Genoprotective role of pembrolizumab liposome in isolated lymphocytes from head and neck squamous cell carcinoma patients compared to those from healthy individuals in vitro.

Pembrolizumab has shown significant anticancer effects against various human cancers. The present study investigated the effects of pembrolizumab liposome and nano (naked) forms in treated lymphocytes from head and neck squamous cell carcinoma (HNSCC) patients compared to healthy individuals. The level of oxidative DNA damage induced by hydrogen peroxide (H2O2) was also investigated. A concentration of 10 µg/ml of pembrolizumab liposome was used to treat the lymphocytes in the Comet and micronucleus assays based on the preliminary dosage optimization tests. To determine the cellular pathways involved in the protective role of pembrolizumab against H2O2, several proteins involved in apoptosis (P53, P21 and Bcl-2) were assessed. Pembrolizumab significantly reduced DNA damage and decreased the number of micronuclei in lymphocytes from HNSCC patients (p < 0.01) compared with healthy individuals. The 10 µg/ml of pembrolizumab liposome significantly reduced the oxidative stress induced by H2O2 and was effective in healthy and HNSCC groups using the Comet and micronucleus assays (p < 0.001). To our knowledge, this is the first report of pembrolizumab in liposome and naked forms exhibiting a protective effect on DNA damage in the treatment of HNSCC patients.

The anticancer/cytotoxic effect of a novel gallic acid derivative in non-small cell lung carcinoma A549 cells and peripheral blood mononuclear cells from healthy individuals and lung cancer patients.

Gallic acid (GA) is a naturally occurring polyphenol with a strong antioxidant capacity. GA stimulates the apoptosis of cancer cells, thereby suppressing cancer cell invasion. However, the low oral permeability of GA limits its therapeutic use. In order to enhance the antioxidant capacity and oral permeability of GA, a series of compounds analogous to GA were synthesized: 4-methoxybenzenesulfonamide (MBS), 3,4-dimethoxybenzenesulfonamide (DMBS) and 3,4,5-trimethoxybenzenesulfonamide (TMBS). In the new compounds, hydroxyl groups were replaced with various numbers of methoxy groups (stronger electron-donating groups), to increase hydrophobicity and oral permeability compared to GA. In addition, the carboxylic group was replaced with a sulfonyl group (a stronger electron-withdrawing group), to increase the molecular polarity and antioxidative activities of the compounds. The cell counting kit-8 (CCK-8) assay was used to detect the effect of GA, MBS, DMBS, and TMBS on cell proliferation and apoptosis in peripheral blood mononuclear cells (PBMCs) from healthy individuals and non-small cell lung carcinoma A549 cells. Additionally, the comet assay was used to assess the genotoxicity of these compounds in PBMCs from healthy individuals, lung cancer patients, and A549 cells. Compared to untreated cells, TMBS reduced DNA damage more effectively than GA in PBMCs from lung cancer patients and healthy donors. Furthermore, in comparison to GA, TMBS was more cytotoxic in A549 cells. Moreover, TMBS was not cytotoxic in healthy PBMCs, suggesting that TMBS demonstrates therapeutic potential in cancer.

Comparing P53 expression and genome-wide transcriptome profiling to Comet assay in lymphocytes from melanoma patients and healthy controls.

This study compared the expression of TP53 in lymphocytes from malignant melanoma (MM) patients with positive sentinel nodes to healthy controls (HCs) following exposure to various doses of UVA radiation. The Lymphocyte Genome Sensitivity (LGS) assay indicated significant differences in DNA damage in lymphocytes between MM patients and HCs. qPCR data demonstrated an overall 3.4-fold increase in TP53 expression in lymphocytes from MM patients compared to healthy controls, following treatment with 0.5 mW/cm2 UVA radiation. Western blotting confirmed that p53 expression was increased in MM lymphocytes following UVA exposure compared to healthy individuals. Genome transcriptome profiling data displayed differences in gene expression between UVA-treated lymphocytes from MM patients and HCs. Peripheral lymphocytes from MM patients are more susceptible to the genotoxic effects of UVA compared to healthy individuals. Our previous studies showed that UVA exposure of various intensities caused significant differences in the levels of DNA damage between lymphocytes from cancer patients compared to HCs through the LGS assay. The present study's results provide further credibility to the LGS assay as a screening test for cancer detection. Peripheral lymphocytes could be a promising blood biopsy biomarker for staging of carcinomas and prevention of carcinoma progression at early stages.

Anticarcinogenic impact of extracellular vesicles (exosomes) from cord blood stem cells in malignant melanoma: A potential biological treatment.

Incidence of Malignant Melanoma has become the 5th in the UK. To date, the major anticancer therapeutics include cell therapy, immunotherapy, gene therapy and nanotechnology-based strategies. Recently, extracellular vesicles, especially exosomes, have been highlighted for their therapeutic benefits in numerous chronic diseases. Exosomes display multifunctional properties, including inhibition of cancer cell proliferation and initiation of apoptosis. In the present in vitro study, the antitumour effect of cord blood stem cell (CBSC)-derived exosomes was confirmed by the CCK-8 assay (p < 0.05) on CHL-1 melanoma cells and improve the repair mechanism on lymphocytes from melanoma patients. Importantly, no significant effect was observed in healthy lymphocytes when treated with the exosome concentrations at 24, 48 and 72 h. Comet assay results (OTM and %Tail DNA) demonstrated that the optimal exosome concentration showed a significant impact (p < 0.05) in lymphocytes from melanoma patients whilst causing no significant DNA damage in lymphocytes of healthy volunteers was 300 µg/ml. Similarly, the Comet assay results depicted significant DNA damage in a melanoma cell line (CHL-1 cells) treated with CBSC-derived exosomes, both the cytotoxicity of CHL-1 cells treated with CBSC-derived exosomes exhibited a significant time-dependent decrease in cell survival. Sequencing analysis of CBSC exosomes showed the presence of the let-7 family of miRNAs, including let-7a-5p, let-7b-5p, let-7c-5p, let-7d-3p, let-7d-5p and two novel miRNAs. The potency of CBSC exosomes in inhibiting cancer progression in lymphocytes from melanoma patients and CHL-1 cells whilst causing no harm to the healthy lymphocytes makes it a potential candidate as an anticancer therapy.

Interferon-γ liposome: a new system to improve drug delivery in the treatment of lung cancer.

Lung cancer is one of the main causes of death worldwide. Published data show the use of interferons (IFNs) in treating lung tumours. IFNs also have potential for their antiproliferative, antiangiogenic, immunoregulatory and proapoptotic effects. IFN-γ functions as an anticancer agent against various forms of cancer. This study aimed to investigate the effect of IFN-γ liposome (nano) on peripheral lymphocytes from 20 individuals in each group: lung cancer patients compared to healthy individuals. The effectiveness of IFN-γ liposome against oxidative stress was also evaluated in this study. A concentration of 100 U·mL-1 of IFN-γ liposome was used to treat the lymphocytes in the Comet and micronucleus assays based on the preliminary test for the optimal dose. The lymphocytes from lung cancer patients presented with higher DNA damage levels than those of healthy individuals. In healthy individuals, IFN-γ liposome did not cause any DNA damage in the lymphocytes. Also, it caused a significant reduction in DNA damage in the lymphocytes from lung cancer patients in both the Comet and micronucleus assays. The 100 U·mL-1 of IFN-γ liposome significantly reduced the oxidative stress caused by H2O2 and appeared to be effective in both groups using the Comet and micronucleus assays. Results from both Comet and micronucleus assays were consistent. The data obtained indicated that IFN-γ in both forms (IFN-γ bulk and IFN-γ nanoliposome) may potentially be effective for the treatment of lung cancer and showed the ability of IFN-γ liposome to reduce DNA damage more than the bulk form.

Ex vivo/in vitro effects of aspirin and ibuprofen, bulk and nano forms, in peripheral lymphocytes of prostate cancer patients and healthy individuals.

Inhibiting inflammatory processes or eliminating inflammation represents a logical role in the suppression and treatment strategy of cancer. Several studies have shown that anti-inflammatory drugs (NSAIDs) act as anticancer agents while reducing metastases and mortality rate. NSAIDs are seriously limited by their side effects and toxicity, which can become cumulative with their long-term administration for chemoprevention. In the current ex vivo / in vitro study, the genotoxicity mechanisms of NSAIDS in bulk and nanoparticle forms allowed a strategy to prevent and minimise the damage in human lymphocytes. When compared to their bulk forms, acetylsalicylic acid (Aspirin) nano and ibuprofen nano (IBU N), both NSAIDs in 500 µg/mL concentration significantly decreased DNA damage measured by alkaline comet assay. Micronuclei (MNi) frequency also decreased after ASP N (500 µg/mL), ASP B (500 µg/mL) and IBU N (200 µg/mL) in prostate cancer patients and healthy individuals, however, the ibuprofen bulk (200 µg/mL) showed a significant increase in MNi formation in lymphocytes from healthy and prostate cancer patients when compared to the respective untreated lymphocytes. These findings suggest that a reduction in particle size had an impact on the reactivity of the drug, further emphasising the potential of nanoparticles to improve the current treatment options.

Anticancer potential of myricetin bulk and nano forms in vitro in lymphocytes from myeloma patients.

Evading apoptosis and chemo-resistance are considered as very important factors which help tumour progression and metastasis. Hence, to overcome chemo-resistance, there is an urgent requirement for emergence of more effective treatment options. Myricetin, a naturally occurring flavonoid, is present in various plant-derived foods and shows antitumour potential in different cancers. In the present in vitro study, results from the comet assay demonstrated that myricetin bulk (10 µM) and nano (20 µM) forms exhibited a non-significant level of genotoxicity in lymphocytes from multiple myeloma patients when compared to those from healthy individuals. Western blot results showed a decrease in Bcl-2/Bax ratio and an increase in P53 protein levels in lymphocytes from myeloma patients, but not in lymphocytes from healthy individuals. A significant increase in intracellular reactive oxygen species level was also observed, suggesting that regulation of apoptotic proteins triggered by myricetin exposure in lymphocytes from myeloma patients occurred through P53 and oxidative stress-dependent pathways. The potency of myricetin against lymphocytes from myeloma patients marks it a potential candidate to be considered as an alternative to overcome chemo-resistance in cancer therapies.

The Antiviral, Anti-Inflammatory Effects of Natural Medicinal Herbs and Mushrooms and SARS-CoV-2 Infection.

The 2019 novel coronavirus, SARS-CoV-2, producing the disease COVID-19 is a pathogenic virus that targets mostly the human respiratory system and also other organs. SARS-CoV-2 is a new strain that has not been previously identified in humans, however there have been previous outbreaks of different versions of the beta coronavirus including severe acute respiratory syndrome (SARS-CoV1) from 2002 to 2003 and the most recent Middle East respiratory syndrome (MERS-CoV) which was first identified in 2012. All of the above have been recognised as major pathogens that are a great threat to public health and global economies. Currently, no specific treatment for SARS-CoV-2 infection has been identified; however, certain drugs have shown apparent efficacy in viral inhibition of the disease. Natural substances such as herbs and mushrooms have previously demonstrated both great antiviral and anti-inflammatory activity. Thus, the possibilities of natural substances as effective treatments against COVID-19 may seem promising. One of the potential candidates against the SARS-CoV-2 virus may be Inonotus obliquus (IO), also known as chaga mushroom. IO commonly grows in Asia, Europe and North America and is widely used as a raw material in various medical conditions. In this review, we have evaluated the most effective herbs and mushrooms, in terms of the antiviral and anti-inflammatory effects which have been assessed in laboratory conditions.

Ex vivo/in vitro protective effect of myricetin bulk and nano-forms on PhIP-induced DNA damage in lymphocytes from healthy individuals and pre-cancerous MGUS patients.

2-Amino-1-methyl-6-phenylimidazo [4,5-b]pyridine (PhIP) is a central dietary mutagen, produced when proteinaceous food is heated at very high temperatures potentially causing DNA strand breaks. This study investigates the protective potential of a well-researched flavonoid, myricetin in its bulk and nano-forms against oxidative stress induced ex vivo/in vitro by PhIP in lymphocytes from pre-cancerous monoclonal gammopathy of undetermined significance (MGUS) patients and those from healthy individuals. The results from the Comet assay revealed that in the presence of myricetin bulk (10 µM) and myricetin nano (20 µM), the DNA damage caused by a high dose of PhIP (100 µM) was significantly (P < 0.001) reduced in both groups. However, nano has shown better protection in lymphocytes from pre-cancerous patients. Consistent results were obtained from the micronucleus assay where micronuclei frequency in binucleated cells significantly decreased upon supplementing PhIP with myricetin bulk (P < 0.01) and myricetin nano (P < 0.001), compared to the PhIP treatment alone. To briefly determine the cellular pathways involved in the protective role of myricetin against PhIP, we studied gene expression of P53 and ATR kinase (ATM- and Rad3-related), using the real-time PCR technique.

An in vitro investigation into the protective and genotoxic effects of myricetin bulk and nano forms in lymphocytes of MGUS patients and healthy individuals.

The present study investigated the genoprotective and genotoxic effects of myricetin bulk (10 µM) and nano forms (20 µM) in the lymphocytes from pre-cancerous, monoclonal gammopathy of unknown significance (MGUS) patients and healthy individuals using the Comet and micronucleus assays. The study also evaluated the effect of myricetin on P53 expression levels, using the Western blot technique. Results showed that throughout the in-vitro treatment, lymphocytes from the patients group had higher levels of baseline DNA damage compared to the healthy group. Myricetin in both forms induced significant DNA damage, only at higher concentrations (>40 µM). The micronucleus assay showed a significant reduction (P < 0.01) in the frequency of micronuclei in mono-nucleated cells in the patient group treated with the nano form of myricetin at the non-toxic dose of 20 µM. There was a significant increase in both gene and protein P53 levels in lymphocytes isolated from healthy individuals and pre-cancerous patients. These results suggested a protective effect of myricetin and indicated its nutritional supplement potential for protection against cancer development among patients suffering from MGUS.

ROS-induced oxidative damage in lymphocytes ex vivo/in vitro from healthy individuals and MGUS patients: protection by myricetin bulk and nanoforms.

We investigated the protective role of myricetin bulk and nanoforms, against reactive oxygen species (ROS)-induced oxidative stress caused by hydrogen peroxide and tertiary-butyl hydro peroxide in lymphocytes in vitro from healthy individuals and those from pre-cancerous patients suffering with monoclonal gammopathy of undetermined significance (MGUS). The change in intracellular reactive oxygen species was measured once cells were treated with myricetin bulk forms and nanoforms with and without either hydrogen peroxide or tertiary-butyl hydro peroxide co-supplementation. The direct and indirect antioxidant activity of myricetin was spectrofluometrically measured using the fluorescent dye 2',7'-dichlorofluorescin diacetate and using the Comet assay, respectively. Hydrogen peroxide (50 µM) and tertiary-butyl hydro peroxide (300 µM) induced a higher level of reactive oxygen species-related DNA damage and strand breaks. Addition of myricetin nanoform (20 µM) and bulk (10 µM) form could, however, significantly prevent hydrogen peroxide- and tertiary-butyl hydro peroxide-induced oxidative imbalances and the nanoform was more effective. Glutathione levels were also quantified using a non-fluorescent dye. Results suggest that myricetin treatment had no significant effect on the cellular antioxidant enzyme, glutathione. The current study also investigates the effect of myricetin on the induction of double-strand breaks by staining the gamma-H2AX foci immunocytochemically. It was observed that myricetin does not induce double-strand breaks at basal levels rather demonstrated a protective effect.

Using a Modified Lymphocyte Genome Sensitivity (LGS) test or TumorScan test to detect cancer at an early stage in each individual.

Our previous case-control study observed isolated lymphocytes from 208 individuals and determined the differences in the sensitivity to genomic damage of lymphocytes derived from cancer patients, pre/suspect cancer patients and healthy volunteers using the Comet assay (Anderson et al, 2014). We adapted the LGS technique using a slightly different method and examined 700 more blood samples from 598 patients with cancer or suspected cancer and 102 healthy individuals. To help increase the sensitivity of the test and detect cancer at the level of each individual, we joined with the IMSTAR team who analysed our cells with their fully automated Pathfinder™ cell reader-analyser system. With this reading and analysis system 4,000 to 10,000 cells were able to be read per slide. The new test which is called TumorScan is a highly sensitive test to detect any cancer at an early stage through the response of the white blood cells to UV treatment. These patient blood samples have also been collected at the stage before confirming diagnosis and treatment. There were four of these individuals with cancer who had received anti-cancer treatment. The results from these patients showed a reverse pattern compared to non-treated cancer patients and followed the pattern seen in healthy individuals. The results are consistent with the early results as reported in the above 2014 paper. Given the results from these samples were in a particularly challenging subgroup, whose cancer status was difficult to distinguish, the data suggest that the technique using the TumorScan system could exceed the area under the ROC curve >93% obtained in the earlier study on a group basis, whereas this present study was to detect cancer at an early stage in each individual.

DNA damage protection by bulk and nano forms of quercetin in lymphocytes of patients with chronic obstructive pulmonary disease exposed to the food mutagen 2-amino-3-methylimidazo [4,5-f]quinolone (IQ).

Chronic obstructive pulmonary disease (COPD) in humans, describes a group of lung conditions characterised by airflow limitation that is poorly reversible. The airflow limitation usually progresses slowly and is related to an abnormal inflammatory response of the lung to toxic particles. COPD is characterised by oxidative stress and an increased risk of lung carcinoma. The 2-amino-3-methylimidazo [4,5-f]quinoline (IQ) is one of a number of mutagenic/carcinogenic heterocyclic amines found mainly in well-cooked meats which are thus part of the regular diet. Antioxidants are very important in order to protect the cells against oxidative damage. The aim of the present study was to assess the effects of IQ on the level of DNA damage and susceptibility to a potent mutagen in peripheral blood cells of COPD patients. DNA damage and the frequency of micronuclei (MNi) were evaluated using the Comet and micronucleus assays, respectively. Differential expressions of both mRNA and protein of the endogenous antioxidant enzyme catalase were evaluated with quantitative polymerase chain reaction (qPCR) and Western blot analysis, respectively. Furthermore, the effect of bulk and nano forms of quercetin and their combination with IQ were examined. Results of the present study clearly demonstrated that MNi frequency in the peripheral blood lymphocytes exhibited a positive correlation with the DNA damage as evident from the different Comet assay parameters. Increase of the endogenous antioxidant catalase also showed there was a stimulation of this enzyme system by IQ. Whereas, the endogenous antioxidant quercetin significantly reduced oxidative stress in COPD patients and healthy individuals.

TiO2 NPs Induce DNA Damage in Lymphocytes from Healthy Individuals and Patients with Respiratory Diseases-An Ex Vivo/In Vitro Study.

Little is known of the effects of nanoparticles in human systems, let alone in diseased individuals and nanotechnology has preceded nanotoxicology. Therefore, the effects of titanium dioxide (TiO2) nanoparticles in peripheral blood lymphocytes from patients with respiratory diseases [lung cancer, chronic obstructive pulmonary disease (COPD) and asthma] were compared with those in healthy Individuals, to determine differences in sensitivity to nanochemical insult. The Comet assay was performed according to recommended guidelines. The micronucleus assay and ras oncoprotein detection were conducted according to published standard methods. The results showed statistically significant concentration-dependent genotoxic effects of TiO2 NPs in both respiratory patient and control groups in the Comet assay. The TiO2 NPs caused DNA damage in a concentration dependent manner in both groups (respiratory and healthy controls) with the exception of the lowest TiO2 concentration (10 µg/ml) which did not induce significant damage in healthy controls (n.s). When OTM data were used to compare the whole patient group and the control group, the patient group had more DNA damage (p > 0.001) with the exception of 10 µg/ml of TiO2 that caused less significant damage to patient lymphocytes (p < 0.05). Similarly, there was an increase in the pattern of cytogenetic damage measured in the MN assay without statistical significance except when compared to the negative control of healthy individuals. Furthermore, when modulation of ras p21 expression was investigated, regardless of TiO2 treatment, only lung cancer and COPD patients expressed measurable ras p21 levels. Results were achieved in the absence of cytotoxicity.

Aspirin and ibuprofen, in bulk and nanoforms: Effects on DNA damage in peripheral lymphocytes from breast cancer patients and healthy individuals.

Regular use of non-steroidal anti-inflammatory drugs (NSAIDs) may be protective against tumours, including breast cancer. We have studied the effects of ibuprofen and aspirin on DNA damage in lymphocytes obtained from breast cancer patients and healthy female controls. Both nanoparticle (NPs) and bulk formulations were used in the comet and micronucleus (MN) assays. Non-toxic doses (250 ng/ml ibuprofen; 500 ng/ml aspirin) were tested. Aspirin, both bulk and nano formulations, significantly reduced DNA damage measured with the comet and micronucleus assays; the nano formulation was more effective. Ibuprofen was not effective in the comet assay but showed a significant reduction in MN frequency, with the nano formulation being more effective. NPs may have better penetration through the nuclear membrane relative to the bulk formulation. NSAIDs such as aspirin and ibuprofen may have a promising role in cancer prevention and treatment.

Inhibition of survivin expression after using oxaliplatin and vinflunine to induce cytogenetic damage in vitro in lymphocytes from colon cancer patients and healthy individuals.

Chemotherapy drugs usually inflict a lethal dose to tumour cells with the consequence that these cells are being killed by cell death. However, each round of chemotherapy also causes damage to normal somatic cells. The DNA cross-linking agent oxaliplatin (OXP), which causes DNA double-strand breaks, and vinflunine (VFN), which disrupts the mitotic spindle, are two of these chemotherapy drugs which were evaluated in vitro using peripheral lymphocytes from colorectal cancer patients and healthy individuals to determine any differential response. Endpoints examined included micronucleus (MN) induction using the cytokinesis-blocked micronucleus (CBMN) assay and pancentromeric fluorescence in situ hybridisation. Also, survivin expression was monitored since it regulates the mitotic spindle checkpoint and inhibits apoptosis. OXP produced cytogenetic damage (micronuclei in binucleated cells) via its clastogenic but also previously unknown aneugenic action, possibly through interfering with topoisomerase II, whilst VFN produced micronuclei in mononucleated cells because of incomplete karyokinesis. Survivin expression was found to be significantly reduced in a concentration-dependent manner by not only OXP but surprisingly also VFN. This resulted in large numbers of multinucleated cells found with the CBMN assay. As survivin is upregulated in cancers, eliminating apoptosis inhibition might provide a more targeted chemotherapy approach; particularly, when considering VFN, which only affects cycling cells by inhibiting their mitotic spindle, and alongside possibly other pro-apoptotic compounds. Hence, these newly found properties of VFN -the inhibition of survivin expression-might demonstrate a promising chemotherapeutic approach as VFN induces less DNA damage in normal somatic cells compared to other chemotherapeutic compounds.

An evaluation of DNA damage in human lymphocytes and sperm exposed to methyl methanesulfonate involving the regulation pathways associated with apoptosis.

Exposure to DNA-damaging agents produces a range of stress-related responses. These change the expression of genes leading to mutations that cause cell cycle arrest, induction of apoptosis and cancer. We have examined the contribution of haploid and diploid DNA damage and genes involved in the regulation of the apoptotic process associated with exposure, The Comet assay was used to detect DNA damage and quantitative RT-PCR analysis (qPCR) to detect gene expression changes in lymphocytes and sperm in response to methyl methanesulfonate. In the Comet assay, cells were administered 0-1.2 mM of MMS at 37 °C for 30 min for lymphocytes and 32 °C for 60 min for sperm to obtain optimal survival for both cell types. In the Comet assay a significant increase in Olive tail moment (OTM) and % tail DNA indicated DNA damage at increasing concentrations compared to the control group. In the qPCR study, cells were treated for 4 h, and RNA was isolated at the end of the treatment. qPCR analysis of genes associated with DNA stress responses showed that TP53 and CDKN1A are upregulated, while BCL2 is downregulated compared with the control. Thus, MMS caused DNA damage in lymphocytes at increasing concentrations, but appeared not to have the same effect in sperm at the low concentrations. These results indicate that exposure to MMS increased DNA damage and triggered the apoptotic response by activating TP53, CDKN1A and BCL2. These findings of the processing of DNA damage in human lymphocytes and sperm should be taken into account when genotoxic alterations in both cell types are produced when monitoring human exposure.

DNA Damage in Healthy Individuals and Respiratory Patients after Treating Whole Blood In vitro with the Bulk and Nano Forms of NSAIDs.

Non-steroidal anti-inflammatory drugs (NSAIDs) inhibit COX enzyme activity which affects the inflammatory response. Inflammation is associated with increasing cancer incidence. Pre-clinical and clinical studies have shown that NSAID treatment could cause an anti-tumor effect in cancers. In the present study, blood was taken from healthy individuals (n = 17) and patients with respiratory diseases or lung cancer (n = 36). White blood cells (WBC) were treated with either a micro-suspension, i.e., bulk (B) or nano-suspension (N) of aspirin (ASP) or ibuprofen (IBU) up to 500 µg/ml in the comet assay and up to 125 µg/ml in the micronucleus assay. In this study results were compared against untreated lymphocytes and their corresponding treated groups. The results showed, that NSAIDs in their nano form significantly reduced the DNA damage in WBCs from lung cancer patients in bulk and nano compared to untreated lymphocytes. Also, there was a decrease in the level of DNA damage in the comet assay after treating WBCs from healthy individuals, asthma and COPD groups with aspirin N (ASP N) but not with IBU N. In addition, the number of micronuclei decreased after treatment with NSAIDs in their nano form (ASP N and IBU N) in the healthy as well as in the lung cancer group. However, this was not the case for micronucleus frequency in asthma and COPD patients. These data show that lymphocytes from different groups respond differently to treatment with ASP and IBU as measured by comet assay and micronucleus assay, and that the size of the suspended particles of the drugs affects responses.

Zinc oxide nanoparticles affect the expression of p53, Ras p21 and JNKs: an ex vivo/in vitro exposure study in respiratory disease patients.

Zinc oxide (ZnO) nanoparticles are the mostly used engineered metal oxide nanoparticles in consumer products. This has increased the likelihood of human exposure to this engineered nanoparticle (ENPs) through different routes. At present, the majority of the studies concerning ZnO ENPs toxicity have been conducted using in vitro and in vivo systems. In this study, for the first time we assessed the effect of ZnO ENPs on the major cellular pathways in the lymphocytes of healthy individuals as well as in susceptible patients suffering from lung cancer, chronic obstructive pulmonary disease (COPD) and asthma. Using the differential expression analysis, we observed a significant (P < 0.05) dose-dependent (10, 20 and 40 µg/ml for 6h) increase in the expression of tumour suppressor protein p53 (40, 60 and 110%); Ras p21 (30, 52 and 80%); c-Jun N-terminal kinases; JNKs) (28, 47 and 78%) in lung cancer patient samples treated with ZnO ENPs compared to healthy controls. A similar trend was also seen in COPD patient samples where a significant (P < 0.05) dose-dependent increase in the expression of tumour suppressor protein p53 (26, 45 and 84%), Ras p21 (21, 40 and 77%), JNKs (17, 32 and 69%) was observed after 6h of ZnO ENPs treatment at the aforesaid concentrations. However, the increase in the expression profile of tested protein was not significant in the asthma patients as compared to controls. Our results reiterate the concern about the safety of ZnO ENPs in consumer products and suggest the need for a complete risk assessment of any new ENPs before its use.