Browsing by Subject "UPR"
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(2015)Endoplasmic reticulum stress (ER-stress) is the result of accumulation of unfolded and misfolded proteins in the ER. The unfolded proteins activate the unfolded protein response (UPR), which seeks to reduce the protein load in the ER and reduces ER-stress. When ER-stress is prolonged, the UPR will activate apoptosis. Amyotrophic lateral sclerosis (ALS) is a rare, progressive neurodegenerative disease that affects lower and higher motorneurons. The cause of ALS is unknown but ER-stress is known to play a role in the disease progression. CDNF is a new neurotrophic factor, which is known to play a role in protein folding in the ER. CDNF is neuroprotective and neurorestorative in animal models of Parkinson's disease. Thus, CDNF is a potential new drug candidate for treating ALS. The aim of this work was to examine the effect of CDNF on disease state and life span in transgenic SOD1(G93A)-mice. CDNF or PBS was injected into the mouse's ventricle in stereotaxic surgery when the mice were about 90 days old. Clinical status and motor coordination was monitored twice a week throughout the study. The mice were dissected when they reached the end point that was set for the study. Deepfrozen gastrocnemius muscles were stained with antibodies, to examine the integrity of the neuromuscular junctions (NMJ). Quantitative PCR (qPCR) was executed on deepfrozen spinal cord and motor cortex samples to measure the expression of ER-stress genes. The results showed that CDNF improves motor coordination and delays disease progression in SOD1 female mice. The NMJs were notably more damaged in SOD1 mice than in wild type mice, but CDNF did not have any significant effect on NMJ integrity. ER-stress could be observed in the spinal cord and motor cortex of SOD1 mice and CDNF decreased ER-stress in the motor cortex. CDNF did not decrease ER-stress in the spinal cord where the expression of apoptosis related genes was increased. Thus, CDNF is a potential new drug candidate for treating ALS and it should be studied further.
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(2021)Parkinson´s disease (PD) is the second most common neurodegenerative disease in the world after Alzheimer´s disease. There is still no drug that alters the state of the disease. It has been found that Endoplasmic reticulum (ER) stress is one mechanism in PD. ER stress occurs due to accumulation of unfolded proteins. ER stress triggers Unfolded protein response (UPR) that protects against ER stress by decreasing unfolding of proteins. In the beginning, UPR has protective effect, but in prolonged ER stress UPR triggers apoptotic cell death. There are several key mediators in the UPR pathway. Characterisation of ER stress in PD models may be important for the current and future drug development of PD. If ER stress is a significant factor that affects the disease development, it would be important to find a drug that alters these mechanisms and UPR. This may be a way to halt the disease development. Different animal models of PD, like 6-OHDA (6-hydroxydopamine) and MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) model, have similarities in their mechanisms. It has been found that ER stress occurs both in the brain of PD patients and animal models of PD. That is why studying and further characterisation in animal models is relevant. The aim of this study was to characterize ER stress in 6-OHDA rat model. The expression of some key mediators of the UPR were determined in this study. There were male and female Spraque Dawley rats in this experiment. 6-OHDA or saline was injected intrastriatally in 3 spots by stereotaxic surgery. Two weeks after 6-OHDA lesions, amphetamine-induced rotation test was conducted to the rats. The rats were divided into groups based on lesion size according to the results. For this study, the rats were euthanised at week 2 or week 4 post lesion. The rats were euthanised by carbondioxide, and the death was confirmed by decapitation. The brains were collected and stored in -80°C. Striatum and substantia nigra were collected later. Total RNA was isolated from these samples. Part of the RNA sample was used to conduct cDNA synthesis. Finally, the gene expression of Atf4, Ire1α, Xbp1s, Xbp1t, Grp78 and Chop was measured from these cDNA samples by qPCR (quantitative polymerase chain reaction). The qPCR data describes the expression of exact gene. The data was processed prior to statistical analysis. By statistical analysis, it was possible to compare the expression of these genes between 6-OHDA group and vehicle group. In addition, comparison was made between 6-OHDA treated groups at week 2 and 4. According to the results, only Chop expression had increased in 6-OHDA lesioned rats at week 2 compared to the vehicle group. In other genes there were no statistical differences, unlike in several other studies where the expression was found to be increased. Thus, the characterisation of this model requires further studying, possibly by increasing the sample size and studying later time points as well.
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(2024)Multiple sclerosis (MS) is an inflammatory neurodegenerative disease of the central nervous system, that affects over 2 million people worldwide. In MS, there is an autoimmune reaction against the myelin surrounding axons, causing neurological deficits and progressive neurological deterioration over time. Currently, there is no curative treatment for MS; only disease-modifying treatments are available. In this study, the effect of C-MANF (C-terminal mesencephalic astrocyte-derived neurotrophic factor) on unfolded protein response (UPR), neuroinflammation and myelination was investigated in different models of myelination by analyzing gene expression with quantitative polymerase chain reaction (qPCR). C-MANF is a C-terminal fragment of endoplasmic reticulum (ER) located protein MANF, which has been shown to protect cells against ER-stress induced apoptosis. UPR is a cellular defense mechanism against ER stress, but when prolonged, it can direct cells into apoptosis. The effect of C-MANF was investigated in ER-stressed Oli-neu cell cultures, both undifferentiated and differentiated. Cells were treated for 5 hours with thapsigargin to induce ER stress and simultaneously with C-MANF. In differentiated cells, a longer 24 hour exposure to ER stress and C-MANF was also investigated. Organotypic cerebellar brain slices from C57BL/6 P10 mice were demyelinated with lysophosphatidylcholine (LPC) for 17 hours, after which C-MANF was given during 5 day remyelination period. After the experiments were conducted, RNA was isolated and converted to cDNA for quantitative polymerase chain reaction (qPCR) using SYBR green. In addition, spinal cords from experimental autoimmune encephalomyelitis (EAE) animal experiment, where mice were given C-MANF subcutaneously, were included in this study, and qPCR was performed on them. This study showed significant changes in UPR gene expression in differentiated Oli-neu cells and EAE mice, indicating that C-MANF could alleviate UPR signaling and thus protect cells. Neuroinflammation was assessed with cerebellar brain slices and EAE mice; both models showed significant changes in gene expression related to neuroinflammation, indicating reduced neuroinflammation in response to C-MANF treatment. C-MANF did not seem to have an effect on myelination in this study. Large variation in between different groups in cerebellar brain slices may be due to challenges in RNA isolation an may have affected the results. However, these findings suggest the potential of C-MANF as a therapeutic for diseases associated with upregulated UPR signaling and a high degree neuroinflammation, such as MS. Further research and additional methodologies are needed to confirm these findings.
Now showing items 1-3 of 3