Cellular labelling using non-usual magnetic nanoparticles

Mezi běžně používané zobrazovací techniky v klinické praxi patří MRI (zobrazování magnetickou rezonancí). Jedná se o metodu pro pacienta zcela neinvazivní, která umožňuje trojrozměrné zobrazování těla na základě detekce vodíkových atomů molekul vody obsažených v jednotlivých tkáních. Intenzita signálu může být navíc ovlivněna přidáním vhodné kontrastní látky. Před vlastním použitím nově připravených kontrastních agens je nutné provést základní in vitro a in vivo experimenty na buněčných liniích a zvířecích modelech, aby případné nežádoucí účinky těchto nanosond na živé organizmy byly odhaleny ještě dřív, než dojde k jejich zavedení do klinické praxe. Současný rozmach nanotechnologií nabízí širokou škálu nanomateriálů, včetně magnetických nanočástic, určených převážně pro snížení intenzity MRI signálu. Mezi nejběžnější a nejdéle používaná kontrastní agens pro MRI patří superparamagnetické nanočástice oxidů železa (SPIO). V této literární rešerši jsou představeny méně běžné typy magnetických nanočástic, které je také možné použít pro zobrazování magnetickou rezonancí. Klíčová slova: magnetická rezonance, nanočástice, značení buněkMRI (magnetic resonance imaging) belongs to an imaging technique in a clinical practice. It is the method completely non-invasive for a patient, which allows three- dimensional imaging of the body based on the detection of hydrogen atoms of water molecules in individual tissues. Intensity of signal can be further influenced by adding a suitable contrast agent. It is necessary to perform basic in vitro and in vivo experiments on a cell cultures and animal models before a new contrast agents will be introduced into the clinical practice. This is due to potential side effects on living organisms. The current boom in nanotechnology offers a variety of nanomaterials including magnetic nanoparticles for decreasing the intensity of the MRI signal. The most common and longest used contrast agents for MRI are based on superparamagnetic iron oxide nanoparticles (SPIO). In this literature review will be presented uncommon types of magnetic nanoparticles which can also be used for the magnetic resonance imaging. Key words: magnetic resonance imaging, nanoparticle, cellular imagingKatedra buněčné biologieDepartment of Cell BiologyPřírodovědecká fakultaFaculty of Scienc

Magnetic resonance imaging with multimodal T2 contrast agents based on fluorescence-labelled magnetic cores with the spinel structure

Předmětem této diplomové práce byla příprava magnetických fluorescenčně značených nanočástic spinelového typu, konkrétně zinkem dopovaného kobaltnatého feritu, pro zobrazování magnetickou rezonancí a fluorescenční mikroskopii v experimentální praxi. V dřívějších výzkumech vykazovaly různé feritové nanočástice poměrně vysoké transverzální relaxační časy a výrazný negativní T2 kontrast. Tyto vlastnosti byly předpokládány také u nanočástic kobaltnatého feritu dopovaného zinkem. Daný předpoklad byl následně ověřen studiem jejich magnetických a kontrastních vlastností. Dále byly s úspěchem studovány také jejich fluorescenční vlastnosti. V návaznosti na biologické experimenty byla řešena problematika povrchové úpravy magnetických jader připravených nanočástic, problematika týkající se jejich stabilizace ve vodné fázi a v neposlední řadě také toxicita těchto nanočástic v živých systémech. Klíčová slova: nanočástice, ferity, MRI, relaxaceThis diploma thesis was focused on the preparation of magnetic and fluorescently labelled spinel type nanoparticles, specifically nanoparticles of zinc-doped cobalt ferrite intended for application in magnetic resonance imaging and fluorescence microscopy in experimental practice. In previous studies, various ferrite nanoparticles exhibited relatively high transverse relaxation times and strong negative T2 contrast. These properties were also supposed for the zinc-doped cobalt ferrite nanoparticles. This assumption was confirmed by studying their magnetic and contrasting properties. Fluorescence properties of the prepared nanoparticles were also successfully studied. With respect to the intended applications of these particles, the issue of suitable surface modification of magnetic cores, their colloidal stabilization in an aqueous suspension and toxicity in biological systems were studied. Key words: nanoparticles, ferrites, MRI, relaxationKatedra anorganické chemieDepartment of Inorganic ChemistryPřírodovědecká fakultaFaculty of Scienc

Combined Inhibition of Soluble Epoxide Hydrolase and Renin-Angiotensin System Exhibits Superior Renoprotection to Renin-Angiotensin System Blockade in 5/6 Nephrectomized Ren-2 Transgenic Hypertensive Rats with Established Chronic Kidney Disease

Background/Aims: We found recently that increasing renal epoxyeicosatrienoic acids (EETs) levels by blocking soluble epoxide hydrolase (sEH), an enzyme responsible for EETs degradation, shows renoprotective actions and retards the progression of chronic kidney disease (CKD) in Ren-2 transgenic hypertensive rats (TGR) after 5/6 renal ablation (5/6 NX). This prompted us to examine if additional protection is provided when sEH inhibitor is added to the standard renin-angiotensin system (RAS) blockade, specifically in rats with established CKD. Methods: For RAS blockade, an angiotensin-converting enzyme inhibitor along with an angiotensin II type receptor blocker was used. RAS blockade was compared to sEH inhibition added to the RAS blockade. Treatments were initiated 6 weeks after 5/6 NX in TGR and the follow-up period was 60 weeks. Results: Combined RAS and sEH blockade exhibited additional positive impact on the rat survival rate, further reduced albuminuria, further reduced glomerular and tubulointerstitial injury, and attenuated the decline in creatinine clearance when compared to 5/6 NX TGR subjected to RAS blockade alone. These additional beneficial actions were associated with normalization of the intrarenal EETs deficient and a further reduction of urinary angiotensinogen excretion. Conclusion: This study provides evidence that addition of pharmacological inhibition of sEH to RAS blockade in 5/6 NX TGR enhances renoprotection and retards progression of CKD, notably, when applied at an advanced stage

Pharmacological Blockade of Soluble Epoxide Hydrolase Attenuates the Progression of Congestive Heart Failure Combined With Chronic Kidney Disease: Insights From Studies With Fawn-Hooded Hypertensive Rats

An association between congestive heart failure (CHF) and chronic kidney disease (CKD) results in extremely poor patient survival rates. Previous studies have shown that increasing kidney epoxyeicosatrienoic acids (EETs) by blocking soluble epoxide hydrolase (sEH), an enzyme responsible for EETs degradation, improves the survival rate in CHF induced by aorto-caval fistula (ACF) and attenuates CKD progression. This prompted us to examine if sEH inhibitor treatment would improve the outcome if both experimental conditions are combined. Fawn-hooded hypertensive (FHH) rats, a genetic model showing early CKD development was employed, and CHF was induced by ACF. Treatment with an sEH inhibitor was initiated 4 weeks after ACF creation, in FHH and in fawn-hooded low-pressure (FHL) rats, a control strain without renal damage. The follow-up period was 20 weeks. We found that ACF FHH rats exhibited substantially lower survival rates (all the animals died by week 14) as compared with the 64% survival rate observed in ACF FHL rats. The former group showed pronounced albuminuria (almost 30-fold higher than in FHL) and reduced intrarenal EET concentrations. The sEH inhibitor treatment improved survival rate and distinctly reduced increases in albuminuria in ACF FHH and in ACF FHL rats, however, all the beneficial actions were more pronounced in the hypertensive strain. These data indicate that pharmacological blockade of sEH could be a novel therapeutic approach for the treatment of CHF, particularly under conditions when it is associated with CKD

Epoxyeicosatrienoic Acid-Based Therapy Attenuates the Progression of Postischemic Heart Failure in Normotensive Sprague-Dawley but Not in Hypertensive Ren-2 Transgenic Rats

Epoxyeicosatrienoic acids (EETs) and their analogs have been identified as potent antihypertensive compounds with cardio- and renoprotective actions. Here, we examined the effect of EET-A, an orally active EET analog, and c-AUCB, an inhibitor of the EETs degrading enzyme soluble epoxide hydrolase, on the progression of post-myocardial infarction (MI) heart failure (HF) in normotensive Hannover Sprague-Dawley (HanSD) and in heterozygous Ren-2 transgenic rats (TGR) with angiotensin II-dependent hypertension. Adult male rats (12 weeks old) were subjected to 60-min left anterior descending (LAD) coronary artery occlusion or sham (non-MI) operation. Animals were treated with EET-A and c-AUCB (10 and 1 mg/kg/day, respectively) in drinking water, given alone or combined for 5 weeks starting 24 h after MI induction. Left ventricle (LV) function and geometry were assessed by echocardiography before MI and during the progression of HF. At the end of the study, LV function was determined by catheterization and tissue samples were collected. Ischemic mortality due to the incidence of sustained ventricular fibrillation was significantly higher in TGR than in HanSD rats (35.4 and 17.7%, respectively). MI-induced HF markedly increased LV end-diastolic pressure (Ped) and reduced fractional shortening (FS) and the peak rate of pressure development [+(dP/dt)max] in untreated HanSD compared to sham (non-MI) group [Ped: 30.5 ± 3.3 vs. 9.7 ± 1.3 mmHg; FS: 11.1 ± 1.0 vs. 40.8 ± 0.5%; +(dP/dt)max: 3890 ± 291 vs. 5947 ± 309 mmHg/s]. EET-A and c-AUCB, given alone, tended to improve LV function parameters in HanSD rats. Their combination amplified the cardioprotective effect of single therapy and reached significant differences compared to untreated HanSD controls [Ped: 19.4 ± 2.2 mmHg; FS: 14.9 ± 1.0%; +(dP/dt)max: 5278 ± 255 mmHg/s]. In TGR, MI resulted in the impairment of LV function like HanSD rats. All treatments reduced the increased level of albuminuria in TGR compared to untreated MI group, but neither single nor combined EET-based therapy improved LV function. Our results indicate that EET-based therapy attenuates the progression of post-MI HF in HanSD, but not in TGR, even though they exhibited renoprotective action in TGR hypertensive rats

Magnetic resonance imaging with multimodal T2 contrast agents based on fluorescence-labelled magnetic cores with the spinel structure

This diploma thesis was focused on the preparation of magnetic and fluorescently labelled spinel type nanoparticles, specifically nanoparticles of zinc-doped cobalt ferrite intended for application in magnetic resonance imaging and fluorescence microscopy in experimental practice. In previous studies, various ferrite nanoparticles exhibited relatively high transverse relaxation times and strong negative T2 contrast. These properties were also supposed for the zinc-doped cobalt ferrite nanoparticles. This assumption was confirmed by studying their magnetic and contrasting properties. Fluorescence properties of the prepared nanoparticles were also successfully studied. With respect to the intended applications of these particles, the issue of suitable surface modification of magnetic cores, their colloidal stabilization in an aqueous suspension and toxicity in biological systems were studied. Key words: nanoparticles, ferrites, MRI, relaxatio

Cellular labelling using non-usual magnetic nanoparticles

MRI (magnetic resonance imaging) belongs to an imaging technique in a clinical practice. It is the method completely non-invasive for a patient, which allows three- dimensional imaging of the body based on the detection of hydrogen atoms of water molecules in individual tissues. Intensity of signal can be further influenced by adding a suitable contrast agent. It is necessary to perform basic in vitro and in vivo experiments on a cell cultures and animal models before a new contrast agents will be introduced into the clinical practice. This is due to potential side effects on living organisms. The current boom in nanotechnology offers a variety of nanomaterials including magnetic nanoparticles for decreasing the intensity of the MRI signal. The most common and longest used contrast agents for MRI are based on superparamagnetic iron oxide nanoparticles (SPIO). In this literature review will be presented uncommon types of magnetic nanoparticles which can also be used for the magnetic resonance imaging. Key words: magnetic resonance imaging, nanoparticle, cellular imagin

Cellular labelling using non-usual magnetic nanoparticles

MRI (magnetic resonance imaging) belongs to an imaging technique in a clinical practice. It is the method completely non-invasive for a patient, which allows three- dimensional imaging of the body based on the detection of hydrogen atoms of water molecules in individual tissues. Intensity of signal can be further influenced by adding a suitable contrast agent. It is necessary to perform basic in vitro and in vivo experiments on a cell cultures and animal models before a new contrast agents will be introduced into the clinical practice. This is due to potential side effects on living organisms. The current boom in nanotechnology offers a variety of nanomaterials including magnetic nanoparticles for decreasing the intensity of the MRI signal. The most common and longest used contrast agents for MRI are based on superparamagnetic iron oxide nanoparticles (SPIO). In this literature review will be presented uncommon types of magnetic nanoparticles which can also be used for the magnetic resonance imaging. Key words: magnetic resonance imaging, nanoparticle, cellular imagin

Fluorescent magnetic nanoparticles for cell labeling: Flux synthesis of manganite particles and novel functionalization of silica shell

International audienceNovel synthetic approaches for the development of multimodal imaging agents with high chemical stability are demonstrated. The magnetic cores are based on La0.63Sr0.37MnO3 manganite prepared as individual grains using a flux method followed by additional thermal treatment in a protective silica shell allowing to enhance their magnetic properties. The cores are then isolated and covered de novo with a hybrid silica layer formed through the hydrolysis and polycondensation of tetraethoxysilane and a fluorescent silane synthesized from rhodamine, piperazine spacer, and 3-iodopropyltrimethoxysilane. The aminoalkyltrialkoxysilanes are strictly avoided and the resulting particles are hydrolytically stable and do not release dye. The high colloidal stability of the material and the long durability of the fluorescence are reinforced by an additional silica layer on the surface of the particles. Structural and magnetic studies of the products using XRD, TEM, and SQUID magnetometry confirm the importance of the thermal treatment and demonstrate that no mechanical treatment is required for the flux-synthesized manganite. Detailed cell viability tests show negligible or very low toxicity at concentrations at which excellent labeling is achieved. Predominant localization of nanoparticles in lysosomes is confirmed by immunofluorescence staining. Relaxometric and biological studies suggest that the functionalized nanoparticles are suitable for imaging applications

Effects of renal sympathetic denervation on the course of congestive heart failure combined with chronic kidney disease: Insight from studies with fawn-hooded hypertensive rats with volume overload induced using aorto-caval fistula

Background: The coincidence of congestive heart failure (CHF) and chronic kidney disease (CKD) results in poor survival rate. The aim of the study was to examine if renal denervation (RDN) would improve the survival rate in CHF induced by creation of aorto-caval fistula (ACF). Methods: Fawn-hooded hypertensive rats (FHH), a genetic model of spontaneous hypertension associated with CKD development, were used. Fawn-hooded low-pressure rats (FHL), without CKD, served as controls. RDN was performed 4 weeks after creation of ACF and the follow-up period was 10 weeks. Results: We found that intact (non-denervated) ACF FHH exhibited survival rate of 58.8% (20 out of 34 rats), significantly lower than in intact ACF FHL (81.3%, 26/32 rats). In intact ACF FHL albuminuria remained stable throughout the study, whereas in ACF FHH it increased significantly, up to a level 40-fold higher than the basal values. ACF FHL did not show increases in renal glomerular and tubulointerstitial injury as compared with FHL, while ACF FHH exhibited marked increases in kidney injury as compared with FHH. RDN did not improve the survival rate in either ACF FHL or ACF FHH and did not alter the course of albuminuria in ACF FHL. RDN attenuated the albuminuria, but did not reduce the kidney injury in ACF FHH. Conclusions: Our present results support the notion that even modest CKD increases CHF-related mortality. RDN did not attenuate CHF-dependent mortality in ACF FHH, it delayed the progressive rise in albuminuria, but it did not reduce the degree of kidney injury