Research

Cellular mechanisms of neurodegenerative diseases

Neurodegenerative disease is a common term for a range of conditions which primarily affect the neurons in the brain and the spinal cord resulting in their injury and death.

Clinical MRI was used to assess neurodegenerative processes in the hSOD-1 G93A ALS rat model and in the trimethyltin (TMT)-treated model of Alzheimer's-like disease. T2-weighted (T2W) hyperintensive neurodegenerative foci were found in the brainstem of the ALS rat with apparent lateral ventricle dilation. Degenerative processes in these areas were also confirmed by confocal images of GFAP-positive astrogliosis. MRI after i.v.i. of magnetic anti-CD4 antibodies indicated an accumulation of inflammatory cells near dilated ventricles. TMT-treated rats also revealed the dilation of lateral ventricles. Immunocytochemistry could reveal significant redistribution of macro- and microglia in the hippocampus. In both models, Gd-DTPA contrast revealed a compromised blood-brain barrier that could serve as the passage for inflammatory immune cells in the vicinity of dilated lateral ventricles. Moreover, in both models the midbrain region of the dorsal hippocampus was the target of the blood-brain barrier compromise, thus revealing a potentially vulnerable point that can be the primary target of neurodegeneration in the CNS.

Cerebral ischemic injury resulting from either focal or global circulatory arrests in the brain is one of the major causes of death and disability in the adult population. The hippocampus, playing important roles in learning and memory, is selectively vulnerable to ischemic insults. Distinct populations of hippocampal neurons are targeted by ischemia and multiple factors, including excitotoxicity, oxidative stress, and inflammation, are responsible for their damage and death. Modifications of synapses occur very early after ischemia, reflecting related changes in synaptic transmission. These modifications structurally relate to spatial patterns formed by synaptic vesicles, geometry of postsynaptic density, and so forth. Ischemia-induced changes of synaptic contacts can be implicated in the mechanisms leading to delayed neuronal death. We are interested in the structural aspects of ischemic injury in tissue culture and animal models and in the pathways of neurodegeneration common for cerebral ischemia and various neurodegenerative disorders.

Neurogenesis is a process that includes not only generation of new neurons from neural stem and progenitor cells, but functional integration of these new born neurons into cortical circuits as well. The subventricular zone of the lateral ventricles and the subgranular zone of the dentate gyrus in the hippocampus are the sites of active adult neurogenesis in most mammals. However, after pathological stimulation, e.g. ischemic brain insult, neurogenesis can appear even in those areas that were previously considered non-neurogenic, such as the striatum and cerebral cortex. This effect is most likely mediated by microglial cells, the main effector cells in postischemic inflammatory processes. The discovery of active adult neurogenesis opens new possibilities for repair of the adult CNS after injury or neurodegenerative diseases, however, thorough investigation of the influence of the neuroinflammatory environment on the neurogenic niche is needeed to better understand the CNS regenerative capabilities under neurodegenerative circumstances, and to finding more successful ways of using adult neurogenesis as a therapeutic approach.

  • Andjus, P.R., Bataveljić D., Vanhoutte, G., Mitrecic, D., Pizzolante, F., Djogo, N., Nicaise, C., Gankam Kengne, F., Gangitano, C., Michetti, F., Van der Linden, A., Pochet, R., Bačić, G. (2009) Anatom. Rec. 292:1882-1892.
  • Nikonenko, A.G. Radenovic L., Andjus, P.R., Skibo, G.G.(2009) Anatom. Rec. 292:1914-1921.
  • Stamenkovic, S., Sekeljic, V., Radenovic, L., Andjus, P.R (2012) Ischemia and neurogenesis - link between neurodegeneration and repair. In: Neurogenesis Research: New Developments, Eds: GJ Clark and WT Anderson, Nova Publishers, Inc., pp 115-135

Neurobiophysics of dynamic interactions between neurons and glia

Multiple spatial and temporal scales of neurons-glia interactions are closely intertwined and they lie at the core of the macroscopic pathologies.

In our ALS studies immunocytochemistry revealed microglial activation and fusion, possibly phagocytic interactions with neurons in the hippocampus and brainstem.

In our long lived ischemic rat model (according to prof. R. Pluta) immunocytochemistry revealed microglial activation in the hippocampus and striatum, with indications of activation in thalamic lateral dorsal nuclei and the subventricular zone. In the CA1 and CA3 regions, it was noted that microglia/macrophage specific (OX42- and ED1-positive) granules appear in neuronal somata.

Astrocytes can tolerate longer periods of oxygen and glucose deprivation as compared to neurons. The reasons for this reduced vulnerability are not well understood. Particularly, changes in mitochondrial membrane potential in astrocytes, an indicator of the cellular redox state, have not been investigated during reperfusion after extended oxygen glucose deprivation. Our in vitro model studies provide novel information regarding the role of the two main substrates of electron transport chain (glucose and oxygen) and their hyperpolarizing effect on the mitochondrial membrane during substrate deprivation that sheds new light on mechanisms of astrocyte resilience to prolonged ischemic injury. Our studies also show that autophagy and lipolysis are essential for the survival of astrocytes under nutrient deprivation this being related to their role as neuron-supporting cells.

  • Andjus, P.R., Bataveljić D., Vanhoutte, G., Mitrecic, D., Pizzolante, F., Djogo, N., Nicaise, C., Gankam Kengne, F., Gangitano, C., Michetti, F., Van der Linden, A., Pochet, R., Bačić, G. (2009) Anatom. Rec. 292:1882-1892.
  • Sekeljic, V., Bataveljic, D., Stamenkovic, S., Ułamek, M., Jabłoński, M., Radenovic, L., Pluta, R., Andjus, P.R. (2012) Brain Structure and Function, 217: 411-420
  • Korenic A, Boltze J, Deten A, Peters M, Andjus P and Radenovic L. (2014) PLoS ONE 9: e90697.
  • Korenić A, Andjus P, Radenović L, Spasojević I (2015) Neurosci Lett. 595:128-133.

Biomarkers in neurodegenerative processes and conditions

The main goal is finding a reliable in vivo biomarker for diagnosis, prognosis and follow-up of the therapeutic response in neurodegeneration.

Neuroinflammation has gained a particular focus as a key mechanism in ALS. Several studies in vivo and in vitro have nominated IgG isolated from ALS patients as an active contributor to disease onset and progression. We have shown that ALS IgG affects astroglial Ca2+ excitability and induces downstream activation of phosphatidylinositol 3-kinase. These studies were hampered by a lack of knowledge of the pathway of entry of immune factors in the CNS. Our MRI data revealed the blood-brain barrier leakage and T cell infiltration into brain parenchyma in ALS hSODG93A rats. MRI of ultra-small paramagnetic iron oxide (USPIO) nanoparticles labeled T cells revealed CD4+ lymphocyte infiltration in the midbrain-interbrain region while the CD8+ cells were more confined to the brainstem region. By way of Gd-contrast it was also confirmed that the blood-brain barrier was compromised and that the regions of blood-brain barrier breakthrough were congruent with the MRI foci of T cell infiltration.

Since astrocytes ensheath blood vessel wall contributing to blood-brain barrier stability and play an important role in ALS pathogenesis, we have studied astrocytic membrane proteins water channel aquaporin-4 and the inwardly rectifying potassium channel. We study the impaired function of astrocytes in ALS and the implications of membrane proteins expressed on astrocytic endfeet, aquaporin-4, and inwardly rectifying potassium channel. In addition to ALS-specific IgGs, these membrane proteins are proposed as novel biomarkers of the disease.

In addition, microglial activation was studied by immunocytochemistry. In the model of long lived (1 year) ischemic rat (according to prof. R. Pluta) MRI showed: 1) blood-brain barrier leakage in the area of the dorsal hippocampus and brainstem-hindbrain level in basal cerebellum, 2) unlike anti-CD8 magnetic antibodies, anti-CD4 USPIO antibodies revealed infiltrated areas in the brainstem-interbrain region and caudoputamen, and 3) a ventricle dilation in the retrosplenial area. Immunostaining of lymphocytes confirmed the T-cell presence in the hippocampus and striatum. Moreover, in these animals an augmented expression of neurogenesis markers and neuroblast migration was also revealed in the subventricular zone. Thus, a balance of degenerative processes and inflammatory surveillance with neurogenesis could determine the long-term outcome of global ischemia survival or the previously proposed formation of amyloid plaques and Alzheimer's-type dementia.

  • Bataveljić D., Djogo N., Župunski Lj., Bajić, A., Nicaise C., Pochet P., Bačić G., Andjus P.R. (2009) Gen. Physiol. Biophys. 28:212–218
  • Bataveljić, D., Stamenković, S., Bačić, G., Andjus, P.R. (2011) Acta Physiologica Hungarica, 98:27-31.
  • Sekeljic, V., Bataveljic, D., Stamenkovic, S., Ułamek, M., Jabłoński, M., Radenovic, L., Pluta, R., Andjus, P.R. (2012) Brain Structure and Function, 217: 411-420
  • Bataveljić D., Nikolić Lj., Milošević M., Todorović N., Andjus P. (2012) Glia 60:1991-2003.
  • Milošević M., Stenovec M., Kreft M., Petrušić V., Stević Z., Trkov S., Andjus P.R.,Zorec R. (2013) Cell Calcium. 54:17-25.

The role of extracellular matrix in structural plasticity

Our main protein of interest, Tenascin C is a large extracellular matrix glycoprotein up-regulated during embryogenesis, in wound healing, and in tumor tissues, while its cleavage is performed by matrix metalloproteinases thus promoting neuronal structural and functional plasticity. Understanding of the contribution of this extracellular matrix constituent to the major developmental processes such as cell proliferation and migration, axonal guidance, as well as synaptic plasticity, is derived from studies on our Tenascin C - deficient mice. Studies on these mice demonstrated that Tenascin C plays an important role in neuronal plasticity in the cerebral cortex, hippocampus and cerebellum, possibly by modulating the activity of L-type voltage-dependent Ca2+ channels.

Since not much is known about the role of tenascins in neuroinflammation, we are also interested in studying the interplay between neural and immune cells in autoimmune diseases, such as multiple sclerosis and polyneuropathies.

  • Andjus P.R., Bajić A., Zhu L., Schachner, M., Strata P. (2005) Ann. N.Y. Acad. Sci. 1048, 185–197.
  • Šekeljić V & Andjus PR (2012) Int J Biochem Cell Biol 44:825-829.
  • Jakovcevski I., Miljkovic D., Schachner M., Andjus P.R. (2013) Tenascins and inflammation in disorders of the nervous system. Amino Acids 44:1115-1127.
  • Senkov O, Andjus P, Radenovic L, Soriano E, Dityatev A. (2014) Prog Brain Res. 214:53-80.