Leishmania donovani mitogen-activated protein kinases as a host-parasite interaction interface.

Leishmaniasis, a major globally re-emerging neglected tropical disease, has a restricted repertoire of chemotherapeutic options due to a narrow therapeutic index, drug resistance, or patient non-compliance due to toxicity. The disease is caused by the parasite Leishmania that resides in two different forms in two different environments: as sessile intracellular amastigotes within mammalian macrophages and as motile promastigotes in sandfly gut. As mitogen-activated protein kinases (MAPKs) play important roles in cellular differentiation and survival, we studied the expression of Leishmania donovani MAPKs (LdMAPKs). The homology studies by multiple sequence alignment show that excepting LdMAPK1 and LdMAPK2, all thirteen other LdMAPKs share homology with human ERK and p38 isoforms. Expression of LdMAPK4 and LdMAPK5 is less in avirulent promastigotes and amastigotes. Compared to miltefosine-sensitive L. donovani parasites, miltefosine-resistant parasites have higher LdMAPK1, LdMAPK3-5, LdMAPK7-11, LdMAPK13, and LdMAPK14 expression. IL-4-treatment of macrophages down-regulated LdMAPK11, in virulent amastigotes whereas up-regulated LdMAPK5, but down-regulated LdMAPK6, LdMAPK12-15, expression in avirulent amastigotes. IL-4 up-regulated LdMAPK1 expression in both virulent and avirulent amastigotes. IFN-γ-treatment down-regulated LdMAPK6, LdMAPK13, and LdMAPK15 in avirulent amastigotes but up-regulated in virulent amastigotes. This complex profile of LdMAPKs expression among virulent and avirulent parasites, drug-resistant parasites, and in amastigotes within IL-4 or IFN-γ-treated macrophages suggests that LdMAPKs are differentially controlled at the host-parasite interface regulating parasite survival and differentiation, and in the course of IL-4 or IFN-γ dominated immune response.

Evaluation of the surface characteristics and antibacterial properties of Titanium dioxide nanotube and methacryloyloxyethylphosphorylcholine (MPC) coated orthodontic brackets-a comparative invitro study.

OBJECTIVES: White spot lesions are the most common iatrogenic effect observed during orthodontic treatment. This study aimed to compare the surface characteristics and antibacterial action of uncoated and coated orthodontic brackets. MATERIALS AND METHODS: Sixty commercially available stainless steel brackets were coated with TiO2 nanotubes and methacryloyloxyethylphosphorylcholine. The sample was divided into Group 1: uncoated orthodontic brackets, Group 2: Stainless steel brackets with TiO2 nanotubes coating, Group 3: Stainless steel brackets with methacryloyloxyethylphosphorylcholine coating, and Group 4: Stainless steel brackets with TiO2 nanotubes combined with methacryloyloxyethylphosphorylcholine coating. Surface characterization was assessed using atomic force microscopy and scanning electron microscopy. Streptococcus mutans was selected to test the antibacterial ability of the orthodontic brackets, total bacterial adhesion and bacterial viability were assessed. The brackets were subjected to scanning electron microscopy to detect the presence of biofilm. RESULTS: The surface roughness was the greatest in Group 1 and least in Group 2 followed by Group 4 and Group 3 coated brackets. The optical density values were highest in Group 1 and lowest in Group 4. Comparison of colony counts revealed high counts in Group 1 and low counts in Group 4. A positive correlation between surface roughness and colony counts was obtained, however, was not statistically significant. CONCLUSIONS: The coated orthodontic brackets exhibited less surface roughness than the uncoated orthodontic brackets. Group 4 coated orthodontic brackets showed the best antibacterial properties. CLINICAL RELEVANCE: Coated orthodontic brackets prevent adhesion of streptococcus mutans and reduces plaque accumulation around the brackets thereby preventing formation of white spot lesions during orthodontic treatment.

Metabolic Characteristics and Discriminative Diagnosis of Growth Hormone Deficiency and Idiopathic Short Stature in Preadolescents and Adolescents.

Growth hormone deficiency (GHD) and idiopathic short stature (ISS) are the most common types of short stature (SS), but little is known about their pathogenesis, and even less is known about the study of adolescent SS. In this study, nuclear magnetic resonance (NMR)-based metabolomic analysis combined with least absolute shrinkage and selection operator (LASSO) were performed to identify the biomarkers of different types of SS (including 94 preadolescent GHD (PAG), 61 preadolescent ISS (PAI), 43 adolescent GHD (ADG), and 19 adolescent ISS (ADI)), and the receiver operating characteristic curve (ROC) was further used to evaluate the predictive power of potential biomarkers. The results showed that fourteen, eleven, nine, and fifteen metabolites were identified as the potential biomarkers of PAG, PAI, ADG, and ADI compared with their corresponding controls, respectively. The disturbed metabolic pathways in preadolescent SS were mainly carbohydrate metabolism and lipid metabolism, while disorders of amino acid metabolism played an important role in adolescent SS. The combination of aspartate, ethanolamine, phosphocholine, and trimethylamine was screened out to identify PAI from PAG, and alanine, histidine, isobutyrate, methanol, and phosphocholine gave a high classification accuracy for ADI and ADC. The differences in metabolic characteristics between GHD and ISS in preadolescents and adolescents will contribute to the development of individualized clinical treatments in short stature.

Comparison study of surface-initiated hydrogel coatings with distinct side-chains for improving biocompatibility of polymeric heart valves.

Polymeric heart valves (PHVs) present a promising alternative for treating valvular heart diseases with satisfactory hydrodynamics and durability against structural degeneration. However, the cascaded coagulation, inflammatory responses, and calcification in the dynamic blood environment pose significant challenges to the surface design of current PHVs. In this study, we employed a surface-initiated polymerization method to modify polystyrene-block-isobutylene-block-styrene (SIBS) by creating three hydrogel coatings: poly(2-methacryloyloxy ethyl phosphorylcholine) (pMPC), poly(2-acrylamido-2-methylpropanesulfonic acid) (pAMPS), and poly(2-hydroxyethyl methacrylate) (pHEMA). These hydrogel coatings dramatically promoted SIBS's hydrophilicity and blood compatibility at the initial state. Notably, the pMPC and pAMPS coatings maintained a considerable platelet resistance performance after 12 h of sonication and 10 000 cycles of stretching and bending. However, the sonication process induced visible damage to the pHEMA coating and attenuated the anti-coagulation property. Furthermore, the in vivo subcutaneous implantation studies demonstrated that the amphiphilic pMPC coating showed superior anti-inflammatory and anti-calcification properties. Considering the remarkable stability and optimal biocompatibility, the amphiphilic pMPC coating constructed by surface-initiated polymerization holds promising potential for modifying PHVs.

Oxidized phospholipid POVPC contributes to vascular calcification by triggering ferroptosis of vascular smooth muscle cells.

Vascular calcification is an actively regulated biological process resembling bone formation, and osteogenic differentiation of vascular smooth muscle cells (VSMCs) plays a crucial role in this process. 1-Palmitoyl-2-(5'-oxo-valeroyl)-sn-glycero-3-phosphocholine (POVPC), an oxidized phospholipid, is found in atherosclerotic plaques and has been shown to induce oxidative stress. However, the effects of POVPC on osteogenic differentiation and calcification of VSMCs have yet to be studied. In the present study, we investigated the role of POVPC in vascular calcification using in vitro and ex vivo models. POVPC increased mineralization of VSMCs and arterial rings, as shown by alizarin red staining. In addition, POVPC treatment increased expression of osteogenic markers Runx2 and BMP2, indicating that POVPC promotes osteogenic transition of VSMCs. Moreover, POVPC increased oxidative stress and impaired mitochondria function of VSMCs, as shown by increased ROS levels, impairment of mitochondrial membrane potential, and decreased ATP levels. Notably, ferroptosis triggered by POVPC was confirmed by increased levels of intracellular ROS, lipid ROS, and MDA, which were decreased by ferrostatin-1, a ferroptosis inhibitor. Furthermore, ferrostatin-1 attenuated POVPC-induced calcification of VSMCs. Taken together, our study for the first time demonstrates that POVPC promotes vascular calcification via activation of VSMC ferroptosis. Reducing the levels of POVPC or inhibiting ferroptosis might provide a novel strategy to treat vascular calcification.

ABC transporter inhibition by beauvericin partially overcomes drug resistance in Leishmania tropica.

Leishmaniasis is a neglected tropical disease infecting the world's poorest populations. Miltefosine (ML) remains the primary oral drug against the cutaneous form of leishmaniasis. The ATP-binding cassette (ABC) transporters are key players in the xenobiotic efflux, and their inhibition could enhance the therapeutic index. In this study, the ability of beauvericin (BEA) to overcome ABC transporter-mediated resistance of Leishmania tropica to ML was assessed. In addition, the transcription profile of genes involved in resistance acquisition to ML was inspected. Finally, we explored the efflux mechanism of the drug and inhibitor. The efficacy of ML against all developmental stages of L. tropica in the presence or absence of BEA was evaluated using an absolute quantification assay. The expression of resistance genes was evaluated, comparing susceptible and resistant strains. Finally, the mechanisms governing the interaction between the ABC transporter and its ligands were elucidated using molecular docking and dynamic simulation. Relative quantification showed that the expression of the ABCG sub-family is mostly modulated by ML. In this study, we used BEA to impede resistance of Leishmania tropica. The IC50 values, following BEA treatment, were significantly reduced from 30.83, 48.17, and 16.83 µM using ML to 8.14, 11.1, and 7.18 µM when using a combinatorial treatment (ML + BEA) against promastigotes, axenic amastigotes, and intracellular amastigotes, respectively. We also demonstrated a favorable BEA-binding enthalpy to L. tropica ABC transporter compared to ML. Our study revealed that BEA partially reverses the resistance development of L. tropica to ML by blocking the alternate ATP hydrolysis cycle.

Leishmania blood parasite dynamics during and after treatment of visceral leishmaniasis in Eastern Africa: A pharmacokinetic-pharmacodynamic model.

BACKGROUND: With the current treatment options for visceral leishmaniasis (VL), recrudescence of the parasite is seen in a proportion of patients. Understanding parasite dynamics is crucial to improving treatment efficacy and predicting patient relapse in cases of VL. This study aimed to characterize the kinetics of circulating Leishmania parasites in the blood, during and after different antileishmanial therapies, and to find predictors for clinical relapse of disease. METHODS: Data from three clinical trials, in which Eastern African VL patients received various antileishmanial regimens, were combined in this study. Leishmania kinetoplast DNA was quantified in whole blood with real-time quantitative PCR (qPCR) before, during, and up to six months after treatment. An integrated population pharmacokinetic-pharmacodynamic model was developed using non-linear mixed effects modelling. RESULTS: Parasite proliferation was best described by an exponential growth model, with an in vivo parasite doubling time of 7.8 days (RSE 12%). Parasite killing by fexinidazole, liposomal amphotericin B, sodium stibogluconate, and miltefosine was best described by linear models directly relating drug concentrations to the parasite elimination rate. After treatment, parasite growth was assumed to be suppressed by the host immune system, described by an Emax model driven by the time after treatment. No predictors for the high variability in onset and magnitude of the immune response could be identified. Model-based individual predictions of blood parasite load on Day 28 and Day 56 after start of treatment were predictive for clinical relapse of disease. CONCLUSION: This semi-mechanistic pharmacokinetic-pharmacodynamic model adequately captured the blood parasite dynamics during and after treatment, and revealed that high blood parasite loads on Day 28 and Day 56 after start of treatment are an early indication for VL relapse, which could be a useful biomarker to assess treatment efficacy of a treatment regimen in a clinical trial setting.

Unmasking the Mechanism behind Miltefosine: Revealing the Disruption of Intracellular Ca2+ Homeostasis as a Rational Therapeutic Target in Leishmaniasis and Chagas Disease.

Originally developed as a chemotherapeutic agent, miltefosine (hexadecylphosphocholine) is an inhibitor of phosphatidylcholine synthesis with proven antiparasitic effects. It is the only oral drug approved for the treatment of Leishmaniasis and American Trypanosomiasis (Chagas disease). Although its precise mechanisms are not yet fully understood, miltefosine exhibits broad-spectrum anti-parasitic effects primarily by disrupting the intracellular Ca2+ homeostasis of the parasites while sparing the human hosts. In addition to its inhibitory effects on phosphatidylcholine synthesis and cytochrome c oxidase, miltefosine has been found to affect the unique giant mitochondria and the acidocalcisomes of parasites. Both of these crucial organelles are involved in Ca2+ regulation. Furthermore, miltefosine has the ability to activate a specific parasite Ca2+ channel that responds to sphingosine, which is different to its L-type VGCC human ortholog. Here, we aimed to provide an overview of recent advancements of the anti-parasitic mechanisms of miltefosine. We also explored its multiple molecular targets and investigated how its pleiotropic effects translate into a rational therapeutic approach for patients afflicted by Leishmaniasis and American Trypanosomiasis. Notably, miltefosine's therapeutic effect extends beyond its impact on the parasite to also positively affect the host's immune system. These findings enhance our understanding on its multi-targeted mechanism of action. Overall, this review sheds light on the intricate molecular actions of miltefosine, highlighting its potential as a promising therapeutic option against these debilitating parasitic diseases.

Successful treatment of recurrent visceral leishmaniasis relapse in an immunocompetent adult female with functional hypopituitarism in Bangladesh.

BACKGROUND: Currently available treatment options are mostly effective in achieving long-term cure in visceral leishmaniasis (VL) patients. However, there have been reports of recurrence of this illness in both immunosuppressed and immunocompetent patients. CASE PRESENTATION: We report the first case of recurrent VL relapse in a 19-year-old immunocompetent female with functional hypopituitarism (hypogonadotropic hypogonadism with central hypothyroidism) from Bangladesh, who has been treated three times previously with optimal dosage and duration- liposomal amphotericin B (LAmB) alone and in combination with miltefosine. We treated the patient successfully with a modified treatment regimen of 10 mg/kg body weight LAmB for two consecutive days along with oral miltefosine for seven days as loading dose. For secondary prophylaxis, the patient received 3 mg/kg body weight LAmB along with oral miltefosine for seven days monthly for five doses followed by hormonal replacement. The patient remained relapse free after 12 months of her treatment completion. CONCLUSION: In the absence of protective vaccines against Leishmania species and standard treatment regimen, this modified treatment regimen could help the management of recurrent relapse cases.

Bioinspired zwitterionic triblock copolymers designed for colloidal drug delivery: 2 - Biological evaluation.

In this work, poly(lactide) nanoparticles were equipped with a bioinspired coating layer based on poly[2-(methacryloyloxy)ethyl phosphorylcholine] and then evaluated when administered to the lungs and after intravenous injection. Compared to the plain counterparts, the chosen zwitterionic polymer shell prevented the coated colloidal formulation from aggregation and conditioned it for lower cytotoxicity, protein adsorption, complement activation and phagocytic cell uptake. Consequently, no interference with the biophysical function of the lung surfactant system could be detected accompanied by negligible protein and cell influx into the bronchoalveolar space after intratracheal administration. When injected into the central compartment, the coated formulation showed a prolonged circulation half-life and a delayed biodistribution to the liver. Taken together, colloidal drug delivery vehicles would clearly benefit from the investigated poly[2-(methacryloyloxy)ethyl phosphorylcholine]-based polymer coatings.

Vascular Endothelial Cell-Derived Exosomal Sphingosylphosphorylcholine Attenuates Myocardial Ischemia-Reperfusion Injury through NR4A2-Mediated Mitophagy.

Cardiomyocyte survival is a critical contributing process of host adaptive responses to cardiovascular diseases (CVD). Cells of the cardiovascular endothelium have recently been reported to promote cardiomyocyte survival through exosome-loading cargos. Sphingosylphosphorylcholine (SPC), an intermediate metabolite of sphingolipids, mediates protection against myocardial infarction (MI). Nevertheless, the mechanism of SPC delivery by vascular endothelial cell (VEC)-derived exosomes (VEC-Exos) remains uncharacterized at the time of this writing. The present study utilized a mice model of ischemia/reperfusion (I/R) to demonstrate that the administration of exosomes via tail vein injection significantly diminished the severity of I/R-induced cardiac damage and prevented apoptosis of cardiomyocytes. Moreover, SPC was here identified as the primary mediator of the observed protective effects of VEC-Exos. In addition, within this investigation, in vitro experiments using cardiomyocytes showed that SPC counteracted myocardial I/R injury by activating the Parkin and nuclear receptor subfamily group A member 2/optineurin (NR4A2/OPTN) pathways, in turn resulting in increased levels of mitophagy within I/R-affected myocardium. The present study highlights the potential therapeutic effects of SPC-rich exosomes secreted by VECs on alleviating I/R-induced apoptosis in cardiomyocytes, thereby providing strong experimental evidence to support the application of SPC as a potential therapeutic target in the prevention and treatment of myocardial infarction.

Monitoring the Long-Term Effectiveness of Miltefosine in Indian Post-Kala-Azar Dermal Leishmaniasis.

Post-kala-azar dermal leishmaniasis (PKDL), the dermal sequel to visceral leishmaniasis (VL), is characterized by hypopigmented macules (macular) and/or papules and nodules (polymorphic). Post-kala-azar dermal leishmaniasis plays a significant role in disease transmission, emphasizing the need for monitoring chemotherapeutic effectiveness. Accordingly, this study aimed to quantify the parasite burden in PKDL patients after treatment with miltefosine by a quantitative polymerase chain reaction (qPCR). A Leishmania kinetoplastid gene-targeted qPCR was undertaken using DNA from skin biopsy specimens of patients with PKDL at three time points, i.e., at disease presentation (week 0, n = 157, group 1), upon completion of treatment (week 12, n = 39, group 2), and at any time point 6 months after completion of treatment (week ≥36, n = 54, group 3). A cycle threshold (Ct) <30 was considered the cutoff for positivity, and load was quantified as the number of parasites/µg genomic DNA (gDNA); cure was considered when samples had a Ct >30. The parasite load at disease presentation (group 1) was 10,769 (1,339-80,441)/µg gDNA (median [interquartile range]). In groups 2 and 3, qPCR results were negative in 35/39 cases (89.7%) and 48/54 cases (88.8%), respectively. In the 10/93 (10.8%) qPCR-positive cases, the parasite burdens in groups 2 and 3 were 2,420 (1,205-5,661)/µg gDNA and 22,195 (5,524-100,106)/µg gDNA, respectively. Serial monitoring was undertaken in 45 randomly selected cases that had completed treatment; all cases in groups 2 or 3 had a Ct >30, indicating cure. Overall, qPCR confirmed an 89.2% cure (as 83/93 cases showed parasite clearance), and the persistent qPCR positivity was attributed to nonadherence to treatment or unresponsiveness to miltefosine and remains to be investigated.

Leishmania and HIV co-infection: first naturally Leishmania strain presenting decreased susceptibility to miltefosine, recovered from a patient in Portugal.

BACKGROUND: In Europe, up to 70% of visceral leishmaniasis (VL) cases occurring in adults living with HIV. People living with HIV with VL co-infection often display persistent parasitemia, requiring chronic intermittent anti-Leishmania therapies. Consequently, frequent VL relapses and higher mortality rates are common in these individuals. As such, it is of paramount importance to understand the reasons for parasite persistence to improve infection management. METHODS: To outline possible causes for treatment failure in the context of HIV-VL, we followed a person living with HIV-VL co-infection for nine years in a 12-month period. We characterized: HIV-related clinicopathological alterations (CD4+ T counts and viremia) and Leishmania-specific seroreactivity, parasitemia, quantification of pro-inflammatory cytokines upon stimulation and studied a Leishmania clinical isolate recovered during this period. RESULTS: The subject presented controlled viremia and low CD4+ counts. The subject remained PCR positive for Leishmania and also seropositive. The cellular response to parasite antigens was erratic. The isolate was identified as the first Leishmania infantum case with evidence of decreased miltefosine susceptibility in Portugal. CONCLUSION: Treatment failure is a multifactorial process driven by host and parasite determinants. Still, the real-time determination of drug susceptibility profiles in clinical isolates is an unexplored resource in the monitoring of VL.

Phosphocholine-induced energy source shift alleviates mitochondrial dysfunction in lung cells caused by geospecific PM2.5 components.

Fine particulate matter (PM2.5) is globally recognized for its adverse implications on human health. Yet, remain limited the individual contribution of particular PM2.5 components to its toxicity, especially considering regional disparities. Moreover, prevention solutions for PM2.5-associated health effects are scarce. In the present study, we comprehensively characterized and compared the primary PM2.5 constituents and their altered metabolites from two locations: Taiyuan and Guangzhou. Analysis of year-long PM2.5 samples revealed 84 major components, encompassing organic carbon, elemental carbon, ions, metals, and organic chemicals. PM2.5 from Taiyuan exhibited higher contamination, associated health risks, dithiothreitol activity, and cytotoxicities than Guangzhou's counterpart. Applying metabolomics, BEAS-2B lung cells exposed to PM2.5 from both cities were screened for significant alterations. A correlation analysis revealed the metabolites altered by PM2.5 and the critical toxic PM2.5 components in both regions. Among the PM2.5-down-regulated metabolites, phosphocholine emerged as a promising intervention for PM2.5 cytotoxicities. Its supplementation effectively attenuated PM2.5-induced energy metabolism disorder and cell death via activating fatty acid oxidation and inhibiting Phospho1 expression. The highlighted toxic chemicals displayed combined toxicities, potentially counteracted by phosphocholine. Our study offered a promising functional metabolite to alleviate PM2.5-induced cellular disorder and provided insights into the geo-based variability in toxic PM2.5 components.

Bioinspired zwitterionic triblock copolymers designed for colloidal drug delivery: 1 - Synthesis and characterization.

This study describes the synthesis and characterization of triblock copolymers composed of poly[2-(methacryloyloxy)ethyl phosphorylcholine]-block-poly(propylene glycol)-block-poly[2-(methacryloyloxy)ethyl phosphorylcholine] (PMPC-b-PPG-b-PMPC) intended for, but not limited to, applications in colloidal drug delivery. Atom transfer radical polymerization led to a library of well-defined PMPC-b-PPG-b-PMPC triblock copolymers with varying overall molecular weight (ranging from ∼5 to ∼25 kDa) and composition (weight fraction of the hydrophobic PPG block ranged from ∼10 to ∼50 wt%). The properties of the synthesized triblock copolymers were linked to the PPG to bioinspired PMPC block(s) ratio, where the more hydrophilic species showed adequate aqueous solubility, surface activity and biocompatibility (non-toxicity) in in vitro cell culture. Their amphiphilic nature makes them adsorb efficiently onto polymer nanoparticles, what improves colloidal stability under stress conditions and, furthermore, depletes proteins from unwanted adsorption to the underlying surface. The current findings strengthen our insights into structure-function relationships of PMPC-based coatings leading to protecting shells on relevant polymer nanoparticle formulations. PMPC-b-PPG-b-PMPC triblock copolymers composed of a hydrophobic PPG block of 2-4 kDa flanked by two hydrophilic PMPC blocks each of 5-10 kDa seem to be most promising to enhance colloidal drug delivery vehicles.

Appraisal of Chitosan-Coated Lipid Nano-Combination with Miltefosine and Albendazole in the Treatment of Murine Trichinellosis: Experimental Study with Evaluation of Immunological and Immunohistochemical Parameters.

PURPOSE: Resistance and adverse consequences of albendazole (ABZ) in treating trichinellosis urged demand for secure and effective new drugs. The current study aimed to assess the effect of chitosan-coated lipid nano-combination with albendazole and miltefosine (MFS) in treating experimental murine trichinellosis and evaluating pathological and immunological changes of trichinellosis. MATERIALS AND METHODS: One hundred twenty Swiss albino mice were divided into six groups. Each group was subdivided into a and b subgroups based on the scarification time, which was 7- and 40-days post-infection (PI), respectively. The treatment efficacy was evaluated using parasitological, histopathological, serological (interleukin (IL)-12 and IL-4 serum levels), immunohistochemical (GATA3, glutathione peroxidase1 (GPX1) and caspase-3), and scanning electron microscopy (SEM) methods. RESULTS: The most effective drug was nanostructured lipid carriers (NLCs) loaded with ABZ (G5), which showed the most significant reduction in adults and larval count (100% and 92.39%, respectively). The greatest amelioration in histopathological changes was reported in G4 treated with MFS. GATA3 and caspase-3 were significantly reduced in all treated groups. GPX1 was significantly increased in G6 treated with MFS + NLCs. The highest degenerative effects on adults and larvae by SEM were documented in G6. CONCLUSION: Loading ABZ or MFS on chitosan-coated NLCs enhanced their efficacy against trichinellosis. Although ABZ was better than MFS, their combination should be considered as MFS caused a significant reduction in the intensity of infection. Furthermore, MFS showed anti-inflammatory (↓GATA3) and antiapoptotic effects (↓caspase-3), especially in the muscular phase. Also, when loaded with NLCS, it showed an antioxidant effect (↑GPX1).

Plasma metabolomic profile in orthostatic intolerance children with high levels of plasma homocysteine.

BACKGROUND: Orthostatic intolerance, which includes vasovagal syncope and postural orthostatic tachycardia syndrome, is common in children and adolescents. Elevated plasma homocysteine levels might participate in the pathogenesis of orthostatic intolerance. This study was designed to analyze the plasma metabolomic profile in orthostatic intolerance children with high levels of plasma homocysteine. METHODS: Plasma samples from 34 orthostatic intolerance children with a plasma homocysteine concentration > 9 µmol/L and 10 healthy children were subjected to ultra-high-pressure liquid chromatography and quadrupole-time-of-flight mass spectrometry analysis. RESULTS: A total of 875 metabolites were identified, 105 of which were significantly differential metabolites. Choline, 1-stearoyl-2-linoleoyl-sn-glycero-3-phosphocholine, 1-(1Z-octadecenyl)-2-(4Z,7Z,10Z,13Z,16Z,19Z-docosahexaenoyl)-sn-glycero-3-phosphocholine, histidine, isocitric acid, and DL-glutamic acid and its downstream metabolites were upregulated, whereas 1-palmitoyl-sn-glycero-3-phosphocholine, 1-stearoyl-sn-glycerol 3-phosphocholine, sphingomyelin (d18:1/18:0), betaine aldehyde, hydroxyproline, and gamma-aminobutyric acid were downregulated in the orthostatic intolerance group compared with the control group. All these metabolites were related to choline and glutamate. Heatmap analysis demonstrated a common metabolic pattern of higher choline, 1-stearoyl-2-linoleoyl-sn-glycero-3-phosphocholine, and DL-glutamic acid, and lower sphingomyelin (d18:1/18:0), 1-stearoyl-sn-glycerol 3-phosphocholine, and 1-palmitoyl-sn-glycero-3-phosphocholine in patients with certain notable metabolic changes (the special group) than in the other patients (the common group). The maximum upright heart rate, the change in heart rate from the supine to the upright position, and the rate of change in heart rate from the supine to the upright position of vasovagal syncope patients were significantly higher in the special group than in the common group (P < 0.05). Choline, 1-stearoyl-2-linoleoyl-sn-glycero-3-phosphocholine, and DL-glutamic acid were positively correlated with the rate of change in heart rate from the supine to the upright position in vasovagal syncope patients (P < 0.05). CONCLUSIONS: The levels of choline-related metabolites and glutamate-related metabolites changed significantly in orthostatic intolerance children with high levels of plasma homocysteine, and these changes were associated with the severity of illness. These results provided new light on the pathogenesis of orthostatic intolerance.

Chitosan nanoparticles improve the effectivity of miltefosine against Acanthamoeba.

BACKGROUND: Acanthamoeba keratitis (AK) is a corneal sight-threatening infection caused by the free-living amoebae of the genus Acanthamoeba. Early and appropriate treatment significantly impacts visual outcomes. Mucoadhesive polymers such as chitosan are a potential strategy to prolong the residence time and bioavailability of the encapsulated drugs in the cornea. Regarding the recent administration of miltefosine (MF) for treating resistant AK, in the present study, we synthesized miltefosine-loaded chitosan nanoparticles (MF-CS-NPs) and evaluated them against Acanthamoeba. METHODOLOGY/PRINCIPAL FINDINGS: Chitosan nanoparticles (CNPs) were prepared using the ionic gelation method with negatively charged tripolyphosphate (TPP). The zeta-potential (ZP) and the particle size of MF-CS-NPs were 21.8±3.2 mV and 46.61±18.16 nm, respectively. The release profile of MF-CS-NPs indicated linearity with sustained drug release. The cytotoxicity of MF-CS-NPs on the Vero cell line was 2.67 and 1.64 times lower than free MF at 24 and 48 hours. This formulation exhibited no hemolytic activity in vitro and ocular irritation in rabbit eyes. The IC50 of MF-CS-NPs showed a significant reduction by 2.06 and 1.69-fold in trophozoites at 24 and 48 hours compared to free MF. Also, the MF-CS-NPs IC50 in the cysts form was slightly decreased by 1.26 and 1.21-fold at 24 and 48 hours compared to free MF. CONCLUSIONS: The MF-CS-NPs were more effective against the trophozoites and cysts than free MF. The nano-chitosan formulation was more effective on trophozoites than the cysts form. MF-CS-NPs reduced toxicity and improved the amoebicidal effect of MF. Nano-chitosan could be an ideal carrier that decreases the cytotoxicity of miltefosine. Further analysis in animal settings is needed to evaluate this nano-formulation for clinical ocular drug delivery.

Quantitative Analysis of the Human Semen Phosphorometabolome by 31P-NMR.

Phosphorus-containing metabolites occupy a prominent position in cell pathways. The phosphorometabolomic approach in human sperm samples will deliver valuable information as new male fertility biomarkers could emerge. This study analyzed, by 31P-NMR, seminal plasma and whole semen from asthenozoospermic and normozoospermic samples (71% vs. 27% and 45% vs. 17%, total and progressive sperm motility, respectively), and also ejaculates from healthy donors. At least 16 phosphorus-containing metabolites involved in central energy metabolism and phospholipid, nucleotide, and nicotinamide metabolic pathways were assigned and different abundances between the samples with distinct sperm quality was detected. Specifically, higher levels of phosphocholine, glucose-1-phosphate, and to a lesser degree, acetyl phosphate were found in the asthenozoospermic seminal plasma. Notably, the phosphorometabolites implicated in lipid metabolism were highlighted in the seminal plasma, while those associated with carbohydrate metabolism were more abundant in the spermatozoa. Higher levels of phosphocholine, glucose-1-phosphate, and acetyl phosphate in the seminal plasma with poor quality suggest their crucial role in supporting sperm motility through energy metabolic pathways. In the seminal plasma, phosphorometabolites related to lipid metabolism were prominent; however, spermatozoa metabolism is more dependent on carbohydrate-related energy pathways. Understanding the presence and function of sperm phosphorylated metabolites will enhance our knowledge of the metabolic profile of healthy human sperm, improving assessment and differential diagnosis.