Targeted Anticancer Drug Delivery Using Chitosan, Carbon Quantum Dots, and Aptamers to Deliver Ganoderic Acid and 5-Fluorouracil.

Breast cancer is a malignancy that affects mostly females and is among the most lethal types of cancer. The ligand-functionalized nanoparticles used in the nano-drug delivery system offer enormous potential for cancer treatments. This work devised a promising approach to increase drug loading efficacy and produce sustained release of 5-fluorouracil (5-FU) and Ganoderic acid (GA) as model drugs for breast cancer. Chitosan, aptamer, and carbon quantum dot (CS/Apt/COQ) hydrogels were initially synthesized as a pH-sensitive and biocompatible delivery system. Then, CS/Apt/COQ NPs loaded with 5-FU-GA were made using the W/O/W emulsification method. FT-IR, XRD, DLS, zeta potentiometer, and SEM were used to analyze NP's chemical structure, particle size, and shape. Cell viability was measured using MTT assays in vitro using the MCF-7 cell lines. Real-time PCR measured cell apoptotic gene expression. XRD and FT-IR investigations validated nanocarrier production and revealed their crystalline structure and molecular interactions. DLS showed that nanocarriers include NPs with an average size of 250.6 nm and PDI of 0.057. SEM showed their spherical form, and zeta potential studies showed an average surface charge of +37.8 mV. pH 5.4 had a highly effective and prolonged drug release profile, releasing virtually all 5-FU and GA in 48 h. Entrapment efficiency percentages for 5-FU and GA were 84.7±5.2 and 80.2 %±2.3, respectively. The 5-FU-GA-CS-CQD-Apt group induced the highest cell death, with just 57.9 % of the MCF-7 cells surviving following treatment. 5-FU and GA in CS-CQD-Apt enhanced apoptotic induction by flow cytometry. 5-FU-GA-CS-CQD-Apt also elevated Caspase 9 and downregulated Bcl2. Accordingly, the produced NPs may serve as pH-sensitive nano vehicles for the controlled release of 5-FU and GA in treating breast cancer.

An electrochemical aptasensor for detection of prostate-specific antigen using reduced graphene gold nanocomposite and Cu/carbon quantum dots.

We report a label-free electrochemical aptamer-based biosensor for the detection of human prostate-specific antigen (PSA). The thiolate DNA aptamer against PSA was conjugated to the reduced graphene oxide/Au (RGO-Au) nanocomposite through the self-assembly of Au-S groups. Owing to the large volume to surface ratio, the RGO-Au nanocomposite provides a large surface for aptamer loading. The RGO-Au/aptamer was combined with a Nafion polymer and immobilized on a glassy carbon electrode. The interaction of aptamer with PSA was studied by cyclic voltammetry, square wave voltammetry, and electrochemical impedance spectroscopy. The detection of limit for prepared electrode was obtained about 50 pg/mL at the potential of 0.4 V in potassium hexacyanoferrate [K4 Fe(CN)6 ] medium. To decrease the limit of detection (LOD) and applied potential of the prepared nanoprobe Cu/carbon quantum dots (CuCQD) is introduced as a new redox. The results show that this new electrochemical medium provides better conditions for the detection of PSA. LOD of a nanoprobe in CuCQD media was obtained as 40 pg/mL at the potential of -0.2 V. Under optimal conditions, the aptasensor exhibits a linear response to PSA with a LOD as small as 3 pg/mL. The present aptasensor is highly selective and sensitive and shows satisfactory stability and repeatability.

Investigating the effect of nanoparticle on phenanthrene biodegradation by Labedella gwakjiensis strain KDI.

Polycyclic aromatic hydrocarbons (PAHs), as persistent organic contaminants, are a major source of concern due to their toxic effect on ecosystems and human health. This study attempted to isolate halotolerant PAHs degrading bacteria from saline oil-contaminated soils. Among the isolates, strain KDI with the highest 16S rRNA gene sequence similarity to Labedella gwakjiensis was able to reduce surface tension (ST) from 65.42 to 26.60 mN m-1 and increase the emulsification index to 81.04%, as a result of significant biosurfactant production. Response Surface Methodology (RSM) analysis was applied to optimize the factors, i.e. PAHs concentration and NaCl concentration as well as to determine the effect of these important variables on PAHs biodegradation. The Carbon Quantum Dots. Iron Oxide (CQDs.Fe3O4) nanoparticles were characterized by several popular analytical techniques, after which the effect of CQD.Fe3O4 nanoparticles on biodegradation was examined. PAHs biodegradation rate and efficiency of strain KDI to degrade PHE in the presence of CQD.Fe3O4 nanoparticles was analyzed by GC. According to the results during biodegradation both the concentration of PAHs and the amount of NaCl were effective. The biodegradation rate significantly increased in the presence of CQD.Fe3O4. The highest biodegradation of PHE occurred in the presence of 0.5 g/L of CQD.Fe3O4 which was 63.63% and 81.77% after 48 and 72 h of incubation. To the best of our knowledge, this is the first report on optimization of PAHs concentration and salinity by RSM and nanobioremediation of PHE using a bacterial strain in the presence of CQD.Fe3O4 nanoparticles.

The Effect of Trehalose Coating for Magnetite Nanoparticles on Stability of Egg White Lysozyme.

In this study, the protein stability of hen egg-white lysozymes (HEWL) by Fe3O4 and Fe3O4-coated trehalose (Fe3O4@Tre) magnetic nanoparticles (NPs) is investigated. For this purpose, the co-precipitation method was used to synthesize magnetic NPs. The synthesized NPs were characterized by XRD, FT-IR spectroscopy, FE-SEM, and VSM analysis. In addition, the stability of HEWLs exposed to different NP concentrations in the range of 0.001-0.1 mg mL-1 was investigated by circular dichroism (CD) spectroscopy, fluorescence, and UV-Vis analysis. Based on the results, in the NP concentration range of 0.001-0.04 mg mL-1 the protein structure is more stable, and this range was identified as the range of kosmotropic concentration. The helicity was measured at two concentration points of 0.02 and 0.1 mg mL-1. According to the results, the α-helix at 0.02 mg mL-1 of Fe3O4 and Fe3O4@Tre was increased from 35.5% for native protein to 37.7% and 38.7%, respectively. The helicity decreased to 36.1% and 37.4%, respectively, with increasing the concentration of Fe3O4 and Fe3O4@Tre to 0.1 mg mL-1. The formation of hydrated water shells around protein molecules occurred by using Fe3O4@Tre NPs. Hence, it can be concluded that the trehalose as a functional group along with magnetic NPs can improve the stability of proteins in biological environments.

Chitosan/Gamma-Alumina/Fe3O4@5-FU Nanostructures as Promising Nanocarriers: Physiochemical Characterization and Toxicity Activity.

Today, cancer treatment is an important issue in the medical world due to the challenges and side effects of ongoing treatment procedures. Current methods can be replaced with targeted nano-drug delivery systems to overcome such side effects. In the present work, an intelligent nano-system consisting of Chitosan (Ch)/Gamma alumina (γAl)/Fe3O4 and 5-Fluorouracil (5-FU) was synthesized and designed for the first time in order to influence the Michigan Cancer Foundation-7 (MCF-7) cell line in the treatment of breast cancer. Physico-chemical characterization of the nanocarriers was carried out using X-ray diffraction (XRD), Fourier-transform infrared spectroscopy (FTIR), vibrating sample magnetometry (VSM), dynamic light scattering (DLS), and scanning electron microscopy (SEM). SEM analysis revealed smooth and homogeneous spherical nanoparticles. The high stability of the nanoparticles and their narrow size distribution was confirmed by DLS. The results of the loading study demonstrated that these nano-systems cause controlled, stable, and pH-sensitive release in cancerous environments with an inactive targeting mechanism. Finally, the results of MTT and flow cytometry tests indicated that this nano-system increased the rate of apoptosis induction on cancerous masses and could be an effective alternative to current treatments.

A novel delivery of curcumin by the efficient nanoliposomal approach against Leishmania major.

Several side effects and drug resistance accompany the current therapies for Leishmaniasis. Nanoliposomal curcumin is applied as a new therapy approach instead of current therapy. In this study, nanoliposomal curcumin was prepared using thin-film hydration method and characterized based on encapsulation efficiency, size, and zeta potential. Curcumin was successfully loaded into nanoliposomes with an encapsulation efficiency of 92%. The surface charge of the nanoparticle was neutral, and the size of nanoparticle was 176.5 nm. Nanoliposomal curcumin is in spherical shape without any agglomeration. Cell viability assay was performed on HFF cell line to show biocompatibility of liposome nanoparticles. Anti-Leishmanial effect of different concentrations of liposomal curcumin (0.05-30 µg mL-1) and amphotericin B (25 µg mL-1) were studied on Leishmania major [MRHO/IR/75/ER] at various hours (24, 48, and 72) using hemocytometer technique. Nanoliposomal curcumin inhibitory concentration (IC50) at hours 24, 48, and 72 were 6.41, 3.8, and 2.33 µg mL-1, respectively. As prepared nanoliposomal curcumin showed a significant antileishmanial effect and induced a better and more tangible effect on the survival of L. major promastigotes and could be suitable candidates for further investigations.

Detection of Microorganisms Using Graphene-Based Nanobiosensors.

Having an insight into graphene and graphene derivatives such as graphene oxide, reduced graphene oxide and graphene quantum dots is necessary since it can help scientists to detect possible properties and features that could be useful when using these carbon materials in preparation of a nanocomposites. In recent years, graphene and its derivatives have attracted a lot of attention and been extensively applied in biosensors due to fascinating properties, such as large surface area, optical and magnetic properties, and high elasticity for the detection of microorganisms as they can be modified with some other materials such as macromolecules, oxide metals and metals to improve the electrochemical behaviour of the biosensor. In this review paper, biosensor design strategies based on graphene and its derivatives (graphene-based nanocomposites in biosensors) are described. Then their application for the detection of microorganisms including prions, viroids, viral and bacterial cells as well as fungi, protozoa, microbial toxins and even microbial sources of antibiotics is reviewed.

The Effect of Calcium/Magnesium Ratio on the Biomass Production of a Novel Thermoalkaliphilic Aeribacillus pallidus Strain with Highly Heat-Resistant Spores.

Hot springs are fascinating extreme environments for the isolation of polyextremophilic microorganisms with extraordinary characteristics. Since polyextremophilic bacterial growth are not as high as routine bacteria, the objective of this study was to investigate the effect of some environmental factors on biomass and metabolites productions in the newly isolated strain, from Larijan hot spring in Iran. The strain was identified as Aeribacillus pallidus Lhs-10 and deposited as CCUG 72355 and IBRC-M 11202 in Sweden and Iran, respectively. This thermoalkaliphilic strain can grow best at 50 °C, pH 8 and in the presence of 25 g/l NaCl. The physiological characterization of this strain show that [Ca/Mg] ratio affect its growth and biomass production with the best results obtained at the ratio of 2.5. Moreover, lactic and acetic acids production by this strain was affected by pH, aeration, and temperature, where a metabolic shift was detected from lactate to acetate production when the culture was aerated. Besides, its spores could tolerate heating at 80, 85, 90, 95 and 98 °C for 30 min without any reduction in the initial spore population, whereas D-value was defined 50 min at 98 °C. This newly lactic acid-producing strain of A. pallidus can be a promising strain that can be used in the harsh conditions in industrial processes.

Novel microfluidic graphene oxide-protein amperometric biosensor for detecting sulfur compounds.

Sulfur compounds are essential for many industries and organisms; however, they cause serious respiratory problems in human beings. Therefore, determination of sulfur concentration is of paramount importance. The research approach in the field of detecting contaminants has led to smaller systems that provide faster and more effective ways for diagnosis purposes. In this study, a novel portable amperometric graphene oxide-protein biosensor platform is investigated. The main characteristic of this structure is the implementation of a microfluidic configuration. With albumin metalloprotein as the biorecognition element, graphene oxide was synthesized and characterized by transmission electron microscopy and Fourier-transform infrared spectroscopy (FTIR). Albumin protein was stabilized on the surface of graphene oxide by the application of the N-(3-dimethylamionpropyl)-N-ethylcarbodiimide hydrochloride/N-hydroxysuccinimide method. The stabilization was confirmed by FTIR and electrochemistry analyses. The calibration curve of sulfur concentration was determined. When the graphene oxide-protein complex was stabilized by nephion on the surface of the microfluidic system, the response time reduced to 50 Sec, which is a relatively faster response among the similar studies and validated the significant effect of the microfluidic system. The nanosystem had an optimized pH of 7.4 and exhibited high sensitivity in determining sulfide. The results confirm that the portable graphene oxide-protein nanosystem has a fast and accurate response in detecting sulfide.

Graphene oxide-l-arginine nanogel: A pH-sensitive fluorouracil nanocarrier.

Nowadays, putting forward an accurate cancer therapy method with minimal side effects is an important topic of research. Nanostructures, for their ability in controlled and targeted drug release on specific cells, are critical materials in this field. In this study, a pH-sensitive graphene oxide-l-arginine nanogel was synthesized to carry and release 5-fluorouracil. Optimized conditions using statistical analysis, based on the maximum relative viscosity of nanogel, were evaluated: 5.489 for the concentration of l-arginine and 2.404 for pH. The prepared nanogels were characterized using scanning electron microscope and transmission electron microscope images and Fourier-transform infrared spectroscopic analysis. Cytotoxicity was assessed using the sulforhodamine B (SRB) assay on MCF-7 breast cancer cells. The fluorouracil release was measured by the dialysis bag method, UV spectrophotometry, and fluorouracil calibration diagram. Results proved the successful controlled release of fluorouracil at pH 5.4 and the beneficial role of graphene-oxide- l-arginine- fluorouracil nanogel in eliminating cancer cells.

A glassy carbon electrode modified with reduced graphene oxide and gold nanoparticles for electrochemical aptasensing of lipopolysaccharides from Escherichia coli bacteria.

An electrochemical aptasensor is described for the voltammetric determination of lipopolysaccharide (LPS) from Escherichia coli 055:B5. Aptamer chains were immobilized on the surface of a glassy carbon electrode (GCE) via reduced graphene oxide and gold nanoparticles (RGO/AuNPs). Fast Fourier transform infrared, X-ray diffraction and transmission electron microscopy were used to characterize the nanomaterials. Cyclic voltammetry, square wave voltammetry and electrochemical impedance spectroscopy were used to characterize the modified GCE. The results show that the modified electrode has a good selectivity for LPS over other biomolecules. The hexacyanoferrate redox system, typically operated at around 0.3 V (vs. Ag/AgCl) is used as an electrochemical probe. The detection limit is 30 fg·mL-1. To decrease the electrochemical potential for detection of LPS, Mg/carbon quantum dots were used as redox active media. They decrease the detection potentialto 0 V and the detection of limit (LOD) to 1 fg·mL-1. The electrode was successfully used to analyze serum of patients and healthy persons. Graphical abstractSchematic representation of the modification of reduced graphene oxide gold nanoparticles with aptamer chains to immobilize on the glassy carbon electrode surface for electrochemical detection of lipopolysaccharides.

Optimizing the nanostructure of graphene oxide/silver/arginine for effective wound healing.

In this study, we introduce a novel graphene oxide/silver/arginine (GO/Ag/Arg) nanohybrid structure, which can act as an angiogenesis promoter and provide antibacterial nanostructure for improving the wound healing process. GO/Ag nanostructure has been optimized in terms of the GO/Ag mass ratio and pH values using central composite design and the response surface method to increase the Ag loading efficiency. Then, Arg was chemically introduced to the surface of GO/Ag nanostructure. Electrospun polycaprolactone (PCL)-GO/Ag/Arg nanocomposite was successfully fabricated and characterized. The synthesized nanocomposite demonstrated not only a great antibacterial effect on both Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) bacterial species, but appropriate biocompatibility against L929 fibroblastic cell lines. The results demonstrated that the preparation of the PCL-GO/Ag/Arg nanocomposite at a concentration of 1.0 wt% GO/Ag/Arg possessed the best biological and mechanical features. In vivo experiments also revealed that the use of optimized PCL-GO/Ag/Arg nanocomposite, after 12 d of treatment, led to significant increase in the healing process and also regeneration of the wound via reconstruction of a thickened epidermis layer on the wound surface, which was confirmed by histological analysis. In conclusion, the proposed approach can introduce a novel notion for preparing antibacterial material that significantly promotes angiogenesis.

Normalization of doxorubicin release from graphene oxide: New approach for optimization of effective parameters on drug loading.

Graphene oxide (GO) has been recently introduced as a suitable anticancer drug carrier, which could be loaded with doxorubicin (DOX) as a general chemotherapy agent. Herein, the attempts were made to optimize the effective parameters on both loading and release of DOX on GO. GO and GO-DOX were characterized using transition electron microscopy , zeta potential, Raman spectroscopy, UV-visible spectroscopy, and Fourier transform infrared spectroscopy. In addition, loading and releasing behaviors of DOX on GO were studied in terms of different temperature and pH values. The primary optimized values of pH and temperature for best-loaded amount of DOX were 8.9 and 309 K, respectively. Moreover, we found that the smallest amount of released DOX, in pH of cancer microenvironment (5.4), occurs when DOX had been previously loaded in pH 7.8 and 310 K. Although the highest amount of loaded DOX was in basic pH, the results of efficient release of DOX from the GO-DOX complex and also cellular toxicity assay revealed that the best pH for loading of DOX on GO was 7.8. Therefore, in addition to optimization of parameters for efficient loading of DOX on GO, this study suggested that normalization of a released drug compared with the amount of a loaded drug could be a new approach for optimization of drug loading on nanocarriers.

Optimization of an anti-HER2 nanobody expression using the Taguchi method.

Despite being widely used in immunotherapy of cancer, whole antibodies are limited by several disadvantages. This has led to the advent of novel biomolecules such as nanobodies. Taguchi method is a statistical experimental design to study the effect of multiple variables in biological processes. In an effort to overexpress a recombinant anti-human epidermal growth factor receptor type 2 (HER2) nanobody, we performed a detailed study to find optimal condition of temperature, induction, culture media, vector, and host strain, using Taguchi methodology. A total of 16 various experiments were designed. Total protein of the formulated cultures were assessed by Bradford test and sodium dodecyl sulfate-polyacrylamide gel electrophoresis, followed by size exclusion high performance liquid chromatography to quantify the relative concentration of the nanobody in different expression settings. Western blotting was performed to confirm the expression of the anti-HER2 nanobody. When, individually, optimum parameters determined by Taguchi were applied, including SHuffle strain cultured in LB medium, induced with 0.4 mM isopropyl-ß-D-thio-galactoside for 18 h at 24°C, production yield further increased by about 9% (25.4 mg/L), compared to the highest expression setting. Flow cytometry and enzyme-linked immunosorbent assay result indicated improved protein binding in optimized conditions. Overall, our findings provide a basis for further investigations on economical production of recombinant nanobodies to improve production yield and activity.

Production of nanocellulose in miniature-bioreactor: Optimization and characterization.

Bacterial cellulose (BC) is a very fascinating microbial biopolymer which is mainly produced by Gluconacetobacter xylinum. Optimization of BC production by G. xylinum was performed based on scale-down studies in miniature-bioreactor and response surface methodology in which the optimum pH value (6.5) and shaking rate (50 rpm) were obtained. The static culture condition for BC production has newly been defined. Nanostructure of BC includes nanofibers up to (60 nm) and nanoporosity up to (265 nm) was observed by scanning electron microscopy. By Fourier transform infrared spectroscopy study, the most expected BC interaction is nucleophilic interaction. MTT assay showed high biocompatibility. Appropriate mechanical strength (0.37 MPa) and Young's modulus (3.36 MPa) evinced BC scaffold utilization for skin tissue. The results indicate that BC sheets can be utilized in biomedical application and nanotechnology approaches.

Use of sulfur-oxidizing bacteria as recognition elements in hydrogen sulfide biosensing system.

Four sulfur-oxidizing bacteria (Thiobacillus thioparus, Acidithiobacillus thiooxidans PTCC1717, Acidithiobacillus ferrooxidans PTCC1646, and Acidithiobacillus ferrooxidans PTCC1647) were used as biorecognition elements in a hydrogen sulfide biosensing system. All the experiments were performed in 0.1 M phosphate buffer solution containing 1-20 ppm H2S with optimum pH and temperature for each species. Although H2 S was applied to the biosensing system, the dissolved O2 content decreased. Dissolved O2 consumed by cells in both free and immobilized forms was measured using a dissolved oxygen sensor. Free bacterial cells exhibit fast response (<200 Sec). Immobilization of the cells on polyvinyl alcohol was optimized using an analytical software. Immobilized A. ferrooxidans and A. thiooxidans retained more than 50% of activity after 30 days of immobilization. According to the data, A. thiooxidans and A. ferrooxidans are appropriate species for hydrogen sulfide biosensor.

Preparation and characterization of 3D bioprinted gelatin methacrylate hydrogel incorporated with curcumin loaded chitosan nanoparticles for in vivo wound healing application.

This study developed a biomimetic composite bioink consisting of gelatin methacrylate (GelMA) /chitosan nanoparticles (CSNPs) for extrusion-based 3D bioprinting. Additionally, curcumin(Cur)-loaded nanoparticles were incorporated which increased the proliferation and antibacterial activity of biomimetic skin constructs. The hydrogel, curcumin-loaded NPs, and the biocomposite was characterized chemically and physically. The results indicated proper modified gelatin with tunable physical characteristics, e.g., swelling ratio and biodegradability up to 1200 % and 25 days, respectively. In addition, the characterized CSNPs showed good distribution with a size of 370 nm and a zeta potential of 41.1 mV. We investigated the mechanical and cytocompatibility properties of chitosan nanoparticles encapsulated in hydrogel for emulating an extracellular matrix suitable for skin tissue engineering. CSNPs entrapped in GelMA (15 % w/v) exhibited controlled drug release during 5 days, which was fitted into various kinetic models to study the mass transfer mechanism behavior. Also, the composite hydrogels were effective as a barrier against both gram-positive and gram-negative bacteria at a concentration of 50 µg/ml nanoparticles in GelMA 15 %. Furthermore, the biocomposite was applied on Wistar rats for wound healing. As a result, this study provides a GelMA-NP50-Cur3 scaffold that promotes cell proliferation and decreases microbial infections in wounds.

A green approach for preparation of chitosan/hydroxyapatite/graphitic carbon nitride hydrogel nanocomposite for improved 5-FU delivery.

Reducing the side effects of cancer treatment methods is an important issue. The loading efficiency and sustained release of 5-Fluorouracil (5-FU) have been significantly improved by creating a new method. A nanocarrier with pH sensitivity has been developed through the w/o/w emulsification method. It is loaded with 5-FU and comprises of chitosan (CS), hydroxyapatite (HAp), and graphitic carbon nitride (g-C3N4). g-C3N4 nanosheets were incorporated in CS/HAp hydrogel to improve the entrapment and loading efficiency. Drug loading efficiency and entrapment efficiency reached 48 % and 87 %, respectively, and the FTIR and XRD tests verified evidence of the formation of chemical bonds among the drug and nanocarrier. Structural analysis was done using FE-SEM. DLS and zeta potential were employed to obtain average size distribution and surface charge. The release profile of 5-FU in various conditions shows the nanoparticles' pH dependence, and the nanocomposite's controlled release is consistent with the Korsmeyer-Peppas kinetic model. Cell apoptosis and cytotoxicity were evaluated in vitro using flow cytometry and MTT analysis. The biocompatibility of CS/HAp/g-C3N4 against MCF-7 cells was shown by the MTT method and confirmed by flow cytometry. CS/HAp/g-C3N4@5-FU led to the highest apoptosis rate in MCF-7 cells, indicating the nanocarrier's efficiency in killing cancer cells. These data indicate that the designed CS/HAp/g-C3N4@5-FU can be a potential drug for treating cancer cells.

Breast cancer detection based on cancer antigen 15-3; emphasis on optical and electrochemical methods: A review.

Cancer antigen 15-3 (CA 15-3) is a crucial marker used in the diagnosis and monitoring of breast cancer (BC). The demand for early and precise cancer detection has grown, making the creation of biosensors that are highly sensitive and specific essential. This review paper provides a thorough examination of the progress made in optical and electrochemical biosensors for detecting the cancer biomarker CA 15-3. We focus on explaining their fundamental principles, sensitivity, specificity, and potential for point-of-care applications. The performance attributes of these biosensors are assessed by considering their limits of detection, reaction times, and operational stability, while also making comparisons to conventional methods of CA 15-3 detection. In addition, we explore the incorporation of nanomaterials and innovative transducer components to improve the performance of biosensors. This paper conducts a thorough examination of recent studies to identify the existing obstacles. It also suggests potential areas for future research in this fast progressing field.The paper provides insights into their advancement and utilization to enhance patient outcomes. Both categories of biosensors provide significant promise for the detection of CA 15-3 and offer distinct advantages compared to conventional analytical approaches.

Alginate hydrogel-PCL/gelatin nanofibers composite scaffold containing mesenchymal stem cells-derived exosomes sustain release for regeneration of tympanic membrane perforation.

Exosomes are among the most effective therapeutic tools for tissue engineering. This study demonstrates that a 3D composite scaffold containing exosomes can promote regeneration in rat tympanic membrane perforation (TMP). The scaffolds were characterized using scanning electron microscopy (SEM), degradation, PBS adsorption, swelling, porosity, and mechanical properties. To confirm the isolation of exosomes from human adipose-derived mesenchymal stem cells (hAMSCs), western blot, SEM, and dynamic light scattering (DLS) were performed. The Western blot test confirmed the presence of exosomal surface markers CD9, CD81, and CD63. The SEM test revealed that the isolated exosomes had a spherical shape, while the DLS test indicated an average diameter of 82.5 nm for these spherical particles. MTT assays were conducted to optimize the concentration of hAMSCs-exosomes in the hydrogel layer of the composite. Exosomes were extracted on days 3 and 7 from an alginate hydrogel containing 100 and 200 µg/mL of exosomes, with 100 µg/mL identified as the optimal value. The optimized composite scaffold demonstrated improved growth and migration of fibroblast cells. Animal studies showed complete tympanic membrane regeneration (TM) after five days. These results illustrate that a scaffold containing hAMSC-exosomes can serve as an appropriate tissue-engineered scaffold for enhancing TM regeneration.