What Does This Mean
This study suggests that nearly two thirds of non-diabetic people with PD may be insulin resistant, despite having normal blood sugar, and in some cases, despite being lean. Thus, the big take-away from this study is that people with PD may want to have a more sensitive insulin test conducted, such as the HOMA-IR, to see if they have undiagnosed insulin resistance. There are several other IR tests as well. Which of these tests, or combinations thereof, might be best to evaluate a persons IR is something to be discussed with ones health care provider.
Mitochondrial Quality Control Mechanisms In Healthy And Pd Neurons
The deubiquitination system tightly counteracts the PINK1-Parkin pathway and contributes to mitochondrial homeostasis. Thus, the deubiquitinase USP30 is localized at the mitochondrial compartment where it deubiquitinates Parkin substrates. By removing the ubiquitin moieties from proteins, USP30 exerts negative effects on Parkin-mediated mitophagy. Interestingly, genetic downregulation of USP30 in flies carrying PD-linked mutations of Parkin or PINK1 reversed the mitophagy defects and restored mitochondrial integrity. Moreover, USP30 knockdown waived the dopaminergic defects in the treated flies, including motor defects and organismal survival . This exemplifies a sophisticated mechanism operating in mitochondria that finely regulates the ubiquitin system to control the homeostatic processes underlying metabolism, respiration, and oxidative stress.
Differential Expression Of Genes In Pd Skin
Hierarchical clustering demonstrating the expression profiles of the top 50 differentially regulated genes in Parkinsons disease versus normal skin . Gene expression levels are presented as colour variations from dark blue to pale green for each individual sample and for each gene
Table 1 Top 10 functional network functions affected in Parkinsons disease versus normal skin as analysed by Ingenuity Pathway Analysis. The first number in the score column reflects the number of genes in a reference gene set and the second reflects the number of focus genes altered in our study
Er Stress And Unfolded Protein Response
In eukaryotes, the UPR is highly conserved and comprises three parallel branches, each of them initiated by a specific ER stress sensor. Such sensor is represented by an ER resident protein, which is sensitive to ER perturbations and signals this information to the cytosol and the nucleus. There are three ER stress sensors: the inositol-requiring enzyme 1 ; the double-stranded RNA-activated protein kinase -like ER kinase ; the activating transcription factor-6 .
Figure 1. The unfolded protein response cascade. Stressful conditions due to starvation, infections, oxidative damage, and changes in ER Ca2+ concentration can lead to accumulation of misfolded proteins in the ER. Induction of the unfolded proteins response through the activation of its three independent arms counteracts the build-up of misfolded proteins and improves the ER folding capacity.
Besides its protective function, the UPR has been recently implicated in memory and synaptic plasticity as it has been shown that the PERK-eIF2 branch or XBP1 can regulate gene expression of proteins implicated in long-term potentiation, memory formation and synapsis remodeling . Thus it appears that the UPR is not only a rescue mechanism in case of stressful conditions in the ER but also a way to modulate normal cellular function and homeostasis.
Production Of Atp13a2 Knockdown/over
siRNA targeting PARK9 was purchased from Dhamacon . For the knockdown of ATP13A2, shRNA against ATP13A2 was cloned into the lentiviral vector pLL5.0 to produce the plasmid pAC1085. For the over-expression of ATP13A2, full-length ATP13A2 was sub-cloned into pLenti6/V5-Dest with gateway technology . The empty pLL5.0 vector backbone was used create vector control cells. For lentivirus production, constructs and lentiviral packaging plasmids were transfected into HEK293 cells using Lipofectamine 2000 .
For the knockdown of PARK9, SHSY5Y cells were transfected with PARK9 siRNA by Amaxa electroporation nucleofection . siRNA-mediated PARK9 knockdown was assessed by immunoblotting. For the creation of stable cell lines, lentivirus was collected and concentrated using Amicon column centrifugation. SHSY5Y and SHSY5Y-aSyn cells were infected with lentivirus for 24 h. Stably infected cells were selected for using 2 µg/ml blasticidin. Fluorescence-activated cell sorting confirmation and western blots were conducted to assess PARK9 knockdown/over-expression.
Understanding The Neurologic Control Of The Cardiac System
Before we explore this issue, lets first learn a bit about the autonomic nervous system and about the cardiac systems place within it. The ANS is part of the peripheral nervous system, a network of nerves throughout the body. The ANS exerts control over functions that are not under conscious direction such as respiration, heart function, blood pressure, digestion, urination, sexual function, pupillary response, and much more. The ANS is further subdivided into the parasympathetic nervous system and the sympathetic nervous system. Both the parasympathetic and sympathetic nervous systems regulate most major organs. Often, they have opposite effects, with the sympathetic nervous system activating a system and the parasympathetic system calming it down.
One of the systems controlled by the ANS is cardiac regulation. Blood pressure sensors, known as baroreceptors, reside in the heart as well as in the carotid artery, the major artery in the neck. If the baroreceptors sense a change in the blood pressure, a signal is sent to particular areas in the brain. From there, the autonomic nervous system sends signals to the heart to control heart rate and cardiac output. Signals are also sent to the blood vessels to change the size of their diameter, thereby regulating blood pressure.
What Is Alzheimer’s Disease
Alzheimer’s disease , the most common form of dementia among older adults, is an irreversible degeneration of the brain that causes disruptions in memory, cognition, personality, and other functions that eventually lead to death from complete brain failure. Genetic and environmental factors including diet, activity, smoking, traumatic brain injury, diabetes, and other medical diseases contribute to the risk of developing this form of the disease. The hallmarks of Alzheimer’s disease are the accumulation of beta-amyloid plaques between nerve cells in the brain and neurofibrillary tangles, which are twisted fibers found inside the brain’s cells). These tangles consist primarily of a protein called tau.
Atp13a2/ Patient Hons Accumulate Zinc
Further evidence of zinc dyshomeostasis in ATP13A2/ patient hONs was sought through the use of synchrotron X-ray fluorescence microscopy , which examines total intracellular element content . ATP13A2/ patient hONs had very similar Zn2+ and Mn2+ levels to those of control hONs . However, when challenged with 100 µm Zn2+, ATP13A2/ patient hONs displayed a 60% increase in total intracellular Zn2+ content compared with control hONs , suggesting that ATP13A2 may contribute to Zn2+ efflux from the cell. Consistent with the lack of sensitivity to Mn2+ , treatment of ATP13A2/ patient hONs with 1 mm Mn2+ did not result in increased manganese accumulation relative to controls .
Lrrk2 Ko Macrophages Are Defective In Oxidative Phosphorylation And Glycolysis
Metabolic reprogramming is becoming increasingly appreciated as a critical contributor to macrophage polarization and transcriptional output . We hypothesized that mitochondrial defects may render Lrrk2 KO macrophages incapable of meeting metabolic demands. To test this idea, we manipulated levels of sodium pyruvate, an intermediate metabolite of glycolysis and the TCA cycle, in the media of Lrrk2-deficient cells plus their respective controls. The presence of pyruvate places additional demands on the mitochondria by pushing cells towards oxidative metabolism rather than glycolysis. Remarkably, addition of as little as 1 mM sodium pyruvate to the growth media increased the already high basal levels of type I IFN in macrophages and MEFs lacking LRRK2 , suggesting that increasing metabolic demands on mitochondria promotes further leakage of mtDNA into the cytosol in these cells.
Lrrk2 KO macrophages are defective in oxidative phosphorylation and glycolysis.
What Are The Symptoms Of Parkinson’s Disease
The symptoms of Parkinson’s disease include tremors or trembling ; difficulty maintaining balance and coordination; trouble standing or walking; stiffness; and general slowness.
Over time, a person with Parkinson’s may have trouble smiling, talking, or swallowing. Their faces may appear flat and without expression, but people with Parkinson’s continue to have feelings even though their faces don’t always show it. Sometimes people with the disease can have trouble with thinking and remembering too.
Because of problems with balance, some people with Parkinson’s fall down a lot, which can result in broken bones. Some people with Parkinson’s may also feel sad or and lose interest in the things they used to do.
The symptoms of Parkinson’s disease appear gradually and get worse over time. But because Parkinson’s disease usually develops slowly, most people who have it can live a long and relatively healthy life.
Animals And Stereotaxic Surgery
Adult male and female Lewis rats were obtained from Envigo . Rats were singly housed in temperature-controlled conditions under a 12:12 light-dark cycle with ad libitum access to food and water. All experiments were approved by the University of Pittsburgh Institutional Animal Care and Use Committee.
Stereotaxic surgery was performed to deliver virus to the substantia nigra . Animals were deeply anesthetized under isoflurane for the entirety of the surgery. Two microliter of virus was infused over a 10-min period . Animals were closely monitored on a heated surface during recovery of surgery. Animals received analgesia in the form of ketoprofen with the first dose presurgery and 2×/day for 3 days following surgery. Animals were sacrificed by transcardial perfusion with phosphate buffered saline followed by 4% paraformaldehyde. The brains were harvested and sectioning was performed on a freezing-stage microtome at 35µm. Sections were maintained in cryoprotectant at 20°C until analysis.
Lrrk2 Ko Mice Can Control Mtb Replication But Have Exacerbated Infection
Previous reports have linked SNPs in LRRK2 with susceptibility to mycobacterial infection and excessive inflammation in humans . Our studies demonstrate that LRRK2 plays a key role in homeostasis of macrophages, the first line of defense and replicative niche of Mtb. Therefore, we sought to understand how LRRK2 deficiency influences Mtb pathogenesis in macrophages ex vivo and during an in vivo infection. Lrrk2 HET and KO BMDMs were infected with Mtb , and colony forming units were measured over the course of five days. We observed a significant increase in CFUs recovered at 5 days following infection , suggesting that while Lrrk2 KO macrophages can control Mtb replication early after infection, they are more permissive once the bacteria have established a niche. These results are consistent with a recent report demonstrating that defective IFNAR signaling in BMDMs leads to increased Mtb replication .
Bone marrow derived macrophages were differentiated from bone marrow cells isolated by washing mouse femurs with 10 ml DMEM. Cells were then centrifuged for 5 min at 1000 rpm and resuspended in BMDM media . BM cells were counted and plated at 5 × 106 in 15 cm non-TC treated dishes in 30 ml complete media and fed with an additional 15 ml of media on Day 3. On Day 7, cells were harvested with 1x PBS-EDTA.
Signalling Pathways And Tumour Genes Influenced In Pd Skin
We observed a basal deregulation of cellular signalling pathways and suppression of several oncogenes and tumorsupressors in PD skin . Several of the central growth and survival pathways were also altered in PD skin, as highlighted by suppression of Ras signalling by decreased expression of central Ras-effectors. Also small-GTPase family genes and G-protein signalling were affected. In addition, central effectors of Wnt signalling and several regulators of NOTCH signalling showed altered levels in PD skin. PD also impacts fibroblast growth factor, insulin-like growth factor, transforming growth factor-, nuclear factor-, and other central signalling proteins, such as carcinoembryonic antigen proteins, epidermal growth factor and vascular endothelial growth factor family proteins.
Living With Parkinson’s Disease
As Parkinson’s develops, a person who has it may slow down and won’t be able to move or talk quickly. Sometimes, speech therapy and occupational therapy are needed. This may sound silly, but someone who has Parkinson’s disease may need to learn how to fall down safely.
If getting dressed is hard for a person with Parkinson’s, clothing with Velcro and elastic can be easier to use than buttons and zippers. The person also might need to have railings installed around the house to prevent falls.
If you know someone who has Parkinson’s disease, you can help by being a good friend.
Lrrk2 Ko Macrophages Exhibit Blunted Type I Ifn Induction In Response To Cytosolic Nucleic Acid Agonists
Lrrk2 KO macrophages exhibit blunted type I IFN expression in response to cytosolic nucleic acid agonists.
We next tested whether loss of LRRK2 impacted the ability of cells to respond to activators of the type I IFN response outside of the cytosolic DNA sensing cascade. To this end, we treated Lrrk2 KO and HET BMDMs with transfected poly , LPS , and CpG and CL097 . Interestingly, while we observed a defect in Ifnb induction in Lrrk2 KO BMDMs stimulated with poly, we saw no difference in the ability of Lrrk2 KO BMDMs to express type I IFNs following treatment with LPS, CL097, or CpG , suggesting that TLR responses are intact in the absence of LRRK2 but cytosolic DNA and RNA sensing pathways are perturbed. We observed similar phenotypes for PEMs and MEFs treated with LPS and poly . Lrrk2 KO BMDMs were, however, defective in ISG expression following recombinant bioactive IFN- treatment .
Increased Basal Type I Ifn In Lrrk2 Ko Macrophages Is Dependent On Cytosolic Dna Sensing Through Cgas
Because both IFN- blockade and loss of Ifnar normalized basal ISG expression in Lrrk2 KO macrophages, we hypothesized that Lrrk2 contributes to basal type I IFN expression upstream of cytosolic RNA or DNA sensing, two nucleic acid sensing pathways that are interconnected between positive and negative feedback loops . To directly test the involvement of cGAS in generating elevated resting levels of type I IFN in Lrrk2 KO macrophages, we crossed Lrrk2 KO and cGas KO mice and compared type I IFN transcript levels in double KO BMDMs with those of littermate controls. Although basal Isg15 expression differences between Lrrk2 KO and HET BMDMs were more modest in this experiment, loss of cGAS significantly reduced basal ISG expression in Lrrk2 KO BMDMs . With lowered resting type I IFN levels, cGas/Lrrk2 double KOs were able to respond normally to IFN/ISG-generating innate immune stimuli like DMXAA, which bypasses cGAS and stimulates STING directly , and poly transfection . Consistent with the ability of cGas ablation to rescue Lrrk2 KO baseline and induction defects, western blot analysis showed that levels of STAT1 phosphorylation were restored in cGas/Lrrk2 double KOs . Together, these results support a model where high basal levels of type I IFN and ISGs in Lrrk2 KO macrophages are due to chronic engagement of the cGAS-dependent DNA sensing pathway.
Cytosolic mtDNA drives basal type I IFN expression in Lrrk2 KO macrophages.
Histology And Proximity Ligation Assay
Immunohistochemistry was performed on free-floating sections before mounting on glass slides for imaging. Primary antibodies used for immunohistochemical analysis are listed in Table .
Proximity ligation assay was analyzed utilizing PLA probes and visualized with orange and far-red kits.
For quantification of immunofluorescence, confocal images of both immunohistochemistry and proximity ligation assay were taken on an Olympus IXB1 confocal. Intensity measurements were obtained in Fluoview software by circling ROIs around TH-positive neurons in the nontransduced hemisphere and TH- and GFP-positive neurons in the transduced hemisphere while blinded to the protein of interest. Intensity values of the protein of interest were compared between transduced and nontransduced TH-positive neurons. Number of objects analysis was performed in Nikon Elements software by circling ROIs around TH-positive neurons in the nontransduced hemisphere and TH- and GFP-positive neurons in the transduced hemisphere while blinded to the protein of interest. Within ROIs, a threshold was applied to the protein of interest in order to achieve unbiased counts of objects. Images to visualize DAB immunoreactivity were taken on an Olympus BX61VS microscope and analyzed for pixel saturation in CellSens software.
Impaired Skin Homeostasis Nuclear Processes And Tumorigenic Pathwaysthe Mechanistic Link For Predisposition To Skin Cancer In Pd Patients
The second focal point of our study enhances the understanding of the mechanistic association between skin cancer and PD, as demonstrated by a basal defect in skin homeostasis, deregulated nuclear processes, as well as dysbalanced cellular signalling, tumorigenic pathways and inflammatory processesall these alterations possibly contributing to the specific vulnerability of PD skin to mutagenic hazards , which can provide the basis for the mechanistic link to the increased risk of skin cancer in this patient population.
And lastly, our study indicates dysregulation of immune pathways in PD skin. The interplay of pro- and anti-inflammatory signalling, and the discrimination of causal and effector changes in the context of both PD and cancer is complex, however chronic inflammation has been shown to be one of the main factors fostering all stages of neoplasia, but also one of the pathogenic processes in PD progression, thus the basal inflammatory dysfunction associated with PD can thereby contribute to increasing the risk of cancer development in these patients.
Rab3a And Rab8a Protect Against
The elongated, highly ramified architecture of neuronal cells and the special demands of regulated dopamine release prompted us to ask whether other Rabs might be more important in combating -syn toxicity in dopaminergic neurons than they are in yeast. RAB8A, the human homolog of yeast Sec4, is the closest paralog to human RAB1A . RAB8A, like yeast Sec4, is localized on post-Golgi vesicles where it facilitates targeting to areas of polarized membrane growth. Given the neuronal specificity of RAB3A expression and the presynaptic terminal localization of -syn, RAB3A was also an attractive candidate to test.
To evaluate the ability of these Rabs to modify -syn-induced toxicity in DA neurons we used two very different models. The nematode Caenorhabditis elegans offers the advantage of testing DA neurons wired in their natural context to other neurons in a whole animal. Rat primary midbrain cultures lack this advantage but are evolutionarily much closer to human neurons and offer the additional advantage of detecting differential sensitivity of DA neurons and selective rescue of those neurons that are most affected by -syn pathology .
Autophagy And Redox Homeostasis In Parkinsons: A Crucial Balancing Act
Natalia Jimenez-MorenoJon D. Lane
1Edinburgh Cancer Research UK Centre, MRC Institute of Genetics and Molecular Medicine, University of Edinburgh, Edinburgh EH4 2XR, UK
2Cell Biology Laboratories, School of Biochemistry, University of Bristol, Bristol BS8 1TD, UK
1. Introduction: Autophagy Forms, Roles, and Regulation
There are three types of autophagy, each with a distinct mechanism for delivery of substrates to the lysosome. These are microautophagy, chaperone-mediated autophagy , and macroautophagy. In microautophagy, the cargo is directly engulfed into lysosomes through lysosomal invaginations or protrusions . CMA is a highly selective type of autophagy, where cargoes containing KFERQ-like motifs and/or proteins that have been posttranslationally modified to generate KFERQ-like motifsbecoming de novo CMA substrates are selectively targeted via heat shock cognate 71kDa protein and cochaperones, and internalized to the lysosome lumen through the lysosome-associated membrane protein 2 receptor for their degradation . Thus, CMA plays an important role in the degradation of altered and aggregated proteins, and impairments in this process have been linked to numerous diseases, including neurodegenerative diseases. For example, accumulation of CMA substrates such as -synuclein and tau are hallmarks, respectively, of Parkinsons and tauopathies .
1.4. General Features and Properties of Autophagy in Neurons
3. Redox Homeostasis
Statistics And Data Reporting
Data are presented as individual data points corresponding to an individual animal and n equal to the number of animals unless otherwise specified. Graphs demonstrate mean and standard error of the mean. Paired t-tests were used to evaluate within animal protein immunoreactivity, neurochemical measurements, neuronal counts, and behavior. Unpaired t-tests were used to compare viral treatment groups. One-way ANOVA with Tukeys post-hoc test was performed to evaluate mRNA.
Nuclear Processes Influenced In Pd Skin
A large set of differentially regulated genes in PD skin play a role in nuclear processes and epigenetic regulation 14 genes participating in different aspects of the cell cycle were supressed in PD, including cyclins, cyclin dependent kinases and other cell division cycle associated genes. Also 14 regulators of basal transcription of RNA were downregulated, with different RNA polymerase II polypeptide associated factors, activators of basal transcription and elongation factors being supressed. Another important category supressed in PD is related to the reparation and degradation of nuclear and mitochondrial DNA, with 11 genes including endo- and exo-nucleases, dnases and growth-arrest associated molecules down-regulated. A large group of transcription factors were deregulated in PD skin, several of these controlling various aspects of skin homeostasis, proliferation and differentiation, but others regulating energy metabolism, cellular signalling and immune responses. In addition, a large set of altered genes can be associated to epigenetic regulation. This is highlighted by the down-regulation of achaete-scute homolog 2, which is known to regulate various aspects of epigenetics. Also genes related to chromatin remodelling and DNA binding, transcriptional/post-transcriptional modification and RNA splicing were differentially regulated, as well as a large group of miRNAs, snRNPRs and snoRNAs.
Lrrk2 And Basal Mitophagy
Coding variants in the leucine-rich repeat kinase 2 gene cause an autosomal dominant form of PD and are responsible for the majority of familial cases . LRRK2 is a large multidomain protein consisting of an armadillo domain, an ankyrin domain , a leucine-rich repeat domain , a Ras of complex GTPase domain , a C-terminal of ROC domain , a kinase domain, and a WD40 domain . Coding variants in LRRK2 leading to PD segregate to its catalytic core, such as in the GTPase domain and in its kinase domain . The G2019S mutation being the most frequent genetic cause of PD, representing 45% of familial cases and about 1% of sporadic cases . All the known pathogenic mutations of LRRK2 lead to an increased kinase activity . Given the track record of small molecule kinase inhibitors as effective drugs for other diseases, LRRK2 kinase inhibitors offer considerable and obvious potential. Indeed, small molecule LRRK2 kinase inhibitors, as well as an antisense LRRK2 oligo, are currently in clinical trials for PD indications . In addition, other types of inhibitors have been developed, either targeting the G-protein cycle by inhibiting its GTPase activity or by inhibiting LRRK2’s proteinprotein interactions . The proliferation in LRRK2 inhibitor development highlights that pharmacologically targeting LRRK2 could be a highly potent solution to diminish pathogenic LRRK2 effects.
Mitophagy Defects In Pd
Currently, no core autophagy-essential genes have been directly implicated in PD. However, several PD-linked genes have been identified that modulate mitophagy, which suggests this pathway may be more relevant to PD pathology compared with autophagy in general. As we will describe in the following paragraphs, these PD-related genes could potentially affect mitophagy at different steps and act independently of one another.
How Parkinsons Disease Affects The Autonomic Nervous System And The Heart
In PD, there are two major reasons why the automatic control of the cardiac system is impaired. First, areas of the brain that control this system often contain Lewy bodies and have undergone neurodegeneration. In addition, the autonomic nervous system itself is directly affected by Lewy body-like accumulations and neurodegeneration. This means, when the baroreceptors in the heart and carotid artery sense a drop in blood pressure and try to generate a signal to the heart and blood vessels to increase the blood pressure, the message may not get through. This results in neurogenic orthostatic hypotension , or drops in blood pressure upon standing due to autonomic nervous system dysfunction. There are no medications that can cure nOH by restoring the autonomic nervous system in PD. nOH however, can be treated. Read more about nOH and its treatments .
Structural problems of the heart such as coronary artery disease or cardiomyopathy are not thought to be part of the pathology of PD, although of course, could co-exist with PD.
How Is A Diagnosis Made
Because other conditions and medications mimic the symptoms of PD, getting an accurate diagnosis from a physician is important. No single test can confirm a diagnosis of PD, because the symptoms vary from person to person. A thorough history and physical exam should be enough for a diagnosis to be made. Other conditions that have Parkinsons-like symptoms include Parkinsons plus, essential tremor, progressive supranuclear palsy, multi-system atrophy, dystonia, and normal pressure hydrocephalus.