The Turkevich method was optimized to prepare gold nanoparticles (AuNP) stabilized by polyethyleneglycol (PEG) for μCT. Using various independent modalities, we thoroughly characterized the optimized PEG-AuNPs. Here, we show that PEG-AuNPs are retained in the blood and provide a high contrast in the high-resolution μCT imaging of blood vessels and inner organs. The biodistribution is characterized by prolonged circulation in the blood and accumulation in the liver, spleen and skin. The accumulation of AuNP in the skin resulted in the blue discoloration of eyes and the whole skin. In vitro experiments using a leukemic monocyte THP-1 cell line model expressing high levels of NLRP3 demonstrated that the NLRP3inflammasome was not activated by PEG AuNP. Over 9 months, the mice were scanned by μCT and were in good health. Scans in mice using PEG-stabilized AuNPs in this study were sharper, with a higher contrast, when compared to a commercial contrasting agent at the same dose. The PEG-AuNPs were morphologically and chemically stable for at least two years when stored in the refrigerator.
- Publikační typ
- časopisecké články MeSH
Liposomes are one of the most important drug delivery vectors, nowadays used in clinics. In general, polyethylene glycol (PEG) is used to ensure the stealth properties of the liposomes. Here, we have employed hydrophilic, biocompatible and highly non-fouling N-(2-hydroxypropyl) methacrylamide (HPMA)-based copolymers containing hydrophobic cholesterol anchors for the surface modification of liposomes, which were prepared by the method of lipid film hydration and extrusion through 100 nm polycarbonate filters. Efficient surface modification of liposomes was confirmed by transmission electron microscopy, atomic force microscopy, and gradient ultracentrifugation. The ability of long-term circulation in the vascular bed was demonstrated in rabbits after i.v. application of fluorescently labelled liposomes. Compared to PEGylated liposomes, HPMA-based copolymer-modified liposomes did not induce specific antibody formation and did not activate murine and human complement. Compared with PEGylated liposomes, HPMA-based copolymer-modified liposomes showed a better long-circulating effect after repeated administration. HPMA-based copolymer-modified liposomes thus represent suitable new candidates for a generation of safer and improved liposomal drug delivery platforms.
- MeSH
- akrylamidy chemie MeSH
- aktivace komplementu účinky léků MeSH
- cholesterol chemie krev MeSH
- hydrofobní a hydrofilní interakce * MeSH
- králíci MeSH
- lékové transportní systémy MeSH
- lidé MeSH
- liposomy * MeSH
- myši MeSH
- polyethylenglykoly * chemie MeSH
- polymery chemie MeSH
- povrchové vlastnosti * MeSH
- zvířata MeSH
- Check Tag
- králíci MeSH
- lidé MeSH
- mužské pohlaví MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Micro-computed tomography (micro-CT) is an exceptional imaging modality which is limited in visualizing soft biological tissues that need pre-examination contrasting steps, which can cause serious deformation to sizeable specimens like engorged ticks. The aim of this study was to develop a new technique to bypass these limitations and allow the imaging of fed ticks in their natural state. To accomplish this, adult Ixodes ricinus females were allowed to engorge in vitro on blood supplemented with PEGylated gold nanoparticles (PEG-AuNPs). In total, 73/120 females divided into 6 groups engorged on blood enriched with 0.07-2.16 mg PEG-AuNPs per ml of blood. No toxic effect was observed for any of the tested groups compared to the control group, in which 12/20 females engorged on clear blood. The ticks were scanned on a Bruker micro-CT SkyScan 1276. The mean radiodensity of the examined ticks exceeded 0 Hounsfield Units only in the case of the two groups with the highest concentration. The best contrast was observed in ticks engorged on blood with the highest tested concentration of 2.16 mg/mL PEG-AuNPs. In these ticks, the midgut and rectal sac were clearly visible. Also, the midgut lumen volume was computed from segmented image data. The reduction in midgut volume was documented during the egg development process. According to this pilot study, micro-CT of ticks engorged on blood supplemented with contrasting agents in vitro may reveal additional information regarding the engorged ticks' anatomy.
- MeSH
- klíště * MeSH
- kovové nanočástice * MeSH
- krev MeSH
- rentgenová mikrotomografie metody MeSH
- stravovací zvyklosti MeSH
- zlato * MeSH
- zvířata MeSH
- Check Tag
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Background and aims: The majority of colorectal cancers arise from detectable adenomatous or serrated lesions. Here we demonstrate how deregulated alternative splicing of CD44 gene in diseased colon mucosa results in downregulation of standard isoform of CD44 gene (CD44s) and upregulation of variant isoform CD44v8-10. Our aim is to show that upregulation of CD44v8-10 isoform is a possible marker of precancerous lesion in human colon. Methods: We analysed pairs of fresh biopsy specimen of large intestine in a cohort of 50 patients. We studied and compared alternative splicing profile of CD44 gene in colon polyps and adjoined healthy colon mucosa. We performed end-point and qRT PCR, western blotting, IHC staining and flow cytometry analyses. Results: We detected more than five-fold overexpression of CD44v8-10 isoform and almost twenty-fold downregulation of standard isoform CD44s in colon polyps compared to adjoined healthy tissue with p = 0.018 and p < 0.001 in a cohort of 50 patients. Our results also show that aberrant splicing of CD44 occurs in both biologically distinct subtypes of colorectal adenoma possibly in ESRP-1 specific manner. Conclusion: 92% of the colon polyp positive patients overexpressed CD44v8-10 isoform in their colon polyps while only 36% of them had positive fecal occult blood test which is currently a standard non-invasive screening technique. Impact: We believe that our results are important for further steps leading to application of CD44v8-10 isoform as a biomarker of colorectal precancerosis in non-invasive detection. Early detection of colon precancerosis means successful prevention of colorectal carcinoma.
- MeSH
- antigeny CD44 genetika metabolismus MeSH
- kolon metabolismus patologie MeSH
- kolorektální nádory diagnóza genetika metabolismus MeSH
- lidé MeSH
- nádorové biomarkery genetika metabolismus MeSH
- polypy tlustého střeva metabolismus patologie MeSH
- prognóza MeSH
- protein - isoformy MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- MeSH
- alergeny aplikace a dávkování imunologie MeSH
- aplikace slizniční MeSH
- imunologická tolerance MeSH
- lékové transportní systémy MeSH
- ovalbumin aplikace a dávkování imunologie MeSH
- prasata MeSH
- sublinguální imunoterapie metody MeSH
- ústní spodina MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- dopisy MeSH
- práce podpořená grantem MeSH
For group of 281 oocytes obtained from 43 stimulated donors and cryopreserved by vitrification protocol using Cryotop and Kitazato medium we determined important parameters of oocytes collection and vitrification processes which strongly affect the probability that warmed oocytes will produce high-quality embryos for transfer. The probability to obtain high-quality embryos for transfer from vitrified and warmed oocytes was highest when two conditions were fulfilled: 1. oocytes were incubated before vitrification for 7-10 h and 2. stimulated ovaries of donors in one cycle produced a smaller number of oocytes (<7 oocytes from one donor per stimulated cycle). The probable reasons for these observations were: 1. early vitrification (less than 7 h) before final oocyte metaphase II maturation negatively affected the crucial process of post-warm remodelling of spindles and chromosomes, which reduced the fertilization and utilization rates, 2. the evaluated vitrification protocol amplifies negative impact of membrane defects of oocytes of those cohorts containing more than 6 oocytes - freezing places great demands on the integrity and elasticity of the cell membranes. The fact that cryopreservation influences a complex state of oocytes was confirmed by confocal microscopy.
One of the proposed strategies for the development of a more efficient HIV-1 vaccine is based on the identification of proteins binding to a paratope of chosen broadly neutralizing antibody (bNAb) that will mimic cognate HIV-1 Env (glyco)protein epitope and could be used as potent immunogens for induction of protective virus-neutralizing antibodies in the immunized individuals. To verify this "non-cognate ligand" concept, we developed a highly complex combinatorial library designed on a scaffold of human myomesin-1 protein domain and selected proteins called Myomedins specifically binding to variable regions of HIV-1 broadly neutralizing antibody 10E8. Immunization of mice with these Myomedin variants elicited the production of HIV-1 Env-specific antibodies. Hyperimmune sera bound to Env pseudotyped viruses and weakly/moderately neutralized 54% of tested clade A, B, C, and AE pseudotyped viruses variants in vitro. These results demonstrate that Myomedin variants have the potential to mimic Env epitopes and could be used as potential HIV-1 vaccine components.
- MeSH
- epitopy MeSH
- genové produkty env - virus lidské imunodeficience genetika MeSH
- HIV infekce * prevence a kontrola MeSH
- HIV protilátky MeSH
- HIV-1 * genetika MeSH
- myši MeSH
- neutralizující protilátky MeSH
- pseudotypování virů MeSH
- široce neutralizující protilátky MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
