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mitochondria

Molecular gatekeepers that regulate calcium ions key to muscle function

mitochondria

Controlled entry of calcium ions into the mitochondria, the cell’s energy powerhouses, makes the difference between whether muscles grow strong or easily tire and perish from injury, according to research published in Cell Reports.

Calcium ions are essential to how muscles work effectively, playing a starring role in how and when muscles contract, tap energy stores to keep working and self-repair damage. Not only are calcium ions vital for the repair of injured muscle fibers, their controlled entry into the mitochondria, the cell’s energy powerhouses, spells the difference between whether muscles will be healthy or if they will easily tire and perish following an injury, according to research published Oct. 29, 2019, in Cell Reports.

“Lack of the protein mitochondrial calcium uptake1 (MICU1) lowers the activation threshold for calcium uptake mediated by the mitochondrial calcium uniporter in both, muscle fibers from an experimental model and fibroblast of  a patient lacking MICU1,” says Jyoti K. Jaiswal, MSc, Ph.D., a principal investigator in the Center for Genetic Medicine Research at Children’s National Hospital and one of the paper’s corresponding authors. “Missing MICU1 also tips the calcium ion balance in the mitochondria when muscles contract or are injured, leading to more pronounced muscle weakness and myofiber death.”

Five years ago, patients with a very rare disease linked to mutations in the mitochondrial gene MICU1 were described to suffer from a neuromuscular disease with signs of muscle weakness and damage that could not be fully explained.

To determine what was going awry, the multi-institutional research team used a comprehensive approach that included fibroblasts donated by a patient lacking MICU1 and an experimental model whose MICU1 gene was deleted in the muscles.

Loss of MICU1 in skeletal muscle fibers leads to less contractile force, increased fatigue and diminished capacity to repair damage to their cell membrane, called the sarcolemma. Just like human patients, the experimental model suffers more pronounced muscle weakness, increased numbers of dead myofibers, with greater loss of muscle mass in certain muscles, like the quadriceps and triceps, the research team writes.

“What was happening to the patient’s muscles was a big riddle that our research addressed,” Jaiswal adds. “Lacking this protein is not supposed to make the muscle fiber die, like we see in patients with this rare disease. The missing protein is just supposed to cause atrophy and weakness.”

Patients with this rare disease show early muscle weakness, fluctuating levels of fatigue and lethargy, muscle aches after exercise, and elevated creatine kinase in their bloodstream, an indication of cell damage due to physical stress.

“One by one, we investigated these specific features in experimental models that look normal and have normal body weight, but also show lost muscle mass in the quadriceps and triceps,” explains Adam Horn, Ph.D., the lead researcher in Jaiswal’s lab who conducted this study. “Our experimental model lacking MICU1 only in skeletal muscles responded to muscle deficits so similar to humans that it suggests that some of the symptoms we see in patients can be attributed to MICU1 loss in skeletal muscles.”

Future research will aim to explore the details of how the impact of MICU1 deficit in muscles may be addressed therapeutically and possible implications of lacking MICU1 or its paralog in other organs.

In addition to Jaiswal and Horn, Children’s National Hospital Center for Genetic Medicine Research co-authors include Marshall W. Hogarth and Davi A. Mazala. Additional co-authors include Lead Author Valentina Debattisti, Raghavendra Singh, Erin L. Seifert, Kai Ting Huang, and Senior Author György Hajnóczky, all from Thomas Jefferson University; and Rita Horvath, from Newcastle University.

Financial support for research described in this post was provided by the National Institutes of Health under award numbers R01AR55686, U54HD090257 and RO1 GM102724; National Institute of Arthritis and Musculoskeletal and Skin Diseases under award number T32AR056993; and Foundation Leducq.

sketch of muscle cells

Losing muscle to fat: misdirected fate of a multipotent stem cell drives LGMD2B

Fibro/adipogenic precursors (FAPs) control the onset and severity of disease in limb-girdle muscular dystrophy type 2 (LGMD2B)

Fibro/adipogenic precursors (FAPs) control the onset and severity of disease in limb-girdle muscular dystrophy type 2 (LGMD2B). a) Healthy and/or pre-symptomatic LGMD2B muscle contains resident FAPs. b) After myofiber injury, inflammatory cells invade and trigger FAP proliferation. c) In symptomatic LGMD2B muscle, there is a gradual accumulation of extracellular AnxA2, which prolongs the pro-inflammatory environment, causing excessive FAP proliferation. d) Blocking aberrant signaling due to AnxA2 buildup blocks FAP accumulation and thus preventing adipogenic loss of dysferlinopathic muscle. Credit: “Fibroadipogenic progenitors are responsible for muscle loss in limb girdle muscular dystrophy 2B.” Published online June 3, 2019, in Nature Communications. Marshall W. Hogarth, Aurelia Defour, Christopher Lazarski, Eduard Gallardo, Jordi Diaz Manera, Terence A. Partridge, Kanneboyina Nagaraju and Jyoti K. Jaiswal. https://rdcu.be/bFu9U.

Research led by faculty at Children’s National published online June 3, 2019, in Nature Communications shows that the sudden appearance of symptoms in limb-girdle muscular dystrophy type 2 (LGMD2B) is a result of impaired communication between different cell types that facilitate repair in healthy muscle. Of particular interest are the fibro/adipogenic precursors (FAPs), cells that typically play a helpful role in regenerating muscle after injury by removing debris and enhancing the fusion of muscle cells into new myofibers.

LGMD2B is caused by mutations in the DYSF gene that impair the function of dysferlin, a protein essential for repairing injured muscle fibers. Symptoms, like difficulty climbing or running, do not appear in patients until young adulthood. This late onset has long puzzled researchers, as the cellular consequences of dysferlin’s absence are present from birth and continue through development, but do not impact patients until later in life.

The study found that in the absence of dysferlin, muscle gradually increases the expression of the protein Annexin A2 which, like dysferlin, facilitates repair of injured muscle fiber. However, increasing Annexin A2 accumulates outside the muscle fiber and drives an increase in FAPs within the muscle as well as encourages these FAPs to differentiate into adipocytes, forming fatty deposits. Shutting down Annexin A2 or blocking the adipocyte fate of FAPs using an off-the-shelf medicine arrests the fatty replacement of dysferlinopathic muscle.

“We propose a feed-forward loop in which repeated myofiber injury triggers chronic inflammation which, over time, creates an environment that promotes FAPs to accumulate and differentiate into fat. This, in turn, contributes to more myofiber damage,” says Jyoti K. Jaiswal, MSc, Ph.D., a principal investigator in the Center for Genetic Medicine Research at Children’s National and the study’s senior author.

“Adipogenic accumulation becomes the nucleating event that results in an abrupt decline in muscle function in patients. This new view of LGMD2B disease opens previously unrealized avenues to intervene,” adds Marshall Hogarth, Ph.D., the study’s lead author.

Joyti Jaiswal

“We propose a feed-forward loop in which repeated myofiber injury triggers chronic inflammation which, over time, creates an environment that promotes FAPs to accumulate and differentiate into fat. This, in turn, contributes to more myofiber damage,” says Jyoti K. Jaiswal, MSc, Ph.D.

A research team led by Jaiswal collaborated with Eduard Gallardo and Jordi Diaz Manera, of Hospital de la Santa Creu in Barcelona, Spain, to examine muscle biopsies from people with LGMD2B who had mild to severe symptoms. They found that adipogenic deposits originate in the extracellular matrix space between muscle fibers, with the degree of accumulation tied to disease severity. They found a similar progressive increase in lipid accumulation between myofibers predicted disease severity in dysferlin-deficient experimental models. What’s more, this process can be accelerated by muscle injury, triggering increased adipogenic replacement in areas that otherwise would be occupied by muscle cells.

“Accumulation and adipogenic differentiation of FAPs is responsible for the decline in function for dysferlinopathic muscle. Reversing this could provide a therapy for LGMD2B, a devastating disease with no effective treatment,” predicts Jaiswal as the team continues research in this field.

Promising off-the-shelf drugs include batimastat, an anti-cancer drug that inhibits the extracellular matrix enzyme matrix metalloproteinase. This drug reduces FAP adipogenesis in vitro and lessens injury-triggered lipid formation in vivo. In experimental models, batimastat also increases muscle function.

In addition to Jaiswal, Hogarth, Gallardo and Diaz Manera, other study co-authors include Aurelia Defour, Christopher Lazarski, Terence A. Partridge and Kanneboyina Nagaraju, all of Children’s National.

Financial support for research described in this post was provided by the Muscular Dystrophy Association under awards MDA477331 and MDA277389, the National Institute of Arthritis and Musculoskeletal and Skin Diseases under award R01AR055686 and the National Institutes of Health under awards K26OD011171, R24HD050846 and P50AR060836.

dystrophin protein

Experimental drug shows promise for slowing cardiac disease and inflammation

dystrophin protein

Duchenne muscular dystrophy (DMD) is caused by mutations in the DMD gene, which provides instructions for making dystrophin, a protein found mostly in skeletal, respiratory and heart muscles.

Vamorolone, an experimental medicine under development, appears to combine the beneficial effects of prednisone and eplerenone – standard treatments for Duchenne muscular dystrophy (DMD) – in the heart and muscles, while also showing improved safety in experimental models. The drug does so by simultaneously targeting two nuclear receptors important in regulating inflammation and cardiomyopathy, indicates a small study published online Feb. 11, 2019, in Life Science Alliance.

DMD is a progressive X-linked disease that occurs mostly in males. It is characterized by muscle weakness that worsens over time, and most kids with DMD will use wheelchairs by the time they’re teenagers. DMD is caused by mutations in the DMD gene, which provides instructions for making dystrophin, a protein found mostly in skeletal, respiratory and heart muscles.

Cardiomyopathy, an umbrella term for diseases that weaken the heart, is a leading cause of death for young adults with DMD, causing up to 50 percent of deaths in patients who lack dystrophin. A collaborative research team co-led by Christopher R. Heier, Ph.D., and Christopher F. Spurney, M.D., of Children’s National Health System, is investigating cardiomyopathy in DMD. They find genetic dystrophin loss provides “a second hit” for a specific pathway that worsens cardiomyopathy in experimental models of DMD.

“Some drugs can interact with both the mineralocorticoid receptor (MR) and glucocorticoid receptor (GR) since these two drug targets evolved from a common ancestor. However, we find these two drug targets can play distinctly different roles in heart and skeletal muscle. The GR regulates muscle inflammation, while the MR plays a key role in heart health,” says Heier, an assistant professor at Children’s National and lead study author. “In our study, the experimental drug vamorolone safely targets both the GR to treat chronic inflammation and the MR to treat the heart.”

After gauging the efficacy of various treatments in test tubes, the study team looked at whether any could mitigate negative impacts of the MR on heart health. Wild type and mdx experimental models were implanted with pumps that activated the MR. These models also received a daily oral MR antagonist (or inhibitor) drug, and either eplerenone, spironolactone or vamorolone. Of note:

  • MR activation increased kidney size and caused elevated blood pressure (hypertension).
  • Treatment with vamorolone maintained normal kidney size and prevented hypertension.
  • MR activation increased mdx heart mass and fibrosis. Vamorolone mitigated these changes.
  • MR activation decreased mdx heart function, while vamorolone prevented declines in function.
  • Daily prednisone caused negative MR- and GR-mediated side effects, such as hyperinsulinemia, whereas vamorolone safely improved heart function without these side effects.

“These findings have the potential to help current and future patients,” Heier says. “Clinicians already prescribe several of these drugs. Our new data support the use of MR antagonists such as eplerenone in protecting DMD hearts, particularly if patients take prednisone. The experimental drug vamorolone is currently in Phase IIb clinical trials and is particularly exciting for its unique potential to simultaneously treat chronic inflammation and heart pathology with improved safety.”

In addition to Heier and senior author Spurney, study co-authors include Qing Yu, Alyson A. Fiorillo, Christopher B. Tully, Asya Tucker and Davi A. Mazala, all of Children’s National; Kitipong Uaesoontrachoon and Sadish Srinivassane, AGADA Biosciences Inc.; and Jesse M. Damsker, Eric P. Hoffman and Kanneboyina Nagaraju, ReveraGen BioPharma.

Financial support for research described in this report was provided by Action Duchenne; the Clark Charitable Foundation; the Department of Defense under award W81XWH-17-1-047; the Foundation to Eradicate Duchenne; the Intellectual and Developmental Disabilities Research Center under award U54HD090257 (through the National Institutes of Health’s (NIH) Eunice Kennedy Shriver National Institute of Child Health and Human Development); and the NIH under awards K99HL130035, R00HL130035, L40AR068727 and T32AR056993.

Financial disclosure:  Co-authors employed by ReveraGen BioPharma were involved in creating this news release.

Preemie Baby

Getting micro-preemie growth trends on track

Preemie Baby

According to Children’s research presented during the Institute for Healthcare Improvement 2018 Scientific Symposium, standardizing feeding practices – including the timing for fortifying breast milk and formula with essential elements like zinc and protein – improves growth trends for the tiniest preterm infants.

About 1 in 10 infants is born before 37 weeks gestation. These premature babies have a variety of increased health risks, including deadly infections and poor lung function.

Emerging research suggests that getting their length and weight back on track could help. According to Children’s research presented during the Institute for Healthcare Improvement 2018 Scientific Symposium, standardizing feeding practices – including the timing for fortifying breast milk and formula with essential elements like zinc and protein – improves growth trends for the tiniest preterm infants.

The quality-improvement project at Children’s National Health System targeted very low birth weight infants, who weigh less than 3.3 pounds (1,500 grams) at birth. These fragile infants are born well before their internal organs, lungs, brain or their digestive systems have fully developed and are at high risk for ongoing nutritional challenges, health conditions like necrotizing enterocolitis (NEC) and overall poor development.

The research team measured progress by tracking the micro-preemies’ mean delta weight Z-score for weight gain, which measures nutritional status.

“In this cohort, mean delta weight Z-scores improved by 43 percent, rising from -1.8 to the goal of -1.0, when we employed an array of interventions. We saw the greatest improvement, 64 percent, among preterm infants who had been born between 26 to 28 weeks gestation,” says Michelande Ridoré, MS, Children’s NICU quality-improvement program lead who presented the group’s preliminary findings. “It’s very encouraging to see improved growth trends just six months after introducing these targeted interventions and to maintain these improvements for 16 months.”

Within Children’s neonatal intensive care unit (NICU), micro-preemies live in an environment that mimics the womb, with dimmed lighting and warmed incubators covered by blankets to muffle extraneous noise. The multidisciplinary team relied on a number of interventions to improve micro-preemies’ long-term nutritional outcomes, including:

  • Reducing variations in how individual NICU health care providers approach feeding practices
  • Fortifying breast milk (and formula when breast milk was not available), which helps these extra lean newborns add muscle and strengthen bones
  • Early initiation of nutrition that passes through the intestine (enteral feeds)
  • Re-educating all members of the infants’ care teams about the importance of standardized feeding and
  • Providing a decision aid about feeding intolerance.

Dietitians were included in the daily rounds, during which the multidisciplinary team discusses each infant’s care plan at their room, and used traffic light colors to describe how micro-preemies were progressing with their nutritional goals. It’s common for these newborns to lose weight in the first few days of life.

  • Infants in the “green” zone had regained their birth weight by day 14 of life and possible interventions included adjusting how many calories and protein they consumed daily to reflect their new weight.
  • Infants in the “yellow” zone between day 15 to 18 of life remained lighter than what they weighed at birth and were trending toward lower delta Z-scores. In addition to assessing the infant’s risk factors, the team could increase calories consumed per day and add fortification, among other possible interventions.
  • Infants in the “red” zone remained below their birth weight after day 19 of life and recorded depressed delta Z-scores. These infants saw the most intensive interventions, which could include conversations with the neonatologist and R.N. to discuss strategies to reverse the infant’s failure to grow.

Future research will explore how the nutritional interventions impact newborns with NEC, a condition characterized by death of tissue in the intestine. These infants face significant challenges gaining length and weight.

Institute for Healthcare Improvement 2018 Scientific Symposium presentation

  • “Improved growth of very low birthweight infants in the neonatal intensive care unit.”

Caitlin Forsythe, MS, BSN, RNC-NIC, NICU clinical program coordinator, Neonatology, and lead author; Michelande Ridoré, MS, NICU quality-improvement program lead; Victoria Catalano Snelgrove, RDN, LD, CNSC, CLC, pediatric clinical dietitian; Rebecca Vander Veer, RD, LD, CNSC, CLC, pediatric clinical dietitian; Erin Fauer, RDN, LD, CNSC, CLC, pediatric clinical dietitian; Judith Campbell, RNC, IBCLC, NICU lactation consultant; Eresha Bluth, MHA, project administrator; Anna Penn, M.D., Ph.D., neonatologist; Lamia Soghier, M.D., MEd, Medical Unit Director, Neonatal Intensive Care Unit; and Mary Revenis, M.D., NICU medical lead on nutrition and senior author; all of Children’s National Health System.