Małgorzata Frontczak-Baniewicz

Synaptic degeneration in rat brain after prolonged oral exposure to silver nanoparticles.

Neurotoxicity of silver nanoparticles has been confirmed in both in vitro and in vivo studies. However, the mechanisms of the toxic action have not been fully clarified. Since nanoparticles are likely to have the ability to enter the brain and significantly accumulate in this organ, it is important to investigate their neurotoxic mechanisms. Here we examine the effect of prolonged exposure of rats to small (10nm) citrate-stabilized silver nanoparticles (as opposed to the ionic silver) on synapse ultrastructure and specific proteins. Administration of both nanosilver and ionic silver over a two-week period resulted in ultrastructural changes including blurred synapse structure and strongly enhanced density of synaptic vesicles clustering in the center of the presynaptic part. Disturbed synaptic membrane leading to liberation of synaptic vesicles into neuropil, which testifies for strong synaptic degeneration, was characteristic feature observed under AgNPs exposure. Also a noteworthy finding was the presence of myelin-like structures derived from fragmented membranes and organelles which are associated with neurodegenerative processes. Additionally, we observed significantly decreased levels of the presynaptic proteins synapsin I and synaptophysin, as well as PSD-95 protein which is an indicator of postsynaptic densities. The present study demonstrates that exposure of adult rats to both forms of silver leads to ultrastructural changes in synapses. However, it seems that small AgNPs lead to more severe synaptic degeneration, mainly in the hippocampal region of brain. The observations may indicate impairment of nerve function and, in the case of hippocampus, may predict impairment of cognitive processes.

Absence of Glial α-Dystrobrevin Causes Abnormalities of the Blood-Brain Barrier and Progressive Brain Edema

Background: Functional blood-brain barrier requires interactions between endothelia and astrocytes, but molecules involved in these contacts are not known. Results: Absence of glial α-dystrobrevin protein causes leaky blood-brain barrier, water retention, and progressive brain edema. Conclusion: Glial α-dystrobrevin is essential for endothelium-astrocyte interactions required for blood-brain barrier functions. Significance: Pathologies altering α-dystrobrevin might lead to blood-brain barrier abnormalities. The blood-brain barrier (BBB) plays a key role in maintaining brain functionality. Although mammalian BBB is formed by endothelial cells, its function requires interactions between endotheliocytes and glia. To understand the molecular mechanisms involved in these interactions is currently a major challenge. We show here that α-dystrobrevin (α-DB), a protein contributing to dystrophin-associated protein scaffolds in astrocytic endfeet, is essential for the formation and functioning of BBB. The absence of α-DB in null brains resulted in abnormal brain capillary permeability, progressively escalating brain edema, and damage of the neurovascular unit. Analyses in situ and in two-dimensional and three-dimensional in vitro models of BBB containing α-DB-null astrocytes demonstrated these abnormalities to be associated with loss of aquaporin-4 water and Kir4.1 potassium channels from glial endfeet, formation of intracellular vacuoles in α-DB-null astrocytes, and defects of the astrocyte-endothelial interactions. These caused deregulation of tight junction proteins in the endothelia. Importantly, α-DB but not dystrophins showed continuous expression throughout development in BBB models. Thus, α-DB emerges as a central organizer of dystrophin-associated protein in glial endfeet and a rare example of a glial protein with a role in maintaining BBB function. Its abnormalities might therefore lead to BBB dysfunction.

Is the Poly (L- Lactide- Co– Caprolactone) Nanofibrous Membrane Suitable for Urinary Bladder Regeneration?

The purpose of this study was to compare: a new five-layered poly (L–lactide–co–caprolactone) (PLC) membrane and small intestinal submucosa (SIS) as a control in rat urinary bladder wall regeneration. The five-layered poly (L–lactide–co–caprolactone) membrane was prepared by an electrospinning process. Adipose tissue was harvested from five 8-week old male Wistar rats. Adipose derived stem cells (ADSCs) were seeded in a density of 3×106 cells/cm2 onto PLC membrane and SIS scaffolds, and cultured for 5-7 days in the stem cell culture medium. Twenty male Wistar rats were randomly divided into five equal groups. Augmentation cystoplasty was performed in a previously created dome defect. Groups: (I) PLC+ 3×106ADSCs; (II) SIS+ 3×106ADSCs; (III) PLC; (IV) SIS; (V) control. Cystography was performed after three months. The reconstructed urinary bladders were evaluated in H&E and Massons trichrome staining. Regeneration of all components of the normal urinary bladder wall was observed in bladders augmented with cell-seeded SIS matrices. The urinary bladders augmented with SIS matrices without cells showed fibrosis and graft contraction. Bladder augmentation with the PLC membrane led to numerous undesirable events including: bladder wall perforation, fistula or diverticula formation, and incorporation of the reconstructed wall into the bladder lumen. The new five-layered poly (L–lactide–co–caprolactone) membrane possesses poorer potential for regenerating the urinary bladder wall compared with SIS scaffold.

SERPENTINE-SMECTITE INTERSTRATIFIED MINERALS FROM LOWER SILESIA (SW POLAND)

Interstratified serpentine-sm ectite was found in the fine-grained fraction of altered metasomatic contact biotite-schists developed between serpentinite and granite-type rocks (Lower Silesia ophiolite sequence, Poland). Ni-rich serpentine-smectite is R0-interstratified lizardite (0.80)-stevensite (0.15)-vermiculite-like (0.05), with a coherent scattering domain (csd) of 5 layers (mean value). The Mg-rich variety of serpentine-smectite is R1 lizardite (0.80)-stevensite (0.20) with a csd size of 7 layers (mean value). A transmission electron microscope study revealed complex layer relationships, with zones composed of various serpentine-smectite packets having lizardite/smectite ratios of 3:1, 2:1, 1:1, 4:1 and scarce serpentine segregations. In both cases, the serpentine-smectites appear to be late products of alteration of the parent biotite-schist.Experimental and calculated positions and intensities of reflections of the ethylene glycol-saturated, heated (250°C, thermal stage), and air-dried samples are in good agreement. Calculated X-ray diffraction patterns for interstratified glycolated and anhydrous serpentine-smectite are included in the Appendix.

The experimental squalene encephaloneuropathy in the rat

To investigate the influence of squalene on the nervous system, adult male Wistar rats were injected with squalene subcutaneously with 20 g/kg of the body weight for 4 consecutive days. After 7 or 30 days from the initiation of the experiment, brain and ischiadic nerves were harvested for electron microscopy. Squalene affected mostly PNS targeting Schwann cells and myelin sheaths. Accumulation of lipid-like droplets in the myelin sheaths in the PNS and in the neurons in the brain cortex, hypertrophy of endothelium, and sometimes endothelial apoptosis in blood vessels, and increased synthesis of collagen in the ischiadic nerve were characteristic for developed squalene encephaloneuropathy.

Repeated exposure of adult rats to Aroclor 1254 induces neuronal injury and impairs the neurochemical manifestations of the NMDA receptor-mediated intracellular signaling in the hippocampus.

Aroclor 1254 is a mixture of polychlorinated biphenyls (PCBs), a class of environmental toxins which cause a wide spectrum of neurotoxic effects. Learning and memory deficits are the profound effects of PCBs which may be related to hippocampal dysfunction. To get insight into the underlying neurochemical mechanisms, we employed the microdialysis technique to investigate the effect of repeated exposure of adult male Wistar rats to Aroclor 1254 (10mg/kg b.w., daily, ig., for 14days), on the neurochemical parameters of NMDA receptor-mediated glutamatergic signaling in the hippocampus in vivo assessed using the microdialysis technique. The results demonstrated that exposure to Aroclor 1254, which was associated with substantial neuronal damage and loss in the hippocampus, markedly decreased the NMDA-induced extracellular accumulation of newly loaded (45)CaCl(2), cGMP and glutamate, and reduced the basal content of the NO precursor, arginine, indicating inhibition of the NMDA/NO/cGMP pathway. Aroclor 1254 exposure also decreased the basal microdialysate content of glutamate and glutamine, which may cause inadequate supply of the neurotransmitter glutamate, while the level of two other neuroactive amino acids, aspartate or taurine was not affected by the exposure. The results underscore neuronal lesion and inhibition of NMDA receptor-mediated glutamatergic signaling in hippocampus as a potential major contributor to the cognitive deficits associated with exposure to PCB.

Ureter regeneration-the proper scaffold has to be defined.

The aim of this study was to compare two different acellular scaffolds: natural and synthetic, for urinary conduit construction and ureter segment reconstruction. Acellular aortic arch (AAM) and poly(L-lactide-co-caprolactone) (PLCL) were used in 24 rats for ureter reconstruction in both tested groups. Follow-up period was 4 weeks. Intravenous pyelography, histological and immunohistochemical analysis were performed. All animals survived surgical procedures. Patent uretero-conduit junction was observed only in one case using PLCL. In case of ureter segment reconstruction ureters were patent in one case using AAM and in four cases using PLCL scaffolds. Regeneration of urothelium layer and focal regeneration of smooth muscle layer was observed on both tested scaffolds. Obtained results indicates that synthetic acellular PLCL scaffolds showed better properties for ureter reconstruction than naturally derived acellular aortic arch.

Perinatal exposure to lead (Pb) induces ultrastructural and molecular alterations in synapses of rat offspring

Lead (Pb), environmentally abundant heavy-metal pollutant, is a strong toxicant for the developing central nervous system. Pb intoxication in children, even at low doses, is found to affect learning and memorizing, with devastating effects on cognitive function and intellectual development. However, the precise mechanism by which Pb impairs synaptic plasticity is not fully elucidated. The purpose of this study was to investigate the effect of pre- and neonatal exposure to low dose of Pb (with Pb concentrations in whole blood below 10μg/dL) on the synaptic structure and the pre- and postsynaptic proteins expression in the developing rat brain. Furthermore, the level of brain-derived neurotrophic factor (BDNF) was analyzed. Pregnant female Wistar rats received 0.1% lead acetate (PbAc) in drinking water from the first day of gestation until weaning of the offspring, while the control animals received drinking water. During the feeding of pups, mothers from the Pb-group were continuously receiving PbAc. Pups of both groups were weaned at postnatal day 21 and then until postnatal day 28 received only drinking water. 28-Day old pups were sacrificed and the ultrastructural changes as well as expression of presynaptic (VAMP1/2, synaptophysin, synaptotagmin-1, SNAP25, syntaxin-1) and postsynaptic (PSD-95) proteins were analyzed in: forebrain cortex, cerebellum and hippocampus. Our data revealed that pre- and neonatal exposure to low dose of Pb promotes pathological changes in synapses, including nerve endings swelling, blurred and thickened synaptic cleft structure as well as enhanced density of synaptic vesicles in the presynaptic area. Moreover, synaptic mitochondria were elongated, swollen or shrunken in Pb-treated animals. These structural abnormalities were accompanied by decrease in the level of key synaptic proteins: synaptotagmin-1 in cerebellum, SNAP25 in hippocampus and syntaxin-1 in cerebellum and hippocampus. In turn, increased level of synaptophysin was noticed in the cerebellum, while the expression of postsynaptic PSD-95 was significantly decreased in forebrain cortex and cerebellum, and raised in hippocampus. Additionally, we observed the lower level of BDNF in all brain structures in comparison to control animals. In conclusion, perinatal exposure to low doses of Pb caused pathological changes in nerve endings associated with the alterations in the level of key synaptic proteins. All these changes can lead to synaptic dysfunction, expressed by the impairment of the secretory mechanism and thereby to the abnormalities in neurotransmission as well as to the neuronal dysfunction.Lead (Pb), environmentally abundant heavy-metal pollutant, is a strong toxicant for the developing central nervous system. Pb intoxication in children, even at low doses, is found to affect learning and memorizing, with devastating effects on cognitive function and intellectual development. However, the precise mechanism by which Pb impairs synaptic plasticity is not fully elucidated. The purpose of this study was to investigate the effect of pre- and neonatal exposure to low dose of Pb (with Pb concentrations in whole blood below 10μg/dL) on the synaptic structure and the pre- and postsynaptic proteins expression in the developing rat brain. Furthermore, the level of brain-derived neurotrophic factor (BDNF) was analyzed. Pregnant female Wistar rats received 0.1% lead acetate (PbAc) in drinking water from the first day of gestation until weaning of the offspring, while the control animals received drinking water. During the feeding of pups, mothers from the Pb-group were continuously receiving PbAc. Pups of both groups were weaned at postnatal day 21 and then until postnatal day 28 received only drinking water. 28-day old pups were sacrificed and the ultrastructural changes as well as expression of presynaptic (VAMP1/2, synaptophysin, synaptotagmin-1, SNAP25, syntaxin-1) and postsynaptic (PSD-95) proteins were analyzed in: forebrain cortex, cerebellum and hippocampus. Our data revealed that pre- and neonatal exposure to low dose of Pb promotes pathological changes in synapses, including nerve endings swelling, blurred and thickened synaptic cleft structure as well as enhanced density of synaptic vesicles in the presynaptic area. Moreover, synaptic mitochondria were elongated, swollen or shrunken in Pb-treated animals. These structural abnormalities were accompanied by decrease in the level of key synaptic proteins: synaptotagmin-1 in cerebellum, SNAP25 in hippocampus and syntaxin-1 in cerebellum and hippocampus. In turn, increased level of synaptophysin was noticed in the cerebellum, while the expression of postsynaptic PSD-95 was significantly decreased in forebrain cortex and cerebellum, and raised in hippocampus. Additionally, we observed the lower level of BDNF in all brain structures in comparison to control animals. In conclusion, perinatal exposure to low doses of Pb caused pathological changes in nerve endings associated with the alterations in the level of key synaptic proteins. All these changes can lead to synaptic dysfunction, expressed by the impairment of the secretory mechanism and thereby to the abnormalities in neurotransmission as well as to the neuronal dysfunction.

Influence of a low dose of silver nanoparticles on cerebral myelin and behavior of adult rats.

Nanoscale particles have large surface to volume ratio that significantly enhances their chemical and biological reactivity. Although general toxicity of nano silver (nanoAg) has been intensively studied in both in vitro and in vivo models, its neurotoxic effects are poorly known, especially those of low-dose exposure. In the present study we assess whether oral administration of nanoAg influences behavior of exposed rats and induces changes in cerebral myelin. We examine the effect of prolonged exposure of adult rats to small (10nm) citrate-stabilized nanoAg particles at a low dose of 0.2mg/kg b.w. (as opposed to the ionic silver) in a comprehensive behavioral analysis. Myelin ultrastructure and the expression of myelin-specific proteins are also investigated. The present study reveals slight differences with respect to behavioral effects of Ag(+)- but not nanoAg-treated rats. A weak depressive effect and hyperalgesia were observed after Ag(+) exposure whereas administration of nanoAg was found to specifically increase body weight and body temperature of animals. Both nanoAg and Ag(+) induce morphological disturbances in myelin sheaths and alter the expression of myelin-specific proteins CNP, MAG and MOG. These results suggest that the CNS may be a target of low-level toxicity of nanoAg.

The role of invasive diagnostics and its impact on the treatment of dilated cardiomyopathy: A systematic review

BACKGROUND Dilated cardiomyopathy is one of the most frequent causes of non-ischemic heart failure. Many factors including genetic disorders, infectious agents, toxins, drugs and autoimmune disorders might take part in the development of dilated cardiomyopathy. Diagnosis of left ventricular dilatation is most often limited to performing echocardiography and excluding ischemic etiology (coronary angiography). Since many pathologies take place at the cellular and subcellular level the only way to clarify the etiology of the disease is to examine the myocardium itself (endomyocardial biopsy). METHODS A systematic literature search was conducted for studies published between September 2000 and September 2015 using the PubMed database. RESULTS Of 7104 studies identified, 73 studies were included in this review. Controversies raised by opponents of the endomyocardial biopsy collide with the low percentage of serious complications confirmed in several single-center registries. Based on the available data the overall complication rate varies from 1% to about 3%, with 0.5% risk of serious complications. According to the current recommendations of the European and American scientific societies endomyocardial biopsy should be performed in most cases of left ventricular dilatation and heart failure of non-ischemic etiology. Endomyocardial biopsy allows for making the diagnosis and providing prognostic information especially in patients with familial dilated cardiomyopathy, diabetic cardiomyopathy with dilated phenotype, alcoholic cardiomyopathy, peripartum cardiomyopathy, iron overload cardiomyopathy, as well as inflammatory and viral cardiomyopathy. Iron overload cardiomyopathy, peripartum cardiomyopathy, inflammatory and viral cardiomyopathy are potentially treatable and reversible. CONCLUSIONS Targeted therapies are more effective when started early before myocardial injury becomes irreversible. Unfortunately, non-invasive techniques are not precise enough to decide if and which targeted therapy is required. Therefore endomyocardial biopsy should be mainly recognized as the essential diagnostic tool and should not be postponed.