1,721,004 research outputs found
Effect of liposomal resveratrol on sperm cell motility
Get PDFBackground:
Resveratrol is found to have antioxidative properties and is reported as a protective pharmacological agent against several diseases. It is however a lipophilic and unstable substance because of its photosensitivity. In order to enhance the drug stability, solubility and to deliver the drug in the targeted area, it can be incorporated in a nano-sized vesicle of liposome formulations. The aim of this study was to identify whether the antioxidants such as resveratrol liposomal formulation has some in vitro effects on the motility and viability in human spermatozoa. Well known antioxidants such as vitamin C and vitamin E, were also included for comparison.
Materials and methods:
The liposomal formulation of resveratrol was prepared in collaboration with the Department of Pharmacy. Four drug samples, resveratrol-loaded liposomes, empty liposomes, vitamin C and vitamin E were used at three concentrations (1, 10, and 100 ug/mL). Seventeen semen samples obtained from the IVF clinic, University Hospital of Northern Norway (UNN) were studied. After purification of the semen samples, sperm cells were treated with specified drug concentration in a 96 well plate. The motility of the sperm cells was observed with the microscope after 24 and 48 hours incubation at 5% CO2 and 37C. The current work was approved by regional ethic committee (REK), REK-2014/032 REK Nord.
Results:
Among more than 90% motile spermatozoa immediately after purification decreased to 35.5% after 24 hours under the normal in vitro conditions. Sperm cells containing antioxidants were found to be significantly less motile and significantly more immotile as compared to controls. No significant differences were seen between empty liposomes and control groups. After 48 hours of incubation, in the control group, 9.3% of spermatozoa were motile. Sperm cells containing the drug samples (empty liposomes, resveratrol, vitamin C and vitamin E) were significantly less motile and significantly more immotile especially at the highest concentration (100 ug/mL) as compared to the control.
Conclusion:
The direct antioxidant treatment on sperm cells did not improve the motility or viability of spermatozoa in vitro. Although we did not study the use of antioxidants for the IVF outcome, the use of such antioxidant substances in the semen sample purification media should be avoided in the IVF procedure
In vitro effects of alcohol and oxidative stress on sperm cell motility and viability
Get PDFThe deteriorating semen quality is one of the increasing problems of male reproductive health worldwide and the exact cause of this has not been fully understood yet. Many stress factors may affect semen quality such as sperm cell motility and viability and thus impair the fertilization outcome. Among the different stress factors, alcohol can inhibit steroidogenesis, suppresses LH release from the pituitary gland, decrease the levels of testosterone, induce apoptosis of sperm cells and reduce the anti-oxidant defence systems. However, it is not clear so far that the chronic and low dose effects of alcohol and oxidative stress on the fate of sperm cell motility and viability. A method of identification of alcohol or oxidative stress affected sperm cells by using artificial intelligence technique could be the useful tool for the treatment of assisted reproductive technology (ART). Such technology has not been available for the clinical application so far. The main objective of the study is to observe the effects of various concentrations of alcohol and hydrogen peroxide on the motility and viability of sperm cells at different time periods
Promising Pharmaceuticals
Get PDFFrom the dawn of civilization, humans have been dreaming of happy, healthy and long-life. Our life expectancy is twice longer than 100 years ago. We know more about the diseases. Therefore we have developed new drugs to fight against them. The demand for drugs was so high that we developed Pharma industries. Although Pharma industries took responsibility of producing the needed drugs and gave us a quality of life, misuse of drugs brought further complication. Therefore, discovery, production, distribution, and the phase of administration of patients' quality assurance has to be controlled with a technological procedure and tight regulations to make the system as effective as possible for the benefit of human health. Our book provides selected but vital information on the sources, tools, technologies and regulations regarding the current status of medicine development
Advanced methods in reproductive medicine: Application of optical nanoscopy, artificial intelligence-assisted quantitative phase microscopy and mitochondrial DNA copy numbers to assess human sperm cells
Get PDFDeclined fertility rate and population is a matter of serious concern, especially in the developed nations. Assisted Reproductive Technologies (ART), including in vitro fertilization (IVF), have provided great hope for infertility treatment and maintaining population growth and social structure. With the help of ART, more than 8 million babies have already been born so far. Despite the worldwide expansion of ART, there is a number of open questions on the IVF success rates. Male factors for infertility contribute equally as female factors, however, male infertility is primarily focused on the “semen quality”. Therefore, the search of new semen parameters for male fertility evaluation and the exploration of the optimal method of sperm selection in IVF have been included among the top 10 research priorities for male infertility and medically assisted reproduction. The development of imaging systems coupled with image processing by Artificial Intelligence (AI) could be the revolutionary step for semen quality analysis and sperm cell selection in IVF procedures.
For this work, we applied optical nanoscopy technology for the analysis of human spermatozoa, i.e., label-based Structured Illumination Microscopy (SIM) and non-invasive Quantitative Phase Microscopy (QPM). The SIM results demonstrated a prominent contrast and resolution enhancement for subcellular structures of living sperm cells, especially for mitochondria-containing midpiece, where features around 100 nm length-scale were resolved. Further, non-labeled QPM combined with machine learning technique revealed the association between gradual progressive motility loss and the morphology changes of the sperm head after external exposure to various concentrations of hydrogen peroxide. Moreover, to recognize healthy and stress-affected sperm cells, we applied Deep Neural Networks (DNNs) to QPM images achieving an accuracy of 85.6% on a dataset of 10,163 interferometric images of sperm cells. Additionally, we summarized the evidence from published literature regarding the association between mitochondrial DNA copy numbers (mtDNAcn) and semen quality.
To conclude, we set up the high-resolution imaging of living human sperm cells with a remarkable level of subcellular structural details provided by SIM. Next, the morphological changes of sperm heads resulting from peroxidation have been revealed by QPM, which may not be explored by microscopy currently used in IVF settings. Besides, the implementation of DNNs for QPM image processing appears to be a promising tool in the automated classification and selection of sperm cells during IVF procedures. Moreover, the results of our meta-analysis showed an association of mtDNAcn in human sperm cells and semen quality, which seems to be a relevant sperm parameter for routine clinical practice in male fertility assessment
Optimisation of high-resolution fluorescence microscopy of normal- and preeclamptic placentas
Get PDFPreeclampsia (PE) affects 3-5 % of pregnant women and may lead to maternal and/or fetal death. The main theory of PE is placental ischemia, leading to a dysfunctional placenta and clinical signs as hypertension and proteinuria in the mother. The primary aim of the thesis was to implement and optimise a method for high-resolution microscopy of placental cryo-sections. Secondary aims were to compare the morphology, total antioxidant capacity (TAC) and the oxidative stress between normal- and preeclamptic placentas. Placental tissue from the fetal and maternal side were collected from three normal pregnant women and three preeclamptic women. For each patient; eight cryo-sections were prepared, four from each side of the placenta. Two were used as negative controls investigated for autofluorescence and two were used as positive controls labelled for morphological analysis. Positive controls were labelled with CellMaskTM Orange, staining cell membranes and 4’,6 diamidino 2-phenylidole, dihydrochloride, staining nuclei. The TAC was determined by comparing the measured 3 ethylbenzothiazoline 6 sulphonic acid radical scavenging activity to an ascorbic acid standard curve. The oxidative stress was determined measuring the malondialdehyde content of the samples. Neither the normal nor the preeclamptic samples had autofluorescence affecting microscopy of the labelled sections. The method allowed visualisation of microscopic placental structures. In preeclamptic sections from the fetal side, there seemed to be more syncytial knots than in fetal sections from normal women. Bright red structures were detected in sections from the fetal side of preeclamptic samples and were not observed in normal sections. Because of their size, they were thought to be extravillous vesicles. The collection-, preservation- and labelling method was successfully implemented and is well suited for high-resolution microscopy. Although there were not found a significant difference in TAC and oxidative stress between normal- and preeclamptic placentas, neither on the fetal- or maternal side, the method is suited for placental tissue
Development of liposomal formulation for green tea catechins targeted for the treatment of vaginal inflammation
Get PDFGreen tea catechins are well known for their antioxidative and anti-inflammatory properties. In this connection, catechins are thought to be potential candidates in prevention and treatment of diseases caused by oxidative damage. In addition, catechins have been shown to be effective in the treatment of genital warts. However, they exhibit low bioavailability and poor solubility, which limits their use. This project focused on development of liposomal delivery system containing catechin and epicatechin, respectively, as mean to circumvent their physiochemical limitations and to enhance their biological activity. The formulation was intended for treatment of genital warts/vaginal inflammation. To become closer to an actual in vivo application, drug-bearing liposomes were incorporated in a mucoadhesive hydrogel. The optimized liposomal formulation for catechin was sonicated for 2 minutes and had an average diameter of 150 nm. These vesicles entrapped approximately 50 % of catechin. The corresponding liposomes prepared with epicatechin were found to have smaller size, however the entrapment was similar. Liposomes containing catechin were further incorporated in a hydrogel vehicle and the hydrogel was found to exhibit satisfactory adhesiveness and cohesiveness. In vitro release of catechin from liposomal preparations (both in a form of suspension and incorporated in hydrogel) was performed on Franz diffusion cells under condition simulating both the vaginal environment and physiological conditions. Liposomal hydrogels were found to provide sustained release of catechin. In conclusion, we can confirm that encapsulation of catechins in liposomes and their subsequent incorporation in hydrogels provides potentials for development of novel drug delivery systems for vaginal therapy with catechin
Can nanomedicine improve the semen quality? The potentials of liposomal curcumin.
Get PDFINTRODUCTION: Worldwide, 10-15 % couples are infertile, and 50 % of the couples seeking assisted reproductive techniques do so because of defective sperm quality. The semen quality can be directly linked to oxidative stress (OS). Due to high metabolic rates and weak anti-oxidant systems, sperm cells are unable to defend against OS produced exogenously and endogenously. Vitamin C and/or vitamin E are anti-oxidants showing promising outcome to reduce OS, but the clinical application has not been fully achieved. Curcumin is an extremely potent lipid-soluble anti-oxidant showing potential in therapy for different human disorders, but its poor bioavailability has limited its wider use. Nanotechnology has been used for the purpose of improving the bioavailability of curcumin. It would be interesting to apply nanotechnology based liposomal delivery of curcumin, a noted anti-oxidant for the enhancement of motility and viability of human sperm cells in vitro.
OBJECTIVES: With this study, the aim was to prove anti-oxidative activities of curcumin. We also studied if liposomal formulation of curcumin could maximize the anti-oxidative activities and hopefully enhance motility and viability of sperm cells in vitro.
METHODS: Curcumin loaded liposomes were made by the film method followed by extrusion. Anti-oxidative activities of free curcumin were tested with ABTS and DPPH assay methods and compared with vitamin C and vitamin E. The effect of empty liposomes, liposomal curcumin, free curcumin, vitamin C and vitamin E was determined by observing the motility and viability of sperm cells in vitro.
RESULTS: Liposomes with curcumin had an average size of 147.82 ±17.3 nm and entrapment efficiency of 88.84 ± 2.3 %. The EC50 for curcumin was 5.82 µM in ABTS and EC50 9.13 µM in DPPH assay, respectively. Anti-oxidants such as curcumin, vitamin C and vitamin E maintained the motility and viability up to 24 hours, however lipid showed toxicity to sperm cells at 0.5 µg/ml or higher concentrations.
CONCLUSION: The liposomes containing curcumin were uniformly distributed with high entrapment efficiency. We confirmed anti-oxidative activity of curcumin, however, rather unexpectedly that the liposomal lipid has shown negative effect on sperm viability at certain concentrations. Further studies with more replicates are needed to confirm these findings.
Key words: curcumin, liposomes, anti-oxidant, sperm, motility, viabilit
Investigation of inflammatory response of macrophages and trophoblasts using structured illumination microscopy
Get PDFApproximately 800 women die from pregnancy or childbirth-related pregnancy complications around the world every day. One of the major reasons for these complications is inflammation following infection. Approximately 40% cases of preterm births occur due to microbial invasion in the genitourinary track. During inflammation, macrophages present at the feto-maternal junction release an increased amount of nitric oxide (NO) and other important pro-inflammatory cytokines: TNF-alpha and INF-gamma. This can disturb the trophoblast function which can lead to pregnancy complications, such as abortion, pre-eclampsia and birth defects.
Here I aimed to study the cellular and sub-cellular morphological changes in macrophages (RAW264.7) and trophoblasts (HTR-8/SVneo) following externally supplied inflammatory agents: lipopolysaccharides (LPS), concanavalin-A (Con-A) and tumor necrosis factor alpha (TNF-alpha), in vitro.
Nitric oxide (NO) produced by the cells under inflammatory response was measured quantitatively using the Griess reagent test. Morphological changes were examined in response to LPS, Con-A and TNF-alpha challenge on live macrophages and trophoblasts using structured illumination microscopy (SIM) and quantitative phase microscopy (QPM).
LPS-challenged macrophages produced approximately 22 folds more NO as compared to controls, whereas no significant increase was seen after TNF-alpha or Con-A-challenge. No significantly increased amount of NO was detected in trophoblasts following LPS, TNF-alpha or Con-A-challenge. Superoxide produced during respiration by mitochondria may react with NO to produce reactive nitrogen species (RNS) e.g. peroxynitrite (ONOO-), which have strong damaging effects on the mitochondria.
SIM imaging showed changes in the morphology of mitochondria and plasma membrane in approximately 50-60% of macrophages following LPS challenge (1μg/ml for 24 h), but no detectable changes in mitochondria or plasma membrane were observed after TNF-alpha (1ng/ml for 24 hr) or Con-A challenge (1μg/ml for 24 hr). QPM revealed that the phase value decreased by approximately 18% in LPS-challenged macrophages after 24 hr as compared to controls. In contrast, mitochondrial morphology appeared different in trophoblasts following TNF-alpha challenge under similar conditions. No effect was seen in trophoblast morphology following either LPS or Con-A challenge. Through SIM and QPM it was found that the effect on the morphology of macrophages can be detected following 2hr of incubation with LPS, whereas The Griess method was unable to detect increased amounts of NO before 4hr.
Our results suggest that the cells which are responsible for the mother-fetus crosstalk, especially macrophages and trophoblasts, respond differently to various inflammatory agents. Additionally, SIM and QPM are shown to be live-cell friendly, useful tools to evaluate sub-cellular mechanisms associated with inflammation mediated pregnancy complications
Antioxidant activity of selected natural medicine used in Nepal
Get PDFDiminished antioxidant defence or increased production of reactive oxygen and nitrogen species in the biological system can result into oxidative stress which can cause damage to deoxyribonucleic acid (DNA), proteins, lipids and as a result different disease states arise like cancer, neurodegenerative diseases, rheumatoid arthritis. Antioxidants from different plant resources can significantly delay or prevent oxidation of the substrate and hence prevents from various diseases. Therefore, present research was focused in search of potent natural antioxidants. For the study, methanolic extracts of twenty-five common natural medicines, mostly spices were screened using 1,1–diphenyl2–picrylhydrazyl (DPPH) radical for their antioxidative activities. Among them, extracts of Chebulae Fructus, Terminalia Billericae Fructus, Phyllanthi Fructus, Cinnamomi Cortex, Arecae Semen, Pericarpium Punicae Granati, Syzygiae Fructus, Rhei Rhizoma, Pterocarpi Lignum and Santali Lignum Albi showed potent antioxidative activity with EC50 values being 1.5, 2.1, 1.4, 2, 1.5, 1.45, 2.7, 2.9, 3, 3.8 μg/mL, respectively.
Ascorbic acid (EC50: 2.6 μg/mL) was used as positive control. Therefore, consumers can increase their intake of foods rich in antioxidant compounds that can lower the risk of chronic health problems
- …
