Ramifications of protease-based liquefaction of camel semen on physical, kinematic and surface glyco-pattern of cryopreserved spermatozoa.

The efficiency of incorporating different proteases in the diluent for reducing camel semen viscosity, and subsequent ramifications on morpho-functional and glycan surface properties of cryopreserved spermatozoa were investigated. Ejaculates (n = 48) were collected from three adult camels, Camelus dromedarius, during the breeding season (January - March). A portion of each raw ejaculate was evaluated for sperm physical and morphological traits, whereas the other portion was divided into three aliquots assigned for the following liquefaction treatments: control (untreated), 0.1 mg/mL papain or 5 U/mL bromelain. All samples were diluted with Tris-lactose diluent containing the anti-enzyme E-64 to neutralize both proteases before being processed for cryopreservation. Post-thaw physical and kinematic properties of spermatozoa were analyzed using a computer-assisted sperm analysis (CASA) system. The sperm surface glycocalyx pattern was evaluated with a panel of 14 fluorescent lectins. Although bromelain was more effective in elimination of semen viscosity, there was a negative correlation between bromelain supplementation and values for the variables: normal sperm, intact acrosome and intact sperm cell membrane. Bromelain supplementation, compared to papain-treated and control samples, was positively correlated with secondary sperm abnormalities, increased straight-line velocity (VSL, µm/s) and straightness (%) of spermatozoa. Results from the glycan analysis indicated that both proteases did not affect the N-linked glycan content of the entire sperm surface, whereas the treatment with proteases induced little change in N-acetylgalactosamine and fucose terminating glycans in the tail region of the sperm. Functional studies are needed to evaluate the sperm fertility rates of bromelain- and papain-treated semen for application in camel assisted reproductive technologies.

Sustained broad-spectrum antibacterial effects of nanoliposomes loaded with silver nanoparticles.

BACKGROUND: The emergence of microbial resistance to antibiotics warrants the search for effective broad-spectrum antibacterial agents. Silver nanoparticles (AgNPs) have been used as antimicrobial agents. AgNPs encapsulated in nanolipososmes have been developed as effective antimicrobial agents. MATERIALS & METHODS: Nanoliposomes (<50 nm) were prepared using a modified reverse-phase evaporation method, and spherical, dextrose-capped AgNPs were synthesized. The prepared liposome AgNPs (LAgNPs) were characterized, and tested for their antibacterial effects. RESULTS: The size of LAgNPs is 25-80 nm. The release of AgNPs from nanoliposomes was sustained over 10 h. Complete growth inhibition of Eschericia coli, Salmonella enterica, Pseudomonas aeruginosa and Staphylococcus aureus was achieved using 180, 200, 160 and 120 µM, respectively, of LAgNPs. LAgNPs exhibited sustained broad-spectrum antibacterial effects compared with free AgNPs. CONCLUSION: Nanoliposomes loaded with AgNPs are potentially effective broad-spectrum antimicrobial agents. This new formula, which can be further fortified by encapsulation of additional established antibacterial agents, may be effective against antibiotic-resistant bacteria and also promote wound healing.

Controlled synthesis and characterization of hollow flower-like silver nanostructures.

BACKGROUND: The synthesis of anisotropic silver nanoparticles is a time-consuming process and involves the use of expensive toxic chemicals and specialized laboratory equipment. The presence of toxic chemicals in the prepared anisotropic silver nanostructures hindered their medical application. The authors have developed a fast and inexpensive method for the synthesis of three-dimensional hollow flower-like silver nanostructures without the use of toxic chemicals. METHODS: In this method, silver nitrate was reduced using dextrose in presence of trisodium citrate as a capping agent. Sodium hydroxide was added to enhance reduction efficacy of dextrose and reduce time of synthesis. The effects of all four agents on the shape and size of silver nanostructures were investigated. RESULTS: Robust hollow flower-like silver nanostructures were successfully synthesized and ranged in size from 0.2 µm to 5.0 µm with surface area between 25-240 m(2)/g. Changing the concentration of silver nitrate, dextrose, sodium hydroxide, and trisodium citrate affected the size and shape of the synthesized structures, while changing temperature had no effect. CONCLUSION: The proposed method is simple, safe, and allows controlled synthesis of anisotropic silver nanostructures, which may represent promising tools as effective antimicrobial agents and for in vitro diagnostics. The synthesized hollow nanostructures may be used for enhanced drug encapsulation and sustained release.

The pharmacokinetics and metabolism of meloxicam in camels after intravenous administration.

The pharmacokinetics and metabolism of meloxicam was studied in camels (Camelus dromedarus) (n = 6) following intravenous (i.v.) administration of a dose of 0.6 mg·kg/body weight. The results obtained (mean ± SD) were as follows: the terminal elimination half-life (t(1/2ß) ) was 40.2 ± 16.8 h and total body clearance (Cl(T) ) was 1.94 ± 0.66 mL·kg/h. The volume of distribution at steady state (V(SS)) was 92.8 ± 13.7 mL/kg. One metabolite of meloxicam was tentatively identified as methylhydroxy meloxicam. Meloxicam and metabolite were excreted unconjugated in urine. Meloxicam could be detected in plasma 10 days following i.v. administration in camels using a sensitive liquid chromatography tandem mass spectrometry (LC/MS/MS) method.

Some biomedical applications of Balanites aegyptiaca grown naturally in radioactive area, Southeastern Desert, Egypt.

Balanites aegyptiaca is a naturally grown desert plant at some radioactive places in Wadi El-Gemal area, Southeastern Desert. The aim of the present study was to highlight on the B. aegyptiaca species grown naturally at radioactive places in Wadi El-Gemal area (fruit part) on the ability of using the fruit in some biomedical application (glucose, cholesterol, triglycerides, HDL-cholesterol, LDL-cholesterol and diabetes). The investigated plant was collected from different location at Wadi El-Gemal area. The uranium content was determined previously and different concentrations from the fruit with highest uranium content were used to examine the effect of B. aegyptiaca (fruit part) on the glucose, triglycerides, total cholesterol (HDL and LDL-cholesterol) levels using experimental rats. Different analysis techniques were used in order to determine different parameters. The obtained data suggest the beneficial role of B. aegyptiaca fruit as an anti-diabetic and hypo-lipidimic agent.

Impact of CD34 subsets on engraftment kinetics in allogeneic peripheral blood stem cell transplantation.

Our objective was to evaluate, probably for the first time, the impact of CD34 subsets on engraftment kinetics in allogeneic PBSC transplantation (PBSCT). PBSC graft components were analyzed in 62 cases for the absolute count/kg of total CD34+ and the following subsets: DR- and +, CD71+/-, CD38+/-, CD33+/- and CD61+/-. Time to ANC >0.5 and >1 x 10(9)/l and platelets >20 and >50 x 10(9)/l was reported. The median value for each parameter was used to discriminate rapid from slow engraftment. Four parameters showed significant predictive power of early neutrophil engraftment, namely CD34+ /DR- (P = 0.002), CD34+/38- (P = 0.02), CD34+/CD61- (P = 0.04) and total CD34+ cell dose (P = 0.04). Four parameters showed significant predictive power of early platelet engraftment, namely CD34+/CD61+ (P = 0.02), CD34+ /CD38- and total CD34+ cell dose (P = 0.04) and CD34+ /CD71- (P = 0.05). Comparing patients who received > to those who received < the threshold dose(s), only CD34+ /CD38- lost its significance for neutrophil engraftment; and only CD34+ /CD61+ retained its significance for platelet engraftment (P = 0.03); furthermore, the former group required significantly fewer platelet transfusions (P = 0.018). We concluded that in allogeneic PBSCT, the best predictor of early neutrophil engraftment is the absolute CD34+ /DR- and for early platelet engraftment is the absolute CD34+ /CD61+ cell dose.

Transplacental chemical exposure and risk of infant leukemia with MLL gene fusion.

Infant acute leukemia (IAL) frequently involves breakage and recombination of the MLL gene with one of several potential partner genes. These gene fusions arise in utero and are similar to those found in leukemias secondary to chemotherapy with inhibitors of topoisomerase II (topo-II). This has led to the hypothesis that in utero exposures to chemicals may cause IAL via an effect on topo-II. We report a pilot case-control study of IAL across different countries and ethnic groups. Cases (n = 136) were population-based in most centers. Controls (n = 266) were selected from inpatients and outpatients at hospitals serving the same populations. MLL rearrangement status was derived by Southern blot analysis, and maternal exposure data were obtained by interviews using a structured questionnaire. Apart from the use of cigarettes and alcohol, very few mothers reported exposure to known topo-II inhibitors. Significant case-control differences were apparent for ingestion of several groups of drugs, including herbal medicines and drugs classified as "DNA-damaging," and for exposure to pesticides with the last two being largely attributable, respectively, to one nonsteroidal anti-inflammatory drug, dipyrone, and mosquitocidals (including Baygon). Elevated odds ratios were observed for MLL+ve (but not MLL-ve) leukemias (2.31 for DNA-damaging drugs, P = 0.03; 5.84 for dipyrone, P = 0.001; and 9.68 for mosquitocidals, P = 0.003). Although it is unclear at present whether these particular exposures operate via an effect on topo-II, the data suggest that specific chemical exposures of the fetus during pregnancy may cause MLL gene fusions. Given the widespread use of dipyrone, Baygon, and other carbamate-based insecticides in certain settings, confirmation of these apparent associations is urgently required.

A simple strategy for breakpoint fragment determination in chronic myeloid leukemia.

Molecular characterization is considered a part of the routine work-up of chronic myeloid leukemia (CML) cases. Southern blot analysis using the universal BCR (UBCR) probe on BglII-digested DNA samples is the most commonly used technique, while employing the human 3' bcr probe (PR-1) is usually considered a complementary tool. In this study, we tried to develop a simple and economic strategy for molecular characterization of CML using the 3' probe as it has been shown to be the one capable of locating the breakpoint site. Seventy-eight cases of CML were studied. Molecular analysis was performed using the Southern blot technique. DNA was digested with Bam HI, BglII, EcoRI, and XbaI. Hybridization was performed using the human 3' bcr (PR-1) probe. BamHI and BglII could differentiate fragment 1 (F1) showing rearrangement (R) with Bam HI and germline configuration (G) with BglII; F2/3 showing R with both, and F4 showing R with BamHI and G with BglII. F2/3 cases were further divided by HindIII enzyme into F2 showing (G) and F3 showing (R). Fragment 0 showed G with both, but R with EcoRI and/or XbaI, while 3' deletion gave G with all four enzymes. Our results showed a relative incidence of 6.4% for F0, 20.5% for F1, 32.1% for F2, 19.2% for F3, 15.4% for F4, and 6.4% for 3' deletion. Sixty cases were evaluated clinically and hematologically and were followed up for disease evolution and survival. They included 32 cases in early chronic phase, 24 in late chronic phase, two in acceleration, and two in blastic crisis. No significant correlation was encountered between the breakpoint site and any of the clinical and hematological data except those patients with 3' deletion who showed a very short survival. The study emphasizes Southern blotting as the method of choice for molecular characterization of CML and offers a simple and economic strategy for diagnosis and determination of breakpoint fragment.

Effects of mobile-phase additives, solution pH, ionization constant, and analyte concentration on the sensitivities and electrospray ionization mass spectra of nucleoside antiviral agents.

The effects of various mobile-phase additives, solution pH, pKa, and analyte concentration on electrospray ionization mass spectra of a series of purine and pyrimidine nucleoside antiviral agents were studied in both positive and negative ion models. The use of 1% acetic acid resulted in good HPLC separation and the greatest sensitivity for [M + H]+ ions. In the negative ion mode, 50 mM ammonium hydroxide gave the greatest sensitivity for [M - H]- ions. The sensitivities as [M + H]+ ions were significantly larger than the sensitivities as [M - H]- ions for purine antiviral agents. Vidarabine monophosphate and pyrimidine antiviral agents, however, showed comparable or greater sensitivities as [M - H]- ions. The sensitivity as [M + H]+ showed no systematic variation with pH; however, the sensitivity as [M - H]- did increase with increasing pH. At constant pH, the ion intensity of the protonated species increased with increasing pKa. At higher analyte concentrations, dimer (M2H+) and trimer (M3H+) ions were observed. [M + Na]+ adducts were the dominant ions with 0.5 mM sodium salts for these compounds. The spectra of the more basic purine antiviral agents showed no [M + NH4]+ adduct ions, but [M + NH4]+ ions were the major peaks in the spectra of the less basic pyrimidine antiviral agents with ammonium salts. The ammonium adduct ion was formed preferentially when the proton affinity of the analyte was close to that of NH3. Abundant [M + OAc]- ions were observed for all of the antiviral agents except vidarabine monophosphate from solutions with added HOAc, NaOAc, and NH4OAc. The utility of mobile phases containing 1% HOAc or 50 mM NH4OH was demonstrated for chromatographic separations.

Electrospray ionization mass spectrometry of tetracycline, oxytetracycline, chlorotetracycline, minocycline, and methacycline.

The effects of mobile-phase additives and analyte concentration on electrospray ionization mass spectra of a series of tetracyclines were investigated in both positive and negative ion modes. Only [M + H](+) and [M - H](-) ions were observed. The greatest sensitivity as [M + H](+) ions was obtained with 1% acetic acid and the greatest sensitivity as [M - H](-) ions was obtained using 50 mM ammonium hydroxide. Sensitivities in the positive ion mode were greater than those in the negative ion mode. The sensitivity as [M + H](+) showed no systematic variation with pH; however, the sensitivity as [M - H](-) did increase with increasing pH. A larger linear range was observed for [M - H](-) than for [M + H](+) ions. Both [M + Na](+) and [M + H](+) ions were observed with 0.5 mM sodium acetate and sodium iodide, but no adduct ions were observed with ammonium acetate. Some M(2)H(+) ions were observed at higher concentrations. Cluster ions, Na(NaOAc)(n)(+) or Na(NaI)(n)(+), but no sample ions were observed using 5 mM salts. The data suggest that mechanisms in addition to solution ionization are involved in the formation of the ESI sample ions. The utility of mobile phases containing 1% HOAc or 50 mM NH(4)OH was demonstrated for chromatographic separations.

Study of the present status of filariasis in an endemic area in Giza Governorate, Egypt.

In a trial to throw some light on the present status of Filariasis in Giza Governorate, both human and mosquitoes were surveyed in two endemic areas: Kafr-Ghataty and Azizya. In Kafr-Ghataty the clinical cases detected were very rare (0.77%), while no microfilaraemic cases were obtained. The situation was different in Azizya village, where no clinical cases were detected but microfilaraemic cases were relatively higher (8%). The results of the larval survey, carried out only in Kafr-Ghataty, showed that Culex pipiens larvae were predominant (99.57%). They were found in all the breeding places specially canals and seepages, while Theobaldia longiareolata larvae were rare (0.43%) and present only in wells and cesspits. Regarding the adult mosquitoes survey, Culex pipiens was the only species detected in both areas. The results of dissection of adult female mosquitoes showed that individual dissection failed to detect infection among mosquitoes, in contrast to mass dissection which gave positive results (0.27% in Kafr-Ghataty and 2.2% in Azizya). It was also found that infective mosquitoes were 0% in Kafr-Ghataty and 1.1% in Azizya.

Serodiagnosis of giardiasis by counterimmunoelectrophoresis and indirect immunofluorescence tests.

Serodiagnosis of giardiasis by counterimmunoelectrophoresis assay (CIEP) and indirect immunofluorescence test (IFAT) were evaluated versus stool examination. Giardia lamblia antibodies were detected in sera of 46.9% and 53.1% using CIEP and IFAT respectively out of 49 giardiasis-infected children (5-10 years) diagnosed microscopically. Among the control group (22 children) 13.6% and 18.2% were positive by CIEP and IFAT respectively. The findings of the study showed that we can not rely on serology to diagnose giardiasis specially in cases with low excretion rates.

Evaluation of an ELISA test in past and present schistosomiasis.

An enzyme linked immunosorbent assay (ELISA) was evaluated in relation to an indirect haemagglutination (IHA) test in schistosomiasis patients who were classified by clinical, sonographic and direct methods of diagnosis. Sensitivities of ELISA and IHA respectively proved to be 100% and 69.23% in acute simple intestinal schistosomiasis; 95.5% and 90.4% in chronic active schistosomiasis patients; 86.06% and 67.41% in patients with past history of exposure; 80% and 64.28% in patients with hepatosplenomegaly with past history of schistosomiasis, and 96% and 80% in patients with hepatic fibrosis as shown by sonar. It was apparent that ELISA is more sensitive than IHA in acute simple schistosomiasis in patients with past history of exposure, those with bilharzial hepatosplenomegaly and those with hepatic fibrosis. Both tests were nearly equally sensitive in chronic active schistosomiasis.

Naturally occurring toxoplasma antibodies in serum and milk of lactating women.

Serum and milk of lactating women were tested for toxoplasmosis using specific-IgG IFAT. Apparently healthy 70 women were selected: 54 from rural and 16 from urban areas. Serum and milk were simultaneously collected from each one. Sera were positive in 22 (31.4%) of the total 70; including 16 (29.6%) and 6 (37.5%) of rural and urban groups respectively. No statistical significant difference was found for positivity and titre levels between the two groups (P greater than 0.05). Milk was positive in 12 (17.1%) of the 70 women; including 10 (18.5%) and 2 (12.5%) from rural and urban groups respectively, having no statistical significant difference (P greater than 0.05). Comparing serum and milk for positivity and titre levels, also there was no statistical significant difference (P greater than 0.05). It is concluded that relatively low antibody levels in serum could be excreted in milk and may be protective for suckling babies. Occurrence of antibodies in serum and milk are homogeneously distributed between rural and urban inhabitants.

The immune defensive mechanism in parasitic infection with eosinophilia.

This study was attempted to assess the extent of stimulation of the immune system in some patients suffering from parasitic infection with eosinophilia. Eighty-eight cases were selected, they included 10 apparently healthy, 10 suffering from different atopic disorders associated with eosinophilia and free from parasitic infections and 68 patients suffering from different parasitic infections associated with marked eosinophilia. The results showed a highly significant increase in the absolute eosinophil count in the atopic group while the parasitized group showed a moderate increase. The IgE serum level was increased in both atopic and parasitized groups. The cellular immunity expressed by phagocytic power of neutrophils was more depressed in parasitized than atopic group.

Immunopathological studies on the encystation phase of experimental trichinosis after cortisone and cyclophosphamide treatment.

Efficacy of cortisone and cyclophosphamide on the muscle phase of Trichinella spiralis in albino rats, before and after complete encapsulation, was studied by haematoxylin and eosin staining for histopathological changes and by the indirect immunoperoxidase test for the extent of antigen deposition. Results showing beneficial effects of cortisone and hazardous action of cyclophosphamide are discussed in details. This study indicates that the use of cortisone in the treatment of trichinosis remains to be substantiated particularly when administered before complete encystation; the time at which the diagnosis is usually made.

Effects of some trace elements on platelet function in rats.

The present study portrays the effects of some elements, namely: iron, zinc, copper, magnesium and gold, on platelet count, PCV and platelet aggregation, 60 minutes following administration of the metal salts. Marked thrombocytopenia was encountered in rats treated with ferrous sulphate while the platelet count was significantly changed with the other elements tested. The PCV was significantly increased following treatment with ferrous sulphate and large dose of gold chloride, but was insignificantly altered with the other elements. As regards platelet aggregation, all metals tested, with the exception of magnesium caused significant inhibition of platelet aggregation was only significantly impaired following treatment with iron and gold, but was insignificantly altered following treatment with zinc and copper. On the other hand, treatment with magnesium resulted in enhancement of both ADP- and collagen-induced aggregation. The mechanisms underlying these effects are discussed.

In vitro effects of trace elements on blood clotting and platelet function. A--Iron, copper, and gold.

The present in vitro study of the effects of iron on the blood coagulation mechanism in rats showed that addition of ferrous sulphate to pooled rat plasma resulted in inhibition of blood coagulation, as shown by prolongation of the clotting parameters tested, an effect which was dose-dependent. In vitro addition of ferrous sulphate to rat PRP in doses of 2-5 mg/ml significantly decreased platelet aggregation in response to ADP, while collagen-induced aggregation was significantly diminished in presence of the higher doses of ferrous sulphate (4-5 mg/ml). Also, preincubation of ferrous sulphate with thrombin or with pure fibrinogen indicated that iron could produce decrease of thrombin activity as well as impairment of fibrinogen clottability. In vitro addition of copper sulphate (300-1000 micrograms/ml) elicited an anticoagulant effect, though thrombin time was markedly shortened with all tested concentrations of copper sulphate. Addition of copper sulphate to PRP produced inhibition of platelet aggregation in response to PRP produced inhibition of platelet aggregation in response to ADP and to collagen. Preincubation of copper sulphate with thrombin resulted in slight enhancement of thrombin activity followed by inhibition, while preincubation of copper sulphate with pure fibrinogen caused only minimal impairment of fibrinogen clottability. Also, addition of gold chloride in doses of 50-500 micrograms/ml to plasma in vitro produced a dose-dependent progressive prolongation of all clotting parameters tested, the effects reaching a maximum after 30 min. incubation. Further the in vitro addition of gold chloride to rat PRP resulted in marked inhibition of platelet aggregation in response to both ADP and collagen. In addition, preincubation of gold chloride with thrombin or with pure fibrinogen showed that gold exerted an antithrombin action and prolonged the fibrinogen clotting time indicating impaired fibrinogen clottability.

In vitro effects of trace elements on blood clotting and platelet function. B--Zinc and magnesium.

The in vitro effects of zinc and magnesium salts on blood coagulation mechanism and platelet aggregation were studied on rat plasma. Addition of zinc sulphate to pooled rat plasma in a range of concentrations (0.3-1 mg/ml) caused a dose dependent significant prolongation of recalcification, prothrombin and partial thromboplastin times. These effects reached a peak after 30 minutes while the thrombin clotting time was not significantly altered and was even shortened in the presence of highest concentration of zinc tested (1 mg/ml). Incubation of thrombin with zinc sulphate (150 micrograms/ml) for up to 30 minutes did not affect significantly the action of thrombin. Incubation of the same concentrations of zinc sulphate with fibrinogen produced non clotting of fibrinogen after 0-minutes. Addition of rising concentrations of zinc sulphate to rat PRP produced inhibition of ADP-induced platelet aggregation. On the other hand, collagen-induced aggregation was insignificantly inhibited in the presence of zinc. In contrast, in vitro additions of rising concentrations of magnesium sulphate (2-5 mg/ml) to pooled rat plasma exerted no effect on recalcification time immediately after addition (0-minutes), but after 5 minutes following incubation it produced significant shortening of recalcification time in all the doses tested. The prothrombin time showed a general trend of shortening, maximal after 5-minutes incubation. The results of partial thromboplastin times revealed clotting before addition of calcium chloride. The thromboplastin time also showed progressive shortening with rising concentrations of magnesium sulphate. When thrombin solution was exposed to magnesium sulphate (2.5 mg/ml) no effect on the activity of thrombin was seen for up to 30 minutes. Fibrinogen solution similarly exposed to the same concentration of magnesium sulphate did not show any significant effect on its clottability with thrombin for up to 30 minutes. Magnesium sulphate in the range of doses tested significantly enhanced platelet aggregation of PRP in response to both ADP and collagen, and the responses observed were not dose dependent. The mechanisms underlying the effects of these two metals on blood clotting and platelet aggregation are discussed.