Bioremediation of hexavalent chromium by transformation of Escherichia coli DH5α with chromate reductase (ChrR) genes of Pseudomonas putida isolated from tannery effluent.

AIMS: Hexavalent chromium Cr(VI), a toxic heavy metal, is a serious pollutant of tannery effluent, and its accumulation in soil and water causes severe environmental concerns of increasing public health issues. The present study focus on the isolation and identification of chromium-reducing bacteria collected from the tannery industry in Dindigul, Tamil Nadu. Chromium-reducing bacteria Pseudomonas putida were identified by 16S rRNA sequencing followed by BLAST search. The plasmid with Cr(VI) reductase gene was isolated from Pseudomonas putida and transferred to E. coli DH5α for further studies. METHODS AND RESULTS: The bacterial cultures were kept under controlled conditions for 72 h to observe the growth rates and bacterial resistance to chromium. When strains wild type and transformant E. coli DH5α were grown in chromium supplemented media revealed significant growth, but strains cured type Pseudomonas putida and E. coli DH5α were minimum growth. The Cr(VI) reduction employed by transformant E. coli DH5α and wild Pseudomonas putida was 42.52 ± 1.48% and 44.46 ± 0.55%, respectively. The culture supernatant of the wild Pseudomonas putida and transformant E. coli DH5α showed an increased reduction of Cr(VI) compared to cell extract supernatant and cell debris due to the extracellular activity of chromium reductase has been responsible for Cr(VI) reduction. Besides, the chromium reductase gene was confirmed in the isolated Pseudomonas putida and transformant E. coli DH5α. CONCLUSIONS: Transformant bacteria could employ an alternative method for heavy metal detoxification in contaminated environments like tannery effluent and mining processes.

Bioremediation of hexavalent chromium by transformation of Escherichia coli DH5α with chromate reductase (ChrR) genes of Pseudomonas putida isolated from tannery effluent.

AIMS: Hexavalent chromium (Cr(VI)), a toxic heavy metal, is a serious pollutant from tannery effluent, and its accumulation in soil and water causes severe environmental concerns and increasing public health issues. The present study focuses on the isolation and identification of chromium-reducing bacteria collected from the tannery industry in Dindigul, Tamil Nadu. Chromium-reducing bacteria Pseudomonas putida were identified by 16S rRNA sequencing followed by BLAST search. The plasmid with Cr(VI) reductase gene was isolated from Ps. putida and transferred to Escherichia coli DH5α for further studies. METHODS AND RESULTS: The bacterial cultures were kept under controlled conditions for 72 h to observe the growth rates and bacterial resistance to chromium. When strains wild-type and transformant E. coli DH5α were grown in chromium-supplemented media, they revealed significant growth, but strains cured type Ps. putida and E. coli DH5α recorded minimum growth. The Cr(VI) reduction employed by transformant E. coli DH5α and wild Ps. putida was 42.52 ± 1.48% and 44.46 ± 0.55%, respectively. The culture supernatant of the wild Ps. putida and transformant E. coli DH5α showed an increased reduction of Cr(VI) compared with cell extract supernatant and cell debris due to the extracellular activity of chromium reductase being responsible for Cr(VI) reduction. Besides, the chromium reductase gene was confirmed in the isolated Ps. putida and transformant E. coli DH5α. CONCLUSIONS: Transformant bacteria could employ an alternative method for heavy metal detoxification in contaminated environments like tannery effluent and mining processes. SIGNIFICANCE AND IMPACT OF STUDY: High Cr(VI) concentration resistance and high Cr(VI) reducing the strain's ability make it suitable for bioremediation. These possible horizontal gene transfer events indicated in this study may have enabled transformant E. coli DH5α as a good candidate for reducing the heavy metal pollution.

Ligand-conjugated mesoporous silica nanorattles based on enzyme targeted prodrug delivery system for effective lung cancer therapy.

Epidermal growth factor receptor antibody (EGFRAb) conjugated silica nanorattles (SNs) were synthesized and used to develop receptor mediated endocytosis for targeted drug delivery strategies for cancer therapy. The present study determined that the rate of internalization of silica nanorattles was found to be high in lung cancer cells when compared with the normal lung cells. EGFRAb can specifically bind to EGFR, a receptor that is highly expressed in lung cancer cells, but is expressed at low levels in other normal cells. Furthermore, in vitro studies clearly substantiated that the cPLA2α activity, arachidonic acid release and cell proliferation were considerably reduced by pyrrolidine-2 loaded EGFRAb-SN in H460 cells. The cytotoxicity, cell cycle arrest and apoptosis were significantly induced by the treatment of pyrrolidine-2 loaded EGFRAb-SN when compared with free pyrrolidine-2 and pyrrolidine-2 loaded SNs in human non-small cell lung cancer cells. An in vivo toxicity assessment showed that silica nanorattles and EGFRAb-SN-pyrrolidine-2 exhibited low systemic toxicity in healthy Balb/c mice. The EGFRAb-SN-pyrrolidine-2 showed a much better antitumor activity (38%) with enhanced tumor inhibition rate than the pyrrolidine-2 on the non-small cell lung carcinoma subcutaneous model. Thus, the present findings validated the low toxicity and high therapeutic potentials of EGFRAb-SN-pyrrolidine-2, which may provide a convincing evidence of the silica nanorattles as new potential carriers for targeted drug delivery systems.

Green synthesis of multifunctional carbon quantum dots: An approach in cancer theranostics.

Carbon quantum dots (CQDs) have gained significant growing attention in the recent past due to their peculiar characteristics including smaller size, high surface area, photoluminescence, chemical stability, facile synthesis and functionalization possibilities. They are carbon nanostructures having less than 10 nm size with fluorescent properties. In recent years, the scientific community is curiously adopting biomass precursors for the preparation of CQDs over the chemical compounds. These biomass sources are sustainable, eco-friendly, inexpensive, widely available and convert waste into valuable materials. Hence in our work the fundamental understating of diverse fabrication methodologies of CQDs, and the types of raw materials employed in recent times, are all examined and correlated comprehensively. Their unique combination of remarkable properties, together with the ease with which they can be fabricated, makes CQDs as promising materials for applications in diverse biomedical fields, in particular for bio-imaging, targeted drug delivery and phototherapy for cancer treatment. The mechanism for luminescence is of considerable significance for leading the synthesis of CQDs with tunable fluorescence emission. Therefore, it is aimed to explore and provide an updated review on (i) the recent progress on the different synthesis methods of biomass-derived CQDs, (ii) the contribution of surface states or functional groups on the luminescence origin and (iii) its potential application for cancer theranostics, concentrating on their fluorescence properties. Finally, we explored the challenges in modification for the synthesis of CQDs from biomass derivatives and the future scope of CQDs in phototherapy for cancer theranostics.

Bisphenol-A alters hematopoiesis through EGFR/ERK signaling to induce myeloblastic condition in zebrafish model.

Experimental evidence from the etiology of cancer studies suggests that a correlation between Bisphenol-A (BPA) exposure and alterations in hematopoiesis leads to blood cancer. In our study zebrafish were used to assess the lethality, developmental effect, embryonic apoptosis and changes in transcription factor of hematopoiesis through EGFR/ERK signaling pathways in response to BPA. The in silico interaction of EGFR and BPA was analysed by molecular dynamic simulation. According to our results, BPA induced a significant lethal effect in hatching retardation, reduction in heart rate and teratogenic effects on zebrafish embryos and larvae at three different concentrations 100, 500 and 2500 µg/L. The mortality of adult zebrafish exposed to the acute toxicity of BPA from 5 to 30 mg/L concentrations was determined for 96 h. The peripheral blood cells and vital organs such as kidney, liver and spleen from BPA exposed fish showed predominantly abnormal myeloid blast cells along with severe morphological changes in erythrocytes at sublethal concentration 245 µg/L. The BPA showed the highest binding affinity to zebrafish EGFR with a docking score of -7.5 kcal/mol with an RMSD of 3.0 nm during MD simulation. We found that EGFR/ERK overexpression leads to induce hematopoietic cell proliferation and impaired differentiation, which enhances the myeloid repopulating activity and the accumulation of immature myeloblast cells. BPA also caused a corresponding increase in expression of hematopoietic transcription factor c-MYB and RUNX-1 leading to polychromasia, poikilocytosis, acanthocytes and anisocytosis and promoted myeloblastosis by inhibiting GATA-1 expression. These morphological changes often resulted in the prior condition of acute myeloid leukemia (AML). Comprehensively, our data suggest that BPA can trigger the malignancy of AML cells by alteration of respective hematopoietic transcription factors via EGFR/ERK signaling in the zebrafish model.

Co-delivery of Diverse Therapeutic Compounds Using PEG-PLGA Nanoparticle Cargo against Drug-Resistant Bacteria: An Improved Anti-biofilm Strategy.

Controlling biofilms of bacteria is a challenging aspect because of their drug-resistance potentials against a range of antibiotics, demanding the development of active anti-biofilm agents. Rutin (R), a natural antioxidant, and benzamide (B), a synthetic antibacterial agent, have several pharmacological and antibacterial abilities. Herein, we developed PEG-PLGA NPs that synergistically carried rutin and benzamide as drug candidates, while displaying therapeutic and anti-biofilm  functions. These drug delivery NPs were synthesized by the oil-in-water emulsion (O/W) solvent evaporation technique. The obtained NPs were characterized by UV-vis, FT-IR, SEM, TEM, and DLS measurements. Confocal laser scanning microscopy was employed to evaluate the anti-biofilm capabilities against Staphylococcus aureus and Pseudomonas aeruginosa and further quantified the levels of residual biofilm constituents such as protein and exopolysaccharide (EPS). Drug release experiments showed the controlled release of rutin-benzamide (RB) for several days. Antibacterial analyses showed that the minimum inhibitory concentration (MIC) of NPs was at least two times lower than that of the free drugs. RB-PEG-PLGA NPs revealed that they targeted biofilm-forming bacteria through the disruption of the membrane and biofilm surface and were observed to be nontoxic when tested using human erythrocytes and human cell lines. In vivo evaluations in zebrafish showed that the NPs did not alter the antioxidant functions and histological features of tissues. On the basis of results obtained, it is substantiated that the rutin-benzamide-loaded nanocarrier offers potential anti-biofilm therapy due to its high anti-biofilm activity and biocompatibility.

Synergistic effect of chemo-photothermal for breast cancer therapy using folic acid (FA) modified zinc oxide nanosheet.

Modern therapies for malignant breast cancer in clinics are not efficacious and often result in deprived patient compliance owing to squat therapeutic effectiveness and strong systemic side effects. In order to overcome this, we combined chemo-photothermal targeted therapy of breast cancer within one novel multifunctional drug delivery system. Folic Acid-functionalized polyethylene glycol coated Zinc Oxide nanosheet (FA-PEG-ZnO NS), was successfully synthesized, characterized and introduced to the drug delivery field for the first time. A doxorubicin (DOX)-loaded FA-PEG-ZnO NS based system (DOX-FA-PEG-ZnO NS) showed stimulative effect of heat, pH responsive and sustained drug release properties. Cytotoxicity experiments confirmed that combined therapy mediated the maximum rate of death in breast cancer cells compared to that of single chemotherapy or photothermal therapy. In vivo toxicity evaluation showed that the DOX-FA-PEG-ZnO NS contains minimum systemic toxicity in the mice model system. The findings of the present study provided an ideal drug delivery system for breast cancer therapy due to the advanced chemo-photothermal synergistic targeted therapy and good drug release properties of DOX-FA-PEG-ZnO NS, which could effectively avoid frequent and invasive dosing and improve patient compliance. Thus, functionalized-ZnO NS could be used as a novel nanomaterial for selective chemo-photothermal therapy.

Effects of the inhibition of cytosolic phospholipase A(2)α in non-small cell lung cancer cells.

PURPOSE: The aim of this study was to investigate the expression of cPLA(2)α in non-small lung cancer cell lines and tissues, and we sought to determine the in vitro effects of the pyrrolidine-2 inhibitor on cPLA(2)α sensitivity in three different non-small lung cancer cell lines. METHODS: The expression of cPLA(2)α was determined in lung cancer cells by Western blot. Cytotoxicity, cell growth and inhibition of cPLA(2)α activity were determined in relation to the concentration of pyrrolidine-2. Finally, this study investigated immunohistochemical expressions of cPLA(2)α in 23 species of human non-small lung cancer and 5 species of human normal lung to assess their clinicopathological relevance. RESULTS: cPLA(2)α is expressed in A549 and H460, however, no expression in H661 cells. Pyrrolidine-2 demonstrated a dose-dependent inhibitory effect on cell growth and its significantly inhibited BrdU incorporation of human non-small lung cancer cells. Inhibition with pyrrolidine-2 results in reduction in cPLA(2)α activity in A549 and H460 lung cancer cells by 50% when present at IC(50) concentration in arachidonoyl thio-PC assay. Immunohistochemistry of human lung tissue revealed that cPLA(2)α is increased in lung cancer tissues. CONCLUSIONS: Pyrrolidine-2 is a more potent and specific cPLA(2)α inhibitor than MAFP and AACOCF3 and represents an excellent pharmacological tool to investigate the biosynthesis and the biological roles of cancer. The present study suggests that pyrrolidine-2 could be a potential therapeutic agent for cancer therapy.

Curtailing overexpression of E2F3 in breast cancer using siRNA (E2F3)-based gene silencing.

BACKGROUND AND AIMS: The E2F3 transcription factor claims its role in controlling cell cycle progression. As reported earlier, nuclear E2F3 overexpression leads to development of bladder and prostate cancer in humans. Accordingly, the present investigation has been designed to assess to what extent E2F3 would be overexpressed in breast cancer. The aim of this study was to emphasize that the levels of E2F3 are increased in breast cancer and highlights the efficacy of siRNA targeted to E2F3. METHODS: To investigate the expression level of E2F3 and the progression of breast tumors, quantitative real-time PCR analysis was carried out. Western blotting analysis was performed to measure its counterparts, namely, E2F3a and E2F3b. RESULTS: In the novel axis of E2F3, a large set of 11 breast cancer cell lines were identified to have the property of overexpression. Furthermore, the small interfering RNA (siRNA) developed against E2F3 significantly blocked the expression of the E2F3 in the selected breast cancer cell lines. Thus, the present findings authenticate the efficiency of siRNA (E2F3) to fight against breast cancer; hence, the siRNA mediated E2F3 gene silencing knockdown the E2F3. CONCLUSIONS: This in vitro study demonstrates that E2F3 is a newly identified diagnostic and potential therapeutic target in breast cancer. Outcomes of this study affirm that siRNA for E2F3 facilitates the silencing of E2F3 overexpression and fights against breast cancer. Therefore, it plays a vital role as an alternative for diagnosis and clinical outcome for the treatment of breast cancer.

γ-Sitosterol from Acacia nilotica L. induces G2/M cell cycle arrest and apoptosis through c-Myc suppression in MCF-7 and A549 cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Acacia nilotica is widely distributed in Asia. In India, it occupies an important place in the indigenous system of medicine against anti-inflammatory, antioxidant, cancers, and/or tumors. AIM OF THE STUDY: The purpose of this study is to investigate the inhibitory effect of Acacia nilotica leaves extract and γ-Sitosterol on cell proliferation, the apoptotic effect and cell cycle arrest in breast and lung cancer cells. MATERIALS AND METHODS: GC-MS and HPLC were used to determine the chemical constituents of this extract and γ-Sitosterol respectively. Human MCF-7 and A549 cell lines were treated with Acacia nilotica extract and γ-Sitosterol. Cell viability was determined by MTT assay. Cell proliferation was determined by BrdU incorporation assay. Apoptosis was detected by cell morphologic observation through AO/EtBr staining, cell cycle analysis, and immunoblot analysis on the expression of protein associated with cell cycle arrest. RESULTS: Experimental results of bioactive compound analysis indicate that γ-Sitosterol, bioactive ingredients of Acacia nilotica extract. The IC(50) value of extract on MCF-7 and A549 cancer cells was 493.3±15.2 and 696.6±11.5 µg/ml, respectively. Acacia nilotica extract and γ-Sitosterol were inhibited the cell proliferation by 54.34±1.8 and 42.18±3.9% for MCF-7 and 58.26±1.5 and 44.36±3.05% for A549 cells. The percentage of apoptotic cells observed in the MCF-7 and A549 cell lines were increased to 42.46 and 36.8% of extract; 46.68 and 43.24% for γ-Sitosterol respectively. Flow cytometric analysis results demonstrate that cells were arrested at the G2/M phase and decrease the c-Myc expression. CONCLUSIONS: This study demonstrates in vitro results, which support the ethnomedical use of γ-Sitosterol against cancer. Experimental results of this study suggest that γ-Sitosterol exerts potential anticancer activity through the growth inhibition, cell cycle arrest and the apoptosis on cancer cells.

Immunohistochemical expression of cytosolic phospholipase A2α in non-small cell lung carcinoma.

PURPOSE: Cytosolic phospholipase A2α is the main target enzyme for the non steroidal anti-inflammatory drugs that have been shown to suppress carcinogenesis in both experimental model and epidemiologic studies. METHODS: We examined cPLA2α expression in normal, premalignant bronchial epithelial cells and nonsmall cell lung carcinoma (NSCLC) samples using an immunohistochemical staining technique. Included in the current study were 76 NSCLC samples and 52 bronchial biopsy samples obtained. RESULTS: In the normal bronchial epithelium, cPLA2α expression was found to be completely negative whereas positive cPLA2α expression was limited to a few macrophages, inflammatory cells. There were relatively more cPLA2α positive tumors, as defined by positive staining in >10% of tumor cells 24 of 76 tumors (32%). When tumor types were considered, there were more cPLA2α positive adenocarcinomas compared with squamous cell carcinomas (17 of 36 adenocarcinomas (47%) vs. 6 of 34 squamous cell carcinomas (18%); P=0.02). Although smokers tended to have more cPLA2α positive tumors than nonsmokers (23 of 64 tumors in the smokers (36%) vs. 1 of 12 tumors in the nonsmokers (8%); P=0.06). CONCLUSION: The results of the current study suggest that cPLA2α expression may not be a useful intermediate biomarker in bronchial chemoprevention trials. Nevertheless, considering the patterns of cPLA2α expression in tumor cells, cPLA2α expression status may be a useful parameter when designing treatment strategies for a subset of NSCLC patients.