Maternal immune activation results in complex microglial transcriptome signature in the adult offspring that is reversed by minocycline treatment.

Maternal immune activation (MIA) during pregnancy has been linked to an increased risk of developing psychiatric pathologies in later life. This link may be bridged by a defective microglial phenotype in the offspring induced by MIA, as microglia have key roles in the development and maintenance of neuronal signaling in the central nervous system. The beneficial effects of the immunomodulatory treatment with minocycline on schizophrenic patients are consistent with this hypothesis. Using the MIA mouse model, we found an altered microglial transcriptome and phagocytic function in the adult offspring accompanied by behavioral abnormalities. The changes in microglial phagocytosis on a functional and transcriptional level were similar to those observed in a mouse model of Alzheimer's disease hinting to a related microglial phenotype in neurodegenerative and psychiatric disorders. Minocycline treatment of adult MIA offspring reverted completely the transcriptional, functional and behavioral deficits, highlighting the potential benefits of therapeutic targeting of microglia in psychiatric disorders.

Molecular imaging of σ1 receptors in vivo: current status and perspectives.

It is widely accepted that sigma (σ) receptors represent a new and different avenue in the possible pharmacological treatment of cancer and several brain-related disorders. Of the two different σ receptor types the σ1 receptors are assumed to be of major impact for brain diseases. Molecular imaging of brain σ1 receptors with positron emission tomography (PET) or single photon emission computed tomography (SPECT) may provide a significant contribution to the understanding of the cross-talk between σ1 receptors and inter- and intracellular signalling systems. New insights into these functional interrelationships will allow a better diagnosis of brain and cancerous diseases and direct a rational development of new therapeutic concepts.

Alterations of cholinergic receptors and the vesicular acetylcholine transporter after lateral fluid percussion injury in newborn piglets.

AIMS: Traumatic brain injury (TBI) is one of the leading causes of death and disability in children. Adult animal models of TBI showed cholinergic alterations. However, there is no comparable data on immature animals. Therefore, this study investigates cholinergic markers in a large animal model of juvenile TBI. METHODS: Twenty-seven female newborn piglets were subjected to lateral fluid percussion (FP) injury and compared with 12 untreated animals. After 6 h, animals were sacrificed and the brains removed. The hemispheres ipsilateral to FP-TBI from seven piglets and corresponding hemispheres from six control animals were used for autoradiography. Receptor density was determined with [(3)H]epibatidine (nicotinic acetylcholine receptors) or [(3)H]QNB (muscarinic acetylcholine receptors). The density of the vesicular acetylcholine transporter (vAChT) was assessed with (-)-[(3)H]vesamicol. Cerebral blood flow was measured by coloured microsphere method. RESULTS: Cerebral blood flow and brain oxygen delivery were transiently reduced early after FP-TBI (P < 0.05). TBI caused reductions of muscarinic acetylcholine receptor density (fmol/mg) in the basal forebrain (sham: 10797 +/- 1339, TBI: 8791 +/- 1031), while nicotinic acetylcholine receptor remained stable. Significant increases in vAChT density (fmol/mg) were observed in the basal forebrain (sham: 2347 +/- 171, TBI: 2884 +/- 544), putamen (sham: 2276 +/- 181, TBI: 2961 +/- 386), cortex (sham: 1928 +/- 262, TBI: 2377 +/- 294), thalamic areas (sham: 2133 +/- 272, TBI: 2659 +/- 413), hippocampus (sham: 2712 +/- 145, TBI: 3391 +/- 501) and hypothalamus (sham: 2659 +/- 139, TBI: 3084 +/- 304). CONCLUSIONS: Cholinergic markers are altered after mild-to-moderate TBI in the immature brain. Whereas the ACh receptors are stable in almost any brain region after TBI, vAChT expression increases after trauma at the employed severity of this specific trauma model.

Carbon-11 labeled indolylpropylamine analog as a new potential PET agent for imaging of the serotonin transporter.

The synthesis and structure-activity relationship of a new class of indole derivatives with low-nanomolar affinity for the SERT and high selectivity versus the 5-HT1A receptor were recently reported. Based on their chemical structure, four new indolylpropylamine derivatives which contain atoms to afford future labeling with PET isotopes, were synthesized and evaluated as SERT ligands. The chemistry of these novel derivatives, their biological evaluation, the general method of preparing the precursor indole for labeling, and the C-11 labeling of the most promising indole derivative, are described herein.

Norchloro-fluoro-homoepibatidine (NCFHEB) - a promising radioligand for neuroimaging nicotinic acetylcholine receptors with PET.

Cholinergic neurotransmission depends on the integrity of nicotinic acetylcholine receptors (nAChRs), and impairment of both is characteristic for various neurodegenerative diseases. Visualization of specific receptor subtypes by positron emission tomography (PET) has potential to assist with diagnosis of such neurodegenerative diseases and with design of suitable therapeutic approaches. The goal of our study was to evaluate in vivo the potential of (18)F-labelled (+)- and (-)-norchloro-fluoro-homoepibatidine ([(18)F]NCFHEB) in comparison to 2-[(18)F]F-A-85380 as PET tracers. In the brains of NMRI mice, highest levels of radioactivity were detected at 20 min post-injection of (+)-[(18)F]NCFHEB, (-)-[(18)F]NCFHEB, and 2-F-[(18)F]-A-85380 (7.45, 5.60, and 3.2% ID/g tissue, respectively). No marked pharmacological adverse effects were observed at 25 mug NCFHEB/kg. Uptake studies in RBE4 cells and in situ perfusion studies suggest an interaction of epibatidine and NCFHEB with the carrier-mediated choline transport at the blood-brain barrier. The data indicate that (+)- and (-)-[(18)F]NCFHEB have potential for further development as PET tracers.

Glycoxidative stress creates a vicious cycle of neurodegeneration in Alzheimer's disease--a target for neuroprotective treatment strategies?

Accumulation of Advanced Glycation Endproducts (AGEs) in the brain is a feature of ageing and degeneration, especially in Alzheimer's disease (AD). Increased AGE levels explain many of the neuropathological and biochemical features of AD such as extensive protein crosslinking (beta-amyloid and MAP-tau), glial activation, oxidative stress and neuronal cell death. Oxidative stress and AGEs initiate a positive feedback loop, where normal age-related changes develop into a pathophysiological cascade. Combined intervention using antioxidants, anti-inflammatory drugs and AGE-inhibitors may be a promising neuroprotective strategy.

Advanced glycation endproducts co-localize with inducible nitric oxide synthase in Alzheimer's disease.

Advanced glycation endproducts (AGEs), protein-bound oxidation products of sugars, have been shown to be involved in the pathophysiological processes of Alzheimer's disease (AD). AGEs induce the expression of various pro-inflammatory cytokines and the inducible nitric oxide synthase (iNOS) leading to a state of oxidative stress. AGE modification and resulting crosslinking of protein deposits such as amyloid plaques may contribute to the oxidative stress occurring in AD. The aim of this study was to immunohistochemically compare the localization of AGEs and beta-amyloid (Abeta) with iNOS in the temporal cortex (Area 22) of normal and AD brains. In aged normal individuals as well as early stage AD brains (i.e. no pathological findings in isocortical areas), a few astrocytes showed co-localization of AGE and iNOS in the upper neuronal layers, compared with no astrocytes detected in young controls. In late AD brains, there was a much denser accumulation of astrocytes co-localized with AGE and iNOS in the deeper and particularly upper neuronal layers. Also, numerous neurons with diffuse AGE but not iNOS reactivity and some AGE and iNOS-positive microglia were demonstrated, compared with only a few AGE-reactive neurons and no microglia in controls. Finally, astrocytes co-localized with AGE and iNOS as well as AGE and were found surrounding mature but not diffuse amyloid plaques in the AD brain. Our results show that AGE-positive astrocytes and microglia in the AD brain express iNOS and support the evidence of an AGE-induced oxidative stress occurring in the vicinity of the characteristic lesions of AD. Hence activation of microglia and astrocytes by AGEs with subsequent oxidative stress and cytokine release may be an important progression factor in AD.

Advanced glycation endproducts change glutathione redox status in SH-SY5Y human neuroblastoma cells by a hydrogen peroxide dependent mechanism.

The reaction of proteins with reducing sugars leads to the formation of 'advanced glycation endproducts' (AGEs). They accumulate in Alzheimer's disease brain in the vicinity of beta-amyloid plaques. AGEs are cytotoxic by a mechanism involving reactive oxygen species, which implies that they could compromise glutathione redox status. In this study, we show that AGEs (BSA-AGE and beta-amyloid-AGE) persistently increase the ratio of oxidized to reduced glutathione in a dose- and time-dependent manner in SH-SY5Y neuroblastoma cells. The level of oxidized glutathione accounted to 10-14% and persisted for up to 24 h in the presence of added AGEs. In contrast, the unmodified beta-amyloid peptides A beta (1-40) and A beta (25-35) had no significant effect on glutathione redox status. The AGE-induced increase in oxidized glutathione could be prevented by the radical scavengers N-acetylcysteine, alpha-lipoic acid and 17beta-estradiol or by application of catalase, indicating that superoxide and hydrogen peroxide production precedes the AGE-mediated depletion of reduced glutathione.

Differences in the modulating potential of advanced glycation end product (AGE) peptides versus AGE proteins.

Differences in the modulating potential of advanced glycation end product (AGE) peptides versus AGE proteins. Advanced glycation end products (AGEs), identified as irreversible products of a complex reaction of carbonyl groups of reducing sugars with free protein amino groups, are characterized by resistance to proteolytic degradation. The incomplete digestion of AGEs results in low molecular weight AGEs accumulating in the blood of diabetic and uremic patients. We hypothesized that the accumulation of these compounds may contribute to the dysfunction and/or degeneration of tubular epithelial cells. Our study examined whether low-molecular-weight AGE peptides and high-molecular-weight AGE proteins affect the functional cellular properties of two tubular epithelial cell lines: immortalized human kidney tubular epithelial (IHKE) and immortalized rat renal proximal tubular cells (IRPTCs). Parameters of cellular damage and growth behavior were cell counting, analysis of the cellular metabolic activity (MTT assay), as well as cellular proliferation (3[H]-thymidine-incorporation). IHKE treated with bovine serum albumin-AGE (BSA-AGE 50) or BSA-AGE-Pep 50 revealed a decrease in cellular metabolic activity as compared with controls after 48 hours of incubation (73 +/- 9% for BSA-AGE 50 and 62 +/- 11% for BSA-AGE-Pep 50 vs. 89 +/- 8% for BSA Co 50). Low molecular weight BSA-AGE-Pep 50 induced a significantly greater cellular damage in IRPTCs as compared with high molecular weight BSA-AGE 50 after 144 hours of incubation (59 +/- 15% for BSA-AGE 50 vs. 31 +/- 13% for BSA-AGE-Pep 50). The decrease in metabolic activity correlated well with a decrease in cellular proliferation. The results suggest a higher toxic potential of low molecular weight AGE peptides compared with high molecular weight AGE proteins in IRPTC and IHKE. This may provide evidence that low molecular weight degradation products of AGE-modified proteins have an important risk potential.

Protein "AGEing"--cytotoxicity of a glycated protein increases with its degree of AGE-modification.

Non-enzymatic glycation of proteins with reducing sugars and subsequent transition metal-catalyzed oxidations leads to the formation of protein-bound "advanced glycation endproducts" (AGEs). They accumulate on long-lived proteins including on and in the vicinity of the beta-amyloid plaques in Alzheimer's disease (AD). Since the AGE modification of a protein increases with time, and such a "long-term incubation" might also occur in the AD brain, we investigated whether an increase in the cytotoxic effects of an AGE-modified model protein occurs over time. Bovine serum albumin (BSA) was modified by glucose for defined time periods, and the viability of SH-SY5Y neuroblastoma cells, incubated with the differentially AGE-modified BSA samples, was measured with the MTT assay. Cytotoxicity of the AGE-modified BSAs increased in correlation to the incubation time with glucose. Among the AGE-specific markers, browning (OD 400) correlated best with cytotoxicity, followed by AGE-specific fluorescence and the defined AGE, carboxymethyllysine. Since AGEs accumulate in AD over time, they may be one of the "age-related" factors contributing to neuronal cell death in Alzheimer's disease.

In vitro-prepared advanced glycation end-products and the modulating potential of their low-molecular weight degradation products in IRPTC-A rat proximal-tubular derived kidney epithelial cell line.

Low-molecular advanced glycation end-products (AGEs)-degradation products resulting from a proteolysis of tissue or circulating AGEs represent up to 80% of AGE plasma immunoreactivity. These AGE peptides contribute to the dramatic increase in AGE levels in end-stage renal disease even in the absence of diabetes. Because glomerular filtered AGE-degradation products may accumulate within intracellular compartments of proximal tubular epithelial cells, we investigated whether there is a pathway potentially mediating damaging effects of AGE-degradation products by perturbation of the function of the tubuloepithelium. Proximal tubular-derived rat kidney cells (IRPTC) were incubated with high-molecular AGEs highly modified by incubation of bovine serum albumin (BSA) with glucose for 50 days in vitro, and with low-molecular AGE-degradation products derived from proteolytic cleavage and isolated in the molecular range between 1 and 30 kDa. The proliferation of IRPTC (3H-thymidine incorporation) was reduced to 89+/-1% and 69+/-2% after 24 hr of incubation with BSA-AGE and BSA-AGE-degradation products, respectively. The cell viability of IRPTC was reduced significantly to 59+/-15% and 31+/-13% after 144 hr of incubation with BSA-AGE and BSA-AGE-degradation products, respectively. Conditioned media obtained from IRPTC incubated for 72 hr with BSA-AGE and its degradation products increased the proliferation rate of renal fibroblasts (RFb) to 222+/-24% and 449+/-40%, respectively. Incubation of IRPTC with BSA-AGE-degradation products increased the expression of endothelin-1 (ET-1) mRNA to 210% after 1 hr; the expression of platelet-derived growth factor-B (PDGF-B) mRNA reached 184% after 2 hr. Regarding the toxicity of AGEs to the kidney, low-molecular weight AGE-degradation products possibly form an individual fraction with a comparatively higher toxic potential.

Immunochemical detection of imidazolone in uremia and rheumatoid arthritis.

The advanced glycation end-product imidazolone is formed by reaction of arginine with 3-deoxyglucosone (3-DG), a reactive intermediate of the Maillard reaction, whose formation is non-oxidative. Using an antibody specific to this 3-DG-derived AGE, we demonstrated the presence of imidazolone-modified proteins in vivo in the urine and dialysate of patients with chronic renal failure, in the synovial fluid of patients with rheumatoid arthritis, as well as in vitro in human serum and human serum albumin incubated with glucose. Furthermore, we could show that in uremic patients the dimeric form of beta(2)-microglobulin is more susceptible to imidazolone modification than the monomeric one. Thus, the immunochemical detection of imidazolone may be a good marker for 3-DG-derived AGE modification in vivo and in vitro permitting a differentiation between the oxidative and the non-oxidative pathway of AGE generation.

Time course of cytokine mRNA expression in kidneys of rats with unilateral ureteral obstruction.

The development of renal interstitial fibrosis (RIF) is related to the expression and excretion of cytokines and growth factors. Thus, we investigated the time course of mRNA expression of cytokines known as causative factors in a model of RIF in rats before and on day 10 after unilateral ureteral obstruction (UUO), when first signs of fibrosis were visible, as well as during progressive RIF. UUO causes a fivefold increase in mRNA expression of monocyte chemoattractant protein 1 15 days after surgery as compared with contralateral kidneys. The level remains elevated about three-fold up to day 25. The mRNA of the fibrogenic cytokine transforming growth factor beta 1 (TGF-beta1) is increased two- to threefold during the time course, whereas the mRNAs of platelet-derived growth factor B chain (PDGF-B) and its receptor beta (PDGF-Rbeta) increase after UUO, reaching their maxima on days 10-15. PDGF-B mRNA increase up to day 15, marking the onset of fibrosis, and decreases thereafter, whereas the expression of the PDGF-Rbeta mRNA remains elevated more than threefold over the entire study period. Incubation of cultured renal fibroblasts with TGF-beta1 and/or PDGF-B suggests that their specific action on cell growth and proliferation is maintained even when they are used in combination. The sustained elevation of TGF-beta1 and PDGF-B/PDGF-Rbeta mRNA levels confirms the assumption of a particular involvement of these cytokines in the pathogenesis of RIF. The mRNA expression of the gap junctional protein connexin 43 in ureteral ligated kidneys is increased sixfold already 5 days after UUO. In this way, the increased connexin 43 mRNA levels indicate a possible function in the remodeling of the kidney tissue after tubular damage and fibrosis.