Neurodegeneration Could Be Slowed by Restoring Copper Ion Balance in Glial Cells

Studies in the roundworm model organism Caenorhabditis elegans, headed by scientists at Florida Atlantic University’s Schmidt College of Medicine and FAU Stiles-Nicholson Brain Institute, have resulted in discoveries that may one day lead to the development of new treatments for human neurodegenerative disorders. Research lead Randy D. Blakely, PhD, and colleagues, linked function of the C. elegans gene swip-10 to the control of copper (Cu), a vital micronutrient that plays several essential roles in all cells, including those of the human brain.

Their studies found that expression of the C. elegans swip-10 gene in glial cells plays an essential role in homeostasis of the Cu(I) form of copper and associated metabolic functions. Deleting swip-10 negatively impacted on systemic levels of Cu(I), leading to deficits in mitochondrial respiration and ATP production, increased oxidative stress, and neurodegeneration. Conversely, the scientists found that increasing Cu(I) levels or restoring swip-10 expression in glia limited these changes.

“Copper is required for the function of mitochondria, the powerhouse of cells, and its production of the energy-storing molecule ATP, which fuels hundreds of vital body functions such as muscle contraction, digestion and heart function as well as the signaling of brain neurons that allows us to think and feel,” said Blakely, the David J.S. Nicholson Distinguished Professor in Neuroscience at FAU. “Copper also helps protect cells from harmful molecules termed reactive oxygen species, or ROS, which in excess can damage proteins and DNA, ultimately driving cell death, including neurons that die in Parkinson’s and Alzheimer’s disease.”

Reporting on their studies in a paper in PNAS “Glial swip-10 controls systemic mitochondrial function, oxidative stress, and neuronal viability via copper ion homeostasis,” the authors stated that their collective studies “… reveal a glial-expressed pathway that supports systemic mitochondrial function and neuronal health via regulation of Cu(I) homeostasis, a mechanism that may lend itself to therapeutic strategies to treat devastating neurodegenerative diseases.”

“Copper (Cu) is an essential micronutrient involved in numerous fundamental aspects of cell physiology including mitochondrial respiration, ATP production, suppression of oxidative stress, redox-dependent biosynthetic pathways, and metal ion-supported cell signaling,” the authors commented. A large amount of dietary copper—about 6%—is ultimately stored in the brain, they continued, which is second only to the liver and muscle in Cu concentrations.

In biological systems copper exists mainly in two forms: cuprous copper, termed Cu(I), and cupric copper, termed Cu(II). These two forms are managed by different proteins in the body and can be converted from one to the other to support various chemical reactions vital to human health. Scientists are still studying how the body maintains the right balance between these two copper forms, which is important, as too much or too little of either can wreak havoc on cells, particularly neurons.

The genetic disorders, Menke’s disease and Wilson’s disease, which are characterized by systemic diminished, and excess copper concentrations, respectively, both result in neurodegeneration, the team further pointed. “Altered CNS Cu homeostasis has also been linked to more common neurodegenerative diseases (NDDs), including Alzheimer’s disease (AD) and Parkinson’s disease (PD).” However, the researchers added, “despite the critical roles that require control of Cu ion homeostasis, the mechanisms that dictate the balance between Cu(I) and Cu(II) levels, particularly in the brain, remain an area of investigation.”

Blakely and colleagues, led by Andrew Hardaway, PhD, had reported their identification of the swip-10 gene in 2015, following a screen for molecules needed to control the activity of worm dopamine (DA) neurons, specifically those that control their ability to swim. “Worms with a damaging mutation in swip-10 initially swim normally, but unlike normal worms whose swimming continues for 30 minutes or more, in less than a minute, the mutants display swimming-induced paralysis or Swip,” said Blakely. “We tracked the paralysis to an excess activity of their dopamine neurons and published what we thought was a fairly complete story.”

Further studies by another past graduate student in the Blakely lab, Chelsea Gibson, PhD, showed that the overactive dopamine neurons in swip-10 mutants exhibit degeneration much earlier in life than normal worms, as seen in Parkinson’s disease (PD). Other types of neurons in swip-10 mutant worms besides those making dopamine also demonstrate degeneration, suggesting to Blakely’s team that links to brain disease might mirror other neurodegenerative disorders besides PD.

A clue to such disorders came with decoding of the swip-10 gene sequence, where Blakely’s team found that humans possess a gene highly related to swip-10, termed MBLAC1. “Amino acid sequence alignment of SWIP-10 to human proteins revealed strongest identity to the protein MBLAC1 …” the team explained in their newly reported paper. Then, in 2019, geneticist Iris Broce, PhD, at the University of California, San Francisco, pointed to MBLAC1 as a risk factor for a particular form of Alzheimer’s disease (AD), one accompanied by cardiovascular disease (AD-CDV).

Importantly, studies also showed a significant reduction in MBLAC1 expression in the frontal cortex of humans with AD-CDV, suggesting a role for MBLAC1 in supporting the health of both the brain and peripheral organs such as the heart. “Studies by Broce and colleagues who associated a risk for AD with cardiovascular disease (CVD) comorbidity through genome-wide association studies and postmortem cortical MBLAC1 mRNA expression studies, along with the death of DA neurons in swip-10 mutants, further encouraged our efforts to identify this activity,” the Blakely and colleagues stated in their newly released paper.

So where is the copper link? “It turns out that MBLAC1 encodes an enzyme key to the production of another class of proteins, termed histones, well known to compact long strands of DNA so they can form chromosomes,” said Blakely. But certain histones possess an additional, surprising activity, the ability to convert Cu(II) to Cu(I). When mutations in these proteins were generated by Narsis Attar, MD, PhD, at the University of California, Los Angeles, these cells show much lower production of Cu(I), higher amounts of ROS, their mitochondria function poorly, and they fail to thrive.

Connecting the dots across the years, Peter Rodriguez Jr., a current graduate student and lead scientist on the study in the Blakely lab, reasoned that swip-10 mutants also would fail to produce the requisite histones, leading to a loss of Cu(I), mitochondrial dysfunction, and an elevation of ROS, which could be a major reason the worm’s dopamine neuron die. “… we hypothesized that SWIP-10 might also contribute to Cu(I) production and homeostasis,” they wrote.

Through their studies described in PNAS Rodriguez Jr, and collaborators showed that this is indeed the case. “… through metabolic, biochemical, imaging, and genetic studies, we provide evidence that glial-expressed swip-10 provides essential, systemic support for Cu(I) homeostasis and phenotypes dependent on the micronutrient’s availability, including mitochondrial respiration, suppression of oxidative stress, and DA neuron viability,” they commented.  The investigators further demonstrated that it’s possible to rescue ATP production, reduce ROS, and promote survival of dopamine neurons by supplementing the C. elegans diet with Cu(I) or by exposing them to a drug known to increase Cu(I) levels in cells.

“Surprisingly, the impact of loss of swip-10 on Cu(I), worm bioenergetics, and oxidative stress is not just an impact felt by dopamine neurons,” said Rodriguez Jr. “Rather, Cu(I) levels, and these good things that Cu(I) does, are greatly diminished body-wide. Another striking finding is that though changes occur with Cu(I) and its actions across the body, these deficits arise from the loss of swip-10 from a small number of cells in the head of the animal known as glia, which make up only 5% of the cells in the animal’s body.”

Glial cells are well known to support the signaling and health of neurons in many organisms. Indeed, in the worm, Rodriguez Jr. could restore the health of worms, as well as whole body Cu(I) levels, by expressing a normal copy of the swip-10 gene only in glial cells. “The powerful control of Cu(I) exerted by swip-10 points to a novel opportunity to sustain neuronal health,” said Blakely. The authors further noted, “Our work implicates swip-10 and orthologs as key determinants of Cu(I) homeostasis that may be exploitable to treat neurodegenerative diseases and their metabolic comorbidities.”

Interestingly, the antibiotic ceftriaxone, which the Blakely lab found to bind MBLAC1 protein, has been reported by multiple groups to be neuroprotective in vitro and in animal models, though its mechanism of action is currently unclear. Blakely’s team believes ceftriaxone’s action relates to modulating copper homeostasis.

“Ceftriaxone isn’t a particularly powerful drug, doesn’t get into the brain very well compared to other medications, and can cause antibiotic resistance and other side-effects. So it’s not surprising that it hasn’t proved useful in the clinic,” said Blakely. “Perhaps now that we have a better idea as to what swip-10 and MBLAC1 do, we think that we may be able to design a truly useful medication to treat neurodegenerative disease.” Their work, they noted, “… provides support for a hypothesis that metabolic changes induced by MBLAC1-dependent Cu(I) homeostasis may contribute to PD risk and/or serve as a target for therapeutics in this and other neurodegenerative disorders.”