Diagnosis of visceral leishmaniasis: comparative potential of amastigote antigen,recombinant antigen and PCR

Development of simple, economical and non-invasive tests for the early diagnosis of visceral leishmaniasis (VL) or kala-azar (KA) remains a challenge, and serological studies based on antigen prepared from the amastigote stage of Leishmania donovani, the stage that causes infection, are lacking. In the present study, circulating antibodies to total antigen isolated from the promastigote and amastigote stages of the parasite, as well as to recombinant K39 (rK39) antigen, are measured by enzyme-linked immunosorbent assay (ELISA) and the results compared with a polymerase chain reaction (PCR) test for KA diagnosis. In 116 samples of KA examined, the amastigote antigen gave significantly higher mean absorbance values in ELISA than did the promastigote antigen. The sensitivity for KA detection was significantly higher using the amastigote antigen (94%) than the promastigote antigen (90.5%). Analysis in 91 controls showed that specificity was higher with amastigote antigen (92.3%) than with promastigote antigen (86.8-89.0%). Reliability of ELISA diagnosis with amastigote antigen was only marginally lower than that with rK39 ELISA or with the PCR test. Easy availability and low cost of indigenous amastigote antigen, together with the simplicity of ELISA compared with PCR, make ELISA based on amastigote antigen a promising choice for the diagnosis of KA.     

Depression of long term potentiation in gerbil hippocampus following postischemic hypothermia

To investigate the mechanism of chronic cell death following postischemic hypothermia, the change of N-methyl- -aspartate receptor (NMDAR) were examined by immunohistochemistry of NMDAR1 and long-term potentiation (LTP) in the CA1 subfield of the gerbil hippocampus. At 1 week following postischemic hypothermia (32°C×4 h), all CA1 neurons survived; however, immunoreactivity of NMDAR1 increased in neuronal perikarya whereas decreased in dendrites in the CA1 neurons. The abnormality was still observed in remaining CA1 neurons at 1 month after hypothermia. LTP was also significantly depressed at 1 week after hypothermia. These results suggest that some abnormalities in the glutamate receptor may be caused by ischemia; such abnormality would persist in spite of hypothermia treatment, resulting in the depression of LTP.     

Biomarker associations with insomnia and secondary sleep outcomes in persons with and without HIV in the POPPY-Sleep substudy: a cohort study

Study ObjectivesWe investigated associations between inflammatory profiles/clusters and sleep measures in people living with HIV and demographically-/lifestyle-similar HIV-negative controls in the Pharmacokinetic and clinical Observations in PeoPle over fiftY (POPPY)-Sleep substudy.MethodsPrimary outcome was insomnia (Insomnia Severity Index [ISI]>15). Secondary sleep outcomes included 7-day actigraphy (e.g. mean/standard deviation of sleep duration/efficiency), overnight oximetry (e.g. oxygen desaturation index [ODI]) and patient-reported measures (Patient-Reported Outcomes Measurement Information System (PROMIS) sleep questionnaires). Participants were grouped using Principal Component Analysis of 31 biomarkers across several inflammatory pathways followed by cluster analysis. Between-cluster differences in baseline characteristics and sleep outcomes were assessed using Kruskal–Wallis/logistic regression/Chi-squared/Fisher’s exact tests.ResultsOf the 465 participants included (74% people with HIV, median [interquartile range] age 54 [50–60] years), only 18% had insomnia and secondary sleep outcomes suggested generally good sleep (e.g. ODI 3.1/hr [1.5–6.4]). Three clusters with distinct inflammatory profiles were identified: “gut/immune activation” (n = 47), “neurovascular” (n = 209), and “reference” (relatively lower inflammation; n = 209). The “neurovascular” cluster included higher proportions of people with HIV, obesity (BMI>30 kg/m²), and previous cardiovascular disease, mental health disorder, and arthritis of knee/hip relative to the other two clusters. No clinically relevant between-cluster differences were observed in proportions with insomnia (17%, 18%, 20%) before (p = .76) or after (p = .75) adjustment for potential confounders. Few associations were observed among actigraphy, oximetry, and PROMIS measures.ConclusionsAlthough associations could exist with other sleep measures or biomarker types not assessed, our findings do not support a strong association between sleep and inflammation in people with HIV.     

Novel flavonoid-based biodegradable nanoparticles for effective oral delivery of etoposide by P-glycoprotein modulation: an in vitro,ex vivo and in vivo investigations

Abstract

A receptor level interaction of etoposide with P-glycoprotein (P-gp) and subsequent intestinal efflux has an adverse effect on its oral absorption. The present work is aimed to enhance the bioavailability of etoposide by co-administering it with quercetin (a P-gp inhibitor) in dual-loaded polymeric nanoparticle formulation. Poly-lactic-co-glycolic acid (PLGA) nanoparticles were optimized for various parameters like o/w phase volume ratio, poly-vinyl alcohol concentration, PLGA concentration and sonication time. The cytotoxicity studies (MTT assay) revealed a 9- and 11-fold decrease in the IC 50 values for etoposide-loaded nanoparticles (ENP) and etoposide?+?quercetin dual-loaded nanoparticles (EQNP) when compared to that of free etoposide, respectively, and the results were further supported by florescent-activated cell sorter studies. The confocal imaging of the intestinal sections treated with ENP and EQNP containing fluorescent probe (rhodamine) showed the superiority of the EQNP to permeate deeper. Furthermore, pharmacokinetic studies on rats revealed that EQNP exhibited a 2.4-fold increase in bioavailability of etoposide than ENP with no quercetin. The developed loaded nanoparticles have the high potential to enhance the bioavailability of the etoposide and sensitize the resistant cells.     

Topical metronidazole in seborrheic dermatitis--a double-blind study

OBJECTIVE: To evaluate the role of topical metronidazole gel in the treatment of seborrheic dermatitis. METHODS: Forty-four patients with seborrheic dermatitis were enrolled in the study. All topical treatments were stopped for at least 2 weeks before the patients were allocated at random to receive either metronidazole 1% gel or placebo for 8 weeks. The severity score was measured at the initial evaluation, and the patients were followed up at 2-week intervals for 8 weeks. A global evaluation of improvement was done at 8 weeks. RESULTS: Thirty-eight patients completed the study; 21 patients in the metronidazole group and 17 patients in the placebo group could be evaluated. There was a statistically significant decrease of the mean score even at week 2; the difference became highly significant at 8 weeks (p < 0.001). On the final evaluation at 8 weeks, 14 patients in the metronidazole group showed marked improvement to complete clearance as compared to only 2 patients having moderate improvement in the placebo group (p < 0.001). CONCLUSION: The present trial has demonstrated the effectiveness of topical 1% metronidazole gel in seborrheic dermatitis.     

Controlling room temperature ferromagnetism and band gap in ZnO nanostructured thin films by varying angle of implantation

The defects in the host lattice play a major role in tuning the surface roughness, optical band gap and the room temperature ferromagnetism (RTFM) of ZnO thin films. Herein, we report a novel approach to tailor the band gap and RTFM of a ZnO nanostructure by varying the angle of implantation of 60 keV N ions keeping the ion fluence of 1 × 10¹⁶ ions per cm² and the beam size of 3 mm constant. The implantation was performed by changing the thin films'' orientations at 30°, 60° and 90° with respect to the incident beams. Remarkably, an enhancement of ∼6 times in RTFM, tuning in band gap from 3.27 to 3.21 eV and ∼60% reduction in surface roughness were noticed when the ion implantation was done at 60° to the normal. This novel technique may be suitable for tuning the physical properties of nanostructures for their application in the spintronics, semiconductor and solar cell industries.

The defects in the host lattice play a major role in tuning the surface roughness, optical band gap and the room temperature ferromagnetism of ZnO thin films.     

The pituitary-adrenal axis of fetal rats after maternal dexamethasone treatment

Elevated glucocorticoid level in the gravid female circulation affects number of endocrine functions in fetuses and offspring. In this research female rats were injected with dexamethasone (Dx) in three consecutive daily doses of 1.0, 0.5, 0.5 mg/kg body weight, starting from day 16 of pregnancy. The influence of this treatment on the pituitary adrenocorticotrophic (ACTH) cells and adrenal glands of 19-day-old fetuses was examined immunocytochemically and by morphometric analysis. Moreover, the proliferative activity of adrenocortical cells was estimated after application of the mitotic inhibitor Oncovine. Administration of Dx to pregnant rats induced a decline of fetal ACTH cell immunopositivity and significant decreases of ACTH cell volume (23%, p<0.05), volume density (41%, p<0.05), and its number per unit area (17%, p<0.05) in comparison to the control 19-day-old fetuses. Reduced proliferative activity of adrenocortical cells (31%; p<0.05) in zona glomerulosa, as well as the volume of this zone were detected. The volume and number of fetal adrenocortical cells in the inner zone and chromoblasts were not significantly reduced after Dx treatment of pregnant rats. These results show that maternal Dx administration in the period when the fetal hypothalamo-pituitary-adrenal (PA) axis begins its function inhibited the PA axis. Reduced ACTH cell function and mitotic activity led to suppression of adrenocortical cell multiplication in zona glomerulosa, the region of the adrenal cortex where most proliferating cells were found in control 19-day-old fetuses. Thus, increased glucocorticoid levels during late pregnancy caused developmental modifications involving the fetal PA axis, which could be the basis of the altered endocrine responsiveness in adult life.     

A synthetic porphyrin as an effective dual antidote against carbon monoxide and cyanide poisoning

Simultaneous poisoning by carbon monoxide (CO) and hydrogen cyanide is the major cause of mortality in fire gas accidents. Here, we report on the invention of an injectable antidote against CO and cyanide (CN^–) mixed poisoning. The solution contains four compounds: iron(III)porphyrin (Fe^IIITPPS, F), two methyl-β-cyclodextrin (CD) dimers linked by pyridine (Py3CD, P) and imidazole (Im3CD, I), and a reducing agent (Na₂S₂O₄, S). When these compounds are dissolved in saline, the solution contains two synthetic heme models including a complex of F with P (hemoCD-P) and another one of F with I (hemoCD-I), both in their iron(II) state. hemoCD-P is stable in its iron(II) state and captures CO more strongly than native hemoproteins, while hemoCD-I is readily autoxidized to its iron(III) state to scavenge CN^– once injected into blood circulation. The mixed solution (hemoCD-Twins) exhibited remarkable protective effects against acute CO and CN^– mixed poisoning in mice (~85% survival vs. 0% controls). In a model using rats, exposure to CO and CN^– resulted in a significant decrease in heart rate and blood pressure, which were restored by hemoCD-Twins in association with decreased CO and CN^– levels in blood. Pharmacokinetic data revealed a fast urinary excretion of hemoCD-Twins with an elimination half-life of 47 min. Finally, to simulate a fire accident and translate our findings to a real-life scenario, we confirmed that combustion gas from acrylic cloth caused severe toxicity to mice and that injection of hemoCD-Twins significantly improved the survival rate, leading to a rapid recovery from the physical incapacitation.

Fire accidents frequently occur around the world and the consequent inhalation of combustion gases represents the leading cause of death under these circumstances (1–10). Typically, combustion gases consist, among others, of carbon dioxide, water (H₂O), carbon monoxide (CO), hydrogen cyanide (HCN), hydrochloric acid, nitrogen oxide, and sulfur oxide. Their amounts depend on the materials of combustion (3–10) and, among them, CO and HCN are the most dangerous due to their severe toxic effects in humans (3–4, 5–15). CO is generated during incomplete combustion of carbon materials while HCN derives from carbon- and nitrogen-containing materials. Thus, CO and HCN gases are simultaneously released when synthetic materials such as plastics, acrylic cloths, and urethanes are burned in buildings. Once inhaled, CO strongly binds to ferrous iron(II) heme proteins such as hemoglobin (Hb) in erythrocytes and myoglobin in muscles (16–19), thus compromising oxygen (O₂) transport/storage in the blood circulation and tissues. On the contrary, HCN binds as cyanide ion (CN^–) to ferric iron(III) heme, preferentially targeting cytochrome c oxidase (CcO) in mitochondria and interrupting O₂-dependent energy production (11, 20–22). Likewise, the toxic effects of CO on mitochondrial respiration are ascribed to its ability to bind to cytochrome c oxidase (23–25). Therefore, both CO and HCN can inhibit aerobic respiration mediated by several heme proteins leading to anoxia-induced lethal toxicity. As the molecular mechanisms of CO and HCN toxicity are closely related, additional or synergistic deleterious effects can be expected when these two gases are produced and inhaled at the same time (26–31). Therapeutic strategies to treat either CO or HCN poisoning have been developed independently (11, 13, 22, 32–37). However, to our knowledge, there is no medical intervention at present to neutralize simultaneously CO and HCN with an effective treatment in vivo.In the present study, we have developed an injectable antidote for CO and HCN mixed poisoning. This agent contains supramolecular compounds, termed hemoCDs, composed of a water-soluble iron(II/III)porphyrin and two cyclodextrin (CD) dimers. Based on our previous research on the synthesis and characterization of heme protein model structures (38–40), the gas-binding ability and the redox status of the iron center of the porphyrin can be regulated by the linker structure of the CD dimers englobing the porphyrin. The porphyrin complexed with a CD dimer having a pyridine linker (hemoCD-P, Fig. 1 A, Left) is stable in the iron(II) state in vivo and shows much higher CO-binding affinity compared to the affinities reported for native heme proteins (41–45). On the contrary, the porphyrin complexed with a CD dimer having an imidazole linker (hemoCD-I, Fig. 1 A, Right) is stable in the iron(III) state and shows a higher binding affinity to CN^– than native ferric met-hemoglobin (met-Hb) (46, 47). These two complexes do not bind to plasma proteins in the circulation when injected separately and are quickly excreted in urines without any chemical decomposition. Therefore, these compounds have the ability to capture either CO or CN^– to their iron(II/III) centers in vivo and expel these toxic ligands from the organism (41–47). Here, we have developed hemoCD-Twins that contains both hemoCD-P and hemoCD-I in saline for simultaneous removal of CO and HCN in vivo. This paper describes the primary pharmacological properties of hemoCD-Twins and its detoxification effects in animals intoxicated with CO and CN^–. To simulate a real-life fire accident, the burning of acrylic cloth with consequent release of combustion gases was also used as an experimental approach for antidotal tests. We found that hemoCD-Twins exhibits the following features: 1) The synthetic compounds are storable at room temperature over a year thanks to their chemical stability; 2) the solution can be quickly prepared without the need of special handlings; 3) a single administration of hemoCD-Twins shows immediate dual antidotal effect against CO and CN^–; and 4) the injected solution is quickly eliminated from the body without significant side effects. We are envisioning a scenario whereby a person who is accidentally exposed to fire gases containing CO and HCN can be promptly treated at the site by infusion with hemoCD-Twins.Open in a separate windowFig. 1.hemoCD-Twins as an antidote system against CO and CN^– mixed poisoning. (A) Biomimetic heme protein model compounds hemoCD-P and hemoCD-I that are used as CO and CN^– scavengers in vivo. (B) Chemical structures of the four components P, I, F, and S contained in hemoCD-Twins. (C) Preparation of hemoCD-Twins. Three powder compounds, F, P, and I, were dissolved in PBS in a 2:1:1 molar ratio, where the solution contains hemoCD-P and hemoCD-I in ferric iron(III) states. The solution is stable and storable at room temperature. The reducing agent S is added just before use. (D) Schematic illustration for the simultaneous removal of CO and CN^– by a single injection of hemoCD-Twins in vivo. CO and CN^– are removed from living organisms and excreted in the urine with hemoCD-P and hemoCD-I in ferrous iron(II) and ferric iron(III) complexes, respectively.