On the genetic diversity of spiny mice (genus Acomys) and gerbils (genus Gerbillus) in the Arabian Peninsula

Using non-destructive sampling we provide further genetic characterisations for spiny mice (Acomys dimidiatus/cahirinus) and gerbils (Gerbillus sp.) in three regions in Saudi Arabia. All individuals were sequenced for a fragment of the cytochrome b gene, and compared against available conspecifics and closely related taxa. We confirm the existence of a second Acomys dimidiatus/cahirinus lineage specific to the Arabian Peninsula as seen previously. The Arabian Gerbillus nanus is shown to group with Middle Eastern rather than African conspecifics. A second cryptic Gerbillus lineage was also sampled across multiple locations, which may be an uncharacterised G. dasyurus.     

Assessment of the potential iridology for diagnosing kidney disease using wavelet analysis and neural networks

Alternative or complementary medicine emphasizes therapies that are claimed to improve quality of life, prevent disease, and address conditions that conventional medicine has limited success in curing. There are many techniques which are prevalent in many countries and these can cause harm if not scientifically evaluated. The objective of this paper is to validate the use of iridology to diagnose kidney abnormalities. Two subject groups were evaluated: one was 168 subjects free from kidney disease and the other was 172 subjects with chronic renal failure. The procedure to acquire, process and classify the iris images was designed in such a way that avoids any dependency on the iridologists by using wavelet analysis and Adaptive Neuro-Fuzzy Inference System. The results show a correct classification for both subjects with kidney problems and normal subjects of 82% and 93%, respectively. The proposed technique conducted on a systemic disease with ocular manifestations showed encouraging results. However, it is necessary to perform extensive studies with diseases that do not have ocular manifestations according to conventional medicine in order to validate iridology as a valid scientific technique.     

Solid lipid nanoparticles for transdermal delivery of avanafil: optimization,formulation, in-vitro and ex-vivo studies

Context: Avanafil (AVA) is used in the treatment of erectile dysfunction, but is reported for its poor aqueous solubility. Solid lipid nanoparticles (SLNs) are lipid carriers that can greatly enhance drug solubility and bioavailability.

Objective: This work was aimed to formulate and optimize AVA SLNs with subsequent loading into hydrogel films for AVA transdermal delivery.

Materials and methods: AVA SLNs were prepared utilizing homogenization followed by ultra-sonication technique. The prepared SLNs were characterized for particle size, charge, surface morphology and drug content. The optimized SLNs formulation was incorporated into transdermal films prepared using HPMC and chitosan. Hydrogel films were evaluated for ex-vivo rat skin permeation using automated Franz diffusion cells. The permeation parameters and the release mechanism were evaluated. The transdermal permeation of the prepared AVA SLNs through the skin layers was studied using confocal laser scanning microscope.

Results: Lipid concentration and % of oil in lipid had a pronounced effect on particle size while, entrapment efficiency was significantly affected by lipid concentration and % of cholesterol. The optimized AVA SLNs showed particle size and entrapment efficiency of 86?nm and 85.01%, respectively. TEM images revealed spherecity of the particles. High permeation parameters were observed from HPMC films loaded with AVA SLNs. The release data were in favor of Higuchi diffusion model. The prepared AVA SLNs were able to penetrate deeper in skin layers.

Conclusion: HPMC transdermal film-loaded AVA SLNs is an effective and alternative to per-oral drug administration.     

Hermetia illucens L. larvae–associated intestinal microbes reduce the transmission risk of zoonotic pathogens in pig manure

Black soldier fly (BSF) larvae are considered a promising biological reactor to convert organic waste and reduce the impact of zoonotic pathogens on the environment. We analysed the effects of BSF larvae on Staphylococcus aureus and Salmonella spp. populations in pig manure (PM), which showed that BSF larvae can significantly reduce the counts of the associated S. aureus and Salmonella spp. Then, using a sterile BSF larval system, we validated the function of BSF larval intestinal microbiota in vivo to suppress pathogens, and lastly, we isolated eight bacterial strains from the BSF larval gut that inhibit S. aureus. Results indicated that functional microbes are essential for BSF larvae to antagonise S. aureus. Moreover, the analysis results of the relationship between the intestinal microbiota and S. aureus and Salmonella spp. showed that Myroides, Tissierella, Oblitimonas, Paenalcalignes, Terrisporobacter, Clostridium, Fastidiosipila, Pseudomonas, Ignatzschineria, Savagea, Moheibacter and Sphingobacterium were negatively correlated with S. aureus and Salmonella. Overall, these results suggested that the potential ability of BSF larvae to inhibit S. aureus and Salmonella spp. present in PM is accomplished primarily by gut-associated microorganisms.     

Synthesis of novel 4- and 5-substituted benzyl ether derivatives of vesamicol and in vitro evaluation of their binding properties to the vesicular acetylcholine transporter site

Detection of the central cholinergic deficits, a consistent feature of Alzheimer's disease, is essential to allow preventive measures and/or symptomatic treatment already at a very early stage of the disease. The vesicular acetylcholine transporter (VAChT) represents an appropriate target to establish PET radiotracer that are adequate for brain imaging the loss of cholinergic terminals. Here we describe the synthesis and binding characteristics of novel derivatives of vesamicol, known to represent a specific antagonist of VAChT sites. Novel benzyl ether derivatives of vesamicol either 4- or 5-substituted at the cyclohexylring have been synthesized by different regioselective ring opening reactions of a same epoxide precursor. The affinity and selectivity of the novel compounds to VAChT sites were analyzed by competitive radioligand binding studies in rat brain and liver membrane preparations using tritium labeled radioligands. The 4-substituted fluorobenzylether of vesamicol 10b was shown to exhibit a high affinity to VAChT sites (K(i)-value(10b)=10.7+/-1.7 nM), but demonstrated also binding capacities to sigma receptors (K(i-)value(10b)=18.5+/-6.9 nM, [(3)H]DTG; K(i)-value(10b)=30.6+/-9.6 nM, [(3)H]haloperidol). The data suggest the potential of vesamicol derivatives to design appropriate radiotracer for PET imaging of central cholinergic deficits.     

Novel branched poly(ethylenimine)-cholesterol water-soluble lipopolymers for gene delivery

A novel water-soluble lipopolymer was synthesized by linking cholesteryl chloroformate to the secondary amino groups of branched poly(ethylenimine) (PEI) of 1,800 and 10,000 Da. Conjugation through PEI secondary amines gives this newly synthesized lipopolymer (abbreviated as PEI-Chol) special advantage over our previously synthesized lipopolymers, which utilized the primary amino groups for conjugation, as the primary amino groups have a significant role in DNA condensation. Also, significantly, only one cholesterol molecule was grafted onto each PEI molecule (confirmed by (1)H NMR and MALDI-TOF mass spectrometry), leaving enough space for the steric interactions of the PEI's primary amines with the DNA. The PEI-Chol lipopolymer was characterized for the critical micellar concentration (cmc), buffer capacity, DNA condensation (by band retardation and circular dichroism), in vitro transfection efficiency, and cell viability. The cmcs of PEI-Chol 1,800 and PEI-Chol 10,000 were 496.6 and 1,330.5 microg/mL, respectively. The acid-base titration indicated high buffering capacity of the polymers around the pH range of 5-7, which indicated their potential for buffering in the acidic pH environment of the endosomes. The band retardation studies indicated that efficient condensation of the plasmid DNA could be achieved using these lipopolymers. The circular dichroism spectra indicated a change in DNA conformation and adoption of lower energy state upon condensation with these lipopolymers when an N/P ratio of 2.5/1 or above was formulated. The mean particle size of these complexes was in the range 110-205 nm, except for the complexes prepared using PEI of 1,800 Da, which had a mean particle size of 384 +/- 300 nm. The zeta potential of DNA complexes prepared using PEI-Chol 1,800, PEI-Chol 10,000 and PEI of 1,800, 10,000, and 25,000 Da at an N/P ratio of 15/1 was in the range 23-30 mV and was dependent on the N/P ratios. The in vitro transfection of PEI-Chol/pCMS-EGFP complexes in Jurkat cells showed high levels of expressed Green Fluorescent Protein (GFP) with little toxicity as determined by flow cytometry. These novel water-soluble lipopolymers provided good transfection efficiency with other desirable characteristics such as water solubility, free primary amino groups for efficient DNA condensation and high buffer capacity that indicated the possibility of efficient endosomal release.     

Effects of benzo[a]pyrene on tissue activities of metabolizing enzymes and antioxidant system in normal and protein-malnourished rats

The effects of benzo[a]pyrene (B[a]P) on some drug-metabolizing and antioxidant systems in liver, lung, and stomach were investigated in normal and protein malnutrition (PM) rats. PM significantly inhibited tissue glutathione (GSH) content and increased hepatic lipid peroxidation. Cytochrome P450 isoform CYP1A1 was significantly increased in various tissues (42-73%). Also, lung glutathione S-transferase (GST) activity was significantly decreased (19%) in PM rats. On the other hand, B[a]P significantly induced tissue GSH of control and PM rats. Also, hepatic lipid peroxidation were significantly increased in control rats treated with B[a]P. Superoxide dismutase (SOD) activity was decreased by B[a]P treatment in PM rat stomach. B[a]P significantly induced both quinone reductase (QR) (in all tissues) and hepatic GST of control and PM rats. GST activity in PM rat liver was significantly higher than that of control rat liver after B[a]P treatment. Also, B[a]P induced hepatic CYP1A1 by 32-fold and 27-fold (P < or = 0.05) in control and PM rats, respectively. Stomach and hepatic UDP-glucuronosyltransferase activities were significantly decreased (34%) and increased (74%), respectively by B[a]P in PM rats. The results suggest that PM status has a modifying effect on the response of some antioxidant and metabolizing systems to a well-known carcinogen risk.     

Antibody-Dependent Cellular Cytotoxicity Directed against Cells Expressing Human Immunodeficiency Virus Type 1 Envelope of Primary or Laboratory-Adapted Strains by Human and Chimpanzee Monoclonal Antibodies of Different Epitope Specificities

The characteristics of antibody-dependent cellular cytotoxicity (ADCC) directed by a panel of human and chimpanzee antienvelope (anti-Env) monoclonal antibodies (MAbs) of different epitope specificities were studied; this was accomplished by using target cells expressing human immunodeficiency virus type 1 (HIV-1) Envs of either primary or laboratory-adapted strains. Human MAbs of similar apparent affinities (1 × 10⁹ to 2 × 10⁹ liters/mol) against either a “cluster II”-overlapping epitope of gp41 or against the CD4 binding site, V3 loop, or C5 domain of gp120 directed substantial and comparable levels of specific lysis against targets infected with laboratory-adapted strains of HIV-1. As expected, those MAbs specific for relatively conserved regions of Env generally exhibited ADCC activity against a broader range of HIV-1 strains than those directed against variable epitopes. Significant ADCC activities of selected MAbs against primary isolate Env-expressing cells were demonstrated. In addition, a new ADCC epitope in the V2 domain of gp120 was defined. CD56⁺ cells were demonstrated to be the effector cells in these studies by fluorescence-activated cell sorting followed by ADCC assays. Notably, all anti-Env MAbs tested in this study, including MAbs directed against each of the known neutralization epitope clusters in gp120, directed significant levels of ADCC against targets expressing Env of one or more HIV-1 strains. These results imply that many, if not most, HIV-1-neutralizing human Abs of high affinity (≥3 × 10⁸ liters/mol in these studies) and of the immunoglobulin G1 (IgG1) subclass (i.e., the predominate IgG subclass) are capable of directing ADCC. Since neutralizing Abs have been associated with long-term survival following HIV-1 infection, this suggests that ADCC activity may be beneficial in vivo.The in vivo role(s) of antibodies (Abs) that can direct antibody-dependent cellular cytotoxicity (ADCC) against human immunodeficiency virus type 1 (HIV-1) Env-expressing cells in vitro remains unclear. In ADCC, anti-Env Abs direct effector cells to kill target cells bearing HIV-1 envelope on their surfaces; this is accomplished via specific binding of the Abs’ antigen-binding sites to Envs and their Fc regions to Fc receptors on the effector cells. Broadly strain reactive, ADCC-directing Abs arise early in the immune response to HIV-1 infection in vivo (14) and may be partially responsible for the initial clearance of viremia.Earlier in the HIV-1 epidemic, concerns were raised that shed soluble gp120 in HIV-1-infected individuals might bind to CD4⁺ cells, including uninfected ones, and could target these cells for “innocent bystander” killing by ADCC (6). However, effector cells armed with serum Abs able to direct ADCC in vitro against either innocent bystanders or HIV-1-infected cells were found at highest frequency in asymptomatic, seropositive individuals; patients with AIDS-related complex and AIDS showed progressively diminished reactivities (20). Furthermore, in a recent study (1), the ability of monoclonal Abs (MAbs) against three distinct gp120 epitopes to direct ADCC against uninfected CD4⁺ cells to which rgp120_SF2 had been adsorbed (i.e., innocent bystanders) was demonstrated to be less efficient by at least an order of magnitude than their ability to direct ADCC against HIV-1-infected cells.The existing data from in vivo studies (reviewed in reference 1) supports the efficacy, rather than the pathogenicity, of ADCC-directing Abs against HIV-1. Consistent with this data is our recent characterization of two MAbs, 42F and 43F, isolated from a long-term survivor of HIV-1 infection (1); these MAbs directed significant levels of ADCC and defined a new, conserved ADCC epitope in the C5 domain of HIV-1 gp120. Preliminary evidence indicated that concentrations of 42F- and 43F-like Abs in the serum of the donor were in the range required to direct high levels of ADCC, and these MAbs were shown to bind both oligomeric primary-isolate and laboratory-adapted Env efficiently (1).Because of the potential importance of ADCC-directing Abs against HIV-1, in this study we have evaluated ADCC directed against cells expressing HIV-1 Envs of primary or laboratory-adapted strains by a panel of human and chimpanzee anti-Env MAbs of different epitope specificities. Significant ADCC activities of selected MAbs against primary-isolate Env-expressing cells were demonstrated, and a new ADCC epitope in the V2 domain of gp120 was defined. Finally, a MAb’s ability to direct ADCC against a specific target cell type was shown to be dependent on additional factors beyond its ability to efficiently bind antigen on the target cell and its possession of an Fc region of the appropriate isotype to engage FcγR on effector cells.     

Effect of plant growth regulators on in vitro fiber development from unfertilized and fertilized Egyptian cotton ovules

Unfertilized and fertilized ovules of Gossypium barbadense Giza 45 (extra long staple variety) were used to study the effect of plant growth substances (auxins, gibberellins and cytokinins) on in vitro fiber initiation and development. Kinetin, alone did not increase total fiber unit (TFU) of unfertilized ovules, while an increase in TFU value occurred when a constant level of IAA and GA₃ were used either separately or in combination in the liquid medium. GA₃ used alone, produced a higher TFU value than that produced by IAA, whilst, IAA with a constant level of GA₃ (5 M) produced the highest value of TFU. GA₃ with a constant level of IAA (5 M) produced a lower TFU value. Kinetin reduced the stimulatory effect of IAA and GA₃ on TFU value when used in combination with either substance. In fertilized ovules, the highest level of TFU was reached when IAA, with a constant level of GA₃, was added to the medium, whilst its lowest level was obtained when IAA was used alone. Estimation of in vitro fiber production, as well as the effect of growth substances used in different concentrations on in vitro fiber initiation and development from unfertilized and fertilized ovules of Egyptian cotton varieties Gossypium barbadense Giza 45 are discussed.