An insight into the binding and inhibition of eye ζ-crystallin by the environmental toxin arsenic: implications in eye diseases.

Arsenic contamination is highly prevalent in food chain, soil and groundwater. Continuous exposure to elevated levels of this environmental toxin is a global concern. Studies have reported enriched accumulation of arsenic in the eyes compared to other body organs leading to various eye diseases. Here, the impact of arsenic exposure on the enzymatic eye ζ-crystallin has been investigated. Arsenic inhibited the activity of the enzyme with an IC50 value of 35 µM. It decreased the free thiol group content of ζ-crystallin due to protein oxidation. The binding of arsenic with ζ-crystallin was explored using biophysical and computational tools. The enzyme undergoes some conformational changes upon arsenic binding. The binding constant (Kb) was determined to be 1.2 × 102 M-1. Thermodynamic parameters were determined by isothermal titration calorimetry (ITC) and the binding energy (ΔG) was calculated to be -3.52 kcal/mol. Molecular docking studies helped in visualizing the amino acid residues (especially Cys165) of the enzyme involved in binding with arsenic. Continuous arsenic exposure is expected to increase the eye crystallin-related abnormalities, elevating the risk of cataractogenesis. Therefore, proper measures need to be taken by authorities to control the contamination of arsenic in the environment and groundwater.Communicated by Ramaswamy H. Sarma.

Elucidation of kinetic and structural properties of eye lens ζ-crystallin: an in vitro and in silico approach.

The Arabian Camelus dromedarius contains significant concentration of eye lens ζ-crystallin. This enzyme is also present in other life forms including humans, however in lower catalytic amounts. The recombinant camel ζ-crystallin was expressed in the E. coli BL21 (DE3) pLysS strain and purified using HisTrap column. The Km of the enzyme for 9,10-phenanthrenequinone (9,10-PQ) substrate and NADPH cofactor was determined to be 11.66 and 50.93 µM, respectively. The Vmax for 9,10-PQ and NADPH was obtained as 23.19 and 19.98 µM min-1, respectively. The optimum activity of the purified enzyme was found to be at pH 6.0 and at 55 °C. Different physico-chemical parameters were analysed including instability index (II), aliphatic index (AI) and the GRAVY index to establish proper characterization. The sequence of the recombinant ζ-crystallin was subjected to homology modelling using SWISS-MODEL webserver followed by validation of the modelled target structure. The evaluation of the modelled ζ-crystallin was performed by several parameters including Ramachandran plot, Z-score values followed by molecular dynamics (MD) simulation. The cumulative analysis of the physico-chemical, quantitative, qualitative and the essential dynamics of simulation of ζ-crystallin and its complexes with 9,10-PQ and NADPH helped in verifying the acceptable quality and stability of the ζ-crystallin structure.Communicated by Ramaswamy H. Sarma.

Glycosphingolipids (GSLs) from Sphingomonas paucimobilis Increase the Efficacy of Liposome-Based Nanovaccine against Acinetobacter baumannii-Associated Pneumonia in Immunocompetent and Immunocompromised Mice.

Due to the high propensity of drug resistance in Acinetobacter baumannii, the number of currently available therapeutic drugs has become very limited. Thus, it becomes incredibly important to prepare an effective vaccine formulation capable of eliciting an effective immune response against A. baumannii. In this study, we prepared a liposomal vaccine formulation bearing glycosphingolipids (GSLs) from Sphingomonas paucimobilis and loaded with the whole cell antigen (WCAgs-GSLs-liposomes) of A. baumannii. The immune-stimulating potential and prophylactic efficacy of WCAgs-GSLs-liposomes were compared with those of WCAgs-liposomes (without GSLs) or free WCAgs in both immunocompetent and immunodeficient mice. The efficacy of vaccine formulations was determined by analyzing antibody titer, cytokine levels, and survival studies in the immunized mice. The findings revealed that vaccination with WCAgs-GSLs-liposomes stimulated a greater secretion of antibodies and cytokines, higher lymphocyte proliferation, and increased expression of the co-stimulatory molecules. Anti-sera from WCAgs-GSLs-liposomes-immunized mice remarkably reduced the biofilm formation by A. baumannii. Most importantly, WCAgs-GSLs-liposomes-vaccinated mice demonstrated a higher defiance against the pathogen, as compared to the immunizations with WCAgs-liposomes (without GSLs) or free WCAgs. Immunocompetent mice immunized with WCAgs-GSLs-liposomes showed a 100% survival rate, while those immunized with WCAgs-liposomes exhibited a 60% survival rate. The protective effect of WCAgs-GSLs-liposomes was also found to be higher in immunocompromised mice, as the immunized mice showed a 50% survival rate, which was greater than the 20% survival rate of those immunized with WCAgs-liposomes. The survival data was also supported by the findings of bacterial load and histological analysis that substantiated the greatest prophylactic potential of the WCAgs-GSLs-liposomes. These findings recommend that WCAgs-GSLs-liposomes may be reckoned as a prospective vaccine to protect the persons against A. baumannii infection.

Safety and Prophylactic Efficacy of Liposome-Based Vaccine against the Drug-Resistant Acinetobacter baumannii in Mice.

In recent years, the emergence of multidrug-resistant Acientobacter baumannii has greatly threatened public health and depleted our currently available antibacterial armory. Due to limited therapeutic options, the development of an effective vaccine formulation becomes critical in order to fight this drug-resistant pathogen. The objective of the present study was to develop a safe vaccine formulation that can be effective against A. baumannii infection and its associated complications. Here, we prepared liposomes-encapsulated whole cell antigens (Lip-WCAgs) as a vaccine formulation and investigated its prophylactic efficacy against the systemic infection of A. baumannii. The immunization with Lip-WCAgs induced the higher production of antigen-specific antibody titers, greater lymphocyte proliferation, and increased secretion of Th1 cytokines, particularly IFN-γ and IL-12. Antisera from Lip-WCAgs-immunized mice showed the utmost bactericidal activity and potently inhibited the biofilm formation by A. baumannii. Interestingly, Lip-WCAgs-induced immune response was translated in in vivo protection studies as the immunized mice exhibited the highest resistance to A. baumannii infection. Mice in the group immunized with Lip-WCAgs had an 80% survival rate and a bacterial burden of 5464 ± 1193 CFUs per gram of the lung tissue, whereas the mice immunized with IFA-WCAgs had a 50% survival rate and 51,521 ± 8066 CFUs. In addition, Lip-WCAgs vaccinated mice had lower levels of the inflammatory markers, including CRP, IL-6, IL-1ß, and TNF-α. The findings of this study suggest that Lip-WCAgs may be considered a potential vaccine formulation to protect individuals against A. baumannii infection.

Encapsulation of MERS antigen into α-GalCer-bearing-liposomes elicits stronger effector and memory immune responses in immunocompetent and leukopenic mice.

Objectives: Here, we prepared a liposome-based vaccine formulation containing Middle East Respiratory Syndrome Coronavirus papain-like protease (MERS-CoV-PLpro). Methods: A persistent leukopenic condition was induced in mice by injecting cyclophosphamide (CYP) three days before each dose of immunization. Mice were immunized on days 0, 14 and 21 with α-GalCer-bearing MERS-CoV PLpro-encapsulated DPPC-liposomes (α-GalCer-MERS-PLpro-liposomes or MERS-CoV PLpo-encapsulated DPPC-liposomes (MERS-PLpro-liposomes), whereas the antigen emulsified in Alum (MERS-PLpro-Alum) was taken as a control. On day 26, the blood was taken from the immunized mice to analyze IgG titer, whereas the splenocytes were used to analyze the lymphocyte proliferation and the level of cytokines. In order to assess the memory immune response, mice were given a booster dose after 150 days of the last immunization. Results: The higher levels of MERS-CoV-PLpro-specific antibody titer, IgG2a and lymphocyte proliferation were noticed in mice immunized with α-GalCer-MERS-PLpro-liposomes. Besides, the splenocytes from mice immunized with α-GalCer-MERS-PLpro-liposomes produced larger amounts of IFN-γ as compared to the splenocytes from MERS-PLpro-liposomes or MERS- PLpro-Alum immunized mice. Importantly, an efficient antigen-specific memory immune response was observed in α-GalCer-MERS-PLpro-liposomes immunized mice. Conclusions: These findings suggest that α-GalCer-MERS-PLpro-liposomes may substantiate to be a successful vaccine formulation against MERS-CoV infection, particularly in immunocompromised individuals.

Synthesis and Anti-Cancer Activity of New Pyrazolinyl-Indole Derivatives: Pharmacophoric Interactions and Docking Studies for Identifying New EGFR Inhibitors.

Newly designed series of indole-containing pyrazole analogs, pyrazolinylindoles, were synthesized, and their structures were confirmed based on the spectral data of the 1H NMR, 13C NMR, and HR-MS analyses. Preliminary anti-cancer activity testings were carried out by the National Cancer Institute, United States of America (NCI, USA). Compounds HD02, HD05, and HD12 demonstrated remarkable cytotoxic activities against nine categories of cancer types based cell line panels which included leukemia, colon, breast, melanoma, lungs, renal, prostate, CNS, and ovarian cancer cell lines. The highest cytotoxic effects were exhibited by the compounds HD02 [1-(5-(1-H-indol-3-yl)-3-(p-tolyl)-4,5-dihydro-1H-pyrazol-1-yl)-2-phenylethanone], HD05 [1-(3-(4-chlorophenyl)-5-(1H-indol-3-yl)-4,5-dihydro-1H-pyrazol-1-yl)-2-phenoxyethanone], and HD12 [(3-(4-chlorophenyl)-5-(1H-indol-3-yl)-4,5-dihydro-1H-pyrazol-1-yl)(pyridin-4-yl)methanone] against some of the 56 types of NCI-based cell lines in different panels. Compound HD05 showed the maximum range of cancer cell growth inhibitions against all categories of the cell lines in all nine panels. On average, in comparison to the referral standard, imatinib, at a dose level of 10 µM, the HD05 showed significant activity against leukemia in the range of 78.76%, as compared to the imatinib at 9% of cancer cells' growth inhibitions. Molecular docking simulation studies were performed in silico on the epidermal growth factor receptor (EGFR) tyrosine kinase, in order to validate the activity.

Lipid-Based Nanoparticle Formulation of Diallyl Trisulfide Chemosensitizes the Growth Inhibitory Activity of Doxorubicin in Colorectal Cancer Model: A Novel In Vitro, In Vivo and In Silico Analysis.

Garlic's main bioactive organosulfur component, diallyl trisulfide (DATS), has been widely investigated in cancer models. However, DATS is not suitable for clinical use due to its low solubility. The current study seeks to improve DATS bioavailability and assess its chemopreventive and chemosensitizing properties in an AOM-induced colorectal cancer model. The polyethylene glycol coated Distearoylphosphatidylcholine/Cholesterol (DSPC/Chol) comprising DATS-loaded DATSL and doxorubicin (DOXO)-encapsulated DOXL liposomes was prepared and characterized. The changes in the sensitivity of DATS and DOXO by DATSL and DOXL were evaluated in RKO and HT-29 colon cancer cells. The synergistic effect of DATSL and DOXL was studied by cell proliferation assay in the combinations of IC10, IC25, and IC35 of DATSL with the IC10 of DOXL. AOM, DATSL, and DOXL were administered to different groups of mice for a period of 21 weeks. The data exhibited ~93% and ~46% entrapment efficiency of DATSL and DOXL, respectively. The size of sham liposomes was 110.5 nm, whereas DATSL and DOXL were 135.5 nm and 169 nm, respectively. DATSL and DOXL exhibited significant sensitivity in the cell proliferation experiment, lowering their IC50 doses by more than 8- and 14-fold, respectively. However, the DATSL IC10, IC25, and IC35 showed escalating chemosensitivity, and treated the cells in combination with DOXL IC10. Analysis of histopathological, cancer marker enzymes, and antioxidant enzymes revealed that the high dose of DATSL pretreatment and DOXL chemotherapy is highly effective in inhibiting AOM-induced colon cancer promotion. The combination of DATSL and DOXL indicated promise as a colorectal cancer treatment in this study. Intermolecular interactions of DATS and DOXO against numerous cancer targets by molecular docking indicated MMP-9 as the most favourable target for DATS exhibiting binding energy of -4.6 kcal/mol. So far, this is the first research to demonstrate the chemopreventive as well as chemosensitizing potential of DATSL in an animal model of colorectal cancer.

Delivery of MERS antigen encapsulated in α-GalCer-bearing liposomes elicits stronger antigen-specific immune responses.

Alpha-Galactosylceramide (α-GalCer) effectively activates the natural killer T (NKT) cells to secrete remarkable amounts of Th1 and Th2 cytokines and therefore, acts as a potential immunoadjuvant in vaccine formulation. In the present study, we prepared α-GalCer-bearing or α-GalCer-free liposomes and loaded them with Middle East Respiratory Syndrome Coronavirus papain-like protease (α-GalCer-Lip-MERS-CoV PLpro or Lip-MERS-CoV PLpro). These formulations were injected in mice to investigate the antigen-specific humoral and cell-mediated immune responses. The immunisation with α-GalCer-Lip-MERS-CoV PLpro or Lip-MERS-CoV PLpro did not induce any notable toxicity in immunised mice. The results demonstrated that mice immunised with α-GalCer-Lip-MERS-CoV PLpro showed greater antigen-specific antibody titre, switching of IgG isotyping to IgG2a subclass and higher lymphocyte proliferation. Moreover, the splenocytes from α-GalCer-Lip-MERS-CoV PLpro immunised mice secreted greater levels of IFN-γ, IL-4, IL-2 and IL-12. Interestingly, a booster dose induced stronger memory immune responses in mice previously immunised with α-GalCer-Lip-MERS-CoV PLpro. In summary, α-GalCer-Lip-MERS-CoV PLpro may prove to be a promising vaccine formulation to protect the individuals against MERS-CoV infection.

In silico and in vitro studies on the inhibition of laccase activity by Ellagic acid: Implications in drug designing for the treatment of Cryptococcal infections.

In recent years, the increased frequency of drug-resistant strains of Cryptococcus neoformans has depleted our antifungal armory. In the present study, we investigated the inhibitory potential of ellagic acid (EA) against C. neoformans laccase through in silico and in vitro studies. For the first time, a homology modelling was established to model laccase and modelled protein served as a receptor for docking EA. Thermodynamic stability of the docked complex was ascertained by molecular dynamics simulation (MD). The analysis of root mean square deviation and fluctuation of alpha carbons of protein justifies the stability of the bound EA in the binding pocket of laccase. Frontier molecular orbitals of the EA was studied by density functional theory-based optimization by using the Lee-Yang-Parr correlation functional (B3LYP) approach. Negative values of the highest occupied/unoccupied molecular orbitals (HOMO/LUMO) indicated that laccase with EA forms a stable complex. Interestingly, EA inhibited laccase activity both in vitro and in yeast cells of C. neoformans. Moreover, EA treatment remarkably inhibited the proliferation of C. neoformans inside macrophages. The findings of the present study unveil the molecular basis of the interactions of laccase with EA, which may prove to be beneficial for designing laccase inhibitors as potential anti-cryptococcal agents.

The Effect of Liposomal Diallyl Disulfide and Oxaliplatin on Proliferation of Colorectal Cancer Cells: In Vitro and In Silico Analysis.

Diallyl disulfide (DADS) is one of the main bioactive organosulfur compounds of garlic, and its potential against various cancer models has been demonstrated. The poor solubility of DADS in aqueous solutions limits its uses in clinical application. The present study aimed to develop a novel formulation of DADS to increase its bioavailability and therapeutic potential and evaluate its role in combination with oxaliplatin (OXA) in the colorectal cancer system. We prepared and characterized PEGylated, DADS (DCPDD), and OXA (DCPDO) liposomes. The anticancer potential of these formulations was then evaluated in HCT116 and RKO colon cancer cells by different cellular assays. Further, a molecular docking-based computational analysis was conducted to determine the probable binding interactions of DADS and OXA. The results revealed the size of the DCPDD and DCPDO to be 114.46 nm (95% EE) and 149.45 nm (54% EE), respectively. They increased the sensitivity of the cells and reduced the IC50 several folds, while the combinations of them showed a synergistic effect and induced apoptosis by 55% in the cells. The molecular docking data projected several possible targets of DADS and OXA that could be evaluated more precisely by these novel formulations in detail. This study will direct the usage of DCPDD to augment the therapeutic potential of DCPDO against colon cancer in clinical settings.

Catalytic roles, immobilization and management of recalcitrant environmental pollutants by laccases: Significance in sustainable green chemistry.

We are facing a high risk of exposure to emerging contaminants and increasing environmental pollution with the concomitant growth of industries. Persistence of these pollutants is a major concern to the ecosystem. Laccases, also known as "green catalysts" are multi-copper oxidases which offers an eco-friendly solution for the degradation of these hazardous pollutants to less or non-toxic compounds. Although various other biological methods exist for the treatment of pollutants, the fact that laccases catalyze the oxidation of broad range of substrates in the presence of molecular oxygen without any additional cofactor and releases water as the by-product makes them exceptional. They have a good possibility of utilization in various industries, especially for the purpose of bioremediation. Besides this, they have also been used in medical/health care, food industry, bio-bleaching, wine stabilization, organic synthesis and biosensors. This review covers the catalytic behaviour of laccases, their immobilization strategies, potential applications in bioremediation of recalcitrant environmental pollutants and their engineering. It provides a comprehensive summary of most factors to consider while working with laccases in an industrial setting. It compares the benefits and drawbacks of the current techniques. Immobilization and mediators, two of the most significant aspects in working with laccases, have been meticulously discussed.

Liposome-Mediated Delivery of MERS Antigen Induces Potent Humoral and Cell-Mediated Immune Response in Mice.

The advancements in the field of nanotechnology have provided a great platform for the development of effective antiviral vaccines. Liposome-mediated delivery of antigens has been shown to induce the antigen-specific stimulation of the humoral and cell-mediated immune responses. Here, we prepared dried, reconstituted vesicles (DRVs) from DPPC liposomes and used them as the vaccine carrier system for the Middle East respiratory syndrome coronavirus papain-like protease (DRVs-MERS-CoV PLpro). MERS-CoV PLpro emulsified in the Incomplete Freund's Adjuvant (IFA-MERS-CoV PLpro) was used as a control. Immunization of mice with DRVs-MERS-CoV PLpro did not induce any notable toxicity, as revealed by the levels of the serum alanine transaminase (ALT), aspartate transaminase (AST), blood urea nitrogen (BUN) and lactate dehydrogenase (LDH) in the blood of immunized mice. Immunization with DRVs-MERS-CoV PLpro induced greater antigen-specific antibody titer and switching of IgG1 isotyping to IgG2a as compared to immunization with IFA-MERS-CoV PLpro. Moreover, splenocytes from mice immunized with DRVs-MERS-CoV PLpro exhibited greater proliferation in response to antigen stimulation. Moreover, splenocytes from DRVs-MERS-CoV PLpro-immunized mice secreted significantly higher IFN-γ as compared to splenocytes from IFA-MERS-CoV PLpro mice. In summary, DRVs-MERS-CoV PLpro may prove to be an effective prophylactic formulation to prevent MERS-CoV infection.

Antimicrobial, Immunomodulatory and Anti-Inflammatory Potential of Liposomal Thymoquinone: Implications in the Treatment of Bacterial Pneumonia in Immunocompromised Mice.

Acinetobacter baumannii has recently been increasing as an aggressive pathogen in immunocompromised persons. In the present study, we determined the in vitro antibacterial and anti-biofilm activity of thymoquinone (TQ) against A. baumannii. A liposomal formulation of TQ (Lip-TQ) was prepared and its therapeutic potential was investigated in the treatment of A. baumannii infection in immunocompromised mice. Leukopenia was induced in mice by injecting cyclophosphamide (CYP) at a dose of 200 mg/kg and the leukopenic mice were infected with 1 × 106 CFUs of A. baumannii. The effectiveness of free TQ or Lip-TQ against A. baumannii infection was assessed by analyzing the survival rate and bacterial burden. Moreover, the efficacy of Lip-TQ was also studied by examining the systemic inflammatory markers and the histological changes in the lung tissues. The results showed that the mice in the group treated with Lip-TQ at a dose of 10 mg/kg exhibited a 60% survival rate on day 40 post-infection, whereas all the mice treated with free TQ at the same dose died within this duration. Likewise, the lowest bacterial burden was found in the lung tissue of mice treated with Lip-TQ (10 mg/kg). Besides, Lip-TQ treatment remarkably alleviated the infection-associated inflammation, oxidative stress, and histological changes in the lung tissues. Based on the findings of the present study, we recommend considering Lip-TQ as a valuable therapeutic formulation in the treatment of A. baumannii-associated pneumonia in immunocompromised subjects.

A comparative study based on activity, conformation and computational analysis on the inhibition of human salivary aldehyde dehydrogenase by phthalate plasticizers: Implications in assessing the safety of packaged food items.

Phthalate plasticizers are commonly used in various consumer-end products. Human salivary aldehyde dehydrogenase (hsALDH) is a detoxifying enzyme which defends us from the toxic aldehydes. Here, the effect of phthalates [Di-2-ethylhexyl phthalate (DEHP), Diethyl phthalate (DEP) and Dibutyl phthalate (DBP)] on hsALDH has been investigated. These plasticizers inhibited hsALDH, and the IC50 values were 0.48 ± 0.04, 283.20 ± 0.09 and 285.00 ± 0.14 µM for DEHP, DEP and DBP, respectively. DEHP was the most potent inhibitor among the three plasticizers. They exhibited mixed-type linear inhibition with inclination towards competitive-non-competitive inhibition. They induced both tertiary and secondary structural changes in the enzyme. Quenching of intrinsic hsALDH fluorescence in a constant manner was observed with a binding constant (Kb) of 8.91 × 106, 2.80 × 104, and 1.31 × 105 M-1, for DEHP, DEP and DBP, respectively. Computational analysis showed that these plasticizers bind stably in the proximity of hsALDH catalytic site, reciprocating via non-covalent interactions with some of the amino acids which are evolutionary conserved. Therefore, exposure to these plasticizers inhibits hsALDH which increases the risk of aldehyde induced toxicity, adversely affecting oral health. The study has implications in assessing the safety of packaged food items which utilize phthalates.

Role of NKT Cells during Viral Infection and the Development of NKT Cell-Based Nanovaccines.

Natural killer T (NKT) cells, a small population of T cells, are capable of influencing a wide range of the immune cells, including T cells, B cells, dendritic cells and macrophages. In the present review, the antiviral role of the NKT cells and the strategies of viruses to evade the functioning of NKT cell have been illustrated. The nanoparticle-based formulations have superior immunoadjuvant potential by facilitating the efficient antigen processing and presentation that favorably elicits the antigen-specific immune response. Finally, the immunoadjuvant potential of the NKT cell ligand was explored in the development of antiviral vaccines. The use of an NKT cell-activating nanoparticle-based vaccine delivery system was supported in order to avoid the NKT cell anergy. The results from the animal and preclinical studies demonstrated that nanoparticle-incorporated NKT cell ligands may have potential implications as an immunoadjuvant in the formulation of an effective antiviral vaccine that is capable of eliciting the antigen-specific activation of the cell-mediated and humoral immune responses.

Liposomal Ellagic Acid Alleviates Cyclophosphamide-Induced Toxicity and Eliminates the Systemic Cryptococcus neoformans Infection in Leukopenic Mice.

Cryptococcus neoformans infections rose sharply due to rapid increase in the numbers of immunocompromised individuals in recent years. Treatment of Cryptococcosis in immunocompromised persons is largely very challenging and hopeless. Hence, this study aimed to determine the activity of ellagic acid (EA) in the treatment of C. neoformans in cyclophosphamide injected leukopenic mice. A liposomal formulation of ellagic acid (Lip-EA) was prepared and characterized, and its antifungal activity was assessed in comparison to fluconazole (FLZ). The efficacy of the drug treatment was tested by assessing survival rate, fungal burden, and histological analysis in lung tissues. The safety of the drug formulations was tested by investigating hepatic, renal function, and antioxidant levels. The results of the present work demonstrated that Lip-EA, not FLZ, effectively eliminated C. neoformans infection in the leukopenic mice. Mice treated with Lip-EA (40 mg/kg) showed 70% survival rate and highly reduced fungal burden in their lung tissues, whereas the mice treated with FLZ (40 mg/kg) had 20% survival rate and greater fungal load in their lungs. Noteworthy, Lip-EA treatment alleviated cyclophosphamide-induced toxicity and restored hepatic and renal function parameters. Moreover, Lip-EA treatment restored the levels of superoxide dismutase and reduced glutathione and catalase in the lung tissues. The effect of FLZ or EA or Lip-EA against C. neoformans infection was assessed by the histological analysis of lung tissues. Lip-EA effectively reduced influx of inflammatory cells, thickening of alveolar walls, congestion, and hemorrhage. The findings of the present study suggest that Lip-EA may prove to be a promising therapeutic formulation against C. neoformans in immunocompromised persons.

Coadministration of Ginger Extract and Fluconazole Shows a Synergistic Effect in the Treatment of Drug-Resistant Vulvovaginal Candidiasis.

BACKGROUND: Azoles are the most common antifungal drugs used in the treatment of vulvovaginal candidiasis (VVC). The frequency of azole-resistant Candida isolates has increased dramatically in the last two decades. Here, we assessed the antifungal activity of a combination of fluconazole (FLZ) and methanolic extract of ginger (Meth-Gin) against drug-resistant vulvovaginal candidiasis (VVC) in a murine model. METHODS: The in vitro activity of FLZ or a combination of FLZ and Meth-Gin was determined against Candida albicans by the agar well diffusion, macrodilution, time-kill and the biofilm eradication methods. The therapeutic efficacy of the formulations was assessed by analyzing the fungal load, pro-inflammatory cytokines, percent apoptotic cells and the histological changes in the vaginal tissues of the mice. Moreover, the renal toxicity the drug formulation was evaluated by analyzing the levels of the blood urea nitrogen (BUN) and creatinine. RESULTS: The results of in vitro study demonstrated that FLZ did not show any activity against C. albicans, whereas a combination of FLZ and Meth-Gin demonstrated greater activity as shown by the data of the zone of growth inhibition, MIC and time-kill assay. FLZ or Meth-Gin treatment could not completely cure VVC, whereas a combination of FLZ and Meth-Gin was greatly effective in the treatment of VVC. The vaginal tissue from mice of the infected control group had the highest fungal load of 155370 ± 20617 CFUs. Treatment with FLZ at a dose of 40 mg/kg reduced the fungal load to 120863 ± 10723 CFUs. Interestingly, the mice treated with a combination of FLZ (40 mg/kg) and Meth-Gin (200 mg/kg) had a fungal load of 256 ± 152 CFUs. Besides, FLZ and Meth-Gin combination effectively reduced the pro-inflammatory cytokines (IL-1ß, TNF-α and IL-17) and the percentage of apoptotic cells in the vaginal tissues. Likewise, the histological analysis revealed the epithelial necrosis, shedding and ulceration in the vaginal tissue, whereas treatment with FLZ and Meth-Gin combination reversed the histopathological changes in the vaginal epithelium and lamina propria. CONCLUSION: The findings of the current study suggest that the co-administration of Meth-Gin and FLZ may have a potential therapeutic effect in the treatment of azole-resistant candidiasis.

Coadministration of liposomal methylglyoxal increases the activity of amphotericin B against Candida albicans in leukopoenic mice.

In this study, we investigated the therapeutic efficacy of a combination of liposomal amphotericin B (Lip-Amp B) and Methylglyoxal (Lip-MG) against Candida albicans in the leukopoenic mice. The antifungal efficacy of Lip-Amp B or Lip-MG or a combination of Lip-Amp B and Lip-MG was evaluated by the analysis of the survival rate and the fungal load in the treated mice. The immune-stimulatory effect of Lip-MG on macrophages was evaluated by analysing the secretion of proinflammatory cytokines. C. albicans infected mice treated at the doses of 1 and 2 mg/kg of Lip-Amp B showed 20% and 50% survival rates, respectively. Whereas the mice treated with free Amp B at the same doses died within 40 days of treatment. Interestingly, C. albicans infected mice treated with a combination of Lip-Amp B and Lip-MG had 70% survival rate on day 40 postinfection. Moreover, treatment of macrophages with Lip-MG increased their fungicidal activity and the secretion of proinflammatory cytokines, including TNF-α and IL-1ß. These findings suggested that co-treatment with Lip-Amp B and Lip-MG had a synergistic effect and could be effective against C. albicans in immunocompromised subjects.

Potential Implications of Black Seed and its Principal Constituent Thymoquinone in the Treatment of COVID-19 Patients.

BACKGROUND: Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is the causative agent of the current pandemic of Coronavirus Disease-2019 (COVID-19). The progression of COVID-19 is related to an excessive host inflammatory immune response to SARS-CoV-2 infection, which is considered a major cause of disease severity and death. Dysregulated immune response produces huge amounts of pro-inflammatory cytokines and chemokines called "cytokine storm". Moreover, the activation of microthrombi formation plays an important role in multiple organ failure. METHODS: Keeping into consideration the potent anti-inflammatory activity of black seed and its major constituent Thymoquinone (TQ), we hypothesize their potential implication in the treatment of COVID-19 patients. A literature search was performed in PubMed, ScienceDirect, Google Scholar and Scopus electronic databases using the terms, including black seed, N. sativa, thymoquinone, SARSCoV- 2, COVID-19 and inflammatory immune response. RESULTS: Various studies confirmed that Black seed and TQ reduced the thrombus formation, the expression of tissue factor and the immune activation. Furthermore, TQ demonstrated the broad-spectrum antimicrobial activity that may be effective in controlling the secondary infections in COVID-19 patients. CONCLUSION: Keeping into consideration the multi-targeting nature of the black seed and TQ, they may be used as a potential therapeutic formulation or as an adjunct therapy in the treatment of COVID-19 patients.