Multilevel Approach for the Treatment of Giardiasis by Targeting Arginine Deiminase.

Giardiasis represents a latent problem in public health due to the exceptionally pathogenic strategies of the parasite Giardia lamblia for evading the human immune system. Strains resistant to first-line drugs are also a challenge. Therefore, new antigiardial therapies are urgently needed. Here, we tested giardial arginine deiminase (GlADI) as a target against giardiasis. GlADI belongs to an essential pathway in Giardia for the synthesis of ATP, which is absent in humans. In silico docking with six thiol-reactive compounds was performed; four of which are approved drugs for humans. Recombinant GlADI was used in enzyme inhibition assays, and computational in silico predictions and spectroscopic studies were applied to follow the enzyme's structural disturbance and identify possible effective drugs. Inhibition by modification of cysteines was corroborated using Ellman's method. The efficacy of these drugs on parasite viability was assayed on Giardia trophozoites, along with the inhibition of the endogenous GlADI. The most potent drug against GlADI was assayed on Giardia encystment. The tested drugs inhibited the recombinant GlADI by modifying its cysteines and, potentially, by altering its 3D structure. Only rabeprazole and omeprazole decreased trophozoite survival by inhibiting endogenous GlADI, while rabeprazole also decreased the Giardia encystment rate. These findings demonstrate the potential of GlADI as a target against giardiasis.

Multi-targeted Effect of Nicotinamide Mononucleotide on Brain Bioenergetic Metabolism.

Dysfunctions in NAD+ metabolism are associated with neurodegenerative diseases, acute brain injury, diabetes, and aging. Loss of NAD+ levels results in impairment of mitochondria function, which leads to failure of essential metabolic processes. Strategies to replenish depleted NAD+ pools can offer significant improvements of pathologic states. NAD+ levels are maintained by two opposing enzymatic reactions, one is the consumption of NAD+ while the other is the re-synthesis of NAD+. Inhibition of NAD+ degrading enzymes, poly-ADP-ribose polymerase 1 (PARP1) and ectoenzyme CD38, following brain ischemic insult can provide neuroprotection. Preservation of NAD+ pools by administration of NAD+ precursors, such as nicotinamide (Nam) or nicotinamide mononucleotide (NMN), also offers neuroprotection. However, NMN treatment demonstrates to be a promising candidate as a therapeutic approach due to its multi-targeted effect acting as PARP1 and CD38 inhibitor, sirtuins activator, mitochondrial fission inhibitor, and NAD+ supplement. Many neurodegenerative diseases or acute brain injury activate several cellular death pathways requiring a treatment strategy that will target these mechanisms. Since NMN demonstrated the ability to exert its effect on several cellular metabolic pathways involved in brain pathophysiology it seems to be one of the most promising candidates to be used for successful neuroprotection.

Comparative Effects of Alkaloid Extracts from Aframomum melegueta (Alligator Pepper) and Aframomum danielli (Bastered Melegueta) on Enzymes Relevant to Erectile Dysfunction.

Aframomum melegueta (alligator pepper (AP)) and Aframomum danielli (bastered melegueta (BM)) seeds have been known to improve sexual function in folkloric medicine. This study investigates the effects of AP and BM seeds' alkaloid extracts on the activities of enzymes (acetylcholinesterase (AChE), angiotensin-1-converting enzyme (ACE), phosphodiesterase-5 (PDE-5), and arginase) relevant to erectile dysfunction (ED). Alkaloids from the seeds were prepared by the solvent extraction method and their interactions with AChE, ACE, PDE-5, and arginase were assessed. Gas chromatographic (GC) analyses of the extracts were also performed. The results revealed that the extracts inhibited the enzymes in a concentration-dependent manner. However, alkaloid extract from AP seed had higher AChE (IC50 = 5.42 µg/mL) and ACE (IC50 = 12.57 µg/mL) but lower PDE-5 (IC50 = 33.80 µg/mL) and arginase (IC50 = 31.36 µg/mL) inhibitory effects when compared to that of BM extract (AChE, IC50 = 42.00; ACE, IC50 = 60.67, PDE-5, IC50 = 7.24; and arginase, IC50 = 2.53 µg/mL). The GC analyses revealed the presence of senkirkine, angustifoline, undulatine, myristicin, safrole, lupanine, powelle, and indicine-N-oxide, among others. The inhibition of these enzymes could be the possible mechanisms by which the studied seeds were being used in managing ED in folklores. Nevertheless, the seed of AP exhibited higher potentials.

Effects of cold storage and 1-methylcyclopropene treatments on ripening and cell wall degrading in rabbiteye blueberry (Vaccinium ashei) fruit.

The effect of postharvest 1-methylcyclopropene and/or cold storage application on texture quality parameters during storage was determined. The changes in fruit quality (including weight loss, firmness, total soluble solids content, and ethylene production), cell wall material (including water-soluble fraction, ethylenediaminetetraacetic acid-soluble fraction, Na2CO3-soluble fraction, 4% KOH-soluble fraction, and 14% KOH-soluble fraction), and cell wall hydrolase activities (including polygalacturonase, endo-1,4-beta-D-glucanase, pectinesterase, alpha-L-arabinofuranosidase, and beta-galactosidase) were periodically measured up to 25 days after postharvest treatments. The application of cold storage reduced weight loss, ethylene production, and delayed ripening of blueberry fruit. The inhibition of senescence was associated with suppressed increase in cell wall hydrolase activities and retarded solubilization of pectins and hemicelluloses. Furthermore, no obvious differences in firmness, weight loss, ethylene production, and cell wall hydrolase activities between fruits with or without 1-methylcyclopropene application were observed, while significant lower levels of the detected parameters were found in cold storage fruit compared with fruit stored in room temperature. Thus, cold storage can be viewed as an effective means to extend the shelf life of blueberry fruit.

Ameliorative prospective of alpha-mangostin, a xanthone derivative from Garcinia mangostana against beta-adrenergic cathecolamine-induced myocardial toxicity and anomalous cardiac TNF-alpha and COX-2 expressions in rats.

Altered membrane integrity and inflammation play a key role in cardiovascular damage. We investigated the salubrious effect of exogenously administered alpha-mangostin against beta-adrenergic cathecolamine-induced cardiovascular toxicity with special reference to membrane ATPases, lysosomal hydrolases and inflammatory mediators TNF-alpha and Cyclooxygenase-2 (COX-2) expressions in albino rats. Induction of rats with isoproterenol (150mg/kg body wt, i.p.) for 2 days resulted in a significant increase in the activities of serum and cardiac lysosomal hydrolases (beta-d-glucuronidase, beta-d-galactosidase, beta-d-N-acetylglucosaminidase, acid phosphatase and cathepsin-D). A significant increase in cardiac levels of sodium, calcium with a decrease in the level of potassium paralleled by abnormal activities of membrane-bound phosphatases (Na(+)-K(+) ATPase, Ca(2+) ATPase and Mg(2+) ATPase) were observed in the heart of ISO-administered rats. Cardiac TNF-alpha and COX-2 expressions were assessed by Western blotting. Cardiac TNF-alpha and COX-2 expressions were significantly elevated in ISO-intoxicated rats. Pre-co-treatment with alpha-mangostin (200mg/kg body wt.) orally for 8 days significantly attenuated these abnormalities and restored the levels to near normalcy when compared to ISO intoxicated group of rats. In conclusion, alpha-mangostin preserves the myocardial membrane integrity and extenuates anomalous TNF-alpha and COX-2 expressions by mitigating ISO-induced oxidative stress and cellular damage effectively. Restoration of cellular normalcy accredits the cytoprotective role of alpha-mangostin.

Gene expression profiling in spleens of deoxynivalenol-exposed mice: immediate early genes as primary targets.

Exposure to the trichothecene mycotoxin deoxynivalenol (DON) alters immune functions in vitro and in vivo. To gain further insight into DON's immunotoxic effects, microarrays were used to determine how acute exposure to this mycotoxin modulates gene expression profiles in murine spleen. B6C3F1 mice were treated orally with 25mg/kg body weight DON, and 2h later spleens were collected for macroarray analysis. Following normalization using a local linear regression model, expression of 116 out of 1176 genes was significantly altered compared to average expression levels in all treatment groups. When genes were arranged into an ontology tree to facilitate comparison of expression profiles between treatment groups, DON was found primarily to modulate genes associated with immunity, inflammation, and chemotaxis. Real-time polymerase chain reaction was used to confirm modulation for selected genes. DON was found to induce the cytokines interleukin (IL)-1alpha, IL-1beta, IL-6 and IL-11. In analogous fashion, DON upregulated expression of the chemokines macrophage inhibitory protein-2 (MIP-2), cytokine-induced chemoattractant protein-1 (CINC-1), monocyte chemoattractant protein (MCP)-1, MCP-3, and cytokine-responsive gene-2 (CRG-2). c-Fos, Fra-, c-Jun, and JunB, components of the activator protein-1 (AP-1) transcription factor complex, were induced by DON as well as another transcription factor, NR4A1. Four hydrolases were found to be upregulated by DON, including mitogen-activated protein kinase phosphatase 1 (MKP1), catalytic subunit beta isoform (CnAbeta), protein tyrosine phosphatase receptor type J (Ptprj), and protein tyrosine phosphatase nonreceptor type 8 (Ptpn8), whereas three other hydrolases, microsomal epoxide hydrolase (Eph) 1, histidine triad nucleotide binding protein (Hint), and proteosome subunit beta type 8 (Psmb8) were significantly decreased by the toxin. Finally, cysteine-rich protein 61 (CRP61) and heat-shock protein 40 (Hsp40), genes associated with signaling, were increased, while Jun kinase 2 (JNK2) was decreased. Taken together, data suggest that DON upregulated the expression of multiple immediate early genes, many of which are likely to contribute to the complex immunological effects reported for this and other trichothecenes.

In vitro characterization of a phosphate starvation-independent carbon-phosphorus bond cleavage activity in Pseudomonas fluorescens 23F.

A novel, metal-dependent, carbon-phosphorus bond cleavage activity, provisionally named phosphonoacetate hydrolase, was detected in crude extracts of Pseudomonas fluorescens 23F, an environmental isolate able to utilize phosphonoacetate as the sole carbon and phosphorus source. The activity showed unique specificity toward this substrate; its organic product, acetate, was apparently metabolized by the glyoxylate cycle enzymes of the host cell. Unlike phosphonatase, which was also detected in crude extracts of P. fluorescens 23F, phosphonoacetate hydrolase was inducible only in the presence of its sole substrate and did not require phosphate starvation.