Localization of heat shock proteins in cerebral cortical cultures following induction by celastrol.

Hsp70, Hsp32, and Hsp27 were induced by celastrol in rat cerebral cortical cultures at dosages that did not affect cell viability. Pronounced differences were observed in the cellular localization of these heat shock proteins in cell types of cerebral cortical cultures. Celastrol-induced Hsp70 localized to the cell body and cellular processes of neurons that were identified by neuron-specific ßIII-tubulin. Hsp70 was not detected in adjacent GFAP-positive glial cells that demonstrated a strong signal for Hsp27 and Hsp32 in both glial cell bodies and cellular processes. Cells in the cerebral cortex region of the brain are selectively impacted during the progression of Alzheimer's disease which is a "protein misfolding disorder." Heat shock proteins provide a line of defense against misfolded, aggregation-prone proteins. Celastrol is a potential agent to counter this neurodegenerative disorder as recent evidence indicates that in vivo administration of celastrol in a transgenic model of Alzheimer's reduces an important neuropathological hallmark of this disease, namely, amyloid beta pathology that involves protein aggregation.

Biological activity of sym-triazines with acetylcholine-like substitutions as multitarget modulators of Alzheimer's disease.

The bioactivities of two novel compounds (TAE-1 and TAE-2) that contain a sym-triazine scaffold with acetylcholine-like substitutions are examined as promising candidate agents against Alzheimer's disease. Inhibition of amyloid-ß fibril formation in the presence of Aß1-42, evaluated by Thioflavin T fluorescence, demonstrated comparable or improved activity to a previously reported pentapeptide-based fibrillogenesis inhibitor, iAß5p. Destabilization of Aß1-42 assemblies by TAE-1 and TAE-2 was confirmed by scanning electron microscopy imaging. sym-Triazine inhibition of acetylcholinesterase (AChE) activity was observed in cytosol extracted from differentiated human SH-SY5Y neuronal cells and also using human erythrocyte AChE. The sym-triazine derivatives were well tolerated by these cells and promoted beneficial effects on human neurons, upregulating expression of synaptophysin, a synaptic marker protein, and MAP2, a neuronal differentiation marker.

sym-Triazines for directed multitarget modulation of cholinesterases and amyloid-ß in Alzheimer's disease.

Alzheimer's disease (AD) is a complex neurodegenerative disorder marked by numerous causative factors of disease progression, termed pathologies. We report here the synthesis of a small library of novel sym-triazine compounds designed for targeted modulation of multiple pathologies related to AD, specifically human acetylcholinesterase (AChE), butyrylcholinesterase (BuChE), and Aß aggregation. Rational targeting of AChE was achieved by the incorporation of acetylcholine substrate analogues into a sym-triazine core in either a mono-, di-, or trisubstituted regime. A subset of these derivatives demonstrated improved activity compared to several commercially available cholinesterase inhibitors. High AChE/BuChE selectivity was characteristic of all derivatives, and AChE steady-state kinetics indicated a mixed-type inhibition mechanism. Further integration of multiple hydrophobic phenyl units allowed for improved ß-sheet intercalation into amyloid aggregates. Several highly effective structures exhibited fibril inhibition greater than the previously reported ß-sheet-disrupting penta-peptide, iAß5p, evaluated by thioflavin T fluorescence spectroscopy and transmission electron microscopy. Highly effective sym-triazines were shown to be well tolerated by differentiated human neuronal cells, as demonstrated by the absence of adverse effects on cellular viability at a wide range of concentrations. Parallel targeting of multiple pathologies using sym-triazines is presented here as an effective strategy to address the complex, multifactorial nature of AD progression.

Induction of heat shock proteins in cerebral cortical cultures by celastrol.

Alzheimer's disease, Parkinson's disease and amyotrophic lateral sclerosis (ALS) are 'protein misfolding disorders' of the mature nervous system that are characterized by the accumulation of protein aggregates and selective cell loss. Different brain regions are impacted, with Alzheimer's affecting cells in the cerebral cortex, Parkinson's targeting dopaminergic cells in the substantia nigra and ALS causing degeneration of cells in the spinal cord. These diseases differ widely in frequency in the human population. Alzheimer's is more frequent than Parkinson's and ALS. Heat shock proteins (Hsps) are 'protein repair agents' that provide a line of defense against misfolded, aggregation-prone proteins. We have suggested that differing levels of constitutively expressed Hsps (Hsc70 and Hsp27) in neural cell populations confer a variable buffering capacity against 'protein misfolding disorders' that correlates with the relative frequencies of these neurodegenerative diseases. The high relative frequency of Alzheimer's may due to low levels of Hsc70 and Hsp27 in affected cell populations that results in a reduced defense capacity against protein misfolding. Here, we demonstrate that celastrol, but not classical heat shock treatment, is effective in inducing a set of neuroprotective Hsps in cultures derived from cerebral cortices, including Hsp70, Hsp27 and Hsp32. This set of Hsps is induced by celastrol at 'days in vitro' (DIV) 13 when cultured cortical cells reached maturity. The inducibility of a set of neuroprotective Hsps in mature cortical cultures at DIV13 suggests that celastrol is a potential agent to counter Alzheimer's disease, a neurodegenerative 'protein misfolding disorder' of the adult brain that targets cells in the cerebral cortex.

Oxidative stress effect of dopamine on α-synuclein: electroanalysis of solvent interactions.

The interaction of dopamine (DA) and α-synuclein (α-S) can lead to protein misfolding and neuronal death triggered by oxidative stress relevant to the progression of Parkinson's disease (PD). In this study, interfacial properties associated with DA-induced α-S aggregation under various solution conditions (i.e., pH, ionic strength) were investigated in vitro. The electrochemical oxidation of tyrosine (Tyr) residues in α-S was detected in the presence of DA. DA concentration dependence was analyzed and found to significantly affect α-S aggregation pathways. At low pH, DA was shown to be stable and produced no observable difference in interfacial properties. Between pH 7 and 11, DA promoted α-S aggregation. Significant differences in oxidation current signals in response to high pH and ionic strength suggested the importance of initial interactions in the stabilization of toxic oligomeric structures and subsequent off-pathways of α-S. Our results demonstrate the importance of solution interactions with α-S and the unique information that electrochemical techniques can provide for the investigation of α-S aggregation at early stages, an important step toward the development of future PD therapeutics.

Hsp60 protein pattern in coral is altered by environmental changes in light and temperature.

The heat shock protein Hsp60 exhibited marked oscillation during a 12-hour day period when the coral Turbinaria reniformis was maintained in the laboratory under constant conditions of light (200µE) and temperature (27°C). A biphasic pattern of Hsp60 was apparent, punctuated by a low protein level at the midpoint of the 12-hour day period. Oscillation of Hsp60 was also apparent when coral was kept in darkness in lieu of a scheduled light period. The pattern of Hsp60 was altered when coral was exposed to increased light intensity (400µE) or temperature elevation (32°C). These observations suggest that Hsp60 in coral exhibits oscillation that is altered by increased light and temperature elevation.