Blog Category: Redox Biology

Redox proteomics reveals overlapping mechanisms of neurodegeneration in Down syndrome and Alzheimer disease

By: Eugenio Barone and Marzia Perluigi

Read the full articles in Free Radical Biology and Medicine here: Barone E et al. Free Radic Biol Med, 111:262-269, 2017Barone E et al. Free Radic Biol Med, 114:84-93, 2018

Accumulation of oxidative damage is a common feature of neurodegeneration that, together with mitochondrial dysfunction, point to the fact that reactive oxygen species are major contributors to loss of neuronal homeostasis and cell death. Among several targets of oxidative stress, free radical-mediated damage to proteins is particularly important in aging and age-related neurodegenerative diseases. In the majority of cases, oxidative stress mediated post-translational modifications cause non-reversible modifications of protein structure that consistently lead to impaired function.

Figure: The workflow of a redox proteomics analysis is showed in six principal steps.

Redox proteomics methods are powerful tools to unravel the complexity of neurodegeneration, by identifying brain proteins with oxidative post-translational modifications that are detrimental for protein function and pathogenesis (Butterfield DA et al. Biochem J, 463:177-89, 2014). Our published studies show evidence of impaired pathways linked to oxidative stress possibly involved in the neurodegenerative process leading to the development of Alzheimer-like dementia (Di Domenico F et al. Antioxid Redox Signal, 26:364-387, 2017). Recently, we also focused on dysregulated pathways underlying neurodegeneration in aging adults with Down syndrome (DS) (Barone E et al. Free Radic Biol Med, 111:262-269, 2017). Interestingly, DS individuals by the age of 40ys are at increased risk to develop Alzheimer disease (AD) like dementia.

Results obtained by the analysis of human specimens and studies from mouse and cellular models of the disease evince a molecular link between protein oxidation/aggregation, the integrity of protein quality control system [proteasome, unfolded protein response (UPR) and autophagy], dysfunction of energy metabolism and neurodegeneration (Di Domenico F et al. Antioxid Redox Signal, 26:364-387, 2017). Many common pathological hallmarks exist between DS and AD including deposition of b-amyloid plaques, Tau-based neurofibrillary tangles, increased oxidative damage, and impaired mitochondrial function, among others. Intriguingly, we propose that all these processes seem to be joined by a “leitmotif” – oxidative stress - since they are all the cause and/or the consequence of increased free radical burden. If low amounts of reactive oxygen species (ROS) can activate the protective cellular apparatus such as the antioxidant and heat shock responses, cell cycle regulation, DNA repair, UPR and autophagy, then chronic exposure to ROS causes irreversible damage to all intracellular macromolecules (Barone E et al. Free Radic Biol Med, 111:262-269, 2017). Among these, protein oxidation impairs multiple cellular functions by a largely irreversible process that results in altered, mostly reduced, protein activity. It is likely that stressed neurons have the challenge of increasing loads of oxidatively damaged proteins, which overwhelm the ability of the proteostasis network. This, in turn, promotes further accumulation of damaged proteins, increasingly prone to aggregation, ultimately resulting in neuronal death. Alteration of protein homeostasis coupled with increasing demand for protein degradation, and reduced ATP production may produce a vicious cycle that may accelerate the neurodegenerative process (Barone E et al. Free Radic Biol Med, 111:262-269, 2017, Barone E et al. Free Radic Biol Med, 114:84-93, 2018).Therapeutic strategies aimed at preventing/reducing multiple components of processes leading to accumulation of oxidative damage will be critical in future studies.

Laboratory of Redox Biochemistry in Neuroscience, Department of Biochemistry, Sapienza University of Rome, Email: Eugenio.barone@uniroma1.it

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Category: Redox Biology


Thinking about a PhD in Biomedical Science? Some Things You Should Know Before You Get Started.

By Dr. Marcus S. Cooke, Ph.D., FRCPath of Florida International University

Luke Skywalker: “I won't fail you! I'm not afraid.”

Yoda: “Oh! You will be. You will be.”

Similar to Luke’s training by Yoda, the PhD is like an apprenticeship, students are mentored and supported throughout, but this is gradually withdrawn, as they become more accomplished, culminating with being an expert in the field, and an independent researcher. On this basis, selection of the right supervisor for you is as vital as selecting the right project area. The successful pursuit of a doctoral degree is a two-way street that requires good alignment between your objectives and those of your supervisor, as well as your progressive capacity to deal with failure and rejection (it’s a fact of life for researchers, I’m afraid). 

A PhD requires fulltime dedication to the project. Before considering undertaking a PhD a prospective student must be willing to devote themselves to the project, understanding that experiments sometimes require working outside business hours. You may possess a great deal of practical experience, but this experience by itself is not sufficient to earn the degree. The project fails or succeeds based on the effort, and ideas from the student, guided and supported by the supervisor. In order to have the required level of commitment, the chosen project on which you work must really excite you – this will provide the motivation to keep going, particularly when experiments prove challenging (i.e. techniques stop working for no apparent no reason!).  There will be lows, but there will also be highs, for example when your abstract gets accepted for an international meeting, or your first manuscript gets accepted (it feels good whether it’s your first or 81st). This strengthening of your character will surely enrich your career and life.

The kinds of wet lab projects offered in the broad area of Biomedical Sciences do not readily lend themselves to a part-time degree, but that is not to say it is not possible. A PhD in the Biomedical Sciences means 3-4+ years of intensive research in the lab – sounds like plenty of time, right? But the time flies by in which you must have generated a significant amount of novel data, addressing an important research question. Also, don’t think that a PhD is simply an extension of a Master’s degree, only longer. It is completely different, with objectives, timelines and expectations of progress toward scientific independence that are not seen at the Master’s level. 

Good organization of personal time will be needed, such that you can improve skills that are just as important as your experimental work. This includes: keeping up with the literature about the subject, improving scientific writing, cultivating attention to detail, attending seminars and workshops in and outside your institution, and very importantly, mastering your own oral communication skills. Most institutions demand a minimum of two original data manuscripts, and a review article to submit towards your PhD. These will need to survive the scrutiny of peer review, which will train your capacity to receive criticism and to work proactively on improving the quality of your research.

A huge amount of self-motivation is required to withstand the intrinsic challenges of developing a scientific project toward thinking independently. The structured, taught element of the PhD is minor, in comparison to the time spent in the lab, and just provides a general grounding to get improve your knowledge base. The real learning and training comes in the lab where research skills are learnt firsthand, and tested on a regular basis through the experiments required to test your hypothesis.

Still wanting to do a PhD in the Biomedical Sciences? Get in touch with prospective mentors, discuss with them theirs and your research and career expectations, and go for it! If you are intrinsically thrilled to investigate Nature, you will not be hampered by the high level of commitment required in this career path. With the right PhD training (irrespective of the discipline), you will be well prepared for a wide variety of career options, though giving you skills which allow you to think, speak, write, and problem-solve in a particular way that makes you an asset to any organization.

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Categories: Research, Free Radical Biology and Medicine, SfRBM Trainee Council, Redox Biology


Mechanisms in Caloric Restriction

By: Alicia J. Kowaltowski, M.D., Ph.D., Universidade de São Paulo

Obesity enhances the incidence and prevalence of many diseases, but is still increasing worldwide. Our group is focused on understanding mechanisms involved in the protective effects of maintaining a normal body weight, in order to uncover new targets for obesity-related diseases. We do this by studying rodents that eat ad libitum (as much as they want) and develop obesity versus animals on a calorically-restricted diet. Calorically-restricted rodents have been shown to have less oxidative damage and age-related diseases, as well as extended lifespans. 

Many of the effects of caloric restriction, predictably, involve changes in energy metabolism and mitochondria, the central hub of energy metabolism. Recently, while investigating mechanisms in which calorie restriction prevents excitotoxic neuronal death (Amigo et al., 2017), we found that mitochondria from calorically-restricted animals can take up Ca2+ ions faster and in larger quantities. The higher buffering capacity these mitochondria have promotes protection against neuronal death caused by excessive Ca2+ in the cytosol.

Because this is a completely new effect of caloric restriction, we decided to investigate if mitochondrial Ca2+ uptake was also modified in the liver (Menezes-Filho et al., 2017). We found that liver mitochondria from calorie restricted mice could also take up more Ca2+, and that this was related to protection against ischemic damage. Overall, the finding that caloric intake affects mitochondrial Ca2+ uptake is interesting because Ca2+ is a well-known modulator of energy metabolism and mitochondrial oxidant production. Based on these initial findings, we hope to uncover more connections between metabolic regulation, redox state, caloric intake and Ca2+ in futures studies.

While studying calorically-restricted mice, we noticed that they had different fur from obese ad libitum fed animals. We then decided to investigate changes in the skin and fur of calorie restricted animals, and found many modifications, both in the structure and metabolism of the skin. One interesting finding was that the fur in calorically-restricted mice has a higher insulation capacity, and is necessary for them to thrive: shaving calorie restricted animals makes them lose muscle mass. We are not sure how these results relate to humans, who aren´t very furry, but believe this is an evolutionary adaptation to deal with heat loss. Although the skin is our largest organ, it is not often studied, and there are almost no related findings. We hope that with this study and the development of methodology to monitor metabolism in the skin, more interest will arise.

Amigo I, Menezes-Filho SL, Luévano-Martínez LA, Chausse B, Kowaltowski AJ. Caloric restriction increases brain mitochondrial calcium retention capacity and protects against excitotoxicity. Aging Cell. 2017 16:73-81.

Forni MF, Peloggia J, Braga TT, Chinchilla JEO, Shinohara J, Navas CA, Camara NOS, Kowaltowski AJ. Caloric restriction promotes structural and metabolic changes in the skin. Cell Rep. 2017 20:2678-2692.

Menezes-Filho SL, Amigo I, Prado FM, Ferreira NC, Koike MK, Pinto IFD, Miyamoto S, Montero EFS, Medeiros MHG, Kowaltowski AJ. Caloric restriction protects livers from ischemia/reperfusion damage by preventing Ca2+-induced mitochondrial permeability transition. Free Radic Biol Med. 2017 110:219-227.

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Category: Redox Biology


Blending After Hiring: 5 Tips for Matching Projects with Individual Talents

By Luciana Hannibal, Ph.D. 

Team success goes hand in hand with the professional development of its members. New employees are selected according to background education (know-how theory), previous accomplishments (know-how experience) and potential (know-how to grow). These important metrics help to recruit candidates who are best suited for the position, and capture them at a stage of high self-motivation and drive.  Only a few however, will stand out for their achievements, even when the same selection process has been applied to all. Why? A pool of employees will choose to remain in a professional comfort-zone, meeting personal and team demands satisfactorily. Others will strive to further their professional development and to make a difference in the team. Some will go beyond and prepare themselves to lead their own teams in the future. Even after careful recruitment, a population of talented individuals may however derail from successful career development. A mismatch of talents and projects can hamper advancement regardless of individual mindset. Researchers in leading positions are expected to excel at understanding the scientific problem, providing new ideas and solutions, teaching and supervising, cooperating effectively with internal and external colleagues and recruiting copious amounts of extramural funding. Optimizing self-reliance of the team is thus crucial for individual and collective return on investment, and this involves aligning talents with projects. Team leaders must identify individual strengths to effectively assign project roles, and employees must be visible for their most valuable talents.  Self-motivated employees, as they typically land in the new job, constitute an invaluable asset.

Reprinted by permission from Macmillan Publishers Ltd: Nature, "Lifelong learning: Science professors need leadership training," copyright 15 July 2015
Reprinted by permission from Macmillan Publishers Ltd: Nature, "Lifelong learning: Science professors need leadership training," copyright 15 July 2015

Here is a brief guide on how to blend projects with selected talent types:

  1. The hands-on: performs best by being directly involved in practical work. Creativity may not necessarily be a natural strength; therefore, projects with a defined start and end work optimally. These employees are highly reliable in meeting milestones and deadlines. 
  2. The creative: generates ideas and is highly valuable in projects that require theoretical development, but their capacity to carry on ideas into practice may be slim. This type of employee may require additional human resources to execute tasks; encouraging them to recruit them themselves can be a win-win situation for both parties. 
  3. The communicator: skillful in communicating their work and knowledge, performs best in projects where training others is crucial. These employees are highly valuable in tasks involving transfer of knowledge, such as teaching, trainings and external presentations. 
  4. The troubleshooter: has the ability to spot a variety of problems and potential solutions. These employees function optimally in projects that involve troubleshooting and the participation of others. 
  5. The larger-than-life: if you come across an employee who has a balanced combination of the four features above, you are in the presence of a natural leader. Your job is to maximize their professional development inside and outside your team! Be part of their success!

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Categories: Education, Redox Biology, Free Radical Biology and Medicine


Redox Biomarker Could Predict Progression of Epilepsy

Approximately 2.9 million people in the United States suffer from epilepsy, according to the CDC. For patients living with this diagnosis and their doctors it is often difficult to predict the onset or progression of chronic seizures. Thanks to a newly published study from the University of Colorado Skaggs School of Pharmacy and Pharmaceutical Sciences at the Anschutz Medical Campus, that may be changing.

The study, led by Drs. Manisha Patel and Li-Ping Liang of the University of Colorado, was recently published in Redox Biology, a journal of the Society for Redox Biology and Medicine (SFRBM).

The study was designed to determine if the ratio of reduced and oxidized forms of an amino acid, cysteine and cystine respectively, could serve as an accurate biomarker to predict the onset or progression of seizures associated with epilepsy. Using a rat model, it was determined that decreased cysteine/cystine ratio in plasma may serve as a redox biomarker in epilepsy.

Specifically, seizures were chemically-induced in rats, which were then monitored closely for behavioral changes. Plasma was then taken from the rats 48 hours and 12 weeks after treatment to mimic acute and chronic epileptic conditions. It was found that cysteine/cystine ratio was an accurate redox biomarker for epilepsy. Plasma cysteine/cystine was reduced over 60% in rats with acute epileptic responses and over 37% in rats with chronic epilepsy. Interestingly, cysteine/cystine ratio was unaltered in rats also treated with an antioxidant known to prevent epileptic brain injury.

“Currently the field of epilepsy lacks peripheral blood-based biomarkers that could predict the onset or progression of chronic seizures following an epileptogenic injury,” said Dr. Manisha Patel a Professor at the University of Colorado School of Pharmacy and SFRBM Member. “We are confident that this study is a significant step toward changing this, and will one day help those living with temporal lobe epilepsy.”

About Dr. Patel’s lab

The overarching theme of research in Dr. Patel’s laboratory is to understand the role of redox and metabolic mechanisms in epilepsy. Her laboratory has provided compelling evidence for the role of reactive species and mitochondrial dysfunction in animal models of epilepsy. Furthermore, her research has identified reactive species as novel targets for co-morbidities of epilepsy.

About the University of Colorado Skaggs School of Pharmacy and Pharmaceutical Sciences

Since its inception in 1911, the University of Colorado Skaggs School of Pharmacy and Pharmaceutical Sciences has experienced numerous milestones and is ranked in the top 20 percent of all schools of pharmacy in the country, and fourth in the nation for total NIH funding. Committed to pharmaceutical education, research and patient care, the School educates students in the properties of medicinal agents, the biology of disease, the actions of drugs, and best practices for clinical and therapeutic uses of drugs. Located at the Anschutz Medical Campus, the University of Colorado is the only completely new education, research and patient care facility in the nation today.

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Categories: Free Radical Biology and Medicine, Redox Biology, Research