Comprehensive evaluation and prediction of groundwater quality and risk indices using quantitative approaches, multivariate analysis, and machine learning models: An exploratory study.

Assessing and predicting quality of groundwater is crucial in managing groundwater availability effectively. In the current study, groundwater quality was thoroughly appraised using various indexing methods, including the drinking water quality index (DWQI), pollution index of heavy metals (HPI), pollution index (PI), metal index (MI), degree of contamination (Cd), and risk indicators, like hazard quotient (HQ) and total hazard indicator (HI). The assessments were augmented through multivariate analytical techniques, models based on recurrent neural networks (RNNs), and integration of geographic information system (GIS) technology. The analysis measured physicochemical parameters across 48 groundwater wells from El-Menoufia region, revealing distinct water types influenced by ion exchange, rock-water interactions, and silicate weathering. Notably, the groundwater showed elevated levels of certain metals, particularly manganese (Mn) and lead (Pb), exceeding the drinking water limits. The DWQI deemed the bulk of the tested samples suitable for consumption, assigning them to the "good" category, whereas a small number were considered inferior quality. The HPI, MI, and Cd indices indicated significant pollution in the central study region. The PI revealed that Pb, Mn, and Fe were significant contributors to water pollution, falling between classes IV (strongly affected) and V (seriously affected). HQ and HI analyses identified the central area of the study as particularly prone to metal contamination, signifying a high risk to children via oral and dermal routes and to adults through oral exposure alone (non-carcinogenic risk). The adults had no health risks due to dermal contact. Finally, the RNN simulation model effectively predicted the health and water quality indices in training and testing series. For instance, the RNN model excelled in predicting the DWQI, with three key parameters being crucial. The model demonstrated an excellent fit on the training set, achieving an R2 of 1.00 with a very low root mean of squared error (RMSE) of 0.01. However, on the testing set, the model's performance slightly decreased, showing an R2 of 0.96 and an RMSE of 2.73. Regarding HPI, the RNN model performed exceptionally well as the primary predictor, with R2 values of 1.00 (RMSE = 0.01) and 0.93 (RMSE = 27.35) for the training and testing sets, respectively. This study provides a unique perspective for improving the integration of various techniques to gain a more comprehensive understanding of groundwater quality and its associated health risks, with a strong focus on feature selection strategies to enhance model accuracy and interpretability.

Insight into the integration effect of chitosan and ß-cyclodextrin on the properties of zinc-phosphate/hydroxyapatite hybrid as delivery structures for 5-fluorouracil: loading and release profiles.

Zinc-phosphate/hydroxyapatite hybrid form (ZP/HP) in core-shell nanostructure was developed and functionalized with both chitosan (CS@ZP/HP) and ß-cyclodextrin (CD@ZP/HP) as bio-composite of enhanced physicochemical and biological properties. These structures were assessed as potential deliveries of 5-fluorouracil, exhibiting enhanced loading, release, and anti-cancer behaviors. The functionalization strongly prompted the loading effectiveness to be 301.3 mg/g (CS@ZP/HP) and 342.8 mg/g (CD@ZP/HP) instead of 238.9 mg/g for ZP/HP. The loading activities were assessed based on the hypotheses of traditional kinetic and isotherm models, alongside the computational variables of the monolayer model with a single energetic site as an advanced isotherm model. The functionalized versions exhibit much greater loading efficacy compared to ZP/HP as a result of the increment in the density of the existing loading sites [Nm(5-Fu) = 78.85 mg/g (ZP/HP), 93.87 mg/g (CS@ZP/HP), and 117.8 mg/g (CD@ZP/HP)]. Furthermore, the loading energies of approximately 40 kJ/mol, together with the loading potential of each receptor (n > 1) and Gaussian energies of approximately 8 kJ/mol, indicate the physical entrapment of 5-Fu molecules according to a vertical orientation. The materials mentioned verify long-term and continuous release characteristics. Following the modification processes, this behavior became faster as both CS@ZP/HP and CD@ZP/HP displayed complete release within 120 h at pH 1.2. The kinetic studies and diffusing exponent (>0.45) indicate that release characteristics are controlled by both diffusion and erosion processes. These carriers also markedly increase the cytotoxicity of 5-Fu against HCT-116 colorectal cancer cell lines: 5-Fu-ZP/HP (3.2% cell viability), 5-Fu-CS@ZP/HP (1.12% cell viability), and 5-Fu-CD@ZP/HP (0.63% cell viability).

Surface Modification Strategies for Chrysin-Loaded Iron Oxide Nanoparticles to Boost Their Anti-Tumor Efficacy in Human Colon Carcinoma Cells.

Enhancing nanoparticles' anti-cancer capabilities as drug carriers requires the careful adjustment of formulation parameters, including loading efficiency, drug/carrier ratio, and synthesis method. Small adjustments to these parameters can significantly influence the drug-loading efficiency of nanoparticles. Our study explored how chitosan and polyethylene glycol (PEG) coatings affect the structural properties, drug-loading efficiency, and anti-cancer efficacy of Fe3O4 nanoparticles (NPs). The loading efficiency of the NPs was determined using FTIR spectrometry and XRD. The quantity of chrysin incorporated into the coated NPs was examined using UV-Vis spectrometry. The effect of the NPs on cell viability and apoptosis was determined by employing the HCT 116 human colon carcinoma cell line. We showed that a two-fold increase in drug concentration did not impact the loading efficiency of Fe3O4 NPs coated with PEG. However, there was a 33 Å difference in the crystallite sizes obtained from chitosan-coated Fe3O4 NPs and drug concentrations of 1:0.5 and 1:2, resulting in decreased system stability. In conclusion, PEG coating exhibited a higher loading efficiency of Fe3O4 NPs compared to chitosan, resulting in enhanced anti-tumor effects. Furthermore, variations in the loaded amount of chrysin did not impact the crystallinity of PEG-coated NPs, emphasizing the stability and regularity of the system.

Insight into the Morphological Properties of Nano-Kaolinite (Nanoscrolls and Nanosheets) on Its Qualification as Delivery Structure of Oxaliplatin: Loading, Release, and Kinetic Studies.

Natural kaolinite underwent advanced morphological-modification processes that involved exfoliation of its layers into separated single nanosheets (KNs) and scrolled nanoparticles as nanotubes (KNTs). Synthetic nanostructures have been characterized as advanced and effective oxaliplatin-medication (OXAP) delivery systems. The morphological-transformation processes resulted in a remarkable enhancement in the loading capacity to 304.9 mg/g (KNs) and 473 mg/g (KNTs) instead of 29.6 mg/g for raw kaolinite. The loading reactions that occurred by KNs and KNTs displayed classic pseudo-first-order kinetics (R2 > 0.90) and conventional Langmuir isotherms (R2 = 0.99). KNTs exhibit a higher active site density (80.8 mg/g) in comparison to KNs (66.3 mg/g) and raw kaolinite (6.5 mg/g). Furthermore, compared to KNs and raw kaolinite, each site on the surface of KNTs may hold up to six molecules of OXAP (n = 5.8), in comparison with five molecules for KNs. This was accomplished by multi-molecular processes, including physical mechanisms considering both the Gaussian energy (<8 KJ/mol) and the loading energy (<40 KJ/mol). The release activity of OXAP from KNs and KNTs exhibits continuous and regulated profiles up to 100 h, either by KNs or KNTs, with substantially faster characteristics for KNTs. Based on the release kinetic investigations, the release processes have non-Fickian transport-release features, indicating cooperative-diffusion and erosion-release mechanisms. The synthesized structures have a significant cytotoxicity impact on HCT-116 cancer cell lines (KNs (71.4% cell viability and 143.6 g/mL IC-50); KNTs (11.3% cell viability and 114.3 g/mL IC-50). Additionally, these carriers dramatically increase OXAP's cytotoxicity (2.04% cell viability, 15.4 g/mL IC-50 (OXAP/KNs); 0.6% cell viability, 4.5 g/mL IC-50 (OXAP/KNTs)).

Synthesis and Characterization of ß-Cyclodextrin-Hybridized Exfoliated Kaolinite Single Nanosheets as Potential Carriers of Oxaliplatin with Enhanced Loading, Release, and Cytotoxic Properties.

Natural kaolinite was subjected to a successful exfoliation process into separated kaolinite nanosheets (KNs), followed by hybridization with ß-cyclodextrin biopolymer (ß-CD), forming an advanced bio-nanocomposite (ß-CD/KNs). The synthetic products were evaluated as enhanced delivery structures for oxaliplatin chemotherapy (OXAPN). The hybridization of KNs with ß-CD polymer notably enhanced the loading capacity to 355.3 mg/g (ß-CD/KNs) as compared to 304.9 mg/g for KNs. The loading of OXAPN into both KNs and ß-CD/KNs displayed traditional pseudo-first-order kinetics (R2 > 0.85) and a conventional Langmuir isotherm (R2 = 0.99). The synthetic ß-CD/KNs validates a greater occupied effective site density (98.7 mg/g) than KNs (66.3 mg/g). Furthermore, the values of the n steric parameter (4.7 (KNs) and 3.6 (ß-CD/KNs)) reveal the vertical orientation of the loaded molecules and the loading of them by multi-molecular mechanisms. These mechanisms are mainly physical processes based on the obtained Gaussian energy (<8 KJ/mol) and loading energy (<40 KJ/mol). The release profiles of both KNs and ß-CD/KNs extend for about 120 h, with remarkably faster rates for ß-CD/KNs. According to the release kinetic findings, the release of OXAPN displays non-Fickian transport behavior involving the cooperation of diffusion and erosion mechanisms. The KNs and ß-CD/KNs as free particles showed considerable cytotoxicity and anticancer properties against HCT-116 cancer cell lines (71.4% cell viability (KNs) and 58.83% cell viability (ß-CD/KNs)). Additionally, both KNs and ß-CD/KNs significantly enhanced the OXAPN's cytotoxicity (2.04% cell viability (OXAPN/KNs) and 0.86% cell viability (OXAPN/ß-CD/KNs).

Insights into the Effect of Chitosan and ß-Cyclodextrin Hybridization of Zeolite-A on Its Physicochemical and Cytotoxic Properties as a Bio-Carrier for 5-Fluorouracil: Equilibrium and Release Kinetics Studies.

Synthetic zeolite-A (ZA) was hybridized with two different biopolymers (chitosan and ß-cyclodextrin) producing biocompatible chitosan/zeolite-A (CS/ZA) and ß-cyclodextrin/zeolite-A (CD/ZA) biocomposites. The synthetic composites were assessed as bio-carriers of the 5-fluorouracil drug (5-Fu) with enhanced properties, highlighting the impact of the polymer type. The hybridization by the two biopolymers resulted in notable increases in the 5-Fu loading capacities, to 218.2 mg/g (CS/ZA) and 291.3 mg/g (CD/ZA), as compared to ZA (134.2 mg/g). The loading behaviors using ZA as well as CS/ZA and CD/ZA were illustrated based on the classic kinetics properties of pseudo-first-order kinetics (R2 > 0.95) and the traditional Langmuir isotherm (R2 = 0.99). CD/ZA shows a significantly higher active site density (102.7 mg/g) in comparison to CS/ZA (64 mg/g) and ZA (35.8 mg/g). The number of loaded 5-Fu per site of ZA, CS/ZA, and CD/ZA (>1) validates the vertical ordering of the loaded drug ions by multi-molecular processes. These processes are mainly physical mechanisms based on the determined Gaussian energy (<8 kJ/mol) and loading energy (<40 kJ/mol). Both the CS/ZA and CD/ZA 5-Fu release activities display continuous and controlled profiles up to 80 h, with CD/ZA exhibiting much faster release. According to the release kinetics studies, the release processes contain non-Fickian transport release properties, suggesting cooperative diffusion and erosion release mechanisms. The cytotoxicity of 5-Fu is also significantly enhanced by these carriers: 5-Fu/ZA (11.72% cell viability), 5-Fu/CS/ZA (5.43% cell viability), and 5-Fu/CD/ZA (1.83% cell viability).

Functionalized Sulfur-Containing Heterocyclic Analogs Induce Sub-G1 Arrest and Apoptotic Cell Death of Laryngeal Carcinoma In Vitro.

In this study, we speculate that the hydroxyl-containing benzo[b]thiophene analogs, 1-(3-hydroxybenzo[b]thiophen-2-yl) ethanone (BP) and 1-(3-hydroxybenzo[b]thiophen-2-yl) propan-1-one hydrate (EP), might possess antiproliferative activity against cancer cells. Hydroxyl-containing BP and EP show selectivity towards laryngeal cancer cells (HEp2), with IC50 values of 27.02 ± 1.23 and 35.26 ± 2.15 µM, respectively. The hydroxyl group present in the third position is responsible for the anticancer activity and is completely abrogated when the hydroxyl group is masked. BP and EP enhance the antioxidant enzyme activity and reduce the ROS production, which are correlated with the antiproliferative effect in HEp-2 cells. An increase in the BAX/BCL-2 ratio occurs during the BP and EP treatment and activates the caspase cascade, resulting in apoptosis stimulation. It also arrests the cells in the Sub-G1 phase, indicating the induction of apoptosis. The molecular docking and simulation studies predicted a strong interaction between BP and the CYP1A2 protein, which could aid in combinational therapy by enhancing the bioavailability of the drugs. BP and EP possess an antioxidant property with low antiproliferative effects (~5.18 µg/mL and ~7.8 µg/mL) as a standalone drug, therefore, they can be combined with other drugs for effective chemotherapy that might trigger the effect of pro-oxidant drug on healthy cells.

Overview of Physicochemical Properties of Nanoparticles as Drug Carriers for Targeted Cancer Therapy.

The advent of nanotechnology has brought about revolutionary innovations in biological research techniques and medical practice. In recent years, various "smart" nanocarriers have been introduced to deliver therapeutic agents specifically to the tumor tissue in a controlled manner, thereby minimizing their side effects and reducing both dosage and dosage frequency. A large number of nanoparticles have demonstrated initial success in preclinical evaluation but modest therapeutic benefits in the clinical setting, partly due to insufficient delivery to the tumor site and penetration in tumor tissue. Therefore, a precise understanding of the relationships betweenthe physicochemical properties of nanoparticles and their interaction with the surrounding microenvironment in the body is extremely important for achieving higher concentrations and better functionality in tumor tissues. This knowledge would help to effectively combine multiple advantageous functions in one nanoparticle. The main focus of the discussion in this review, therefore, will relate to the main physicochemical properties of nanoparticles while interacting within the body and their tuning potential for increased performance.

The Anticancer Efficacy of Thiourea-Mediated Reduced Graphene Oxide Nanosheets against Human Colon Cancer Cells (HT-29).

The current research focuses on the fabrication of water-soluble, reduced graphene oxide (rGO) employing thiourea (T) using a simple cost-effective method, and subsequently examining its anticancer characteristics. The cytotoxicity caused by graphene oxide (GO) and T-rGO is investigated in detail. Biological results reveal a concentration-dependent toxicity of GO and T-rGO in human colon cancer cells HT-29. A decrease in cell viability alongside DNA fragmentation is observed. Flow cytometry analysis confirms the cytotoxic effects. The novelty in this work is the use of raw graphite powder, and oxidants such as KMNO4, NaNO3, and 98 percent H2SO4 to produce graphene oxide by a modified Hummers method. This study demonstrates a simple and affordable procedure for utilising thiourea to fabricate a water-soluble reduced graphene oxide, which will be useful in a variety of biomedical applications.

Structural Study of Nano-Clay and Its Effectiveness in Radiation Protection against X-rays.

With the increasing applications of nuclear technology, radiation protection has become very important especially for the environment and the personnel close to radiation sources. Natural clays can be used potentially for shielding the X-ray radiations. In this study, the correlation between structural parameters and radiation shielding performance of natural clay extracted from Algerian Sahara (Adrar, Reggan, and Timimoune) was investigated. Phase composition and structural parameters (lattice parameters, average crystallite size, and microstrain) were determined by the Rietveld refinements of X-ray diffraction patterns in the frame of HighScore Plus software. The obtained results showed that the studied clays are nanocrystalline (nano-clay) since the calculated crystallite size was ≈3 nm for the feldspar phase. FTIR spectra confirmed the presence of all phases already detected by XRD analysis besides Biotite (around the band at 3558 cm-1). The remaining bands corresponded to absorbed and adsorbed water (3432 cm-1 and 1629 cm-1, respectively) and atmospheric CO2 (2356 cm-1). The shielding properties (mass absorption coefficient-µ/ρ and radiative attenuation rate-RA) for (green-yellow, green, and red) clays of Adrar, (red, white, and white-red) clays of Reggan, and red clay of Timimoune at same energy level were examined. The results of clay samples were compared with each other. The obtained results indicated that the green clay of Adrar exhibited the superior radiation shielding, i.e., 99.8% and 243.4 cm2/g for radiative attenuation rate and mass absorption coefficient, respectively.

Oxidized Alginate Dopamine Conjugate: In Vitro Characterization for Nose-to-Brain Delivery Application.

BACKGROUND: The blood-brain barrier (BBB) bypass of dopamine (DA) is still a challenge for supplying it to the neurons of Substantia Nigra mainly affected by Parkinson disease. DA prodrugs have been studied to cross the BBB, overcoming the limitations of DA hydrophilicity. Therefore, the aim of this work is the synthesis and preliminary characterization of an oxidized alginate-dopamine (AlgOX-DA) conjugate conceived for DA nose-to-brain delivery. METHODS: A Schiff base was designed to connect oxidized polymeric backbone to DA and both AlgOX and AlgOX-DA were characterized in terms of Raman, XPS, FT-IR, and 1H- NMR spectroscopies, as well as in vitro mucoadhesive and release tests. RESULTS: Data demonstrated that AlgOX-DA was the most mucoadhesive material among the tested ones and it released the neurotransmitter in simulated nasal fluid and in low amounts in phosphate buffer saline. Results also demonstrated the capability of scanning near-field optical microscopy to study the structural and fluorescence properties of AlgOX, fluorescently labeled with fluorescein isothiocyanate microstructures. Interestingly, in SH-SY5Y neuroblastoma cell line up to 100 µg/mL, no toxic effect was derived from AlgOX and AlgOX-DA in 24 h. CONCLUSIONS: Overall, the in vitro performances of AlgOX and AlgOX-DA conjugates seem to encourage further ex vivo and in vivo studies in view of nose-to-brain administration.

Occurrence and characterization of surface sediment microplastics and litter from North African coasts of Mediterranean Sea: Preliminary research and first evidence.

The aim of this research work is to investigate the abundance of macroplastics and microplastics from North African coasts of Mediterranean Sea in the Gulf of Annaba (Algeria). The study areas are well known for the high population and high industrial activities that can contribute to the plastic pollution. In this case, microplastics were sampled from the surface sediments at four different stations representing different local activities. The microplastics were extracted by the density separation method from collected samples. Each particle was identified using Fourier transform infrared spectroscopy (ATR-FTIR). Furthermore, beach litters were obtained according to the adopted methodology - guidance on monitoring of marine litter in European seas. The obtained mean concentrations are 182.66 ± 27.32 and 649.33 ± 184.02 kg-1 sediment (DW), respectively. A detailed spatial and quantitative analysis revealed that their distribution was a function of proximity to populated areas and associated with wastewater effluents, industrial installations, degree of shore exposure and complex tidal flow patterns. Five types of microplastics were identified; the most common were fibers (70%), fragments (21%), pellets (5%), films (2%) and foams (2%). Attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) analysis showed that the main polymers detected in the studied microplastics were polyethylene (48%), polypropylene (16%), polyethylene terephthalate (14%), polystyrene (9%), butyl branham (7%), ethylene propylene (3%) and cellulose tri acetate (3%). The obtained results provided a holistic view of the abundance, distribution, and characteristics of microplastics in the Gulf of Annaba. This study confirmed the presence of microplastics in the Gulf of Annaba, albeit further studies remain necessary for better understanding of sources and fate.

Multiwalled carbon nanotube buckypaper induces cell cycle arrest and apoptosis in human leukemia cell lines through modulation of AKT and MAPK signaling pathways.

MWCNT buckypaper (BP) shows physico-chemical and mechanical properties that make it potentially useful as a substrate in nano-bio interface research including in tissue engineering. When used as a scaffold material, BP comes into contact with host cells and surrounding tissues; therefore it is critical to determine its biocompatibility and interaction with living systems. The aim of this study was to investigate BP effects on cell growth, apoptosis and reactive oxygen species (ROS) production in three human leukemia cell lines HL-60, U-937 and K-562. BP was able to induce both the reduction of cell proliferation, associated with an arrest in G0/G1 phase of cell cycle and the increase of apoptosis in leukemic cell lines, thus exerting both cytostatic and cytotoxic effects. The growth inhibitory effect was likely mediated by the decrease of cyclins D, E, A, B1 levels and CDK4 expression; meanwhile, the apoptotic effect, not mediated by ROS production, was presumably due to the combined action of the survival and pro-apoptotic AKT and MAPK signal transduction pathways. These results raised the issue of biocompatibility of MWCNT BP for the creation of carbon nanotubes based scaffolds to utilize as prostheses in tissue engineering.

Differences in cytotoxic, genotoxic, and inflammatory response of bronchial and alveolar human lung epithelial cells to pristine and COOH-functionalized multiwalled carbon nanotubes.

Functionalized MWCNTs are used in many commercial and biomedical applications, but their potential health effects are not well defined. We investigated and compared cytotoxic, genotoxic/oxidative, and inflammatory effects of pristine and carboxyl MWCNTs exposing human respiratory (A549 and BEAS-2B) cells to 1-40 µg/mL of CNTs for 24 h. Both MWCNTs induced low viability reduction (by WST1 assay) in A549 cells and only MWCNTs-COOH caused high viability reduction in BEAS-2B cells reaching 28.5% viability at 40 µg/mL. Both CNTs induced membrane damage (by LDH assay) with higher effects in BEAS-2B cells at the highest concentrations reaching 20% cytotoxicity at 40 µg/mL. DNA damage (by Fpg-comet assay) was induced by pristine MWCNTs in A549 cells and by both MWCNTs in BEAS-2B cells reaching for MWCNTs-COOH a tail moment of 22.2 at 40 µg/mL versus 10.2 of unexposed cells. Increases of IL-6 and IL-8 release (by ELISA) were detected in A549 cells exposed to MWCNTs-COOH from 10 µg/mL while IL-8 increased in BEAS-2B cells exposed to pristine MWCNTs at 20 and 40 µg/mL. The results show higher cytogenotoxicity of MWCNTs-COOH in bronchial and of pristine MWCNTs in alveolar cells. Different inflammatory response was also found. The findings suggest the use of in vitro models with different end points and cells to study CNT toxicity.

Targeted nanodrugs for cancer therapy: prospects and challenges.

The recent advent of nanomedicine holds potential to revolutionize cancer therapy. This innovative discipline has paved the way for the emergence of a new class of drugs based on nanoengineered particles. These "nanodrugs" are designed to greatly enhance drug therapeutic indices. First-generation nanodrugs consisted of conventional anti-cancer drugs loaded into/onto nanoengineered particles (nanocarriers) devoid of targeting features (non-targeted nanodrugs). Non-targeted nanodrugs have provided the opportunity to carry large amounts of drugs, including poorly water-soluble and/or permeable drugs, to several types of tumors, improving the therapeutic index with respect to comparable free drugs. Although effective, the primary delivery mechanism of non-targeted nanodrugs was through passive tissue accumulation, due to pathophysiological differences between tumor-associated and healthy vessels, and through non-specific targeting of cell subsets, posing the danger of off-target binding and effects. Recently, the therapeutic indices of certain anti-cancer drugs were further improved by attaching targeting ligands to nanodrugs (targeted-nanodrugs). Targeted-nanodrugs selectively bind to cognate receptors expressed on target cells and enter cells more efficiently than non-targeted formulations. Although these advancements have been sufficiently beneficial to place targeted-nanodrugs into clinical development for use in cancer therapy, they also come at a price. The addition of ligands to drug-loaded nanocarriers often leads to additional synthesis steps and costs, and more complex biological performance relative to ligand-devoid nanodrugs. Here, we will discuss the benefits and challenges facing the addition of targeting features to nanodrugs for cancer therapy.

In vivo targeting of intratumor regulatory T cells using PEG-modified single-walled carbon nanotubes.

Recent evidence regarding the role of regulatory T cells (Treg) in tumor development has suggested that the manipulation of Treg function selectively in the tumor microenvironment would be a desirable immunotherapy approach. Targeting intratumor immune populations would reduce side effects on peripheral healthy cells and increase antitumor efficacy of immunotherapies. However, no current approaches are available which enable selective in vivo targeting of intratumor Treg or other immune cell subpopulations. Herein, we investigated the ability of ligands against Treg-specific receptors to drive selective internalization of PEG-modified single-walled carbon nanotubes (PEG-SWCNTs) into Treg residing in the tumor microenvironment. We focused our attention on the glucocorticoid-induced TNFR-related receptor (GITR), as it showed higher overexpression on intratumor vs peripheral (i.e., splenic) Treg compared to other reported Treg-specific markers (folate receptor 4, CD103, and CD39). Ex vivo investigations showed that the Treg targeting efficiency and selectivity of PEG-SWCNTs depended on incubation time, dose, number of ligands per nanotube, and targeted surface marker. In vivo investigations showed that PEG-SWCNTs armed with GITR ligands targeted Treg residing in a B16 melanoma more efficiently then intratumor non-Treg or splenic Treg. The latter result was achieved by exploiting a combination of passive tumor targeting due to enhanced tumor vascular permeability, naturally increased intratumor Treg vs effector T cell (Teff) ratio, and active targeting of markers that are enriched in intratumor vs splenic Treg. We also found that PEG-SWCNTs loaded with GITR ligands were internalized by Treg through receptor-mediated endocytosis and transported into the cytoplasm and nucleus ex vivo and in vivo. This is the first example of intratumor immune cell targeting and we hope it will pave the way to innovative immunotherapies against cancer.

Comparative cyto-genotoxicity assessment of functionalized and pristine multiwalled carbon nanotubes on human lung epithelial cells.

Chemical functionalization extends CNT applications conferring them new functions, but could modify their toxicity. We compared cytotoxicity and genotoxic/oxidative effects of -OH functionalized and pristine MWCNTs to evaluated the influence of the functionalization exposing A549 cells to 1-40µg/ml of both MWCNTs for 2, 4 and 24h. Cytotoxicity was evaluated by MTT and LDH tests and apoptosis induction, direct/oxidative DNA damage by Fpg-modified comet assay. After 24h we found viability reduction significant at 20 and 40µg/ml for both the MWCNTs with a detectable viability reduction already at lower concentrations for MWCNTs. A significant LDH release was found only for MWCNTs. Significant apoptosis induction was found from 10µg/ml of MWCNT-OH. A concentration-dependent increase of direct DNA damage, significant at 40µg/ml of MWCNTs and beginning from 5µg/ml of MWCNT-OH was detected at all exposure times. Oxidative DNA damage was not observed for both CNTs. The results indicate a different cytotoxic mechanism, by membrane damage for MWCNTs and apoptosis for MWCNT-OH, that could be explained by a different cellular uptake. Moreover, we found an earlier genotoxic effect for MWCNT-OH. The findings suggest that further studies on functionalized CNTs are necessary before using them in several applications particularly in biomedical field.

Non-functionalized multi-walled carbon nanotubes alter the paracellular permeability of human airway epithelial cells.

Little information is available upon the effects of carbon nanotubes (CNT) on the airway barrier. Here we study the barrier function of Calu-3 human airway epithelial cells, grown on permeable filters, after the exposure to commercial single-walled or multi-walled CNT, produced through chemical vapour deposition. To assess changes in the paracellular permeability of CNT-treated Calu-3 monolayers, we have measured the trans-epithelial electrical resistance (TEER) and the permeability to mannitol. Multi-walled CNT caused a large decrease in TEER and an increase in mannitol permeability but no substantial alteration in monolayer viability. Single-walled CNT produced much smaller changes of TEER while CNT, synthesized through the arc discharge method, and Carbon Black nanoparticles had no effect. If commercial multi-walled CNT were added during the formation of the tight monolayer, no further increase in trans-epithelial resistance was observed. Moreover, the same nanomaterials, but neither single-walled counterparts nor Carbon Black, prevented the TEER recovery observed after the discontinuation of interleukin-4, a Th2 cytokine that causes a reversible barrier dysfunction in airway epithelia. These findings suggest that commercial multi-walled CNT interfere with the formation and the maintenance of tight junctional complexes in airway epithelial cells.