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Our previous study indicated that nanocomposite packaging material (Nano-PM) containing nano-Ag, nano-TiO 2 , nano-SiO 2 and nanoattapulgite alleviated postharvest senescence of Flammulina velutipes by regulating respiration and energy metabolism. In this study, extracellular ATP (eATP) and programmed cell death (PCD) were employed as critical factors to further investigate the senescence mechanism of postharvest F. velutipes. Results demonstrated that Nano-PM delayed apyrase activity decrease and stimulated critical oxidative phosphorylation-related gene expression to inhibit eATP content increase, which is a crucial signaling molecule related to delaying senescence. The regulation of eATP resulted in alleviating PCD including chromosomal concentration, DNA fragmentation, Ca 2+ influx, high caspase-1 activity and cytochrome c content and leading to high cell viability. Overall, Nano-PM alleviated PCD and postharvest senescence of F. velutipes by regulating extracellular ATP., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Background: Codeine, a 3-methylmorphine, and other related opioids have been implicated in androgen suppression, although the associated mechanisms remain unclear., Aim: Therefore, the objective of the current study was to elucidate the in vivo molecular mechanisms underlying codeine-induced androgen suppression., Methods: This study made use of twenty-one healthy male rabbits, distributed into three groups randomly, control and codeine-treated groups. The control had 1ml of normal saline daily p.o. The codeine-treated groups received either 4mg/kg b.w of codeine or 10mg/kg b.w of codeine p.o. for six weeks. Reproductive hormonal profile, testicular weight, testicular enzymes, oxidative and inflammatory parameters, testicular DNA fragmentation, histological examination and apoptosis marker were evaluated to examine the effects of codeine use., Key Findings: Oral administration of codeine resulted in testicular atrophy and alterations in testicular histomorphology, elevated testicular enzymes, and suppression of circulatory and intra-testicular testosterone. These changes were associated with a marked rise in oxidative markers and decline in the activities of testicular enzymatic antioxidants, as well as oxidative DNA damage, inflammatory response, testicular DNA fragmentation, and caspase-dependent apoptosis (p<0.05)., Significance: In conclusion, chronic codeine use resulted in testicular degeneration and testosterone suppression, which is attributable to TNF-α/nitric oxide-/oxidative stress-mediated caspase-dependent apoptotic testicular cell death and loss of testicular function., Competing Interests: The authors have declared that no competing interests exist.

MoS 2 /α-NiMoO 4 ultra-thin nanoneedle composite was synthesized by microwave hydrothermal process in one step. The nanocomposite revealed the complete destruction of multidrug resistant Staphylococcus aureus (S. aureus) within 150 min under visible light irradiation. According to electron spin resonance measurement and radical trapping experiment, it has been established that O 2 ¯ acts as a major active species for bacterial inactivation in visible light. The bacterial inactivation was further proved by membrane deformities in bacterial cell membrane, DNA fragmentation, and protein destruction. TEM- elemental mapping confirms the inactivation of S. aureus by reactive oxygen species (ROS) but not the toxicity of photocatalyst. Transient photocurrent responses, electrochemical impedance spectroscopy, and cyclic voltammetry measurements reveal the efficient separation of electron-hole pairs in the composite photocatalyst. The composite photocatalyst shows greater ROS production, higher degree of DNA fragmentation and protein degradation, detrimental effects on the morphology of the bacterial cell wall, outstanding transient photocurrent responses, reduction of interfacial charge transfer resistance, superb oxidation/reduction potential, strong visible light absorption, and adequate separation of photogenerated electron-hole pairs as compared to host photocatalyst. The photocatalytic inactivation mechanism was explained. So, this promising composite photocatalyst can be applied for inactivation of multidrug resistant bacteria in biological waste water., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Tebuconazole is an effective systemic fungicide that belongs to the triazoles family. It has been widely used in both agricultural and medical sectors for the control of fungal diseases. Although TEB poses serious threats to mammals health, studies regarding its cardiotoxicity are very limited. Thus, we aimed to evaluate the effects of TEB on some biochemical parameters, the induction of apoptosis and DNA damage in the heart tissue. Male Wistar rats were treated with TEB at varied oral doses for 28 consecutive days. This study demonstrates that TEB decreased cardiac acetylcholinesterase, increased serum marker enzymes such as creatinine phosphokinase (CPK) and lactate dehydrogenase (LDH), and altered the lipid profile by increasing serum levels of total cholesterol (T-CHOL), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C) and reduced high-density lipoprotein cholesterol (HDL-C) levels. Furthermore, TEB increased levels of p53 and Bax/Bcl2 ratio, released the cytochrome c into the cytosol and activated caspase-9 and caspase-3. Besides, our results showed that TEB induced genotoxic effects. TEB induced DNA fragmentation and increased the frequency of micronucleated bone marrow cells. Moreover, TEB treatment developed fibrosis in the myocardium. Our results suggest that TEB exposure may affect myocardial cells normal functioning and triggers apoptosis., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Loss of dopaminergic neurons following Parkinson's disease (PD) diminishes quality of life in patients. The present study was carried out to investigate the protective effects of simultaneous inhibition of dipeptidyl peptidase-4 (DPP-4) and P2X7 purinoceptors in a PD model and explore possible mechanisms. The 6-hydroxydopamine (6-OHDA) was used as a tool to establish PD model in male Wister rats. The expressions of SIRT1, SIRT3, mTOR, PGC-1α, PTEN, P53 and DNA fragmentation were evaluated by ELISA assay. Behavioral impairments were determined using apomorphine-induced rotational and narrow beam tests. Dopamine synthesis and TH-positive neurons were detected by tyrosine hydroxylase (TH) immunohistochemistry. Neuronal density was determined by Nissl staining. OHDA-lesioned rats exhibited behavioral impairments that reversed by BBG, lin and lin + BBG. We found significant reduced levels of SIRT1, SIRT3, PGC-1α and mTOR in both mid brain and striatum from OHDA-lesioned rats that reversed by BBG, lin and lin + BBG. Likewise, significant increased levels of PTEN and P53 were found in both mid brain and striatum from OHDA-lesioned rats that was reversed by BBG, lin and lin + BBG. TH-positive neurons and neuronal density were markedly reduced OHDA-lesioned rats that reversed by BBG, lin and lin + BBG. Collectively, our results showed protective effects of simultaneous inhibition of DPP-4 and P2X7 purinoceptors in a rat model of PD can be linked to targeting SIRT1/SIRT3, PTEN-mTOR pathways. Moreover, our findings demonstrated that simultaneous inhibition of DPP-4 and P2X7 purinoceptors might have stronger effect on mitochondrial biogenesis compared to only one.

The aim of this article was to evaluate the effects of different concentrations of carnosine added during human semen processing. Semen samples from 34 patients were submitted to processing by discontinuous density gradient centrifugation without (control) or with different concentrations of carnosine supplementation as follows: (a) 20 mM of carnosine supplementation on the layers of Percoll; and (b) 50 mM carnosine supplementation. Sperm samples were then washed with human tubal fluid medium and evaluated according to sperm kinetics and functional assessment. For statistical analysis, data were evaluated by a general linear model or a Friedman test, whenever appropriate. The 50 mM carnosine supplementation led to improved sperm mitochondrial activity when compared to untreated samples. Motility variables, such as percentage of motile and progressively motile spermatozoa, average path velocity, straight line velocity, curvilinear velocity and linearity, showed an improvement after semen processing irrespective of carnosine supplementation. Both concentrations of carnosine increased the beat-cross frequency (BCF) when compared to samples before processing. We conclude that carnosine supplementation in semen samples benefits sperm mitochondrial activity and BCF., (© 2019 Blackwell Verlag GmbH.)

Cryopreservation is a useful approach to preserve male fertility for assisted reproduction technique and other evaluations. In spite of extensive development in the cryopreservation field, there are biological and biochemical points including DNA fragmentation and antioxidant which should be illuminated to preserve fertility in safe form. Several molecular markers such as coding and noncoding RNAs are transferred from spermatozoa into oocyte via fertilization. These biomarkers affect fertility potential during the cryopreservation. Despite its impact, sperm cryopreservation has some destructive effect including loss of sperm motility and viability, acrosomal damage, mitochondrial membrane depolarization, plasma membrane permeability alteration even nuclear, and DNA damage. There is a growing concern about the impact of sperm cryopreservation on biological factors which can interrupt successful fertility procedures such as in vitro fertilization. Additionally, cryo-damage can decrease embryonic development extent. Promoting cryopreservation method via investigating the wide range of non-invasive factors may be increasingly important to have access to safe approach of freezing. Therefore, the aim of this study is the assessment of biological factors which can penetrate the fertility potential during the freezing procedure, explicate innovative methods in fertility preservation.

Sperm cryopreservation is a routine method in andrology and IVF laboratory. However, the sperm quality and its fertilizing capacity have been decreased during this process. The purpose of this experiment was to determine the role of myoinositol as a supplement in amelioration of total and progressive sperm motility, DNA fragmentation, total antioxidant capacity (TAC), reactive oxygen species (ROS), and lipid peroxidation after the freezing-thawing process on patients with oligoasthenoteratozoospermia (OAT) syndrome. Semen samples obtained from 40 patients were divided into two aliquots and freezed with simple and 2 mg/mL myoinositol (MYO) supplemented freezing media. All samples were thawed and assessed after one month. Semen parameters were analyzed in terms of the motility by CASA, the level of total ROS by fluorimetry, TAC and MDA by colorimetric assay and finally DNA fragmentation by TUNEL assay. Our results clearly showed that MYO could improve total (37.46 vs. 12.91, p < 0.001) and progressive motility (21.92 vs. 6.49, p < 0.001) in experimental group compared to control group. A higher TAC level was observed in the MYO treated group in comparison to control group (1.11 vs. 0.91, p = 0.05). While MYO supplementation could not be effective on ROS level, it reduced DNA fragmentation of sperm after freeze-thaw process (p = 0.01). Therefore, MYO could be a good supplement for sperm freezing to reduce the detrimental effects of freezing process especially on DNA integrity, which is an important factor in the success of ART, in OAT suffered patients.

The present study was conducted to demonstrate cytotoxicity, apoptosis and hepatic damage induced by gemcitabine in laboratory mice. Animals were treated with a single dose of gemcitabine (415 mg/kg body wt), equivalent to a human therapeutic dose, and sacrificed after 1, 2 and 3 weeks. A significant decrease in mean body weight and absolute liver weight was registered. The levels of alkaline phosphatase (ALP), aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were increased as a result of this induced stress. Various structural changes were observed in the liver tissue of treated mice, as evident in the histological sections. Specifically, gemcitabine exposure was able to induce apoptosis in liver cells, and the incidence of TUNEL positive liver cells was increased compared to the control group. DNA fragmentation appeared on agarose gel and flow cytometry analysis confirmed the induction of apoptosis. These findings in gemcitabine-treated animal tissues suggest that inhibition or disruption of cells' DNA synthesis may be the mechanism by which this drug induces toxicity in the animal body.

Background: Bisphenol A is widely used in the manufacture of polycarbonate plastics and has caused increasing concern over its potential adverse impacts on spermatogenesis. However, the effect of bisphenol A on spermiogenesis is yet to be explored., Objectives: To evaluate whether bisphenol A has adverse effects on DNA integrity and protamination of spermatogenic cell., Materials and Methods: Newborn male mice were subcutaneously injected with bisphenol A (0.1, 5 mg/kg body weight, n = 15) or coin oil (control group, n = 20) daily from post-natal day 1 until 35. At post-natal day 70, epididymis caudal spermatozoa and testes were collected. Sperm count, sperm motility, and sperm morphology were analyzed. The sperm chromatin structure assay was performed to examine the sperm DNA fragmentation. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) method was used to assess apoptosis of spermatogenic cells. The ultrastructural features of testicular sections were examined under a transmission electron microscope. Western blot and RT-PCR were used to detect the expression levels of transition protein (Tnp) 1 and Tnp2, protamine (Prm) 1 and Prm2 protein, and mRNA in mice testes., Results: Bisphenol A significantly reduced sperm counts, impaired sperm motility, and increased the percentage of malformed spermatozoa. Poor sperm chromatin integrity and increased TUNEL-positive spermatogenic cells were also observed in mice exposed to bisphenol A. Ultrastructural analysis of testes showed that bisphenol A exposure caused incomplete chromatin condensation, retention of residual cytoplasm, and abnormal acrosome formation. In addition, the relative expression levels of Tnp2 and Prm2 in mice testes decreased significantly in bisphenol A groups., Discussion and Conclusion: Our findings identified that neonatal bisphenol A exposure may negatively contribute to the sperm quality in adult mice. Mechanistically, we showed that bisphenol A reduced sperm chromatin integrity along with increased DNA damage, which may be due to poor protamination of spermatozoa., (© 2019 American Society of Andrology and European Academy of Andrology.)

Hesperetin is a natural flavonoid with robust antioxidant properties. Our previous study reported that hesperetin can prevent cataract formation. However, an important consideration regarding hesperetin consumption is the limited bioavailability due to its poor solubility. The present study investigated the anti‑cataract effects of α‑glucosyl hesperidin in vivo and in vitro using a selenite‑induced cataract model. SD rats (age, 13 days) were orally administered PBS (0.2 ml) or α‑glucosyl hesperidin (200 mg/kg) on days 0, 1 and 2. Sodium selenite was subcutaneously administered to the rats 4 h after the first oral administration on day 0. Antioxidant levels in the lens and blood were measured on day 6. In vitro, human lens epithelial cells were treated with sodium selenite (10 µM) and/or hesperetin (50 or 100 mM) for 24 h and analyzed for apoptosis markers using sub‑G1 population and Annexin V‑FITC/propidium iodide staining and DNA ladder formation. α‑glucosyl hesperidin treatment significantly reduced the severity of selenite‑induced cataract. The level of antioxidants was significantly reduced in the selenite‑treated rats compared with in the controls; however, they were normalized with α‑glucosyl hesperidin treatment. In vitro, hesperetin could significantly reduce the number of cells undergoing apoptosis induced by sodium selenite in human lens epithelial cell lines. Overall, oral consumption of α‑glucosyl hesperidin could delay the onset of selenite‑induced cataract, at least in part by modulating the selenite‑induced cell death in lens epithelial cells.

The aim of this study was to evaluate the potentials of rutin on 2,5-hexanedione-induced toxicities. Two successive phases were involved using in silico and in vivo approaches. The in silico was adopted for potential oral toxicity and docking. The in vivo was carried-out in two stages for two weeks; the ameliorative (stage 1, first week), preventive, and curative studies (stage 2, extended to second week). In stage 1, rats were divided into four groups of seven each (distilled water, 3% (v/v) 2,5-hexanedione, 10 mg/kg rutin, and co-administration). In stage 2, the experimental groups were given either rutin or 2,5-hexanedione and treated in reverse order. Lipid peroxidation, protein carbonyl, and DNA fragmentation in tissues and bone marrow cells micronucleus were determined. The predicted Median lethal dose (LD 50 ) of >5000 mg/kg and toxicity class of five (5) indicates the safety of rutin when orally administered. 2,5-Hexanedione comfortably docked in to the active sites of SOD (-22.857Kcal/mol; KI  = 0.9621 µM), GPx (-11.2032Kcal/mol; KI  = 0.9813 µM), and CAT (-16.446Kcal/mol; KI  = 0.9726 µM) with strong hydrogen bond and hydrophobic interactions. However, only strong hydrophobic interaction was observed in the case of DNA (-3.3296Kcal/mol; KI  = 0.9944). In vivo findings revealed deleterious effects of 2,5-hexanedione through induction of oxidative and chromosomal/DNA damage characterized by higher level of malondialdehyde, micronuclei formations, and DNA fragmentation. These have invariably, validates the findings from in silico experiments. Furthermore, rutin was able to ameliorate, protect, and reverse these effects, and was relatively non-toxic corroborating toxicity predictions. Rutin exhibited counteractive effects on 2,5-hexanedione-induced oxidative, chromosomal, and DNA damage.

(-)-Epicatechin is a phenolic compound with antioxidant activity that is present in natural food and drinks, such as cocoa and red wine. Evidence suggests that (-)-epicatechin exhibits anticancer activity; however, its mechanism of action is poorly understood. Here, we investigated the anticancer effects of (-)-epicatechin and its mechanism of action in breast cancer cells. We assessed the anticancer activity by cell proliferation assays, apoptosis by DNA fragmentation and flow cytometry. The expression of proteins associated with apoptosis was analyzed by the human apoptosis array. MitoSOX TM Red and biomarkers of oxidative damage were used to measure the effect of (-)-epicatechin on mitochondrial reactive oxygen species (ROS) and cellular damage, respectively. (-)-Epicatechin treatment caused a decreasing in the viability of MDA-MB-231 and MCF-7 cells. This cell death was associated with DNA fragmentation and an apoptotic proteomic profile. Further, (-)-epicatechin in MDA-MB-231 cells upregulated death receptor (DR4/DR5), increased the ROS production, and modulated pro-apoptotic proteins. In MCF-7 cells, (-)-epicatechin did not involve death receptor; however, an increase in ROS and the upregulation of pro-apoptotic proteins (Bad and Bax) were observed. These changes were associated with the apoptosis activation through the intrinsic pathway. In conclusion, this study shows that (-)-epicatechin has anticancer activity in breast cancer cells and provides novel insight into the molecular mechanism of (-)-epicatechin to induce apoptosis., Competing Interests: The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.

trans-Fatty acids (TFAs) are unsaturated fatty acids that contain one or more carbon-carbon double bonds in trans configuration. Epidemiological evidence has linked TFA consumption with various disorders, including cardiovascular diseases. However, the underlying pathological mechanisms are largely unknown. Here, we show a novel toxic mechanism of TFAs triggered by DNA damage. We found that elaidic acid (EA) and linoelaidic acid, major TFAs produced during industrial food manufacturing (so-called as industrial TFAs), but not their corresponding cis isomers, facilitated apoptosis induced by doxorubicin. Consistently, EA enhanced UV-induced embryonic lethality in C. elegans worms. The pro-apoptotic action of EA was blocked by knocking down Sab, a c-Jun N-terminal kinase (JNK)-interacting protein localizing at mitochondrial outer membrane, which mediates mutual amplification of mitochondrial reactive oxygen species (ROS) generation and JNK activation. EA enhanced doxorubicin-induced mitochondrial ROS generation and JNK activation, both of which were suppressed by Sab knockdown and pharmacological inhibition of either mitochondrial ROS generation, JNK, or Src-homology 2 domain-containing protein tyrosine phosphatase 1 (SHP1) as a Sab-associated protein. These results demonstrate that in response to DNA damage, TFAs drive the mitochondrial JNK-Sab-ROS positive feedback loop and ultimately apoptosis, which may provide insight into the common pathogenetic mechanisms of diverse TFA-related disorders.

The nature of the atmospheric particulate matters (PMs) varies depending on their sizes and their origin from different activities in the background environment. These PMs are associated with potentially hazardous elements (PHEs) such as organic compounds (e.g. Polyaromatic Hydrocarbons) that can be harmful to health. The main objective of this work is the identification and investigation of the toxicological aspects of PHEs in PMs during pre-monsoon and post-monsoon season in an urban area of Northeast region (NER) of India. In the course of the study, the 24 -hs average concentrations of PMs were detected to be more than two-times higher than the Indian standard limit (NAAQ, category) which indicates poor air quality in both the seasons around the sampling sites. This study demonstrates that the concentrations of PM-bound PAHs are mutagenic and that the Excess Cancer Risks exceed the USEPA standard limits. PMs cause cytotoxicity and can also induce genotoxicity to human health analyzed by cell culture and gel electrophoresis. This study helps to promote research to evaluate the PMs bound PHEs toxicity in diverse human cell lines and also their relationship with climatic factors as well as quantitative source apportionment for mitigation purposes., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Ischemic heart disease (IHD), a severe condition of myocardium facing impediment in the supply of basic needs for cellular metabolism is caused by atherosclerosis. Though statin drugs could control the use of surgery on IHD patients, the complete rehabilitation or prophylaxis can be achieved through herbal-based medicines viz. either in the form of crude extract or pure phytocompounds. In the present study, pretreatment with leaf extract of Nelumbo nucifera Gaertn. was investigated for cardioprotective activity-in vitro by mitigating H 2 O 2 -induced oxidative stress. Analysis such as estimation of antioxidants, lipid peroxidation, and DNA fragmentation assay revealed significant protective effect of plant extract on cardiomyocytes. Reactive oxygen species detection was done by using 2',7'-dichlorofluorescein diacetate, apoptosis detection with Acridine Orange/Ethidium Bromide and nuclear damage detection by diamidino-2-phenylindole which confirmed the protective effect of N. nucifera extract. In addition, gene expression studies of apoptotic regulatory genes (Bcl2 and Cas-9) resulted in significant protection of nucifera extract pretreated and maintained cells. To conclude, in vitro cardioprotective activity of N. nucifera against H 2 O 2 induced oxidative stress was achieved at the concentration of 50 µg/ml. Therefore, major phytocompounds present in extract could be beneficial in managing cardiac complications in the future.

Malignant melanoma is a high aggressive malignancy in humans and causes 60-80% of deaths from skin cancer. Defect in an intrinsic pathway of apoptosis via overexpression of Mcl-1 is responsible for malignant melanoma development and progression, and also for resistance to chemotherapeutic agents. MIM1 is a specific low molecular Mcl-1 protein inhibitor that is able to induce Mcl-1-dependent cancer cells death. Here, we examined the effect of MIM1 as well as MIM1 and dacarbazine (DTIC) mixture on cell viability, apoptosis, and cell cycle progression in COLO829 melanoma cells. Cell viability was performed by the WST-1 assay. Analysis of apoptosis as well as cell cycle progression was determined by fluorescence image cytometer NucleoCounter NC-3000. The obtained results demonstrated that the MIM1 exhibited high cytotoxicity against melanotic melanoma cells and induced mitochondrial membrane breakdown, GSH depletion, and DNA fragmentation. Additionally, MIM1 enhanced the proapoptotic effect of DTIC toward melanoma cells; furthermore, a mixture of these drugs caused cell cycle arrest at G2/M phase in COLO829 cells. Taken together, these data provide, for the first time, evidence that a low molecular weight Mcl-1 inhibitor-MIM1 may be a promising agent with antitumor and proapoptotic properties toward melanoma cells., (© 2019 Société Française de Pharmacologie et de Thérapeutique.)

Fasciolosis is a neglected tropical disease caused by the liver fluke Fasciola gigantica. The absence of successful vaccine and emerging resistance in flukes against the drug of choice, triclabendazole, has necessitated the search for alternatives including phyto-therapeutic approaches. Curcumin and thymoquinone, the active ingredients of Curcuma longa and Nigella sativa plants respectively, were first screened for their binding affinity with Glutathione-S-transferase (GST) molecule through in silico molecular docking followed by in vitro treatment of worms with varying concentrations of the test compounds. The in silico molecular docking of curcumin and thymoquinone with sigma GST revealed strong hydrogen bonding as well as hydrophobic interactions with high fitness scores but showing inter-specific differences. The in vitro treatment of F. gigantica worms with both curcumin and thymoquinone resulted in a significant increase in the generation of reactive oxygen species (ROS) whereas the level of reduced glutathione, a primary redox regulator, was found to be significantly decreased (p < 0.05). The two compounds not only inhibited the GST activity, which is an important detoxification enzyme and also a key drug/vaccine target for the control of fasciolosis but also significantly inhibited the activity of antioxidant enzymes glutathione peroxidase and glutathione reductase that are vital in maintenance of redox homeostasis. The immunohistochemistry performed using anti sigma GST polyclonal antibodies revealed that both the compounds used in the present study significantly reduced immunofluorescence in the vitellaria, developing eggs present in the ovary and the intestinal caecae indicating inhibition of GST enzyme in these regions of the worms. Further, following treatment with curcumin and thymoquinone, chromatin condensation and DNA fragmentation was also observed in F. gigantica worms. In conclusion, both curcumin and thymoquinone generated oxidative stress in the worms by production of ROS and significantly inhibiting their antioxidant and detoxification ability. The oxidative stress along with induction of apoptotic like events would compromise the survival ability of worms within the host. However, further studies are required to establish their anthelmintic potential alone and in combination with the commonly used anthelmintic drugs under in vivo conditions., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Outbred male rats were repeatedly injected intraperitoneally two-level sub-lethal doses of lead acetate and/or cadmium chloride solutions 3 times a week during 6 weeks. The animals developed explicit, even if moderate, subchronic intoxication characterized by a large number of indices, both common to both metals (including increased DNA fragmentation coefficient) and lead-specific. Special attention was paid to hemodynamic and electrocardiographic effects. The combined action of lead and cadmium was modeled with the help of the Response Surface Methodology to obtain additional support for the previously substantiated postulates of combined toxicity's typological ambiguity. This is dependent on which particular effect comes under consideration, on its level, and on the acting dose ratio. For one and the same toxic combination, the type of combined toxic action can vary from synergistic to contra-directional. In particular, the actions of lead and cadmium on blood pressure were found to be opposite in direction. Furthermore, it is shown once again that the systemic toxic effects of a metal combination, its in vivo genotoxicity included, can be more or less attenuated by background administration of a theoretically justified composition of biologically active agents., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2019. Published by Elsevier Ltd.)

Three asymmetric tridentate acyl hydrazone Schiff base ligands namely L1, L2 and L3 were prepared via condensation of 4-methoxybenzohydrazide with picolinaldehyde, 1-(pyridin-2-yl)ethanone and phenyl(pyridin-2-yl)methanone respectively. Three bio-relevant mononuclear zinc(ii) complexes [Zn(L1)Cl 2 ]·2H 2 O (1), [Zn(L2)Cl 2 ] (2) and [Zn(L3)Cl 2 ] (3) were synthesized by treatment of zinc(ii) chloride with the corresponding Schiff base ligands and characterised by the usual physicochemical techniques. The solid state structures of complexes 1 and 3 were evaluated by single crystal X-ray analysis. All complexes were able to hydrolyse the P-O bond of the phosphate monoester in 90% (v/v) DMSO-water medium using 4-nitrophenylphosphate (4-NPP) as the model substrate and the trend in their activity is 1≈2 > 3. On considering the highly efficient hydrolysis properties, complexes 1-3 were tested as potential therapeutic agents for cancer using HCT116 (human colorectal carcinoma), HepG2 (human hepatocellular carcinoma) and A549 (human non-small lung carcinoma) cells. Complex 2 showed the highest IC 50 values for anti-cancer activity towards all three cell lines among the three complexes and complex 3 showed the least activity as observed in the phosphatase activity study. The internucleosomal degradation of DNA during apoptosis can be detected by cell death detection ELISA. DNA fragmentation that leads to cell death was examined and when induced by complex 2 in HepG2 cells a significant level of DNA fragmentation was observed at regular intervals of time.

Purpose: This study aimed to investigate the protective effect of Gallic acid (GA) on the Cyclophosphamide (CP) toxicity induced in the reproductive system., Materials and Methods: After a pilot study for dose responses of Gallic acid ,Forty adult male NMRI mice were divided into 5 groups (n=8): control, sham (NaCl Serum: 0.2mL per day), CP (15 mg kg-1 per week; IP), GA (12.5 mg kg-1 per day ; IP) and GA (12.5 mg kg-1 per day ; IP) +CP(15 mg kg-1 per week; IP). After treatment, the left testis was detached and used for Histological examination and right testis used for Malondialdehyde (MDA) measures. Left caudal epididymis was placed in the Ham's F10 medium and released spermatozoa were used in order to analyze sperm parameters. Sperm DNA fragmentation was assessed by Sperm Chromatin Dispersion (SCD) method., Results: In the CP group, there was a significant increase in the sperm DNA fragmentation (% 57.89 ± 23.91) compared with control group (% 24.52 ± 10.27). That was significantly improved by GA  (12.5 mg kg-1 per day ; IP)  in GA+CP group (% 28.4 ± 8.85) compared to CP group (p< .001).A significant increase was reported about MDA levels in CP group (6.26 ± 2.59) in compared with the control group (4.30 ± 2.05), But GA (3.24 ± 1.33) decreased it in GA+ CP group (p< .01).  The histopathological investigation revealed marked testicular atrophy in CP group, whereas GA diminished these deviations (P< .05)., Conclusion: Gallic acid can modify the reproductive toxicity of cyclophosphamide in NMRI mice and increase the antioxidant capacity of testis tissue.

Plant proteins are emerging as an alternative to conventional treatments against candidiasis. The aim of this study was to better understand the mechanism of action of Mo-CBP 2 against Candida spp, evaluating redox system activity, lipid peroxidation, DNA degradation, cytochrome c release, medium acidification, and membrane interaction. Anti-candida activity of Mo-CBP 2 decreased in the presence of ergosterol, which was not observed with antioxidant agents. C. albicans treated with Mo-CBP 2 also had catalase and peroxidase activities inhibited, while superoxide dismutase was increased. Mo-CBP 2 increased the lipid peroxidation, but it did not alter the ergosterol profile in live cells. External medium acidification was strongly inhibited, and cytochrome c release and DNA degradation were detected. Mo-CBP 2 interacts with cell membrane constituents, changes redox system enzymes in C. albicans and causes lipid peroxidation by ROS overproduction. DNA degradation and cytochrome c release suggest apoptotic or DNAse activity. Lipid peroxidation and H + -ATPases inhibition may induce the process of apoptosis. Finally, Mo-CBP 2 did not have a cytotoxic effect in mammalian Vero cells. This study highlights the biotechnological potential of Mo-CBP 2 as a promising molecule with low toxicity and potent activity. Further studies should be performed to better understand its mode of action and toxicity., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Testicular cancer is one of the most common malignancy in young men, chemotherapy induced damage in cancerous cells as well as healthy tissue, and we decided to investigate recovery effect of zinc (Zn) on chemotherapy-induced complications in rat chromatin integrity and testicular histomorphometry. The male rats (n = 40) were treated with BEP at appropriate dose levels of BEP (0.75, 7.5, and 1.5 mg/kg) for 9 weeks, with or without Zn; testicular histology, sperm DNA methylation, ubiquitination, DNA fragmentation and protamination were further assessed through immunofluorescence. BEP treatment significantly increased ubiquitination, and DNA fragmentation, considerably reducing global DNA methylation and protamination (P < 0.001), resulting in degenerative changes in testicular structure. Zn restored normal DNA methylation, protamination and structure of male gonads, maintained spermatogonial stem cells, and significantly reduced the mean percentage of ubiquitination and sperm DNA fragmentation as compared with BEP group (P < 0.001). We found that supplementation of Zn following chemotherapy can improve chromatin integrity, testicular organization and spermatogenesis., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Importance: Dietary supplements marketed for male fertility commonly contain folic acid and zinc based on limited prior evidence for improving semen quality. However, no large-scale trial has examined the efficacy of this therapy for improving semen quality or live birth., Objective: To determine the effect of daily folic acid and zinc supplementation on semen quality and live birth., Design, Setting, and Participants: The Folic Acid and Zinc Supplementation Trial was a multicenter randomized clinical trial. Couples (n = 2370; men aged ≥18 years and women aged 18-45 years) planning infertility treatment were enrolled at 4 US reproductive endocrinology and infertility care study centers between June 2013 and December 2017. The last 6-month study visit for semen collection occurred during August 2018, with chart abstraction of live birth and pregnancy information completed during April 2019., Interventions: Men were block randomized by study center and planned infertility treatment (in vitro fertilization, other treatment at a study site, and other treatment at an outside clinic) to receive either 5 mg of folic acid and 30 mg of elemental zinc (n = 1185) or placebo (n = 1185) daily for 6 months., Main Outcomes and Measures: The co-primary outcomes were live birth (resulting from pregnancies occurring within 9 months of randomization) and semen quality parameters (sperm concentration, motility, morphology, volume, DNA fragmentation, and total motile sperm count) at 6 months after randomization., Results: Among 2370 men who were randomized (mean age, 33 years), 1773 (75%) attended the final 6-month study visit. Live birth outcomes were available for all couples, and 1629 men (69%) had semen available for analysis at 6 months after randomization. Live birth was not significantly different between treatment groups (404 [34%] in the folic acid and zinc group and 416 [35%] in the placebo group; risk difference, -0.9% [95% CI, -4.7% to 2.8%]). Most of the semen quality parameters (sperm concentration, motility, morphology, volume, and total motile sperm count) were not significantly different between treatment groups at 6 months after randomization. A statistically significant increase in DNA fragmentation was observed with folic acid and zinc supplementation (mean of 29.7% for percentage of DNA fragmentation in the folic acid and zinc group and 27.2% in the placebo group; mean difference, 2.4% [95% CI, 0.5% to 4.4%]). Gastrointestinal symptoms were more common with folic acid and zinc supplementation compared with placebo (abdominal discomfort or pain: 66 [6%] vs 40 [3%], respectively; nausea: 50 [4%] vs 24 [2%]; and vomiting: 32 [3%] vs 17 [1%])., Conclusions and Relevance: Among a general population of couples seeking infertility treatment, the use of folic acid and zinc supplementation by male partners, compared with placebo, did not significantly improve semen quality or couples' live birth rates. These findings do not support the use of folic acid and zinc supplementation by male partners in the treatment of infertility., Trial Registration: ClinicalTrials.gov Identifier: NCT01857310.

Atractylodes macrocephala is known to exhibit multi-arrays of biologic activity in vitro. However, detail of its anti-tumor activity is lacking. In this study, the effects of atractylenolide I (AT-I), a bio-active compound present in Atractylodes macrocephala rhizome was studied in the human colorectal adenocarcinoma cell line HT-29. The results showed that AT-I induced apoptosis of human colon cancer cells through activation of the mitochondria-dependent pathway. The IC 50 of AT-I was 277.6 μM, 95.7 μM and 57.4 μM, after 24, 48 and 72 h of incubation with HT-29, respectively. TUNEL and Annexin V-FITC/PI double stain assays showed HT-29 DNA fragmentation after cell treatment with various AT-I concentrations. Western blotting analysis revealed activation of both initiator and executioner caspases, including caspase 3, caspase 7, and caspase 9, as well as PARP, after HT-29 treatment with AT-I via downregulation of pro-survival Bcl-2, and upregulation of anti-survival Bcl-2 family proteins, including Bax, Bak, Bad, Bim, Bid and Puma. The studies show for the first time that AT-I is an effective drug candidate towards the HT-29 cell.

Gemcitabine (Gem), busulfan (Bu), and melphalan (Mel) are used for hematopoietic stem cell transplantation. To further improve their efficacy, a preclinical study on their synergism with the histone deacetylase inhibitor panobinostat (Pano) and the BCL2 inhibitor venetoclax/ABT199 was performed. Multiple myeloma cell lines MM.1R and MC/CAR were exposed to ∼IC 20 levels of the drugs. Synergistic cytotoxicity was observed in cells exposed to the five-drug combination as indicated by combination indexes <1, supported by ∼86% inhibition of proliferation and ∼84% annexin V positivity in MM.1R and ∼58% inhibition of proliferation and ∼46% annexin V positivity in MC/CAR cells. Activation of the DNA damage response and apoptosis were suggested by a modest increase in the phosphorylation of ATM and its substrates; significant cleavage of PARP1, caspase 3, and heat shock protein 90; DNA fragmentation; mitochondrial membrane depolarization; and reactive oxygen species production. The five-drug combination significantly decreased the levels of PI3K, AKT, mTOR, RAPTOR, P-P70S6K, and eIF2α, with concomitant increases in P-AMPK and its substrate Tuberin/TSC2, suggesting that the mTOR signaling pathway was compromised. Endoplasmic reticulum stress through activation of the unfolded protein response was also observed as suggested by increases in the levels of calnexin, BiP/GRP78, ERO1-Lα, and protein disulfide isomerase, which may relate to venetoclax-mediated inhibition of BCL2 in the endoplasmic reticulum. This is the first report on the effects of a venetoclax-containing regimen on the unfolded protein response. These results provide a rationale to propose a clinical trial on use of Gem + Bu + Mel + Pano + Venetoclax as part of a conditioning regimen for multiple myeloma patients undergoing autologous hematopoietic stem cell transplantation., Competing Interests: Conflict of interest disclosure All authors declare no competing financial interests., (Published by Elsevier Inc.)

Background: It has been reported that paternal folic acid deficiency is correlated with male infertility and increased birth defects in the offspring. However, there are few data concerning the influence of folic acid supplementation on male-factor infertility with MTHFR gene polymorphisms., Objectives: To evaluate whether folic acid supplementation has a beneficial effect on oligozoospermia with MTHFR gene polymorphisms in Chinese infertility population., Materials and Methods: The infertile men suffering oligozoospermia with MTHFR gene polymorphisms were randomly divided into the folic acid treatment groups receiving folic acid 0.8 mg daily for 3 months and the placebo groups receiving placebo for 3 months. Semen parameters, seminal MDA, and DNA fragmentation were measured. Furthermore, spontaneous pregnancy rate and live birth rate were evaluated., Results: Administration of folic acid for 3 months could significantly improve the seminal parameters in patients with MTHFR 677 TT genotype in comparison with that receiving placebo. Moreover, seminal MDA and sperm DNA fragmentation index in patients with MTHFR 677 TT genotype significantly declined at the end of treatment. Spontaneous pregnancy rate and live birth rate tended to be significantly higher in couples in which the men with MTHFR 677 TT genotype receiving folic acid than that receiving placebo. However, folic acid treatment did not exhibit any advantage in MTHFR 677 CT, 1298 AC, 1298 CC, 1793 GA, or combined 677 CT/1298 AC genotype., Discussion: The anti-oxidation function of folic acid is one of possible mechanisms invovled in improving seminal parameters and pregnancy outcome., Conclusions: Folic acid supplementation has a beneficial effect on oligozoospermia with MTHFR 677 TT genotype in term of seminal parameters, seminal MDA, sperm DNA fragmentation, and pregnancy outcome., (© 2019 American Society of Andrology and European Academy of Andrology.)

Degeneration of apical spikelets and reduced panicle fertility are common reasons for low seed-setting rate in rice (Oryza sativa). However, little is known about the underlying molecular mechanisms. Here, we report a novel degenerated panicle and partial sterility 1 (dps1) mutant that showed panicle apical degeneration and reduced fertility in middle spikelets. dps1 plants were characterized by small whitish anthers with altered cuticle morphology and absence of pollen grains. Amounts of cuticular wax and cutin were significantly reduced in dps1 anthers. Panicles of dps1 plants showed an accumulation of reactive oxygen species (ROS), lower antioxidant activity, and increased programmed cell death. Map-based cloning revealed that DPS1 encodes a mitochondrial-localized protein containing a cystathionine β-synthase domain that showed the highest expression in panicles and anthers. DPS1 physically interacted with mitochondrial thioredoxin proteins Trx1 and Trx20, and it participated in ROS scavenging. Global gene expression analysis in dps1 revealed that biological processes related to fatty acid metabolism and ROS homeostasis were significantly affected, and the expression of key genes involved in wax and cutin biosynthesis were downregulated. These results suggest that DPS1 plays a vital role in regulating ROS homeostasis, anther cuticle formation, and panicle development in rice., (© 2019 The Authors. New Phytologist © 2019 New Phytologist Trust.)

Breast cancer is the most common cancer in females, and the leading cause of cancer-related mortality in the world. Among the available treatment options for cancer, chemotherapy is the therapy for treating a variety of cancer patients. However, the therapeutic efficacy of current agents is minimal and these drugs do not retard the progression of disease pathology. Lack of appropriate therapy may increase the prevalence of disease in world. Hence, more effective strategies and novel therapies must be pursued for altering the progression of the disease acting through different mechanisms. There is a continuing need for new and improved therapy. Hence, Vitamin B17 is suggested a therapeutic potential for treating breast cancer. This study is to evaluate the potential therapy of vitamin B17 (Vit B17, amygdalin) against Ehrlish solid tumors, bearing mice (EST) induced DNA damage, NF-Kb, TNFα and apoptosis. Sixty female mice were randomly divided into four groups: (I, control group; II, VitB17 group; III, EST group; IV, EST+VitB17 group). EST induced group had elevated in the levels of serum ALT, AST, ALP, creatinine, urea, potassium ions, cholesterol, triglycerides, cytokine IFNγ, NF-kb, DNA damage, tumor TNF-α, VEGF expressions and had an associated reduction in serum albumin, total proteins, sodium ions, tumor NF-kb, Bcl2 and survivin expressions. Treatment of EST with vitamin B17 (EST+VitB17) modulates the changes in liver and kidney functions, electrolytes, cytokines, NF-kb and apoptosis in mice bearing EST. Hence, these findings suggest that vitamin B17 can be a reliable and novel therapy for breast cancer, further validate the neoplastic activity of Vitamin B17 as a potential therapy for other types of cancer is needed.

Colorectal cancer (CRC) is the third most common malignancy and the fourth most common cause of cancer-related death worldwide. The treatment of metastatic CRC considered palliative for many years aiming for an improved life, with little hope of a cure, highlighting the need for developing novel targeted therapy for CRC. Human protein kinases constitute a complicated system with complex internal and external interactions, which stimulates various cellular processes such as cell growth, metabolism, survival, and apoptosis. This study investigated the effect of a combination of atypical Protein Kinase C (PKC) inhibitor (either ICA-I or ζ-Stat) and 5-FU (a thymidylate synthase inhibitor) on CRC cells viability concerning cellular DNA damage. In this study, we took multiple approaches such as colony formation assay, flow cytometry, DNA ladder assay, TUNEL assay, etc. to examine the CRC cell viability and apoptosis as a function of combination treatment. Our findings showed that the combination of atypical PKC inhibitor and 5-FU synergistically reduced the viability of CRC cells and induced apoptosis. Additionally, the DNA ladder and TUNEL assays indicated that there was a notable DNA damage and fragmentation because of lack of thymidylate synthase and due to the deactivation of atypical PKC dependent CDK7. These data suggest that the simultaneous knockdown of upstream atypical PKC protein and downstream DNA damage repairing mechanism would be a useful approach to combat CRC and to improve overall patients' survival rate., (Copyright © 2019 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Self-incompatibility (SI) is genetically determined reproductive barrier preventing inbreeding and thereby providing the maintenance of plant species diversity. At present, active studies of molecular bases of SI mechanisms are underway. S-RNAse-based SI in Petunia hybrida L. is a self-/non-self recognition system that allows the pistil to reject self pollen and to accept non-self pollen for outcrossing. In the present work, using fluorescent methods including the TUNEL method allowed us to reveal the presence of markers of programmed cell death (PCD), such as DNA fragmentation, in growing in vivo petunia pollen tubes during the passage of the SI reaction. The results of statistical analysis reliably proved that PCD is the factor of S-RNAse-based SI. It was found that preliminary treatment before self-pollination of stigmas of petunia self-incompatible line with aminooxyacetic acid (AOA), inhibitor of ACC synthesis, led to stimulation of pollen tubes growth when the latter did not exhibit any hallmarks of PCD. These data argue in favor of assumption that ethylene controls the passage of PCD in incompatible pollen tubes in the course of S-RNAse-based SI functioning. The involvement of the hormonal regulation in SI mechanism in P. hybrida L. is the finding observed by us for the first time.

Rapid sample preparation is one of the leading bottlenecks to low-cost and efficient sample component detection. To overcome this setback, a technology known as Lyse-It has been developed to rapidly (less than 60 seconds) lyse Gram-positive and-negative bacteria alike, while simultaneously fragmenting DNA/RNA and proteins into tunable sizes. This technology has been used with a variety of organisms, but the underlying mechanism behind how the technology actually works to fragment DNA/RNA and proteins has hitherto been studied. It is generally understood how temperature affects cellular lysing, but for DNA/RNA and protein degradation, the temperature and amount of energy introduced by microwave irradiation of the sample, cannot explain the degradation of the biomolecules to the extent that was being observed. Thus, an investigation into the microwave generation of reactive oxygen species, in particular singlet oxygen, hydroxyl radicals, and superoxide anion radicals, was undertaken. Herein, we probe one aspect, the generation of reactive oxygen species (ROS), which is thought to contribute to a non-thermal mechanism behind biomolecule fragmentation with the Lyse-It technology. By utilizing off/on (Photoinduced electron transfer) PET fluorescent-based probes highly specific for reactive oxygen species, it was found that as oxygen concentration in the sample and/or microwave irradiation power increases, more reactive oxygen species are generated and ultimately, more oxidation and biomolecule fragmentation occurs within the microwave cavity., Competing Interests: The University of Maryland, Baltimore County (UMBC) has filed and has had issued several patents related to microwave-based lysing and DNA fragmentation. Those patents are licensed to Lyse-It LLC, a Maryland-based biotechnology company, which Professor Geddes currently holds stock. Patent #: US US9500590B2, “Assays for pathogen detection using microwaves for lysing and accelerating metal-enhanced fluorescence.” Patent #: US10294451B2, “Flow and static lysing systems and methods for ultra-rapid isolation and fragmentation of biological materials by microwave irradiation”, Chris D. Geddes. Lyse-It LLC was not involved in the study design of the material within this paper, nor do any of the authors receive any salaries or consultancies from Lyse-It LLC. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

Objective: Isorhamnetin, a derivative of quercetin, exerts antioxidant and anti-inflammatory effects in different diseases, and we examined its protective effects against diabetes-related changes in the brain., Methods: A single dose of a freshly prepared solution of streptozotocin (STZ) (60 mg/kg body weight) was intraperitoneally injected to establish STZ-induced diabetic model in male Wistar rats. The animals were randomly divided into four groups: control, control + isorhamnetin, diabetic, diabetic + isorhamnetin. Isorhamnetin at a dose of 10 mg/kg body weight was intraperitoneally administrated once a day for 12 weeks. Formalin and tail immersion tests were performed to evaluate the severity of pain. Astrogliosis markers such as GFAP and APO-E4, DNA fragments, MDA level, and TNFα expressions were evaluated using ELISA assay. Neuronal density in the hippocampus region was evaluated using Nissl staining. The method of Ellman and fluorescent probe 2, 7-dichlorofluorescein diacetate (DCFH-DA) was used to measure brain acetyl-cholinesterase activity and detect reactive nitrogen and oxygen species (RNS and ROS), respectively., Results: Isorhamnetin reduced pain, blood glucose levels, and increased body weight significantly compared to control. Moreover, isorhamnetin inhibited astroglial activation, acetyl-cholinesterase activity, oxidative stress, apoptosis, and inflammation., Conclusion: These findings suggested that isorhamnetin has potential effects as neuroprotective agents against diabetes-related changes in the brain., (Copyright © 2019 Elsevier B.V. All rights reserved.)

The present study compares a protocol that mimics freezing of ram semen in static nitrogen vapor with two protocols with an initial low cooling rate in the first step, followed by higher cooling rates where ice nucleation occurs. Semen ejaculates, obtained from twelve adults rams, were diluted with TEST-based extender and frozen with either Protocol 1 (three-step decelerating cooling): from +5 °C to -35 °C (40 °C/min), from -35 °C to -65 °C (17 °C/min), and then from -65 °C to -85 °C (3 °C/min); or Protocol 2 (three-step accelerating cooling): from +5 °C to -5 °C (4 °C/min), from -5 °C to -110 °C (25 °C/min), and then from -110 °C to -140 °C (35 °C/min); or Protocol 3 (two-step accelerating cooling), from +5 °C to -10 °C (5 °C/min), and then from -10 °C to -130 °C (60 °C/min). Post-thaw sperm quality was reduced for all protocols (p < .05) compared with fresh semen. Post-thaw percentages of sperm motility characteristics and sperm with intact plasma membrane, intact acrosome, and intact mitochondrial membrane were greater using Protocol 3 than Protocol 2 (p < .05) and Protocol 1 (p < .01). In addition, the post-thaw percentage of sperm with fragmented DNA was lower (p < .05) using Protocol 3 compared with Protocol 1. The present results indicate that a cooling rate of 60 °C/min around and after the time point of ice nucleation provided better post thaw survival and function of ram sperm than lower (and/or decelerating) cooling rates., (Copyright © 2019 The Authors. Published by Elsevier Inc. All rights reserved.)

Cancer therapies are aimed at eliminating the rapidly growing tumor cells by surgery and radiotherapy. The present therapies are only fruitful in early identified cases. The present study involves the preparation and characterization of eudragit S100-coated mini-capsules filled with chitosan nanoparticles-unconjugated and folic acid (FA)-conjugated encapsulating caspase 3 activator (7-hydroxystaurosporine). The formulated nanoparticles were compared for the cancer targeting and curing ability of the same by pre-investigation through drug release in organ-imitated fluids and ex vivo studies (cell viability, DNA fragmentation, caspase 3 activity), and then its confirmation through in vivo studies (tumor regression and distribution). The prepared nanoparticles were nearly spherical in shape, having positive zeta potential. From the cell line studies, it can be concluded that both the conjugated formulations showed better uptake, apoptosis, caspase 3 activation and DNA fragmentation. Stability study was performed according to ICH guidelines and formulation stored at 5° ± 3 °C was found to be most stable. The in vivo studies also supported the findings and showed better comprehensive residence time (23.61 ± 1.75 h), tumor distribution profile than UCN 01 alone. The results of in vitro, ex vivo and in vivo studies lead to the conclusion that the coated minicapsules specifically deliver the drug in the colon showing high therapeutic value and low side effects.

Nanoparticles (NPs) synthesized by laser ablation in distilled water were used to study their biological effect on normal and cancer cells. Parameters such as cell morphology, cell proliferation and viability were examined for treated cell lines, and the effect was represented in terms of cells cytotoxicity using standard procedures. The study reveals the higher cytotoxic effect of nanoparticles on cancerous cells of breast, melanoma and colon origin compared to normal fibroblast cells NIH- 3T3. Furthermore, DNA fragmentation assay results demonstrated the apoptosis mediated cell death in nanoparticle-treated cancer cells. The distinct role of nanoparticles in normal and cancer cells of different origin showed that nanoparticles were specific to cause cytotoxicity in particular cancer cells type. NPs exhibit cytotoxic effects in cancer cells by inducing apoptosis. These studies provide fundamental evidence for the easy, simple and safe mode of nanoparticles synthesis and their application in cancer cells death.

As the largest proportion of male infertility population, asthenozoospermia patients often resort to sperm cryopreservation to preserve fertility as well as to enrich motile sperm for assisted reproductive techniques (ART), although it may cause some cryodamage during the freezing-thawing process. The objective of this study was to investigate whether mitochondrial antioxidant Mito-Tempo was effective in preventing cryodamage of asthenozoospermic spermatozoa. Asthenozoospermic semen samples were collected and cryopreserved in media supplemented with different concentrations (0.0, 1.0, 10 and 100 μM) of Mito-Tempo. We measured sperm motility, viability, membrane integrity, DNA fragmentation, mitochondrial membrane potential, oxidation product, and antioxidant enzymes activities. Supplementation of the cryopreservation media with Mito-Tempo (10 and 100 μM) induced a significant improvement in sperm viability, motility, membrane integrity, mitochondrial membrane potential and chromatin integrity (P < 0.05). Significant enhancement of antioxidant enzymes activities accompanied by the decreased formation of oxidation products (ROS and MDA) was also observed in groups supplemented with Mito-Tempo (10 and 100 μM). It is concluded that mitochondria targeted antioxidant Mito-Tempo alleviates cryodamage by regulating intracellular oxidative metabolism in spermatozoa from asthenozoospermic patients after cryopreservation., (Copyright © 2019. Published by Elsevier Inc.)

Alzheimer's disease (AD) is the most prevalent neurodegenerative amyloid disorder with progressive deterioration of cognitive and memory skills. Despite many efforts, no decisive therapy yet exists for AD. Safranal is the active constituent of saffron essential oil with antioxidant, anti-inflammatory, and anti-apoptotic properties. In this study, the possible beneficial effect of safranal on cognitive deficits was evaluated in a rat model of AD induced by intrahippocampal amyloid beta (Aβ 1-40 ). Safranal was daily given p.o. (0.025, 0.1, and 0.2 ml/kg) post-surgery for 1 week and finally learning and memory were evaluated in addition to assessment of the involvement of oxidative stress, inflammation, and apoptosis. Findings showed that safranal treatment of amyloid β-microinjected rats dose-dependently improved cognition in Y-maze, novel-object discrimination, passive avoidance, and 8-arm radial arm maze tasks. Besides, safranal attenuated hippocampal level of malondialdehyde (MDA), reactive oxygen species (ROS), protein carbonyl, interleukin 1β (IL-1β), interleukin 6 (IL-6), tumor necrosis factor α (TNFα), nuclear factor-kappa B (NF-kB), apoptotic biomarkers including caspase 3 and DNA fragmentation, glial fibrillary acidic protein (GFAP), myeloperoxidase (MPO), and acetylcholinesterase (AChE) activity and improved superoxide dismutase (SOD) activity and mitochondrial membrane potential (MMP) with no significant effect on nitrite, catalase activity, and glutathione (GSH). Furthermore, safranal prevented CA1 neuronal loss due to amyloid β 1-40 . In summary, safranal treatment of intrahippocampal amyloid beta 1-40 -microinjected rats could prevent learning and memory decline via neuronal protection and at a molecular level through amelioration of apoptosis, oxidative stress, inflammation, cholinesterase activity, neutrophil infiltration, and also by preservation of mitochondrial integrity.

Leishmania infantum is responsible for the cutaneous and visceral form of this zoonotic disease, which is potentially lethal for humans and has dogs as natural reservoir. In the light of the antiparasitic properties displayed by several natural products, L. infantum promastigotes were exposed to green tea ( Camellia sinensis ) leaves extract (GTE) and pomegranate ( Punica granatum ) peel extract (PPE). Both extracts, characterized by NMR and HPLC analysis, inhibited parasite proliferation in a dose-dependent manner, as proved by IC 50 evaluation determined by MTT assay.Moreover, the reversibility assay showed that GTE and PPE have an aptotosis-mediated leishmanicidal effect, as evidenced by DNA degradation and confirmed by DNA fragmentation and real-time PCR analyses. Finally, for the first time morphological and ultrastructural alterations induced by a P. granatum extract on Leishmania were shown by the use of light, transmission and scanning electron microscopy.

Male subfertility is a global issue in human reproduction as well as in animal reproduction. Bacterial infection and semen contamination are still widely overlooked. As the collection of ejaculates is not a sterile process, it is necessary to add antimicrobial agents to avoid a possible depreciation of semen samples. As traditionally used antibiotics have been questioned because of an ever-increasing bacterial resistance, natural bioactive molecules could offer an alternative because of their antibacterial and antioxidant properties. As such, we decided to compare the effects of selected natural biomolecules (resveratrol-RES, quercetin-QUE and curcumin-CUR) with routinely used antibiotics in animal biotechnologies (penicillin-PEN, gentamicin-GEN and kanamycin-KAN) on the rabbit sperm vitality in the presence of Enterococcus faecalis . Changes in the sperm structural integrity and functional activity were monitored at 0, 2, 4 and 6 h. Computer-assisted sperm analysis (CASA) was used for the assessment of spermatozoa motility. Production of reactive oxygen species (ROS) was evaluated using chemiluminiscence, while the mitochondrial membrane potential (ΔΨm) was examined using the JC-1 dye. Finally, the sperm chromatin dispersion (SCD) test was used to assess DNA fragmentation, and changes to the membrane integrity were evaluated with the help of annexin V/propidium iodide. The motility assessment revealed a significant sperm motility preservation following treatment with GEN ( p < 0.001), followed by PEN and CUR ( p < 0.01). QUE was the most capable substance to scavenge excessive ROS ( p < 0.001) and to maintain ΔΨm ( p < 0.01). The SCD assay revealed that the presence of bacteria and antibiotics significantly ( p < 0.05) increased the DNA fragmentation. On the other hand, all bioactive compounds readily preserved the DNA integrity ( p < 0.05). In contrast to the antibiotics, the natural biomolecules significantly maintained the sperm membrane integrity ( p < 0.05). The microbiological analysis showed that GEN ( p < 0.001), KAN ( p < 0.001), PEN ( p < 0.01) and CUR ( p < 0.01) exhibited the strongest antibacterial activity against E. faecalis . In conclusion, all selected biomolecules provided protection to rabbit spermatozoa against deleterious changes to their structure and function as a result of Enterococcus faecalis contamination. Therefore, administration of RES, QUE and/or CUR to rabbit semen extenders in combination with a carefully selected antibacterial substance may be desirable.

Tannic acid (TA) portrays a myriad of beneficial properties and has forthwith achieved incessant significance for its cytoprotective qualities in traditional and modern-day medicine. However, TA displays an ambiguous nature demonstrating anti-oxidant and pro-oxidant traits, beckoning further research. Although vast literature on the anti-proliferative effects of TA on cancer cell lines exist, the effects on normal cells remain unchartered. Herein, the cytoproliferative and anti-oxidant effects induced by TA in human embryonic kidney (Hek-293) cells were investigated. Data obtained from the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay demonstrated that TA increased the cell viability and cellular proliferation rate at higher concentrations. Hoechst assay, examining proliferation marker Ki67 supported these findings. DNA fragmentation and oxidative stress-inducers were specifically noted at IC25 and IC50 treatments via biochemical assays. This alluded to TA's pro-oxidant characteristics. However, the countervailing anti-oxidant defence mechanisms as the endogenous anti-oxidants and phase2 detoxification enzymes were significantly upregulated. Luminometry fortified the anti-oxidant capacity of TA, whereby executioner caspase-3/7 were not activated subservient to the activation of initiator caspases-8 and -9. Thus, proving that TA has anti-apoptotic traits, inter alia. Therefore, TA proved to harbour anti-oxidant, anti-apoptotic, and proliferative effects in Hek-293 cells with its partial cytotoxic responses being outweighed by its cytoprotective mechanisms.

This study examines the ability of arabinoxylan rice bran (MGN-3/Biobran) to enhance the anti-cancer effects of fractionated X-ray irradiation of Ehrlich solid tumor-bearing mice. Swiss albino mice bearing tumors were exposed to the following: (i) Biobran treatment (40 mg/kg/day, intraperitoneal injections) beginning on day 11 post-tumor cell inoculation until day 30; (ii) ionizing radiation (Rad) 2 Gy at three consecutive doses on days 12, 14 and 16; or (iii) Biobran + Rad. Final tumor weight was suppressed by 46% for Biobran, 31% for Rad and 57% for the combined treatment (Biobran + Rad) relative to control untreated mice. Biobran and Rad also arrested the hypodiploid cells in the sub-G1-phase, signifying apoptosis by +102% and +85%, respectively, while the combined treatment induced apoptosis by +123%, with similar results in the degree of DNA fragmentation. Furthermore, Biobran + Rad upregulated the relative gene expression and protein level of p53 and Bax in tumor cells, down-regulated Bcl-2 expression, and increased the Bax/Bcl-2 ratio and caspase-3 activity, with the combined treatment greater than for either treatment alone. Additionally, the combined treatment modulated the decrease in body weight, the increase in liver and spleen weight, and the elevation of liver enzymes aspartate aminotransferase, alanine aminotransferase and gamma-glutamyl transferase to be within normal values. We conclude that Biobran enhances radiation therapy-induced tumor regression by potentiating apoptosis and minimizing toxicities related to radiation therapy, suggesting that Biobran may be useful in human cancer patients undergoing radiotherapy and warranting clinical trials., (© The Author(s) 2019. Published by Oxford University Press on behalf of The Japan Radiation Research Society and Japanese Society for Radiation Oncology.)

Acute myeloid leukemia (AML) is a blood cancer characterized by the formation of faulty defective myelogenous cells with morphological heterogeneity and cytogenic aberrations leading to a loss of their function. In an attempt to find an effective and safe AML treatment, vitamin E derivatives, including tocopherols were considered as potential anti-tumor compounds. Recently, other isoforms of vitamin E, namely tocotrienols have been proposed as potential potent anti-cancerous agents, displaying promising therapeutic effects in different cancer types. In this study we evaluated the anti-cancerous effects of γ-tocotrienol, on AML cell lines in vitro . For this purpose, AML cell lines incubated with γ-tocotrienol were examined for their viability, cell cycle status, apoptotic cell death, DNA fragmentation, production of reactive oxygen species and expression of proapoptotic proteins. Our results showed that γ-tocotrienol exhibits time and dose-dependent anti-proliferative, pro-apoptotic and antioxidant effects on U937 and KG-1 cell lines, through the upregulation of proteins involved in the intrinsic apoptotic pathway., Competing Interests: The authors declare no conflicts of interest.

Different antioxidants have been introduced to reduce oxidative stress during the cryopreservation. The main goal of this study was to evaluate the effects of canthaxanthin on human sperm parameters during the freeze-thaw process. This study was performed on 25 normozoospermic semen samples dividing into five groups including 0, 0.1, 1, 10, and 25 µM of canthaxanthin. The prepared spermatozoa were cryopreserved by rapid freezing technique. Sperm motility, viability (eosin-nigrosin), morphology (Papanicolaou), acrosome reaction (double staining), DNA denaturation (acridine orange), chromatin packaging (aniline blue and toluidine blue), and DNA fragmentation (sperm chromatin dispersion test) were evaluated before freezing and after thawing. All sperm parameters after thawing significantly were decreased compared to before freezing. Twenty-five micromolar canthaxanthin could significantly improve the progressive and total motility, viability, normal morphology, chromatin packaging, acrosome integrity and DNA denaturation and fragmentation. Ten micromolar canthaxanthin significantly improved total motility, viability, normal morphology, chromatin packaging, acrosome integrity and DNA denaturation and fragmentation. Whereas, in 1 µM group, there were significant differences only in improvement of acrosome integrity, chromatin packaging (toluidine blue) and DNA denaturation and fragmentation. But, in 0.1 µM group, there were no significant differences in any of measured parameters. It seems that canthaxanthin ameliorates detrimental effects of cryopreservation on human sperm parameters., (© 2019 Blackwell Verlag GmbH.)

Aims: Entamoeba histolytica can induce host cell death through induction of various intracellular signalling pathways. The responses triggered by E. histolytica are closely associated with tissue pathogenesis and immune evasion. Although E. histolytica can induce reactive oxygen species (ROS) in host cells, which NADPH oxidase (NOX) isoform contributes to amoeba-triggered Jurkat T-cell death is unclear. In this study, we investigated the signalling role of NOX4-derived ROS in E. histolytica-induced Jurkat T-cell death process., Methods and Results: In resting-state Jurkat T cells, NOX4 is strongly expressed. When Jurkat T cells were incubated with live E. histolytica trophozoites, intracellular ROS was significantly increased compared to cells incubated with medium alone. E. histolytica-induced ROS production was inhibited by pretreating Jurkat T cells with a NOX inhibitor. In addition, pretreating Jurkat T cells with a NOX inhibitor (Diphenyleneiodonium chloride) effectively blocked E. histolytica-induced phosphatidylserine (PS) exposure and DNA fragmentation of host cells. Moreover, siRNA-mediated knockdown of NOX4 protein expression in Jurkat T cells prevented E. histolytica-induced ROS generation and DNA fragmentation., Conclusion: These results suggest that NOX4 has a critical role in ROS-dependent cell death process in Jurkat T cells induced by E. histolytica., (© 2019 John Wiley & Sons Ltd.)

CGA-N9, a peptide derived from human chromogranin A (CGA), was found to have antimicrobial activity in our previous investigation, but its mechanism of action remains unclear. Herein, the mechanism of action of CGA-N9 was investigated. We found that CGA-N9 induced the depolarization of the cell membrane and uptake of calcium ions into the cytosol and mitochondria. With the disruption of the mitochondrial membrane potential, the generation of intracellular reactive oxygen species (ROS) increased. Accordingly, we assessed apoptotic processes in Candida tropicalis cells post-treatment with CGA-N9 and found cytochrome c leakage, chromatin condensation and DNA degradation. The interaction of CGA-N9 with DNA in vitro showed that CGA-N9 did not degrade DNA but bound to DNA via an electrostatic interaction. In conclusion, CGA-N9 exhibits antifungal activity by inducing apoptosis in C. tropicalis., (© 2019 The Author(s).)

Background: Methylation patterns in circulating cell-free DNA are potential biomarkers for cancer and other pathologies. Currently, bisulfite treatment underpins most DNA methylation analysis methods, however, it is known to fragment DNA. Circulating DNA is already short, and further fragmentation during bisulfite treatment is of concern, as it would potentially reduce the sensitivity of downstream assays., Methods: We used high molecular weight genomic DNA to compare fragmentation and recovery following bisulfite treatment with 2 commercially available kits (Qiagen). The bisulfite treated DNA was visualised on an agarose gel and quantified by qPCR. We also bisulfite treated, visualised and quantitated circulating DNA from plasma., Results: There was no difference in DNA fragmentation between the two kits tested, however, the Epitect Fast kit gave better recovery than the standard Epitect kit, with the same conversion efficiency. We also found that bisulfite treated circulating DNA migrates as distinct bands on agarose gels, suggesting that, in contrast to genomic DNA, it remains largely intact following treatment. Bisulfite treatment of 129 and 234 base PCR products confirmed that this was due to the short length of the circulating DNA fragments. Compared to double stranded DNA, bisulfite treated single stranded DNA gives a very weak signal on gel electrophoresis., Conclusions: DNA fragmentation during bisulfite treatment does not contribute to loss of sensitivity in methylation analysis of circulating DNA. The absence of DNA fragments below approximately 170 bases from agarose gel images of purified circulating DNA raises the possibility that these fragments are single stranded following the DNA extraction step., Competing Interests: I have read the journals competing interests policy and the authors of this manuscript have the following competing interests: KW- stock ownership Guardant Health, Exact Sciences and Epigenomics AG. This does not alter our adherence to PLOS ONE policies of sharing data and materials.

Excessive fluoride mainly causes skeletal lesions. Recently, it has been reported that an appropriate level of calcium can alleviate fluorosis. However, the appropriate concentration and mechanism of calcium addition is unclear. Hence, we evaluated the histopathology and ultrastructure, DNA fragmentation, hormonal imbalances, biomechanical levels, and expression of apoptosis-related genes after treating the rats with 150 mg/L NaF and different concentrations of CaCO 3 . Our results suggested that NaF induced the histopathological and ultrastructural injury, with a concomitant increase in the DNA fragmentation ( P < 0.05) and serum OC (17.5 ± 0.89 pmoL/L) at 120 days. In addition, the qRT-PCR and western blotting results indicated that NaF exposure upregulated the mRNA and protein expression of Bax, Calpain, Caspase 12, Caspase 9, Caspase 7, Caspase 3, CAD, PARP, and AIF while downregulated Bcl-2 ( P < 0.01) and decreased the bone ultimate load by 27.1%, the ultimate stress by 10.1%, and the ultimate deformity by 23.3% at 120 days. However, 1% CaCO 3 supplementation decreased the serum OC (14.7 ± 0.65 pmoL/L), bone F content ( P < 0.01), and fracture and breakage of collagen fibers and changed the expression of endoplasmic reticulum pathway-related genes and proteins at 120 days. Further, 1% CaCO 3 supplementation increased the bone ultimate load by 20.9%, the ultimate stress by 4.89%, and the ultimate deformity by 21.6%. In summary, we conclude that 1% CaCO 3 supplementation alleviated fluoride-induced bone damage by inhibiting endoplasmic reticulum stress and mitochondrial dysfunction.

A neutral bidentate ligand 2-(3-methyl-5-phenyl pyrazol-1-yl) benzthiazole (L) has been synthesized by refluxing equimolar proportions of 2-hydrazino benzthiazole and benzoyl acetone in ethanol. The ligand acts in a NN donor fashion and forms stable mononuclear, MoOX 3 L [L = Ligand, X = Cl (1), Br (2)] and binuclear Mo 2 O 4 X 2 L 2 [L = Ligand, X = Cl (3), Br (4)] complexes with molybdenum(V). The ligand and complexes are thoroughly characterized by elemental analyses, IR, UV-Vis spectroscopy, EPR study, magnetic susceptibility, thermogravimetry and cyclic voltammetry. Magnetic moment measurements reveal that the mononuclear complexes are paramagnetic while the binuclear complexes are diamagnetic in nature. EPR studies also confirm the presence of a mononuclear Mo(V) moiety in the complexes. Relevant Density Functional Theory (DFT) calculations have been carried out to determine the structures of the synthesized compounds. The binding mode and mechanism of interaction of the synthesized compounds with bovine serum albumin (BSA) were studied by concentration dependent absorption and fluorescence titration experiments. The ligand and complexes 1-4 are screened for their potential in vitro anticancer activities against three different human cancer cell lines, namely, cervix adenocarcinoma epithelial cells (HeLa), renal carcinoma cells (SK-RC-45) and breast adenocarcinoma cells (MCF-7). The oxomolybdenum(V) complexes are found to exhibit higher anticancer potency towards the cancer cells than the free ligand. Also, structure activity relationship (SAR) studies of this new series of oxomolybdenum(V) complexes indicate that the anticancer activity is to some extent dependent on the electronic effects of the halogen atom coordinated to the molybdenum centre., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Sunitinib is cardiotoxic, but the mechanisms are not entirely clear. We aimed to enlarge our knowledge about the role of mitochondria in cardiac toxicity of sunitinib in vitro and in vivo. For this reason, we studied the toxicity of sunitinib on cardiac H9c2 cells exposed for 24 h, permeabilized rat cardiac fibers exposed for 15 min and in mice treated orally with sunitinib for 2 weeks (7.5 mg/kg/day). In H9c2 cells exposed for 24 h, sunitinib was more cytotoxic under galactose (favoring mitochondrial metabolism) compared to glucose conditions (favoring glycolysis). Sunitinib dissipated the mitochondrial membrane potential starting at 10 μM under glucose and at 5 μM under galactose conditions. Sunitinib reduced activities of mitochondrial enzyme complexes of the electron transport chain (ETC), increased mitochondrial ROS accumulation and decreased the cellular GSH pool. Electron microscopy revealed swollen mitochondria with loss of cristae. Accordingly, sunitinib caused caspase 3 activation and DNA fragmentation in H9c2 cells. Co-exposure with mito-TEMPO (mitochondrial-specific ROS scavenger) for 24 h prevented ATP and GSH depletion, as well as the increases in H 2 O 2 and caspase 3/7 activity observed with sunitinib. In mice, treatment with sunitinib for two weeks increased plasma concentrations of troponin I and creatine kinase MB, indicating cardiomyocyte damage. The activity of enzyme complexes of the ETCwas decreased, mitochondrial ROS were increased and cleavage of caspase 3 was increased, suggesting cardiomyocyte apoptosis. In conclusion, mitochondrial damage with ROS accumulation appears to be an important mechanism of cardiotoxicity associated with sunitinib, eventually leading to apoptotic cell death., (Copyright © 2019 Elsevier B.V. All rights reserved.)