Viability Test Research Articles

Exploring apoptotic pathways in SH-SY5Y neuroblastoma cells: combined effects of napabucasin and doxorubicin.

Neuroblastoma often begins in infancy and one of the most common types of cancer among children is someone. Napabucasin (NP) (BBI608), a natural naphthoquinone emerging as a novel inhibitor of STAT3, has been found to effectively kill cancer stem-like tumor cells. On the other hand, the effect of Napabucasin on SH-SY5Y cells is currently unclear. The effects and mechanisms of NP and doxorubicin (DX) on human metastatic neuroblastoma cells were investigated. In this study, human neuroblastoma cells line (SHSY-5Y) were used. Apoptotic activation of NP and DX via the Bcl-2/Bax signaling pathway was evaluated by qRT-PCR, western blot and Tali cytometry. It was also detected by MTT, a cell viability test. NP and DX antiproliferative and invasive effected to SH-SY5Y cells. Additionally, NP induced apoptosis by pausing the cell cycle. Moreover, NP treatment inhibited the expression of Bcl-2, which is associated with apoptosis, while it clearly inhibited the expression of Bax and CASP3 genes. As a results showed that NP and DX suppressed the proliferation of neuroblastoma cells and could do this through apoptotic pathways. NP can be used to suppress metastasis of SHSY-5Y cells as an inhibitor of the apoptosis pathway Bcl-2. It is thought that NP, which provides tumor suppression through an apoptotic mechanism, may be an alternative treatment agent in neurological cancers such as neuroblastoma.

Dual targeting of HSP90 and BCL-2 in breast cancer cells using inhibitors BIIB021 and ABT-263.

The incidence of breast cancer has been increasing in recent years, and monotherapy approaches are not sufficient alone in the treatment of breast cancer. In the combined therapy approach, combining two or three different agents in lower doses can mitigate the side effects on living cells and tissues caused by high doses of chemical agents used alone. ABT-263 (navitoclax), a clinically tested Bcl-2 family protein inhibitor, has shown limited success in clinical trials due to the development of resistance to monotherapy in breast cancer cells. This resistance shows that monotherapy approaches are inadequate and more effective treatment strategies are needed. It is the ability of HSP90 inhibitors to destabilize many oncoproteins that are critical for the survival of cancer cells. This study aimed to examine the anticancer activity of the combination of ABT-263 with BIIB021, a new generation HSP90 inhibitor, on two widely used breast cancer cell lines: MCF-7 (ER-positive) and MDA-MB-231 (triple-negative breast cancer, TNBC). These cell lines were selected to represent distinct breast cancer subtypes with different molecular characteristics and clinical behaviors. Single and combined cytotoxic effects of this agents on MCF-7 and MDA-MB-231 breast cancer cell lines were determined using the MTT cell viability test. The combined use of these two agents showed a synergistic effect, and this effect was assigned using the Chou and Talalay method. mRNA and protein levels of apoptosis-related genes Bax, Bcl-2, Casp9, and Heat Shock Proteins HSP27, HSP70, and HSP90 were analyzed using Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR) and Western Blotting, respectively. The cytotoxicity analysis, combined with the application of the Chou-Talalay method, demonstrated that the BIIB021 and ABT-263 combination exhibited significantly greater anticancer activity compared to the individual effects of either BIIB021 or ABT-263 in breast cancer cell lines. The analysis of mRNA and protein levels indicated that the BIIB021+ABT-263 combination may have triggered the intrinsic apoptotic pathway in breast cancer cells. This study showed thatco-administration of ABT-263 and BIIB021 agents exhibited synergistic cytotoxic effects and increased the expression of apoptosis-related genes in breast cancer cell lines.

Viability assessment of Chlamydia trachomatis in men who have sex with men using molecular and culture methods.

Chlamydia trachomatis is the most commonly diagnosed bacterial sexually transmitted infection (STI) worldwide. Diagnosis relies on nucleic acid amplification techniques, such as PCR, which does not distinguish between viable pathogens and residual bacterial DNA, leading to potential overdiagnosis and overtreatment. PCR with confirmation of pathogen viability has not been widely explored in the STI field. Our aim was to establish a C. trachomatis viability PCR (V-PCR) and to apply it to anorectal swabs from men who have sex with men (MSM). We performed validation of a published V-PCR protocol by preparing artificial samples with known ratios of viable and non-viable C. trachomatis. Mock samples were treated with propidium monoazide (PMAxxTM) before DNA extraction and quantitative PCR (qPCR) to detect C. trachomatis. The V-PCR was then applied to C. trachomatis PCR-positive anorectal swabs from MSM. Viability was expressed as the difference in C. trachomatis (CT) copies between PMAxxTM untreated and treated samples (ΔLog10 CT/mL). The anorectal samples were inoculated in cell culture for isolation. Genotyping was performed by examining the ompA gene sequence. Of 236 anorectal swabs, 69 (29.2%) were C. trachomatis PCR-positive, and we obtained V-PCR data from 54. There were 7/54 (12.9%), samples with <1% viable CT (>2.52 ΔLog10 CT/mL) 4/54 (7.4%) samples with 1%-10% viable CT (1.59-2.52), 16/54 (29.6%) with 10.01-50% viable CT (0.86-1.59) and 27/54 (50.0%) with 50.01%-100% viable CT (<0.35-0.86). C. trachomatis was isolated successfully from 39/69 (56.5%) samples in cell culture. Genotypes based on ompA were obtained for 62/69 (89.9%) samples: G (n=15/62), D/Da (n=15/62), J (n=15/62), E (n=11/62), L1 (n=4/62) and L2 (n=2). We successfully implemented a viability test based on PCR, which can distinguish, detect and quantify viable C. trachomatis in anorectal swabs from MSM. Rapid, reliable assessment of C. trachomatis viability could help to improve antimicrobial stewardship.

Cure Efficiency and Biocompatibility of an Iron-Based Coordination Complex as a Photoinitiator for Dental 3D-Printed Resins

Objective: The aim of this study was to evaluate the cure efficiency and biocompatibility of a novel iron-based coordination complex used as a photoinitiator in comparison to conventional ethyl (2,4,6-trimethylbenzoyl) phenylphosphinate (TPO-L) and camphorquinone (CQ) as photoinitiators in dental 3D-printed resins. Materials and Methods: Experimental dental resin formulations were prepared by blending 1:1 ratio of Bis-GMA and TEGDMA, to which 0.2 wt% of either the iron-based coordination complex or CQ were added, along with 0.2 wt% EDAB and 0.4 wt% IOD, and the TPO-L. The degree of conversion (DC) was measured using Fourier transform infrared spectroscopy (FTIR). Biocompatibility was assessed by evaluating the viability of L929 fibroblast-like cells using the MTT assay 24 h post-exposure. Statistical analyses included a two-way ANOVA followed by Tukey’s test for post hoc comparisons, with significance at p &lt; 0.05. Results: The degree of conversion for the iron-based coordination complex (84.54% ± 1.69%) was significantly higher than that for the TPO-L (78.77% ± 1.25%) and CQ-based resins (73.21% ± 0.47%) (p &lt; 0.001). The iron-based coordination complex and TPO-L resins exhibited significantly higher conversion than CQ-based resins (p &lt; 0.001). Regarding biocompatibility, the cell viability test revealed that the iron-based coordination complex demonstrated the highest cell viability at 86.5% ± 10.24%, followed by TPO-L with 80.03% ± 11.07%. CQ showed the lowest cell viability of 51.29% ± 8.44% (p &lt; 0.05). Tukey’s test confirmed significant differences between CQ and other photointiators (p &lt; 0.05), while no significant difference was found between TPO-L and the iron-based coordination complex. Conclusions: This study introduces a novel iron-based coordination complex photoinitiator that demonstrates enhanced cure efficiency and comparable biocompatibility to TPO-L, while significantly reducing the cytotoxicity associated with CQ. Its longer absorption wavelength supports deeper layer curing, making it a promising alternative for dental 3D printing, particularly in bioactive scaffold applications requiring minimized cytotoxicity.

Development of gelatine and alginate based inks for 3D bioprinters and characterization.

Three-dimensional (3D) printing is a rapidly evolving technology. This study focuses on developing biopolymeric inks tailored for Three-dimensional (3D) printing applications, specifically to produce 3D-printed materials for wound dressing. Humic Acid (HA) was incorporated into the ink formulations due to its anti-inflammatory properties. Although alginate-gelatine-based inks are widely available, no existing systems include HA as an additional component, underscoring the originality of this work.This innovation addresses a key challenge in wound care by promoting healing while minimizing inflammatory responses. Alginate(Alg) used in this study was extracted from Cystoseira barbata, a brown algae species collected from Gideros Bay in Kastamonu, located along Turkey's western Black Sea coast. This is the first study to purify alginate from algae sourced from this regio. The extraction process was optimized. Four biopolymeric ink formulations were prepared in the study and were used to print 3D-biomaterials, which underwent characterization focusing on basic physicochemical properties. Biocompatibility was evaluated through cell viability tests (MTT assays), and Immune response analysis was evaluated through inflammatory markers; pro-inflammatory cytokines (IL-1β, TNFα), and the anti-inflammatory cytokine (IL-10). In conclusion, this study successfully purified and characterized alginate from algae collected in a previously unstudied region, Gideros Bay. A novel ink formulation, incorporating humic acid, was developed and 3D-printed for the first time. The results demonstrate that the printed materials in the study possess high potential for use as wound dressing materials, marking a significant contribution to the literature.

The complex role of cardiovascular imaging in viability testing.

Myocardial viability assessment is used to determine if chronically dysfunctional myocardium may benefit from coronary revascularization. Cardiac magnetic resonance with late gadolinium enhancement is the current gold standard for visualizing myocardial scar and provides valuable insight into myocardial viability. Viability assessments can also be made with Cardiac Positron Emission Tomography, Echocardiography, Single Photon Emission Tomography, and Cardiac Computed Tomography with each having advantages and disadvantages. Despite the classical interpretation that viability predicts segmental functional improvement, more recent studies have found that revascularization of viable myocardium has conflicting roles in predicting benefits for patients, especially as it relates to major adverse cardiovascular events, development of heart failure symptoms, and all-cause mortality. This review covers these conflicts along with an in-depth review of the pathophysiologic processes that are fundamental to myocardial viability and the various methods used for determining viability.

A hyaluronic acid-modified cyclodextrin self-assembly system for the delivery of β-carotene in the treatment of dry eye disease.

Dry eye disease (DED) is a multifactorial ocular disease, the core mechanism of which is the tear film instability caused by ocular oxidative stress damage and inflammation. Although various pharmaceutical agents are available for DED treatment, their effectiveness is often limited by the eyes' unique biological barriers, and the long-term use of steroid hormones can lead to several adverse effects. This study reported a nano-supramolecular delivery system consisting of a polycyclodextrin (PCD), hyaluronic acid (HA) and the natural compound β-carotene (BC) for the DED treatment. Our findings indicate that the HA/PCD@BC eye drops effectively distribute on the ocular surface, retain BC, and significantly enhance the corneal penetration of BC. The excellent biocompatibility of HA/PCD@BC was demonstrated through viability testing on different cell lines, the Draize eye test, as well as the hematoxylin-eosin staining (H&E) sections of cornea and conjunctiva. Both in vitro oxidative stress assays and in vivo DED model evaluations demonstrated that the HA/PCD@BC delivery system significantly reduced abnormal oxidative stress levels on the ocular surface, inhibited the secretion of inflammatory factors, and increased the secretion of tear film stabilizing mucin. These effects collectively improved pathological changes in eye tissues and minimized damage to the ocular surface. It is of particular importance to note that HA/PCD@BC eye drops showed superior efficacy in comparison to cyclosporine A (CsA), an FDA-approved first-line drug. To sum up, the HA/PCD@BC nanodelivery system provides a natural, safe and effective therapeutic strategy for the treatment of DED and various ocular diseases.

Printed polylactic acid/akermanite composite scaffolds for bone tissue engineering; development and surface modification

The susceptibility of bone tissues to various factors such as ageing, accidents, and diseases has led to extensive tissue engineering research focusing on bone tissues. Hence, this research also aims to determine the optimal amount of Akermanite (AK) addition to the polylactic acid scaffold for bone tissue engineering applications, as well as the effects of surface modification on its properties. The Akermanite was synthesized using the sol-gel method. Then, composite scaffolds of polylactic acid, including 0, 10, 20, and 30 wt% AK, were printed via the fused deposition modelling (FDM) process. These scaffolds were labelled as PLA, 10 wt% AK, 20 wt% AK, and 30 wt% AK, respectively. The X-ray diffraction analysis confirmed the production of the AK high-purity phase. Cell viability tests on composite scaffolds confirmed non-toxicity, and cell adhesion improved with AK addition. Mechanical testing showed that the compressive strength of composite scaffolds increased by increasing the AK content of the composite. This study recommended the 20 wt% AK scaffold as the optimal composition for bone tissue engineering. The surface-modification of polylactic acid/AK composite scaffolds using sodium hydroxide showed that it can be suitable for advanced tissue structures and medical applications, contributing to advancements in tissue engineering and medical technology for improved bone treatments.

Design, synthesis, and biological evaluation of pyrazole-ciprofloxacin hybrids as antibacterial and antibiofilm agents against Staphylococcus aureus.

In our continued efforts to tackle antibiotic resistance, a new series of pyrazole-ciprofloxacin hybrids were designed, synthesized, and evaluated for their antibacterial activity against Staphylococcus aureus (S. aureus), Pseudomonas aeruginosa (P. aeruginosa), and Mycobacterium tuberculosis (Mtb). Most of the compounds exhibited good to excellent activities against S. aureus, and six compounds (7a, 7b, 7d, 7g, 7k, and 7p) exhibited higher or comparable activity (MIC = 0.125-0.5 μg mL-1) to ciprofloxacin (0.125 μg mL-1). Further, these selected compounds were non-toxic (CC50 ≥ 1000 μg mL-1) when evaluated for cell viability test against the Hep-G2 cell line. Three compounds (7a, 7d, and 7g) demonstrated excellent activity against ciprofloxacin-resistant S. aureus with MIC values ranging from 0.125-0.5 μg mL-1 and good antibiofilm activity. Among them, 7g displayed remarkable antibiofilm activity with an MBIC50 value of 0.02 μg mL-1, which is 50 times lower than ciprofloxacin (MBIC50 = 1.06 μg mL-1). A time-kill kinetics study indicated that 7g showed both concentration and time-dependent bactericidal properties. In addition, 7g effectively inhibited DNA-gyrase supercoiling activity at 1 μg mL-1 (8× MIC). Two compounds 7b and 7d exhibited the highest activity against Mtb with a MIC of 0.5 μg mL-1, while 7c showed the highest activity against P. aeruginosa with a MIC value of 2 μg mL-1. Molecular docking studies revealed that 7g formed stable interactions at the DNA active site.

Promoting Angiogenesis/Osteogenesis by a New Copper/Magnesium Hydroxide Hybrid Nanoparticle: InVitro and InVivo Investigation.

In this study, a new hybrid nanoparticle composed of magnesium hydroxide and copper oxide (Mg(OH)2/CuO) with an optimized ratio of magnesium (Mg) to copper (Cu) was designed and incorporated into a 3D-printed scaffold made of polycaprolactone (PCL) and gelatin. These hybrid nanostructures (MCNs) were prepared using a green, solvent-free method. Their topography, surface morphology, and structural properties were characterized using scanning electron microscopy (SEM), transmission electron microscopy (TEM), X-ray diffraction (XRD), and X-ray photoelectron spectroscopy (XPS). The fabricated 3D-printed PCL/Gelatin/MCN scaffolds were investigated invitro and invivo. Cell viability tests on murine calvarial preosteoblasts (MC3T3-E1) and human umbilical vein endothelial cells (HUVECs) demonstrated that the scaffolds could induce proper cell proliferation. Additionally, the angiogenic and osteogenic properties of the constructs were evaluated using alkaline phosphatase (ALP) activity, osteogenesis-related, and angiogenesis-related gene expression tests. The invivo study was conducted using a rat calvarial defect model, which confirmed the superior angiogenic and osteogenic properties of the PCL/gelatin/MCN scaffolds compared to PCL/Gelatin and PCL/Gelatin/Mg(OH)2 scaffolds. Overall, the PCL/Gelatin/MCN scaffolds showed promising potential for bone regeneration, particularly for critical-sized defects where proper angiogenesis is essential for tissue reconstruction.

Toward improved auricle reconstruction: The role of FDM 3D printing with PCL and TPU materials.

Microtia, along with trauma, represents one of the main causes of external ear malformation. Different clinical techniques were developed for the reconstruction of the auricle, but they all have some drawbacks. This work is focused on the development of an innovative 3D porous scaffold, printed by Fused Deposition Modelling (FDM) and based on laser-scanned images of the healthy contralateral ear of the patient. The scaffold was printed using polycaprolactone (PCL) and thermoplastic polyether urethane (TPU) to mimic the components of the cartilage and adipose tissue, respectively. After the optimization of the printing parameters, the 3D surface obtained as an output of the laser scan was elaborated, sliced and used as input for printing the layered structure. Micro CT investigation confirmed the structure homogeneity and good pore interconnection. Mechanical compression and torsion tests were performed to verify that the mechanical properties of the 3D PCL/TPU structure were adequate. The 3D structure exhibited elastic moduluses comparable to those of the cartilaginous and adipose parts of the auricle. The torsion tests showed the adequacy of the structure without detachment between TPU and PCL printed layers. In vitro cell viability tests confirmed the absence of cytotoxicity in both materials. The PCL/TPU layered scaffold reproduced the anatomy of the patient's healthy contralateral ear, representing a good compromise between flexibility and strength. This, along with the other assessed properties, makes this scaffold a valid alternative for the reconstruction of the external ear.

Formulation of recombinant Lactococcus lactis as an oral vaccine candidate for COVID-19

Background: Lactococcus lactis is a promising oral vaccine carrier. However, to improve the stability of the bacteria, the freeze-drying method and formulation need to be optimised. Objective: To develop a formulation of oral recombinant food-grade Lactococcus lactis as a candidate oral vaccine for coronavirus disease 2019 (COVID-19). Method: To preserve bacteria during storage, a combination of skim milk, trehalose, and sucrose concentrations was adjusted. The impacts of the freeze-drying procedure were investigated using a bacterial viability test, and bacterial morphology was assessed using scanning electron microscopy. Bacteria produced from the freeze-drying technique were combined with excipients to create granules, tablets, and capsules. After one month of storage, the bacterial viability of each product was assessed after one-month storage in 4°C and 25°C, and physicochemical testing was conducted on each product. Results: The cryoprotectant formula containing 8% of skim milk and 7.5% of sucrose protected the bacteria the most from the freeze-drying process. The tablets and capsules complied with current specifications, including disintegration test, tablet hardness, capsule weight uniformity, and bacteria viability. Conclusion: Among the products, tablets stored for one month at 4°C had the best bacterial viability. This study demonstrated the potential to develop and administer an easy-to-use oral COVID-19 vaccine candidate using L. lactis.

HER-2 SMASH.

Human epidermal growth factor-2 (HER-2) targeted drugs are used in only HER-2 overexpressed cancers. However, only a small portion of these cancer types are HER-2 overexpressed. In this study, we aimed to upregulate HER-2 receptors in MCF-7 breast cancer and HT-29 colon cancer cell cultures, which these cells are not HER-2 upregulated in natural status. We used a 10-day non-cytotoxic lapatinib dose to upregulate HER-2 receptors. HER-2 levels of these cell lines were tested with ELISA and immunofluorescence tests before and after 10days of lapatinib administration. After upregulation of HER-2, we administered trastuzumab, and T-DM1 to these cell lines to observe whether there is an increase in anticancer activity. We used a cell viability test to show the cytotoxicity of trastuzumab and T-DM1. Also, we used ELISA and immunofluorescence for HER-2 pathway proteins to understand the mechanism of increased anti-cancer activity. We showed that administration of lapatinib for 10days leads to overexpression of HER-2 receptors on both MCF-7 and HT-29 cells. A significant increase in the cytotoxicity of trastuzumab or T-DM1 was observed after 10days of lapatinib administration. We named this method the smash method, which is the volleyball term. In volleyball, the ball is raised while low and quickly hits the ground again, just like we do with the HER-2 receptor. The smash method can switch HER-2 negative or HER-2 low tumors into HER-2 overexpressed, iatrogenically. Thus, we can use her2-targeted therapies in all cancer patients instead of a small portion.

Advancements in Palynology: Enhancing Plant Breeding through Pollen Studies

Palynology, the study of pollen and spores, has emerged as a vital discipline in plant science, with applications extending into plant breeding. This review explores the role of palynology in understanding reproductive biology, enhancing breeding programs, overcoming hybridization barriers and addressing challenges like male sterility, stress tolerance and species compatibility. Advanced techniques like transcriptomics reveal gene expression profiles, while proteomics and metabolomics provide insights into fertility and stress responses. Pollen-mediated transformation and CRISPR-Cas9 gene editing facilitate targeted trait improvements, with pollen-specific promoters aiding hybrid breeding. Additionally, molecular tools support pollen viability testing, gene flow studies, and stress tolerance research, contributing to hybrid seed production and environmental adaptability. Palynology also ensures biosafety by tracking transgenic pollen, making it an integral component of sustainable crop improvement strategies. Case studies illustrate its impact on cereals, legumes and horticultural crops. The article concludes by highlighting challenges and future directions for integrating palynological studies with emerging technologies to improve plant breeding outcomes. Plant breeding aims to develop superior crop varieties with desirable traits such as higher yields, resistance to biotic and abiotic stresses, improved quality and adaptability to diverse environments. Successful breeding hinges on understanding the reproductive biology of plants, particularly pollen, which serves as the carrier of male gametes.

Green Synthesis of Highly Fluorescent Carbon Dots from Groundnut Shell Biomass for Bioimaging Applications.

Carbon dots from alternative renewable carbon sources are emerging as alternatives to metal-based quantum dots. These nature-derived carbon dots exhibit excellent optical and fluorescent properties, which enable their use in several applications, including bioimaging. This work presents a facile and green approach to synthesizing highly fluorescent carbon dots from groundnut shells (GNS), an abundantly available agricultural residue. HRTEM analysis confirmed the synthesis of Groundnut shell Carbon Dots (GCDs) with a lattice spacing of around 0.22nm, corresponding to low dimensional graphitic structures. The observed intense absorption at around 278nm can be ascribed to the л - л* transitions resulting from the hybridization of sp2/sp3 orbitals in carbon dots. The fluorescence spectroscopy of GCDs displayed pronounced emission characteristics that varied depending on the excitation wavelength, which ranges from 280 to 480nm. The quantum yield of these GCDs was estimated to be 17.1%. The biocompatibility of GCDs is confirmed by the cell viability test, which indicates their suitability for yeast cell imaging.

Mechanical and antimicrobial properties of green and photoactive AgTiO2/poly(3hydroxybutyrate) (PHB) electrospun membranes

AbstractThe rise of antibiotic‐resistant microbes and concerns over non‐biodegradable waste highlight the need for innovative materials with integrated functionalities that address these critical issues. In this study, we synthesized poly(3hydroxybutyrate) (PHB) electrospun nanofibers blended with gelatin (Ge) and loaded with photoactive AgTiO2 nanoparticles with improved mechanical and biological properties. Biological tests revealed excellent antibacterial activity of the prepared membrane, with efficiency exceeding 99% against Escherichia coli and 95% against Staphylococcus epidermidis after 90 and 60 min of exposure to low‐power commercial LED lights, respectively. Filtration studies using a dead‐end stainless‐steel cell reveal that both bacteria are eliminated (&gt;99% after one filtration cycle). Results of the viability test showed that blending PHB with Ge improves the membrane's anti‐biofouling properties. The membranes were also characterized using SEM and EDX mapping techniques for morphological and elemental analysis, DSC and TGA to evaluate thermal properties and crystallinity, and FTIR to confirm chemical structure. Moreover, the electrospun membranes exhibited enhanced mechanical properties with the addition of Ge. PHB/Ge/3 wt% AgTiO2 sample showed 2.2 times better tensile strength and 1.89 times improved Young's modulus compared to PHB membranes. Finally, the breakdown of PHB/Ge/AgTiO2 membranes occurred progressively over only 8 weeks, showing the membranes' green and sustainable nature.Highlights Antibacterial efficacy of 99% against E. coli and 95% against S. epidermidis. Microfiltration efficiency &gt;99% achieved in one filtration cycle. Mechanical strength is enhanced by 2.2 times with the addition of gelatin. Effective anti‐biofouling is confirmed by CLSM and SEM tests. Membranes degraded within 8 weeks in natural soil.

Radiosensitization impact assessment of silica-layered iron oxide nanocomposites with various shell thickness.

Silica shell is considered to be a promising design that enhances nanocomposite stability, cellular internalization, and consequentially therapeutic impacts by overcoming their aggregation under physiological conditions. This study addressed synthesizing silica-layered iron oxide-based nanoparticles (SCINPs) with different shell thicknesses (1-SCINPs, 2-SCINPs, 3-SCINPs, and 4-SCINPs). Also, the impact of shell thickness on the nanoparticle's cellular internalization and the radio-sensitizing effect of prepared nano-formulations were assessed. The physical properties of the synthesized nanoparticles were examined using transmission electron microscopy (TEM), atomic force microscopy (AFM), dynamic light scattering (DLS), vibrating sample magnetometry (VSM), and X-ray diffraction (XRD). Cytotoxicity assay, oxidative stress parameters, and comet assay were used to investigate the radio-sensitizing effect of various nanoformulations. Results revealed that the mean diameter of prepared oxide-based nanoparticles (INPs) was about 12.63±1.36nm, and the shell thickness for 1-SCINPs, 2-SCINPs, 3-SCINPs, and 4-SCINPs was 22.58±3.51, 26.13±1.40, 46.95±3.10 and 60.30±4.30nm, respectively. Interestingly, we found that in cells treated with 40μg/ml of INPs, their viability decreased to 44.6%. Meanwhile, the viability was 41.69% and 39.4% for cells treated with 1-SCINPs and 2-SCINPs, respectively. This means that a thicker silica shell led to a decreased impact on radiosensitization. This was attributed to the influence of surface properties and size of SCINPs on their cellular uptake and the secondary electrons' entrapment within thicker shells upon radiation exposure. Cell viability test, comet assay and oxidative stress parameters show that 2-SCINPs formulations had the most potent radiosensitizing effect (with the highest dose enhancement factor equal to 2.1) when combined with radio-treatment. The results suggest that optimizing the silica shell thickness is critical for maximizing the therapeutic efficacy of SCINPs, with 2-SCINPs showing the highest radiosensitization effect.

Counteractive Effects of Copper Nanoparticles and Betacellulin on Ovarian Cells.

Copper nanoparticles (CuNPs) are known to affect many ovarian cell functions. CuNPs, prepared using a chemical reduction method, were fully characterized by different means (TEM, DLS, XRD, Z potential, XPS, and AES). The resulting colloidal suspension contained needle-like CuNPs aggregates made of a core of metallic copper and an oxidized surface of Cu2O and CuO. The separate and coupled effects of CuNPs and the growth factor betacellulin (BTC) were analyzed on the control of some basic functions of ovarian cells. With this purpose, porcine ovarian granulosa cells, together with CuNPs, BTC, and both (CuNPs + BTC), were cultured. Viability and BrDU tests, quantitative immunocytochemistry, TUNEL, and ELISA were used to evaluate markers of the S-phase (PCNA) and G-phase (cyclin B1) of the cell cycle, cell proliferation (BrDU incorporation), cytoplasmic/mitochondrial apoptosis (bax) and extrinsic (nuclear DNA fragmentation) markers, and the release of estradiol and progesterone. CuNPs were accumulated within the cells and were found to reduce all the markers of proliferation, but promoted all the markers of apoptosis and the release of steroid hormones. When added alone, BTC raised the expression of all cell viability and proliferation markers, depleted the expression of all apoptosis markers, and stimulated the release of both estradiol and progesterone. Furthermore, BTC prevented and even reversed the effect of CuNPs on all the measured parameters, whereas CuNPs mitigated BTC's effect on all the analyzed cell functions. These results support a direct toxic effect of CuNPs and a stimulatory effect of BTC on ovarian cell functions, as well as the capability of BTC to protect against the adverse effects of CuNPs.

A Safe and Convenient Method to Isolate Bone Marrow Mononuclear Cells in Clinical Practice.

Bone marrow mononuclear cells (BMMNCs) are becoming a promising cell therapy in regeneration medicine. BMMNCs are now obtained by density gradient centrifugation (DGC) in clinical practice, which is complicated and greatly influenced by human manipulation. Our objective is to develop a simple and safe method to isolate BMMNCs. Bone marrow was aspirated from nine minipigs. The optimal hypotonic sodium chloride (NaCl) concentration was first investigated based on the BMMNCs viability and lysis efficiency tests. Afterward, three different methods (ammonium chloride (NH4Cl) lysis, hypotonic NaCl lysis, and DGC) were used for BMMNCs isolation. Nucleated cell yield, residual red blood cells (RBCs) level, BMMNCs viability, apoptotic cell percentage, and colony-forming ability were measured in three groups. Cell morphology, cell phenotype, proliferative capacity, and osteogenic, adipogenic, and chondrogenic lineage differentiation potential of the bone marrow mesenchymal stem cells (BMSCs) were compared in three groups. 0.3% NaCl lysis group had optimal cell viability and lysis efficiency and the 0.3% NaCl lysis group had higher cell yield and lower RBCs remaining in BMMNCs compared to the DGC group. The BMSCs harvested from the NH4Cl lysis group had the worst proliferation ability. The NaCl lysis group was not inferior to the other two groups in terms of other biological characteristics of BMMNCs/BMSCs. The optimal concentration for hypotonic NaCl lysis to obtain BMMNCs is 0.3%. Compared with NH4Cl lysis and DGC, the 0.3% NaCl lysis may be a safe, appropriate, and low-cost method for BMMNCs isolation. This journal requires that authors assign a level of evidence to each submission to which Evidence-Based Medicine rankings are applicable. This excludes Review Articles, Book Reviews, and manuscripts that concern Basic Science, Animal Studies, Cadaver Studies, and Experimental Studies. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

Enhancement of Chondrogenic Differentiation in Bone Marrow-Derived Stem Cell Spheroids by Cuminum cyminum Methanolic Extract: Insights into Concentration-Dependent mRNA Expression and Gene Clustering Analysis.

Background/Objectives: Cuminum cyminum L. has been utilized as a medicinal plant for centuries. This research sought to examine the effects of cumin methanolic extract (CMT) on the chondrogenic differentiation of human bone marrow-derived mesenchymal stem cells. Methods: Spheroids were generated using human stem cells and cultured with CMT at concentrations between 0 and 1 µg/mL. Morphological assessments and cell viability tests were conducted on days 1 and 3. Chondrogenic differentiation expression was evaluated through quantitative polymerase chain reaction, Western blot, and RNA sequencing. SOX9, FAM20B, COL2A1, and COL1A1 mRNA expression levels were determined using real-time polymerase chain reaction. Protein expression was analyzed via Western blot. Results: Throughout this study, the spheroids maintained their integrity and shape. No significant variations in spheroid diameter were observed among the groups. CMT treatment enhanced the expression of SOX9 and FAM20B. Conclusions: The methanolic extract of Cuminum cyminum facilitated chondrogenic differentiation in human bone marrow-derived mesenchymal stem cells by modulating SOX9 and FAM20B expression. This indicates its potential application in cartilage tissue engineering.