surrounding tissues. Collagen modified by ferulic acid showed potentially negative effects on hFOB cells as a significantly increased LDH release was found, but all of the studied materials had antimicrobial activity against Staphylococcus aureus and Escherichia coli. It may be assumed that phenolic acids, such as caffeic, ferulic, and gallic acid, are modifiers and provide novel biological properties of collagen-based scaffolds. This paper provides the summarization and comparison of the biological properties of scaffolds based on collagen modified with three different phenolic acids.

Osteoblast and bacterial cell response on RGD peptide-functionalized chitosan coatings electrophoretically deposited from different suspensions on Ti13Nb13Zr alloy.

Metallic materials for long-term load-bearing implants still do not provide high antimicrobial activity while maintaining strong compatibility with bone cells. This study aimed to modify the surface of Ti13Nb13Zr alloy by electrophoretic deposition of a chitosan coating with a covalently attached Arg-Gly-Asp (RGD) peptide. The suspensions for coating deposition were prepared in two different ways either using hydroxyacetic acid or a carbon dioxide saturation process. The coatings were deposited using a voltage of 10 V for 1 min. The prepared coatings were examined using SEM, EDS, FTIR, and XPS techniques. In addition, the wettability of these surfaces, corrosion resistance, adhesion of the coatings to the metallic substrate, and their antimicrobial activity (E. coli, S. aureus) and cytocompatibility properties using the MTT and LDH assays were studied. The coatings produced tightly covered the metallic substrate. Spectroscopic studies confirmed that the peptide did not detach from the chitosan chain during electrophoretic deposition. All tested samples showed high corrosion resistance (corrosion current density measured in nA/cm2 ). The deposited coatings contributed to a significant increase in the antimicrobial activity of the samples against Gram-positive and Gram-negative bacteria (reduction in bacterial counts from 99% to, for CS-RGD-Acid and the S. aureus strain, total killing capacity). MTT and LDH results showed high compatibility with bone cells of the modified surfaces compared to the bare substrate (survival rates above 75% under indirect contact conditions and above 100% under direct contact conditions). However, the adhesion of the coatings was considered weak.

T regulatory cells metabolism: The influence on functional properties and treatment potential.

CD4+CD25highFoxP3+ regulatory T cells (Tregs) constitute a small but substantial fraction of lymphocytes in the immune system. Tregs control inflammation associated with infections but also when it is improperly directed against its tissues or cells. The ability of Tregs to suppress (inhibit) the immune system is possible due to direct interactions with other cells but also in a paracrine fashion via the secretion of suppressive compounds. Today, attempts are made to use Tregs to treat autoimmune diseases, allergies, and rejection after bone marrow or organ transplantation. There is strong evidence that the metabolic program of Tregs is connected with the phenotype and function of these cells. A modulation towards a particular metabolic stage of Tregs may improve or weaken cells' stability and function. This may be an essential tool to drive the immune system keeping it activated during infections or suppressed when autoimmunity occurs.

Effects of Marginal Zn Excess and Thiamine Deficiency on Microglial N9 Cell Metabolism and Their Interactions with Septal SN56 Cholinergic Cells.

Mild thiamine deficiency aggravates Zn accumulation in cholinergic neurons. It leads to the augmentation of Zn toxicity by its interaction with the enzymes of energy metabolism. Within this study, we tested the effect of Zn on microglial cells cultivated in a thiamine-deficient medium, containing 0.003 mmol/L of thiamine vs. 0.009 mmol/L in a control medium. In such conditions, a subtoxic 0.10 mmol/L Zn concentration caused non-significant alterations in the survival and energy metabolism of N9 microglial cells. Both activities of the tricarboxylic acid cycle and the acetyl-CoA level were not decreased in these culture conditions. Amprolium augmented thiamine pyrophosphate deficits in N9 cells. This led to an increase in the intracellular accumulation of free Zn and partially aggravated its toxicity. There was differential sensitivity of neuronal and glial cells to thiamine-deficiency-Zn-evoked toxicity. The co-culture of neuronal SN56 with microglial N9 cells reduced the thiamine-deficiency-Zn-evoked inhibition of acetyl-CoA metabolism and restored the viability of the former. The differential sensitivity of SN56 and N9 cells to borderline thiamine deficiency combined with marginal Zn excess may result from the strong inhibition of pyruvate dehydrogenase in neuronal cells and no inhibition of this enzyme in the glial ones. Therefore, ThDP supplementation can make any brain cell more resistant to Zn excess.

Metabolic and Cellular Compartments of Acetyl-CoA in the Healthy and Diseased Brain.

The human brain is characterised by the most diverse morphological, metabolic and functional structure among all body tissues. This is due to the existence of diverse neurons secreting various neurotransmitters and mutually modulating their own activity through thousands of pre- and postsynaptic interconnections in each neuron. Astroglial, microglial and oligodendroglial cells and neurons reciprocally regulate the metabolism of key energy substrates, thereby exerting several neuroprotective, neurotoxic and regulatory effects on neuronal viability and neurotransmitter functions. Maintenance of the pool of mitochondrial acetyl-CoA derived from glycolytic glucose metabolism is a key factor for neuronal survival. Thus, acetyl-CoA is regarded as a direct energy precursor through the TCA cycle and respiratory chain, thereby affecting brain cell viability. It is also used for hundreds of acetylation reactions, including N-acetyl aspartate synthesis in neuronal mitochondria, acetylcholine synthesis in cholinergic neurons, as well as divergent acetylations of several proteins, peptides, histones and low-molecular-weight species in all cellular compartments. Therefore, acetyl-CoA should be considered as the central point of metabolism maintaining equilibrium between anabolic and catabolic pathways in the brain. This review presents data supporting this thesis.

Properties of New Composite Materials Based on Hydroxyapatite Ceramic and Cross-Linked Gelatin for Biomedical Applications.

The main aim of the research was to develop a new biocompatible and injectable composite with the potential for application as a bone-to-implant bonding material or as a bone substitute. A composite based on hydroxyapatite, gelatin, and two various types of commercially available transglutaminase (TgBDF/TgSNF), as a cross-linking agent, was proposed. To evaluate the impacts of composite content and processing parameters on various properties of the material, the following research was performed: the morphology was examined by SEM microscopy, the chemical structure by FTIR spectroscopy, the degradation behavior was examined in simulated body fluid, the injectability test was performed using an automatic syringe pump, the mechanical properties using a nanoindentation technique, the surface wettability was examined by an optical tensiometer, and the cell viability was assayed by MTT and LDH. In all cases, a composite paste was successfully obtained. Injectability varied between 8 and 15 min. The type of transglutaminase did not significantly affect the surface topography or chemical composition. All samples demonstrated proper nanomechanical properties with Young's modulus and the hardness close to the values of natural bone. BDF demonstrated better hydrophilic properties and structural stability over 7 days in comparison with SNF. In all cases, the transglutaminase did not lead to cell necrosis, but cellular proliferation was significantly inhibited, especially for the BDF agent.

Resveratrol Inhibits Metabolism and Affects Blood Platelet Function in Type 2 Diabetes.

Chronic hyperglycemia contributes to vascular complications in diabetes. Resveratrol exerts anti-diabetic and anti-platelet action. This study aimed to evaluate the effects of resveratrol on metabolism and the function of blood platelets under static and in in vitro flow conditions in patients with type 2 diabetes. Blood obtained from 8 healthy volunteers and 10 patients with type 2 diabetes was incubated with resveratrol and perfused over collagen-coated capillaries. Isolated blood platelets were incubated with resveratrol and activated by collagen to assess platelet function, metabolism, ATP release, TXA2 production, lipid peroxidation, and gluthatione content. In the type 2 diabetes group, plasma glucose and fructosamine concentrations were significantly higher than in the healthy group. In in vitro studies, collagen-induced thrombi formation in the blood of diabetic patients was 33% higher than in the healthy group. Resveratrol reduced thrombi by over 50% in the blood of healthy and diabetic patients. TXA2 production was 47% higher in diabetic platelets than in the healthy group. Resveratrol reduced TXA2 release by 38% in healthy platelets and by 79% in diabetic platelets. Resveratrol also reduced the activities of enzymes responsible for glycolysis and oxidative metabolism in the platelets of both groups. These data indicate that the resveratrol-induced inhibition of platelet metabolism and TXA2 release may lead to a reduction of platelet function and thrombus formation in patients with type 2 diabetes. Therefore, resveratrol may be beneficial to prevent vascular complications as a future complementary treatment in aspirin-resistant diabetic patients.

Protection of Cholinergic Neurons against Zinc Toxicity by Glial Cells in Thiamine-Deficient Media.

Brain pathologies evoked by thiamine deficiency can be aggravated by mild zinc excess. Cholinergic neurons are the most susceptible to such cytotoxic signals. Sub-toxic zinc excess aggravates the injury of neuronal SN56 cholinergic cells under mild thiamine deficiency. The excessive cell loss is caused by Zn interference with acetyl-CoA metabolism. The aim of this work was to investigate whether and how astroglial C6 cells alleviated the neurotoxicity of Zn to cultured SN56 cells in thiamine-deficient media. Low Zn concentrations did not affect astroglial C6 and primary glial cell viability in thiamine-deficient conditions. Additionally, parameters of energy metabolism were not significantly changed. Amprolium (a competitive inhibitor of thiamine uptake) augmented thiamine pyrophosphate deficits in cells, while co-treatment with Zn enhanced the toxic effect on acetyl-CoA metabolism. SN56 cholinergic neuronal cells were more susceptible to these combined insults than C6 and primary glial cells, which affected pyruvate dehydrogenase activity and the acetyl-CoA level. A co-culture of SN56 neurons with astroglial cells in thiamine-deficient medium eliminated Zn-evoked neuronal loss. These data indicate that astroglial cells protect neurons against Zn and thiamine deficiency neurotoxicity by preserving the acetyl-CoA level.

Anticancer Properties of Amino Acid and Peptide Derivatives of Mycophenolic Acid.

BACKGROUND: Although Mycophenolic Acid (MPA) is applied as prodrugs in clinic as an immunosuppressant, it also possesses anticancer activity. MPA acts as Inosine-5'-Monophosphate Dehydrogenase (IMPDH) inhibitor, where the carboxylic group at the end of the side chain interacts with Ser 276 of the enzyme via hydrogen bonds. Therefore, MPA derivatives with other polar groups indicated high inhibition too. On the other hand, potent anticancer agents like dacarbazine and cisplatin give numerous side-effects. OBJECTIVE: Based on the literature data, MPA derivatives should be explored towards anticancer properties. Conversion of the carboxylic group of MPA to amide could maintain antiproliferative activity. Therefore, we decided to investigate several amino acid and peptide derivatives of MPA against chosen cancer cell lines in vitro. METHODS: Amides of MPA hold threonine and arginine amino acid unit. These amino acid derivatives were tested as L and D enantiomers and both in free acid and methyl esters forms. Additionally, MPA was modified with tuftsin or retro-tuftsin as biologically active peptides, which could act as a drug carrier. RESULTS: Amino acid and peptide derivatives of MPA were investigated in vitro as potential anticancer agents on cell lines: Ab melanoma, A375 melanoma and SHSY5Y neuroblastoma. The activity of the tested compounds was compared to parent MPA and known chemotherapeutics: dacarbazine and cisplatin. CONCLUSION: Amino acid moiety and the sequence of amino acids in the peptide part influenced observed activity. The most active amino acid MPA analogues occurred to be D and L-threonine derivatives as methyl esters, probably due to better cell membrane penetration.

Aggravated effects of coexisting marginal thiamine deficits and zinc excess on SN56 neuronal cells.

Objectives: Zinc excitotoxicity and thiamine pyrophosphate deficiency (TD) are known pathogenic signals contributing to mechanism of different encephalopathies through inhibition of enzymes responsible for energy metabolism such as pyruvate dehydrogenase, aconitase or ketoglutarate dehydrogenase. The aim of this work was to investigate whether subclinical Zn excess and TD, frequent in aging brain, may combine yielding overt neuronal impairment.Results: Clonal SN56 cholinergic neuronal cells of septal origin were used as the model of brain cholinergic neurons, which are particularly susceptible to neurodegeneration in the course of Alzheimer's disease, hypoxia and other dementia-linked brain pathologies. Neither subtoxic concentration of Zn (0.10 mM) nor mild 20-25% TD deficits alone caused significant negative changes in cultured cholinergic neurons viability and their acetyl-CoA/acetylcholine metabolism. However, cells with mild TD accumulated Zn in excess, which impaired their energy metabolism causing a loss of neurons viability and their function as neurotransmitters. These negative effects of Zn were aggravated by amprolium which is an inhibitor of thiamine intracellular transport.Conclusion: Our data indicate that TD may amplify otherwise non-harmful border-line Zn excitotoxic signals yielding progress of neurodegeneration.

Chloroacridine derivatives as potential anticancer agents which may act as tricarboxylic acid cycle enzyme inhibitors.

PURPOSE: This paper concerns the cytotoxicity of 9-chloro-1-nitroacridine (1a) and 9-chloro-4-methyl-1-nitroacridine (1b) against two biologically different melanoma forms: melanotic and amelanotic. Melanomas are tumors characterized by high heterogeneity and poor susceptibility to chemotherapies. Among new analogs synthesized by us, compound 1b exhibited the highest anticancer potency. Because of that, in this study, we analyzed the mechanism of action for 1a and its 4-methylated derivative, 1b, against a pair of biological melanoma forms, with regard to proliferation, cell death mechanism and energetic state. METHODS: Cytotoxicity was evaluated by XTT assay. Cell death was estimated by plasma membrane structure changes (phosphatidylserine externalization), caspase activation, and ROS presence. The energetic state of cells was estimated based on NAD and ATP levels, and the activity of tricarboxylic acid cycle enzymes (pyruvate dehydrogenase complex, aconitase, isocitrate dehydrogenase). RESULTS: The chloroacridines affect biological forms of melanoma in different ways. Amelanotic (Ab) melanoma (with inhibited melanogenesis and higher malignancy) was particularly sensitive to the action of the chloroacridines. The Ab melanoma cells died through apoptosis and through death without caspase activation. Diminished activity of TAC enzymes was noticed among Ab melanoma cells together with ATP/NAD depletion, especially in the case of 1b. CONCLUSION: Our data show that the biological forms of the tumors responded to 1a and its 4-methylated analog in different ways. 1a and 1b could be inducers of regulated melanoma cell death, especially the amelanotic form. Although the mechanism of the cell death is not fully understood, 1b may act by interfering with the TAC enzymes and blocking specific pathways leading to tumor growth. This could encourage further investigation of its anticancer activity, especially against the amelanotic form of melanoma.

Synthesis and Biological Evaluation of Acridine/Acridone Analogs as Potential Anticancer Agents.

BACKGROUND: The lack of efficacious therapy for advanced melanoma and neuroblastoma makes new approaches necessary. Therefore, many scientists seek new, more effective, more selective and less toxic anticancer drugs. OBJECTIVE: We propose the synthesis of the new functionalized analogs of 1-nitroacridine/4- nitroacridone connected to tuftsin/retro-tuftsin derivatives as potential anticancer agents. METHODS: Acridine and acridone analogues were prepared by Ullmann condensation and then cyclization reaction. As a result of nucleophilic substitution reaction 1-nitro-9-phenoxyacridine or 1- chloro-4-nitro-9(10H)-acridone with the corresponding peptides, the planned acridine derivatives (10a-c, 12, 17-a-d, 19) have been obtained. The cytotoxic activity of the newly obtained analogs were evaluated against melanotic (Ma) and amelanotic (Ab) melanoma cell lines and neuroblastoma SH-SY5Y by using the XTT method. Apoptosis and cell cycle were analyzed by flow cytometry. RESULTS: Among the investigated analogs compound 12 exhibited the highest potency comparable to dacarbazine action for amelanotic Ab melanoma cells. FLICA test (flurochrome-labeled inhibitors of caspases) showed that this analog significantly increased the content of cells with activated caspases (C+) among both neuroblastoma lines and only Ab melanoma line. Using phosphatidylserine (PS) externalization assay, 12 induced changes in the Ab melanoma plasma membrane structure as the externalization of phosphatidylserine (An+ cells). These changes in neuroblastoma cells were less pronounced. CONCLUSION: Analog 12 could be proposed as the new potential chemotherapeutic against amelanotic melanoma form especially.