Effects of zinc deficiency and pinealectomy on cellular immunity in rats infected with Toxoplasma gondii

The effects of zinc and/or melatonin deficiencies on cellular immunity were investigated in rats infected with Toxoplasma gondii. A total of 50 adult male Sprague-Dawley rats were divided into 5 groups of 10 rats each. In group I, the rats were infected with T. gondii and fed a zinc-deficient diet; in group II, the rats were infected and their pineal gland was surgically removed. Group III included rats that were infected, pinealectomized, and fed a zinc-deficient diet. Group IV consisted of T. gondii-infested rats that received no treatment of any kind, and group V were normal controls. After 3 wk of treatment, all rats were sacrificed and the percentages of CD3, CD4, and CD8 lymphocytes, zinc, and melatonin levels in plasma and the percentage of lymphocyte in blood smears were analyzed. The CD3 ratios of groups I–III were significantly lower than those of groups IV and V (p<0.01). The CD4 lymphocytes were significantly higher in group IV than that in all other groups (p<0.05). In group IV, the CD8 lymphocytes were higher than in groups I–III (p<0.01) and those in group V were higher than for groups I and III (p<0.01). Lymphocyte incidence in group IV was higher than in the other four groups (p<0.01). The plasma zinc and plasma melatonin levels in groups I–III were significantly lower than those in the controls (p<0.01, both cases). These results suggest that zinc and/or melatonin deficiency have a negative influence on cellular immunity in rats with toxoplasmosis.     

Green synthesis and characterization of zinc oxide nanoparticles using Cayratia pedata leaf extract

The synthesis of Zinc oxide nanoparticles using a plant-mediated approach is presented in this paper. The nanoparticles were successfully synthesized using the Nitrate derivative of Zinc and plant extract of the indigenous medicinal plant Cayratia pedata. 0.1 mM of Zn (NO₃)₂.6H₂O was made to react with the plant extract at different concentrations, and the reaction temperature was maintained at 55 °C, 65 °C, and 75 °C. The yellow coloured paste obtained was wholly dried, collected, and packed for further analysis. In the UV visible spectrometer (UV–Vis) absorption peak was observed at 320 nm, which is specific for Zinc oxide nanoparticles. The characterization carried out using Field Emission Scanning Electron Microscope (FESEM) reveals the presence of Zinc oxide nanoparticles in its agglomerated form. From the X-ray diffraction (XRD) pattern, the average size of the nanoparticles was estimated to be 52.24 nm. Energy Dispersive Spectrum (EDX) results show the composition of Zinc and Oxygen, giving strong energy signals of 78.32% and 12.78% for Zinc and Oxygen, respectively. Fourier Transform - Infra-Red (FT-IR) spectroscopic analysis shows absorption peak of Zn–O bonding between 400 and 600 cm^?1. The various characterization methods carried out confirm the formation of nano Zinc oxide. The synthesized nanoparticles were used in the immobilization of the enzyme Glucose oxidase. Relative activity of 60% was obtained when Glucose oxidase was immobilized with the green synthesized ZnO nanoparticles. A comparative study of the green synthesized with native ZnO was also carried out. This green method of synthesis was found to be cost-effective and eco-friendly.     

Green synthesis of zinc oxide nanoparticles using Anoectochilus elatus,and their biomedical applications

Zinc and its derivatives requirement increased to enhance human immunity against the different pandemics, including covid-19. Green synthesis is an emerging field of research. Zinc oxide (ZnO) nanoparticles have been prepared from Anoectochilus elatus and characterized using absorption, vibrational and electron microscope analysis. They were carried for antibacterial, inflammatory control tendency, and potential antioxidant activities. The brine shrimp lethal assay tested the biologically derived nanomaterial toxicity and the lethal concentration (LC₅₀) is 599.79 µg/ml. The inhibition against the important disease-causing pathogens was measured against four-gram negative, gram-positive bacteria and two fungus pathogens. The nanomaterial exposed inhibition zone for gram-positive bacteria between 17 mm and 25 mm. The inhibition zone against gram-negative bacteria exists between 19 mm and 24 mm. The anti-inflammatory activity was assessed by inhibition of protein denaturation and protease inhibitory activity using nanomaterial. The antioxidant activity was examined using four assays for the therapeutic activities. The average size range of 60–80 nm nanoparticles has prepared and exposed the good biological activity between 50 µg/ml and 100 µg/ml. The comparative results of anti-inflammatory and antioxidant assay results with standards such as Aspirin and vitamin C exposed that two to three times higher concentrations are required for the fifty percent of inhibitions. The prepared low-cost nanoparticle has exhibited excellent biological activity without any side effects and may enhance immunity.     

Primary brain cell infection by Toxoplasma gondii reveals the extent and dynamics of parasite differentiation and its impact on neuron biology

Toxoplasma gondii is a eukaryotic parasite that forms latent cysts in the brain of immunocompetent individuals. The latent parasite infection of the immune-privileged central nervous system is linked to most complications. With no drug currently available to eliminate the latent cysts in the brain of infected hosts, the consequences of neurons'' long-term infection are unknown. It has long been known that T. gondii specifically differentiates into a latent form (bradyzoite) in neurons, but how the infected neuron responds to the infection remains to be elucidated. We have established a new in vitro model resulting in the production of mature bradyzoite cysts in brain cells. Using dual, host and parasite RNA-seq, we characterized the dynamics of differentiation of the parasite, revealing the involvement of key pathways in this process. Moreover, we identified how the infected brain cells responded to the parasite infection revealing the drastic changes that take place. We showed that neuronal-specific pathways are strongly affected, with synapse signalling being particularly affected, especially glutamatergic synapse signalling. The establishment of this new in vitro model allows investigating both the dynamics of parasite differentiation and the specific response of neurons to long-term infection by this parasite.     

Phyto-assisted synthesis of zinc oxide nanoparticles using Cassia alata and its antibacterial activity against Escherichia coli

Zinc oxide, an established inorganic metal oxide in nanoparticles form exhibits tremendous anti-bacterial activity. The present study focuses on determining the anti-bacterial activity of green synthesized zinc oxide nanoparticles (ZnO NPs). Results clearly validate the effective synthesis of spherical shaped nanoparticles with average size range of 60–80 nm. SEM and EDAX data buttresses the results obtained by XRD pattern in terms of size and purity. ZnO NPs exhibited dose-dependent anti-bacterial activity against Escherichia coli (E. coli) and the IC₅₀ value was calculated to be around 20 μg/mL. Growth kinetics study was conducted in the presence of nanoparticles which demonstrated the bacteriostatic effect of ZnO NPs. The study recommends the potential use of ZnO NPs in industries like food, pharmaceutical, agriculture, cosmetic industries for its anti-bacterial activity.     

Green synthesis and characterization of gold nanoparticles using endophytic fungi Fusarium solani and its in-vitro anticancer and biomedical applications

The present study aimed to explore the anticancer potentials of the gold nanoparticles (NPs) obtained by green synthesis method using an endophytic strain Fusarium solani ATLOY – 8 has been isolated from the plant Chonemorpha fragrans. The formation of the NPs was analyzed by UV, FTIR, SEM and XRD. The synthesized NPs showed pink-ruby red colors and high peak plasmon band was observed between 510 and 560 nm. It is observed that intensity of absorption steadily increases the wavelength and band stabilizes at 551 nm. The XRD pattern revealed the angles at 19, 38.32, 46.16, 57.50, and 76.81° respectively. Interestingly, the FTIR band absorption noted at 1413 cm⁻¹, 1041 cm⁻¹ and 690 cm⁻¹ ascribed the presence of amine II bands of protein, C-N and C-H stretching vibrations of the nanoparticles. SEM analysis indicated that the average diameter of the synthesized nanoparticles was between 40 and 45 nm. These NPs showed cytotoxicity on cervical cancer cells (He La) and against human breast cancer cells (MCF-7) and the NPs exhibited dose dependent cytotoxic effect. IC50 value was 0.8 ± 0.5 μg/mL on MCF-7 cell line and was found to be 1.3 ± 0.5 μg/mL on MCF-7 cell lines. The synthesized NPs induced apoptosis on these cancer cell lines. The accumulation of apoptotic cells decreased in sub G0 and G1 phase of cell cycle in the MCF-7 cancer cells were found to be 55.13%, 52.11% and 51.10% after 12 h exposure to different concentrations. The results altogether provide an apparent and versatile biomedical application for safer chemotherapeutic agent with little systemic toxicity.     

Green synthesis of silver nanoparticles by Nigella sativa extract alleviates diabetic neuropathy through anti-inflammatory and antioxidant effects

Green synthesis of silver nanoparticles has gained great interest among scientists. In view of this data, we conducted this study to identify the ameliorative effect of green synthesis of silver nanoparticles using Nigella sativa extract in diabetic neuropathy induced experimentally. In this study, 50 adult male albino rats were used and they were randomly divided into five groups; the first group was the healthy control group, the second group were the diabetic neuropathy diabetic neuropathy induced, Groups (3-6) diabetic neuropathy induced group and treated with silver nanoparticles, Nigella sativa extract and green synthesized silver nanoparticles using Nigella sativa extract respectively. Biochemical parameters including diabetic, inflammatory and antioxidant biomarkers were evaluated. Brain histopathology was also performed. Results revealed substantial rise in glucose, AGE, aldose reductase with insulin reduction in diabetic neuropathy induced group as compared to healthy control. Also, inflammatory markers increased significantly in diabetic neuropathy induced group. A remarkable change in oxidative status was observed in the same group. Furthermore, significant decline in nitrotyrosin level was observed. Regarding gene expression, we found significant down regulation in brain TKr A accompanied by upregulation of nerve growth factor in diabetic neuropathy group comparing with healthy control. Several treatments for diabetic neuropathy remarkably ameliorate all the investigated biomarkers. Histological findings are greatly relied on for the results achieved in this study. Therefore, it can be established that green synthesis of silver nanoparticles in combination with Nigella sativa extract could be a newly neuroprotective agents against inflammation and oxidative stress characterizing diabetic neuropathy through their antidiabetic, anti-inflammatory and anti-oxidants effects.     

Biogenic silver nanoparticles reduce Toxoplasma gondii infection and proliferation in RAW 264.7 macrophages by inducing tumor necrosis factor-alpha and reactive oxygen species production in the cells

Owing to the serious adverse effects caused by pyrimethamine and sulfadiazine, the drugs commonly used to treat toxoplasmosis, there is a need for treatment alternatives for this disease. Nanotechnology has enabled significant advances toward this goal. This study was conducted to evaluate the activity of biogenic silver nanoparticles (AgNp-Bio) in RAW 264.7 murine macrophages infected with the RH strain of Toxoplasma gondii. The macrophages were infected with T. gondii tachyzoites and then treated with various concentrations of AgNp-Bio. The cells were evaluated by microscopy, and culture supernatants were collected for ELISA determination of their cytokine concentration. Treatment with 6 μM AgNp-Bio reduced the infection and parasite load in infected RAW 264.7 macrophages without being toxic to the cells. The treatment also induced the synthesis of reactive oxygen species and tumor necrosis factor-alpha (both pro-inflammatory mediators), which resulted in ultrastructural changes in the tachyzoites and their intramacrophagic destruction. Our findings suggest that AgNp-Bio affect T. gondii tachyzoites by activating microbicidal and pro-inflammatory mechanisms and may be a potential alternative treatment for toxoplasmosis.     

Comparison of the effects of zinc deprivation and actinomycin D on ribonucleic acid synthesis by stimulated lymphocytes.

1. EDTA inhibited incorporation of [3H]uridine into RNA of lymphocytes, but did not decrease uptake into the cold-acid-soluble fraction of the cells. The inhibition by EDTA was largely reversible by simultaneous addition of Zn2+. 2. Low concentrations pf actinomycin D (3 ng/ml) added at the time of stimulation of the cells inhibited [3H]uridine incorporation into RNA, but concentrations of 50-100 ng/ml were required to produce the same degree of inhibition if addition of actinomycin D was delayed until just before the incorporation was measured. This difference in sensitivity did not reg within the cells. 3. When added immediately before phytohaemagglutinin, actinomycin D (3 ng/ml) and EDTA produced similar time-courses of inhibition of uridine incorporation. 4. Uridine incorporation at 32h was inhibited when actinomycin D (3 ng/ml) or EDTA was added just before stimulation of the cells, but was only slightly affected when they were added at 32h. At intermediate times the incorporation of uridine remained sensitive to addition of EDTA for longer than it was sensitive to actinomycin D. 5. Polyacrylamide-gel separation of RNA synthesized in EDTA-treated cultures in the presence or absence of added Zn2+ showed that lower availability of Zn2+ resulted in a decreased rate of transfer of radioactivity from 32S to 28S rRNA and decreased survival of 28S rRNA relative to 18S rRNA. 6. Close similarities have been shown to exist between the effects of EDTA and low concentrations of actinomycin D. Not all the effects of EDTA could be explained by postulating that Zn2+ was a constituent of RNA polymerase I, nor were the effects of actinomycin D readily explained by previously suggested mechanisms of action of this antibiotic.     

Biogenic synthesis of gold nanoparticles using Commiphora wightii and their cytotoxic effects on breast cancer cell line (MCF-7)

The current study aimed at developing gold nanoparticles (AuNPs) using the aqueous extract of the medicinal plant Commiphora wightii. The phytosynthesized gold nanoparticles (Cw@AuNPs) were evaluated for their anticancer activity against MCF-7 breast cancer cell model. The formation of AuNPs by Commiphora wightii leaf extract was confirmed by UV–vis spectra where their surface plasmon resonance was found at 533 nm. Further characterization of Cw@AuNPs was done by transmission electron microscopy (TEM), X-ray diffraction (XRD), energy dispersive X-ray (EDX) analysis, and fourier-transform infrared spectroscopy (FTIR) analysis. In vitro anticancer potential of thus obtained AuNPs was evaluated against MCF-7 and where the IC₅₀ was found to be 66.11 μg/mL Further, apoptotic studies were carried out using ethidium bromide dual staining, DNA fragmentation, comet assay, and flow cytometry studies. Results revealed that Cw@AuNPs at higher concentration significantly increased the apoptotic cells when compared to control cells. Cell cycle analysis of MCF-7 cells confirmed the cell cycle arrest at G2/M phase. These results demonstrate that the biosynthesized Cw@AuNPs appear to be promising for therapeutical applications against breast cancer.     

The effects of light quality and intensity on the synthesis of ribulose-1,5-bisphosphate carboxylase and its mRNAs in the green alga Chlorogonium elongatum

In the green alga Chlorogonium elongatum the promoting effect of light on the synthesis of ribulose bisphosphate carboxylase/oxygenase (RuBPCase) is mainly caused by blue light of wavelengths between 430 nm and 510 nm, with a maximum effect at about 460 nm. Blue light also causes an increase in the amounts of the mRNAs for the large and the small subunits of the enzyme. Furthermore, the concentration of RuBPCase is affected by the light energy fluence rate. The rate of synthesis as well as the maximal obtainable concentration of the enzyme are functions of the light energy fluence rate up to 26 W·m^-2. No further increase occurs beyond that intensity. The quantity of irradiation also alters the concentrations of the subunit mRNAs. The results indicate that the changes in the mRNA levels are the major regulatory steps in the light-dependent synthesis of the RuBPCase enzyme.Abbreviations LSU large subunit - pSSU precursor of the small subunit - RuBPCase ribulose bisphosphate carboxylase/oxygenase EC 4.1.1.39 Dedicated to Prof. Dr. E. Bünning on the occasion of his 80 th birthday     

Green synthesis of silver nanoparticles using aqueous rhizome extract of Zingiber officinale and Curcuma longa: In-vitro anti-cancer potential on human colon carcinoma HT-29 cells

This study was aimed to analyze the anti-cancer activity of silver nanoparticles (AgNPs) synthesized using aqueous plant extracts from the rhizome of Curcuma longa and Zingiber officinale. Synergistic aqueous extract of rhizome of C. longa and Z. officinale was used to green synthesis of AgNPs. Characterization of AgNPs was performed using UV–visible spectroscopy, FTIR, X-ray diffraction, TEM, and SEM analyses. Anti-cancer activity of AgNPs against human colon carcinoma (HT-29) cells was tested using MTT assay. UV–Visible spectroscopy analysis indicated the surface plasmon resonance (SPR) sharp peak at 350–430 nm wavelength that corresponds to the production of AgNPs. FTIR analysis reveals that existence of carboxyl (CO) and amine (NH) functional groups in the AgNPs. The X-ray diffraction analysis confirms four spectral peaks at 111, 200, 220, and 311. SEM analysis showed that AgNPs are in a spherical shape with a size of 42–61 nm and TEM analysis showed particle size are ranged between 20–51 nm. Anti-cancer study reveals that AgNPs had shown cytotoxicity against HT-29 cells at the concentrations ranged from 25 to 500 μg/mL and IC₅₀ at 150.8 µg/mL. This study concludes that AgNPs synthesized using rhizome of Z. officinale and C. longa possesses potential anti-cancer activity.     

Kinetic studies on endo-beta-galactosidase by a novel colorimetric assay and synthesis of N-acetyllactosamine-repeating oligosaccharide beta-glycosides using its transglycosylation activity.

Novel chromogenic substrates for endo-beta-galactosidase were designed on the basis of the structural features of keratan sulfate. Galbeta1-4GlcNAcbeta1-3Galbeta1-4GlcNAcbeta-pNP (2), which consists of two repeating units of N-acetyllactosamine, was synthesized enzymatically by consecutive additions of GlcNAc and Gal residues to p-nitrophenyl beta-N-acetyllactosaminide. In a similar manner, GlcNAcbeta1-3Galbeta1-4GlcNAcbeta-pNP (1), GlcNAcbeta1-3Galbeta1-4Glcbeta-pNP (3), Galbeta1-4GlcNAcbeta1-3Galbeta1-4Glcbeta-pNP (4), Galbeta1-3GlcNAcbeta1-3Galbeta1-4Glcbeta-pNP (5), and Galbeta1-6GlcNAcbeta1-3Galbeta1-4Glcbeta-pNP (6) were synthesized as analogues of 2. Endo-beta-galactosidases released GlcNAcbeta-pNP or Glcbeta-pNP in an endo-manner from each substrate. A colorimetric assay for endo-beta-galactosidase was developed using the synthetic substrates on the basis of the determination of p-nitrophenol liberated from GlcNAcbeta-pNP or Glcbeta-pNP formed by the enzyme through a coupled reaction involving beta-N-acetylhexosaminidase (beta-NAHase) or beta-d-glucosidase. Kinetic analysis by this method showed that the value of Vmax/Km of 2 for Escherichia freundii endo-beta-galactosidase was 1.7-times higher than that for keratan sulfate, indicating that 2 is very suitable as a sensitive substrate for analytical use in an endo-beta-galactosidase assay. Compound 1 still acts as a fairly good substrate despite the absence of a Gal group in the terminal position. In addition, the hydrolytic action of the enzyme toward 2 was shown to be remarkably promoted compared to that of 4 by the presence of a 2-acetamide group adjacent to the p-nitrophenyl group. This was the same in the case of a comparison of 1 and 3. Furthermore, the enzyme also catalysed a transglycosylation on 1 and converted it into GlcNAcbeta1-3Galbeta1-4GlcNAcbeta1-3Galbeta1-4GlcNAcbeta-pNP (9) and GlcNAcbeta1-3Galbeta1-4GlcNAcbeta1-3Galbeta1-4GlcNAcbeta1-3Galbeta1-4GlcNAcbeta-pNP (10) as the major products, which have N-acetyllactosamine repeating units.     

Photo-mediated optimized synthesis of silver nanoparticles using the extracts of outer shell fibre of Cocos nucifera L. fruit and detection of its antioxidant,cytotoxicity and antibacterial potential

Presently, photo-mediated optimized synthesis of SNPs (CS-AgNPs) was carried out with the help of aqueous extracts of coconut (Cocos nucifera) outer shell fibre. Green synthesis of CS-AgNPs was undertaken under laboratory light conditions and characterized by several standard techniques such as UV–visible spectrophotometer (UV–Vis), X-ray diffraction pattern (XRD), Fourier transform infrared (FT-IR), and scanning electron microscopy (SEM) images and energy dispersive spectroscopy (EDX). UV–Vis spectra displayed a surface plasmon resonance peak at 468 nm equivalent to CS-AgNPs, and the FT-IR spectra confirmed the association of biological molecules from the extract in the synthesis process. The SEM image data confirmed the round and circular nature of CS-AgNPs. The EDX data presented the elemental configuration with a solid peak at 3 KeV that matched with the Ag. The synthesized CS-AgNPs exhibited substantial cytotoxicity potential against the HepG2 cells with (effective concentration (IC₅₀) value of 15.28 µg/ml along with robust antioxidant potential, with respect to its 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging (IC₅₀ of 96.39 µg/ml) and reducing assay (IC_0.5 of 209.96 µg/ml). The CS-AgNPs demonstrated encouraging antimicrobial potential against four different pathogenic bacteria and one Candida sp. with inhibition zone diameter ranged between 8.87 and 13.07 mm. Overall, the existing investigation suggested that CS-AgNPs can be an attractive, cost-effective, and environment-friendly candidate for its possible uses in the food, cosmetics, and therapeutic fields.     

Biologically synthesis of gold nanoparticles using Cirsium japonicum var. maackii extract and the study of anti-cancer properties on AGS gastric cancer cells

Plant extract-mediated synthesis of metal nanoparticles (NPs) is an eco-friendly and cost-effective biosynthesis method that is more suitable for biological applications than chemical ones. We prepared novel gold NPs (AuNPs), Cirsium japonicum mediated-AuNPs (CJ-AuNPs), using a biosynthetic process involving Cirsium japonicum (Herba Cirsii, CJ) ethanol extract. The physicochemical properties of CJ-AuNPs were characterized using spectrometric and microscopic analyses. The in vitro stability of CJ-AuNPs was studied for 3 months. Moreover, the selective human gastric adenocarcinoma (AGS) cell killing ability of CJ-AuNPs was verified in cancer and normal cells. An in vitro study revealed that CJ-AuNPs trigger oxidative stress and iron-dependent ferroptosis in AGS cells. Mechanistically, CJ-AuNPs induced mitochondrial reactive oxygen species (ROS), Fe²⁺, and lipid peroxidation accumulation, and mitochondrial damage by destroying the glutathione peroxidase-4 (GPX4)-dependent antioxidant capacity. Furthermore, in a xenograft mouse model implanted with AGS cells, treatment with 2.5, 5, and 10 mg/kg CJ-AuNPs for 16 days reduced tumor xenograft growth in a dose dependent manner in vivo without systemic toxicity. These results demonstrate that CJ-AuNPs exert anticancer effects in vitro and in vivo by inducing ferroptosis-mediated cancer cell death. This study, based on green-synthesized nanodrug-induced ferroptosis, provides new insight into potential developments in cancer therapies.     

Location of functional region at N-terminus of polyhydroxyalkanoate (PHA) synthase by N-terminal mutation and its effects on PHA synthesis

Polyhydroxyalkanoate (PHA) synthase PhaC plays a very important role in biosynthesis of microbial polyesters PHA. Compared to the extensively analyzed C-terminus of PhaC, N-terminus of PhaC was less studied. In this paper, the N-terminus of two class I PHA synthases PhaC_Re and PhaC_Ah from Ralstonia eutropha and Aeromonas hydrophila, respectively, and one class II synthase PhaC2_Ps of Pseudomonas stutzeri strain 1317, were investigated for their effect on PHA synthesis. For PhaC_Re, deletion of 2–65 amino acid residues on the N-terminus led to enhanced PHB production with high PHB molecular weight of 2.50 × 10⁶ Da. For PhaC_Ah, the deletion of the N-terminal residues resulted in increasing molecular weights and widening polydispersity accompanied by a decreased PHA production. It was found that 3-hydroxybutyrate (3HB) monomer content in copolyesters of 3-hydroxybutyrate and 3-hydroxyhexanoate (3HHx) increased when the first 2–9 and 2–13 amino acid residues in the N-terminus of PhaC2_Ps were deleted. However, deletion up to the 40th amino acid disrupted the PHA synthesis. This study confirmed that N-terminus in different types of PHA synthases showed significant roles in the PHA productivity and elongation activity. It was also indicated that N-terminal mutation was very effective for the location of functional regions at N-terminus.