Cytotoxicity Experiments Research Articles

Cytotoxic Effects of Aminotriles with Bioactive Potential: An Integrative Review

Aminonitriles are pharmacological-interest bioactive due to their promising antimicrobial and antitumor activity. Since cytotoxicity tests are inherent to the new drug development process, this work aimed to verify reports in the scientific literature on the cytotoxic effects of aminonitriles. The method adopted was an integrative review of works published in the last 10 years in the PubMed, Embase, Web of Science, and Virtual Health Library (VHL) databases. Three articles that matched the selection and eligibility criteria were included in this review. A total of 33 aminonitriles were used in the cytotoxicity experiments, and of the nine molecules based on pyridine, two exerted moderate cytotoxic activity, of the twelve synthesized from benzimidazole, none showed cytotoxic activity, and of the twelve derived from renieramycins, all showed considerable cytotoxic activities. The studies used in this research evaluated the cytotoxic effects of aminonitriles with evident anticancer and antimicrobial activity. The importance of evaluating the cytotoxicity of aminonitriles is emphasized, as well as the need for investigative research that explores other evaluation methods in pre-clinical tests that may corroborate the existing findings, with a view to the development of therapies against emerging health problems.

Release kinetics and binding interactions of curcumin‐encapsulated nanoparticles released from chitosan‐graft‐(methoxy poly(ethylene glycol)‐block‐polycaprolactone) injectable hydrogels

AbstractInjectable hydrogels are alternative materials for drug delivery in biomedical applications. Most injectable hydrogels are derived from stimuli‐responsive polymers that can respond to the changes of physical and/or chemical properties of the environment, such as temperature and pH variation. In this study, chitosan‐graft‐(methoxy poly (ethylene glycol)‐block‐polycaprolactone) injectable hydrogels were loaded with curcumin in different concentrations. The loading capacity and encapsulation efficiency of curcumin in such hydrogels were increased and decreased respectively with increasing curcumin concentrations. The encapsulation efficiency, however, was more than 90% for all formulations. Such hydrogels demonstrated curcumin release that proceeded in a sustained manner. The release profiles followed Korsmeyer‐Peppas and zero‐order kinetic release models which suggested that the release was facilitated by drug diffusion and the erosion of the self‐assembled polymeric networks. The curcumin released from the hydrogels was found as encapsulated core‐shell nanoparticles, the size of which depended on the loading concentration. A combination of fluorescence spectroscopy and thermodynamics analysis showed that van der Waals interactions and hydrogen bonds were major binding forces between curcumin and the copolymers. In addition, the curcumin‐encapsulated nanoparticles showed a progressive reduction in survival of the cervical cancer HeLa cell line, as shown in in vitro cytotoxicity experiments. © 2023 Society of Industrial Chemistry.

Bifunctional electrospun poly (L-lactic acid) membranes incorporating black phosphorus nanosheets and nano-zinc oxide for enhanced biocompatibility and antibacterial properties in catheter materials

For several decades, urinary tract infections caused by catheter-associated devices have negatively impacted not only medical device utilization, but also patient health. As such, the creation of catheter materials with both superior biocompatibility and antibacterial properties has become necessary. This study aimed to produce electrospun membranes based on polylactic acid (PLA) with the incorporation of black phosphorus nanosheets (BPNS) and nano-zinc oxide (nZnO) particles, as well as a mixture of both, in order to design bifunctional membranes with enhanced bioactivity and antibacterial features. The optimum spinning process was determined through examination of various PLA mass concentrations, spinning solution propelling speeds, and receiving drum rotating speeds, with emphasis on the mechanical properties of PLA membranes. Additionally, the antibacterial properties and cytocompatibility of the ZnO-BP/PLA antibacterial membranes were explored. Results demonstrated that the ZnO-BP/PLA antibacterial membranes displayed a rich porous structure, with uniform distribution of nZnO particles and BPNS. With the increase of polylactic acid concentration and the decrease of spinning solution advancing and drum rotation speeds, the mechanical properties of the fiber membrane were significantly improved. Furthermore, the composite membranes exhibited remarkable photothermal therapy (PTT) capabilities when aided by the synergistic effect of BP nanosheets and ZnO. This was achieved through near-infrared (NIR) irradiation, which not only dissipated the biofilm but also enhanced the release capability of Zn2+. Consequently, the composite membrane demonstrated an improved inhibitory effect on both Escherichia coli and Staphylococcus aureus. The results of cytotoxicity and adhesion experiments also indicated good cytocompatibility, with cells growing normally on the surface of the ZnO-BP/PLA antibacterial membrane. Overall, these findings validate the utilization of both BPNS and n-ZnO fillers in the creation of novel bifunctional PLA-based membranes, which possess both biocompatibility and antibacterial properties for interventional catheter materials.

Carbonized human hair derived carbon dots for detection of clozapine

Clozapine (CLZ) is known as the most effective antipsychotic medication for schizophrenia. However, low dosage or over dosage of CLZ is adverse to the treatment of Schizophrenia. Thus, it is necessary to develop effective detection method for CLZ. Recently, due to the advantages such as excellent optical properties, good photobleachability and sensitivity, carbon dots (CDs)-based fluorescent sensors for the detection of target analytes have drawn a great deal of attention. In this work, blue fluorescent CDs (Named as B-CDs) with quantum yield (QY) as high as 38% were obtained by using carbonized human hair as source material through one-step dialysis method for the first time. B-CDs showed obvious graphite-like structure with an average of 1.76 nm, containing abundant functional groups such as -C=O, amino N and C-N on the surface of carbon cores. Optical analysis showed that the B-CDs exhibited excitation-dependent emission property with maximum emission wavelength of 450 nm. Moreover, B-CDs were further applied as a fluorescence sensor to the detection of CLZ. The B-CDs based sensor exhibited a good quenching response by CLZ through the inner filter effect and static quenching mechanism with a limit of detection of 67 ng/mL, which was much lower than the minimal effective concentration in blood (0.35 μg/mL). Finally, to test the practical application value of the developed fluorescence method, the determination of the content of CLZ in tablets and the concentration in blood was carried out. Compared with the results of high-performance liquid chromatography (HPLC) method, it can be found that the constructed fluorescence detection method showed high accuracy and had great application potential in the detection of CLZ. Additionally, the results of cytotoxicity experiment showed that B-CDs had low cytotoxicity, which laid the foundation for the subsequent application of B-CDs in biological systems.

Characterization of a novel therapeutic antibody targeting peptide/HLA-E

Abstract T-cell receptor mimic antibodies (TCRm) have become an attractive therapeutic modality group as they allow for the targeting of intracellular proteins presented by HLA, which may allow for a large potential of new targets. However, the polymorphic nature of classical HLA class I and findings of HLA downregulation in cancer cells might limit the use of these agents for cancer treatment. An alternative approach would be to target peptides presented by HLA-E, a highly monomorphic molecule often found upregulated in cancer cells. Thus, we aimed to generate a TCRm against HLA-E presenting the leader sequence peptide from HLA-G since both HLAs are often expressed in multiple cancer types. An antibody was identified from a naïve human phage library and further affinity matured using yeast display. The final clone, ABX-001, had an affinity constant (K D) of 7.9nM. Specificity studies using both peptide-loaded cells and tumor cell lines showed that ABX-001 only bound when both HLA-E and HLA-G were present. Extensive profiling was done on immune cells to also confirm that they did not bind ABX-001. Finally, ABX-001 was assessed for its ability to direct killing of target-positive tumor cell lines. We observed that cytotoxicity in CD8 +T cells was enhanced with the addition of an ABX-001-CD3 T cell engager. We also observed that ABX-001 can act as a checkpoint inhibitor by blocking the interaction of HLA-E with NKG2A in cytotoxicity experiments using NK cells. These data indicate that we can effectively target peptide/HLA-E and highlights its potential therapeutic applications in cancer treatment.

Green synthesis of Fe3O4@SiO2@Salg particles for organophosphorus pesticides removal: Mechanisms, biosafety, and application

A biosafe magnetic Fe3O4@SiO2@Salg (GMS) was successfully developed via a simple and economic method. Fe3O4@SiO2 particles were selected as the magnetic core, and they directly interacted with sodium alginate, thus mitigating the use of cross-linking agents. The microstructure, properties, and formation mechanism of GMS were examined using FE-SEM, TEM, VSM, TGA, zeta potential, and FT-IR. The as-developed GMS was used as a desirable adsorbent to remove organophosphorus pesticides (OPPs) from water bodies, and the influence of different factors on the removal efficiencies of OPPs was investigated. Optimal preparation conditions were determined, and the excellent removal efficiencies of the 18 OPPs were achieved in this study (80.5%–100% for tap, river, and seawater). After the GMS was recycled eight times, the removal efficiencies of 18 OPPs were maintained at >80%. Adsorption kinetics and isotherm models showed that the adsorption process was accurately fitted with the Langmuir isotherm and pseudo-second-order models. The values of ΔG, ΔH, and ΔS were negative, indicating that the OPPs were adsorbed on the GMS spontaneously via an exothermal reaction, and chemisorption was the dominant adsorption mechanism. Overall, the as-developed GMS was biosafe as revealed by cytotoxicity experiments. GMS had good environmental security, stability, and reusability, and it exhibited excellent potential for removing OPPs from aqueous solutions without causing secondary pollution.

Surface engineering of hollow gold nanoparticle with mesenchymal stem cell membrane and MUC-1 aptamer for targeted theranostic application against metastatic breast cancer

Mesenchymal stem cell membrane (MSCM)-coated biomimetic doxorubicin-loaded hollow gold nanoparticles were fabricated and decorated with MUC1 aptamer in order to provide smart theranostic platform. The prepared targeted nanoscale biomimetic platform was extensively characterized and evaluated in terms of selective delivery of DOX and CT-scan imaging.The fabricated system illustrated spherical morphology with 118 nm in diameter. Doxorubicin was loaded into the hollow gold nanoparticles through physical absorption technique with encapsulation efficiency and loading content of 77%±10 and 31%±4, respectively. The in vitro release profile demonstrated that the designed platform could respond to acidic environment, pH 5.5 and release 50% of the encapsulated doxorubicin during 48 h, while 14% of the encapsulated doxorubicin was released in physiological condition, pH 7.4 up to 48 h. The in vitro cytotoxicity experiments on 4T1 as MUC1 positive cell line illustrated that the targeted formulation could significantly increase mortality at 0.468 and 0.23 µg/ml of equivalent DOX concentration compared to non-targeted formulation while this cytotoxicity was not observed in CHO as MUC1 negative cell line. Furthermore, in vivo experiments showed high tumor accumulation of the targeted formulation even 24 h after intravenous injection which induced effective tumor growth suppression against 4T1 tumor bearing mice. On the other hand, existence of hollow gold in this platform provided CT scan imaging capability of the tumor tissue in 4T1 tumor bearing mice up to 24 h post-administration. The obtained results indicated that the designed paradigm are promising and safe theranostic system for fighting against metastatic breast cancer.

Hierarchical Self‐assembly of G‐Quadruplexes Based Hydrogel Consisting of Guanine and Peptide Epitope†

Comprehensive SummaryGuanosine‐based hydrogels have attracted considerable attention because of their simplicity and easy preparation. However, the sugar moiety limits its further applications because of the necessity of sugar in the hydrogel formation. This work reports a G‐quadruplexes‐based hydrogel consisting of guanine and peptide epitope to form a supramolecular hydrogel in the presence of metal cations. Using the metal ion‐responsive peptide epitope from the ion channel to replace sugar motif at N9 position of guanosine results in a novel nucleopeptide. The results show that the gelation time, the diameter of nanofibers, the anisotropic property, and the mechanical property of the hydrogel can be simply controlled using metal cations. The magnesium and calcium ions direct the alignment of nanofibers to form anisotropic nano‐bundles. The mechanistic studies indicate the formation of G‐quadruplexes in the hydrogel. Compared to the storage modulus of nucleopeptide without the metal cation, adding zinc ions results in an over three‐order increase in mechanical properties. Cytotoxicity experiment indicates the good biocompatibility of our hydrogel. Moreover, we demonstrate that the guanine‐capped peptide could release STING agonist in a controlled manner. This work illustrates a simple way to modulate the property of the nucleopeptide hydrogel to develop soft materials.

Preparation of N-doped carbon dots and application to enhanced photosynthesis

Regulation of photosynthesis rates is one of the key ways to increase crop yields. Carbon dots (CDs), which are low-toxity and biocompatible optical nanomaterials, can be easily prepared and are ideal for improving photosynthesis efficiency. In this study, nitrogen-doped CDs (CNDs) with a fluorescent quantum yield of 0.36 were synthesized via a one-step hydrothermal method. These CNDs can convert a part of ultraviolet light in solar energy to blue light (emission peak at 410 nm) that can be utilized in photosynthesis and that overlaps with the optical absorption spectrum of chloroplasts in the blue light zone. Consequently, chloroplasts can pick up photons excited by the CNDs and transfer them to the photosynthetic system in the form of electrons, thereby accelerating the photoelectron transport rate. These behaviors can reduce ultraviolet light stress on wheat seedlings and improve the efficiency of electron capture and transfer from chloroplasts through optical energy conversion. As a result, various photosynthetic indices and biomass of wheat seedlings are improved. Cytotoxicity experiments have showed that CNDs within a certain concentration range almost do not affect cell survival.

Abstract 4040: Asciminib activates erythroid differentiation in K562 cell line

Abstract Background: Asciminib an allosteric tyrosine kinase inhibitor (TKI) was recently approved by the FDA for managing Philadelphia positive chronic myeloid leukemia (CML) patients in chronic phase who had failed to respond to previous TKIs. In this study, we used human erythroleukemic K562 cell line to investigate the role of asciminib as an inducer of erythroid differentiation. Additionally, imatinib, which has previously been reported as a potential inducer of erythroid differentiation, was also tested on these cells, both independently as well as in combination, in order to investigate the degree of erythroid differentiation in a K562 cell line. Methodology: In this study, we investigated the effect of asciminib (5-20nM) with and without imatinib (50-200nM) in differentiating the erythroleukemic cell line K562 towards erythroid lineage. K562 cells were cultured over a 12-day period in basic (IMDM) media and an EPO-based differentiation media to select optimal medium conditions. In addition, an in-vitro drug cytotoxicity experiment (XTT) was used to determine the preliminary toxicity of asciminib with IMDM media and EPO-based differentiation media CD235a and CD71 surface marker expressions were analyzed by flow cytometry at all time-points to determine the degree of differentiation achieved by each of the conditions Moreover, to analyze and identify hemoglobin producing cells ensuing differentiation, benzidine staining was performed at indicated time points. Results: Our results suggest that asciminib, imatinib or a combination of both drugs in EPO-based differentiation media results in efficient erythroid differentiation with high levels of GPA expression (approx. 90%) and significant upregulation of globin genes (p value = 0.0001). These results were further confirmed by benzidine staining, where drug addition and dose escalation significantly increased benzidine-positive cells. Our results also demonstrated that asciminib was able to achieve maximum differentiation at a much earlier time point as compared to imatinib. Moreover, asciminib, when combined with another TKI such as imatinib, can also aid in lowering the drug concentrations being used, hence resulting in lessening the toxicity of the drug when used alone required at high doses. Conclusion: The results from this study highlighted the novel role of asciminib in erythroid differentiation using an in-vitro K562 model that can give high-levels of surface markers and globin expression. Potential clinical application of asciminib in hematological diseases that are associated specifically with a failure in the expression of globin genes should be evaluated. Keywords:K562, erythroid differentiation, asciminib, imatinib, glycophorin A, transferrin receptor, hemoglobin Citation Format: Hammad Hassan, Sarah Yousuf, Syed Muhammad Ahmed, Karim Ruknuddin, Hadiqa Raees, Sadia Habib, Fizza Iftikhar, Afsar Ali Mian, Elnasir Lalani. Asciminib activates erythroid differentiation in K562 cell line. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4040.

Abstract 3827: Schinus terebinthifolius extracts impair migration of BT549 triple negative breast cancer cells in vitro

Abstract Bark extracts, derived from the Brazilian peppertree (Schinus terebinthifolius) were tested on BT549 triple negative breast cancer cells, a model system for invasive, metastatic cancers. Crude extracts, prepared with a 50% ethanol-50% water solvent, were investigated for cytotoxicity and effects on cell migration in vitro. Extract-treated cells exhibited statistically significant slower migration velocities than untreated control cells. Crude bark extracts, screened at a concentration of 3.15mg/mL in five independent experiments, produced an approximate 80% decrease in average migration velocity (p<0.05 in every experiment). Three independent tetrazolium based MTT cytotoxicity experiments confirmed that cells exposed to the same 3.15 mg/mL crude extract concentrations displayed similar viability as untreated control cells. Interestingly, these data suggest the Brazilian peppertree bark extract harbors bioactivity that impairs BT549 cell migration without cytotoxic side effects. Current experiments aim to discover which bioactive molecules, present in the crude extract, impede cancer cell migration. Silica-based column chromatography was used to separate the crude extract into fractions of increasing polarity. A wet method, which commenced with hexane, followed by dichloromethane, methanol, ethanol, ethyl acetate, and butanol, was employed. Fractions which retained anti-migratory activity were identified. Gas chromatography Mass Spectrometry (GCMS) was performed on crude extracts, samples with strong anti-migratory activity, and samples with weak anti-migratory activity, solubilized with ethanol, methanol, and acetonitrile. Data from two independent, complete sets of experiments, involving column chromatography, cellular assays with fractionated extracts, and GCMS, replicated results. Compounds that were unique to specific samples, as well as compounds that were found in multiples samples were identified. Rstudio software was utilized to organize and visualize data. This work lays down the preliminary foundation for future experiments testing specific purified forms of candidate molecules present in Brazilian peppertree bark extracts for the ability to limit BT549 cancer cell migration in vitro. Furthermore, Western blotting and immunocytochemistry experiments are underway to determine cellular signaling pathways targeted by the extract. Funding was provided by St. Thomas University’s Summer Research Institute and U.S. Dept. of Education STEM grants P03C110190, P031C160161, and P031C210035. Citation Format: Alexis Tapanes-Castillo, Mariacira Di Santi, Abel Sousa, Jonathan M. Brown, Paola M. Gonzalez Rodriguez, Thiago P. Leal, Leticia Saladrigas, Vadym Trokhymchuk, Leonardo M. Soto Chumpitaz, Andrea Peterson, Susana Gutierrez, Rolando J. Petit, Carlos Planchart, Christian Del Corro, Daniela A. Ramos, Valeria Nazaire, Sean Mondesire, Maria Pina. Schinus terebinthifolius extracts impair migration of BT549 triple negative breast cancer cells in vitro. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3827.

Roxarsone Delivery Nanocomposite Based on Nitrite-Functionalized Mesoporous Polydopamine for Multidrug-Resistant Bacterial Infections via Enhanced Chemo-Photothermal Therapy.

Current treatments for infections caused by multidrug-resistant bacteria still remain challenging and therapeutic materials with high efficacy are of demand. Herein, a bactericidal nanocomposite was constructed by loading Roxarsone (ROX) onto nitrosylated mesoporous polydopamine (named mPDA@NO-ROX). The designed nanocomposite exhibited considerable photothermal effect and controlled NO and ROX co-delivery under the irradiation of near-infrared laser (NIR) to achieve enhanced chemo-photothermal antibacterial therapy. The in vitro antibacterial evaluation of the mPDA@NO-ROX demonstrated the effective elimination of the Gram-negative tetracycline-resistant Escherichia coil and Gram-positive methicillin-resistant Staphylococcus aureus under mild NIR irradiation compared to merely ROX loaded unmodified mPDA, indicating the NO enhanced chemo-photothermal therapy. In addition, the cytotoxicity experiments indicated that mPDA@NO-ROX exhibited only 5 % of hemolysis rate and high cell viability at 1 mg mL-1 against mammalian fibroblasts, suggesting the excellent biocompatibility. In conclusion, the mPDA@NO-ROX could be a promising candidate for anti-infection therapy of multidrug-resistant bacteria.

Effective Platform for Enhancing the Bioavailability and Anti-Cancer Efficacy of Norcantharidin: Nanoemulsion Hybrid Lipid Carriers

Novel antitumor drugs and nano-delivery systems for treating liver cancer are becoming a research hotspot, given that the incidence and mortality rates of liver cancer are high. Herein, nanoemulsion hybrid lipid carriers containing norcantharidin (Nor) (NNHLCs) were first prepared by loading water-in-oil nanoemulsions into hybrid solid lipids using nanoemulsion-thin film ultrasonic dispersion method. The in vitro and in vivo kinetic features were determined by in situ unidirectional perfusion and plasma concentration-time curve method, respectively. The activity of NNHLCs against liver cancer HepG2 cells was determined by evaluating in vitro cytotoxicity, clone formation, apoptosis, and cell cycle experiments. Fourier transform infrared (FT-IR) spectroscopy, differential scanning calorimetry (DSC), and X-ray diffraction (XRD) results indicated that Nor exists in NNHLCs in an amorphous state with good encapsulation in the lipid matrix. The absorptive constants and permeabilities of NNHLCs were significantly increased compared with Nor. The relative bioavailability of NNHLCs to Nor was 207.68%. NNHLCs could inhibit the viability and proliferation of HepG2 cells and promote apoptosis. Thus, nanoemulsion hybrid lipid carriers may provide a new option for the efficient delivery of chemotherapeutic drugs.

The effect of molecular weight and chemical structure of cross-linkers on the properties of redox-responsive hyaluronic acid hydrogels

In this work, we investigated the effect of the size and the chemical structure of crosslinkers on the properties of hyaluronic acid-based hydrogels prepared via an inverse electron demand Diels-Alder reaction. Hydrogels having loose and dense networks were designed by cross-linkers with and without polyethylene glycol (PEG) spacers of different molecular weights (1000 and 4000 g/mol). The study showed that the properties of hydrogels such as swelling ratios (20–55 times), morphology, stability, mechanical strength (storage modulus in the range 175–858 Pa), and drug loading efficiency (87 % ~ 90 %) were greatly influenced by the addition of PEG and changing its molecular weight in the cross-linker. Particularly, the presence of PEG chains in redox- responsive crosslinkers increased the doxorubicin release (85 %, after 168 h) and the degradation rate (96 %, after 10 d) of hydrogels in the simulated reducing medium (10 mM DTT). The in vitro cytotoxicity experiments conducted for HEK-293 cells revealed that the formulated hydrogels were biocompatible, which could be a promising candidate for drug delivery applications.

Targeting CD43 optimizes cancer immunotherapy through reinvigorating antitumor immune response in colorectal cancer.

Colorectal cancer (CRC) is one of the most common malignancies worldwide, with dramatically increasing incidence and mortality for decades. However, current therapeutic strategies for CRC, including chemotherapies and immunotherapies, have only demonstrated limited efficacy. Here, we report a novel immune molecule, CD43, that can regulate the tumor immune microenvironment (TIME) and serves as a promising target for CRC immunotherapy. The correlation of CD43 expression with CRC patient prognosis was revealed by public data analysis. CD43 knockout (KO) CRC cell lines were generated by CRISPR-Cas9 technology, and a syngenetic murine CRC model was established to investigate the in vivo function of CD43. The TIME was analyzed via immunohistochemical staining, flow cytometry and RNA-seq.Immune functions were investigated by depletion of immune subsets in vivo and T-cell functional assays in vitro, including T-cell priming, cytotoxicity, and chemotaxis experiments. In this study, we found that high expression of CD43 was correlated with poor survival of CRC patients and the limited infiltration of CD8+ T cells in human CRC tissues. Importantly, CD43 expressed on tumor cells, rather than host cells, promoted tumor progression in a syngeneic tumor model. Loss of CD43 facilitated the infiltration of immune cells and immunological memory in the TIME of CRC tumors. Mechanistically, the protumor effect of CD43 depends on T cells, thereby attenuating T-cell-mediated cytotoxicity and cDC1-mediated antigen-specific T-cell activation. Moreover, targeting CD43 synergistically improved PD-L1 blockade immunotherapy for CRC. Our findings revealed that targeting tumor-intrinsic CD43 could activate the antitumor immune response and provide particular value for optimized cancer immunotherapy by regulating the TIME in CRC patients.

Doxorubicin- and Trastuzumab-Modified Gold Nanoparticles as Potential Multimodal Agents for Targeted Therapy of HER2+ Cancers.

Recently, targeted nanoparticles (NPs) have attracted much attention in cancer treatment due to their high potential as carriers for drug delivery. In this article, we present a novel bioconjugate (DOX-AuNPs-Tmab) consisting of gold nanoparticles (AuNPs, 30 nm) attached to chemotherapeutic agent doxorubicin (DOX) and a monoclonal antibody, trastuzumab (Tmab), which exhibited specific binding to HER2 receptors. The size and shape of synthesized AuNPs, as well as their surface modification, were analyzed by the TEM (transmission electron microscopy) and DLS (dynamic light scattering) methods. Biological studies were performed on the SKOV-3 cell line (HER2+) and showed high specificity of binding to the receptors and internalization capabilities, whereas MDA-MB-231 cells (HER2-) did not. Cytotoxicity experiments revealed a decrease in the metabolic activity of cancer cells and surface area reduction of spheroids treated with DOX-AuNPs-Tmab. The bioconjugate induced mainly cell cycle G2/M-phase arrest and late apoptosis. Our results suggest that DOX-AuNPs-Tmab has great potential for targeted therapy of HER2-positive tumors.

Untargeted screening, quantitative analysis, and toxicity estimation of degradation products of fluazinam in vegetables

Fluazinam exhibits rapid degradation in some vegetable matrixes. The degradation products may pose risk for human health. In the present study, three degradation products of fluazinam in vegetables have been identified, named as sulfhydryl fluazinam, hydroxyl fluazinam, and amine fluazinam. Among them, sulfhydryl fluazinam has not been previously reported. Quantitative/semi-quantitative method for simultaneous analysis of fluazinam and the three degradation products was developed using QuEChERS in combination of HPLC-MS/MS. The proposed analytical method was further applied in monitoring fluazinam and its degradation products in fluazinam-processed vegetables. As a result, sulfhydryl fluazinam was demonstrated to be a characteristic degradation product of fluazinam in the sulphur-rich vegetables with transformation rates of above 10.0% within 60.0 min. In addition, we found for the first time that sulfhydrylation of pesticide occurred in putrid vegetable samples covering various plant families. The result contributed to discovery of the overlooked degradation pathways of pesticides in vegetables. Among fluazinam and the three degradation products, amine fluazinam was predicted to have highest toxicity, followed by sulfhydryl fluazinam and fluazinam, whereas hydroxyl fluazinam displayed the lowest toxicity. Cytotoxicity experiments in BRL-3A cells showed that toxicity of sulfhydryl fluazinam was comparable to that of fluazinam. In short-term oral studies in mice, liver appeared to be a target organ for sulfhydryl fluazinam. The present study not only provides an efficient analysis method, but also inspires further comprehensive risk assessment of fluazinam and its degradation products in vegetables.

Study on the effect of γδ T cells expanded in vitro to kill hepatocellular carcinoma cells.

γδ T cells for tumor cell immunotherapy has recently become a hot topic. To investigate the stimulation of expanded γδ T cells in vitro to kill liver cancer cells and its mechanism, and in vivo validation. Peripheral blood mononuclear cells (PBMCs) were isolated and amplified. The proportion of γδ T cells in T cells was determined using flow cytometry. γδ T cells were selected as effector cells, and HepG2 cells as target cells in the cytotoxicity experiment. NKG2D blocker was used to block effector cells from identifying target cells, and PD98059 was used to block intracellular signaling pathways. The nude mice tumor model was established in two batches, the tumor growth curve was drawn, and the tumor formation effect was tested using small animal imager to verify the killing effect of γδ T cells. The γδ T cells in the three experimental groups exhibited a large amount of amplification (P < 0.01). In the killing experiment, the killing rate of γδ T cells stimulated by zoledronate (ZOL) in the experimental group was significantly higher than that in the HDMAPP group and the Mycobacterium tuberculosis H37Ra strain (Mtb-Hag) group (P < 0.05). The blocking effect of PD98059 is stronger than that of the NKG2D blocker (P < 0.05). Among them, in the HDMAPP group, when the target ratio was 40:1, the NKG2D blocker exhibited a significant blocking effect (P < 0.05). Alternatively, in the ZOL group, when the effect ratio was 10:1, the effector cells were blocked significantly after treatment using PD98059 (P < 0.05). In vivo experiments verified the killing effect of γδ T cells. According to the tumor growth curve, there was a difference between the experimental and control groups after cell treatment (P < 0.05). ZOL has high amplification efficiency and a positive effect on killing tumor cells.

Efficacy and toxicity of anise oil as a potential topical wound healer: a cell culture study.

We studied the cytotoxic effects of topical anise oil on NIH/3T3 fibroblast cells using a cell culture assay. NIH/3T3 fibroblast cells were grown in Dulbecco's Modified Eagle Medium (DMEM) supplemented with fetal bovine serum (10%) and penicillin/streptomycin under standard cell culture conditions in a humidified incubator containing 5% carbon dioxide. For the MTT cytotoxicity experiment, NIH/3T3 cells were plated in triplicate at a concentration of 3x103 per well in 96-well plates and incubated for 24 hours. The cells were treated with anise oil concentrations ranging from 3.13 to 100 μM, and the plates were cultured for 24, 48, and 72 hours under standard cell culture conditions. For assessment by confocal microscopy, NIH/3T3 cells were seeded on sterilized coverslips in 6-well plates at a concentration of 105 cells per well in triplicate. For 24 hours, cells were treated with 100 μM of anise oil. Three wells that were not treated with anise oil served as the control group. The MTT findings demonstrated that anise oil is not cytotoxic to NIH/3T3 fibroblast cells. Anise oil stimulated cell growth and triggered cell division at all three incubation intervals of 24, 48, and 72 hours. The maximum growth was obtained in the applied highest concentration of 100 μM anise oil. At doses of 25, 50, and 100 μM, there was also a statistically significant improvement in cell viability. At 72 hours of incubation, dosages of 6.25 and 12.5 micro of anise oil were shown to be viability-inducing for NIH/3T3 cells. In the confocal microscopy pictures, it was found that anise oil was not cytotoxic on NIH/3T3 cells at the applied maximal dose. The experimental group of NIH/3T3 cells exhibited the same cell morphology as the untreated control group. In both sets of NIH/3T3 cells, the nucleus was round and undamaged, and the cytoskeleton was determined to be compact. Anise oil is not cytotoxic on NIH/3T3 fibroblast cells and initiates cell growth. Anise oil could be used topically to enhance wound healing after surgical procedures if clinical trials will confirm experimental data.

Development of an Fe3O4 Surface-Grafted Carboxymethyl Chitosan Molecularly Imprinted Polymer for Specific Recognition and Sustained Release of Salidroside

The choice of carrier material is critical in the study of natural drug release preparations and glycosylated magnetic molecularly imprinted materials. The stiffness and softness of the carrier material affect the efficiency of drug release and the specificity of recognition. The dual adjustable aperture-ligand in molecularly imprinted polymers (MIPs) provides the possibility of individualized design for sustained release studies. In this study, a combination of paramagnetic Fe3O4 and carboxymethyl chitosan (CC) was used to enhance the imprinting effect and improve drug delivery. A combination of tetrahydrofuran and ethylene glycol was used as a binary porogen to prepare MIP-doped Fe3O4-grafted CC (SMCMIP). Salidroside serves as the template, methacrylic acid acts as the functional monomer, and ethylene glycol dimethacrylate (EGDMA) serves as the crosslinker. Scanning and transmission electron microscopy were used to observe the micromorphology of the microspheres. The structural and morphological parameters of the SMCMIP composites were measured, including the surface area and pore diameter distribution. In an in vitro study, we found that the SMCMIP composite had a sustained release property of 50% after 6 h of release time in comparison to the control SMCNIP. The total amounts of SMCMIP released at 25 °C and 37 °C were 77% and 86%, respectively. In vitro results showed that the release of SMCMIP followed Fickian kinetics, meaning that the rate of release is dependent on the concentration gradient, with diffusion coefficients ranging from 3.07 × 10-2 cm2/s to 5.66 × 10-3 cm2/s. The results of cytotoxicity experiments showed that the SMCMIP composite did not have any harmful effects on cell growth. The survival rates of intestinal epithelial cells (IPEC-J2) were found to be above 98%. By using the SMCMIP composite, drugs may be delivered in a sustained manner, potentially leading to improved therapeutic outcomes and reduced side effects.