Immunization Against Oxidized Elastin Exacerbates Structural and Functional Damage in Mouse Model of Smoke-Induced Ocular Injury

We have a new manuscript out in Investigative Ophthalmology & Visual Science, Immunization Against Oxidized Elastin Exacerbates Structural and Functional Damage in Mouse Model of Smoke-Induced Ocular Injury

Authors: Balasubramaniam Annamalai; Crystal Nicholson; Nathaniel Parsons; Sarah Stephenson; Carl Atkinson; Bryan Jones; and Bärbel Rohrer.

Purpose: Age-related macular degeneration (AMD) is the leading cause of blindness in Western populations. While an overactive complement system has been linked to pathogenesis, mechanisms contributing to its activation are largely unknown. In aged and AMD eyes, loss of the elastin layer (EL) of Bruch’s membrane (BrM) has been reported. Elastin antibodies are elevated in patients with AMD, the pathogenic significance of which is unclear. Here we assess the role of elastin antibodies using a mouse model of smoke-induced ocular pathology (SIOP), which similarly demonstrates EL loss.

Methods: C57BL/6J mice were immunized with elastin or elastin peptide oxidatively modified by cigarette smoke (ox-elastin). Mice were then exposed to cigarette smoke or air for 6 months. Visual function was assessed by optokinetic response, retinal morphology by spectral-domain optical coherence tomography and electron microscopy, and complement activation and antibody deposition by Western blot.

Results: Ox-elastin IgG and IgM antibodies were elevated in ox-elastin immunized mice following 6 months of smoke, whereas elastin immunization had a smaller effect. Ox-elastin immunization exacerbated smoke-induced vision loss, with thicker BrM and more damaged retinal pigment epithelium (RPE) mitochondria compared with mice immunized with elastin or nonimmunized controls. These changes were correlated with increased levels of IgM, IgG2, IgG3, and complement activation products in RPE/choroid.

Conclusions: These data demonstrate that SIOP mice generate elastin-specific antibodies and that immunization with ox-elastin exacerbates ocular pathology. Elastin antibodies represented complement fixing isotypes that, together with the increased presence of complement activation seen in immunized mice, suggest that elastin antibodies exert pathogenic effects through mediating complement activation.

Teleworking In Response to COVID-19

The Marclab for Connectomics is immediately implementing teleworking strategies, moving many functions off campus in response to the COVID-19 outbreak/pandemic.

I am feeling fortunate in the much of our work is computational, but we do have a substantial wet lab component in addition to the ultrastructural data capture component.

We are not shutting down completely, yet. All students and postdocs will be working from home.  We have limited capacity to process tissue as only one person at a time will be in the laboratory proper, sectioning, performing immuno procedures, or capturing ultrastructural data.

I will be working intermittently from home and the laboratory office and working to determine where we are at from a personnel standpoint and how we will be impacted by NIH funding decisions. The good news is that NIH, I believe will honor charging salaries to grants, even if we cannot maintain our planned effort.  This means no job losses for the time being.

The COVID-19 outbreak is getting bigger and bigger.  I have cancelled all of my travel for the year to date, including a trip with friends to Lisbon in two weeks to walk around with cameras that I was planning on being the highlight of my year.  Trips to California, Italy, Sweden, and Germany are also off the table, at least through May/June.  ARVO has been cancelled. FASEB has been cancelled.

Why?

– COVID-19 is far more infectious than the flu.

– COVID-19 is far more deadly than the flu. On the order of 300-1000x more deadly. The difference is that it is not as common as the flu… yet.

-Our current Surgeon General just said on the 9th that the outbreak was already “contained”. I don’t even know where to start with this one as it is gross misinformation.

-I am seeing a level of buildup in the outbreak community and in the military that I’ve never seen before. This buildup is stymied by an administration that keeps talking it down. But I’ve never before seen a military response (calling out National Guard units specialized in biowarfare management) for an outbreak. At the same time, there is no coordination between the services that I can see. Navy is not talking with Army for instance, which is bizarre.

-The President has not invoked the Defense Production Act yet… I have no idea why.

-There is nobody at CDC that is coordinating all the information from the individual states as of today. This infrastructure *used* to exist. I don’t know why it is not there and that scares me, and makes me more angry than ever with the Trump administration.

HHS is estimating that a “moderate” outbreak will have 200,000 hospitalizations required for respiratory distress from COVID-19. We only have 100,000 ICU beds total in the United States, and some percentage of those are already in use, being optimized for markets. That looks to me like Italy’s situation right now, and their advanced medical system (among the best in Europe) is starting to fail which is why Italy has just extended their quarantine to the entire country.

-Per the Johns Hopkins dashboard, there is effectively no screening being done in Africa. No screening being done in Russia, with one backchannel report to me of at least 300 active cases in Moscow, but the doctors are not allowed to report it.

-No screening being done in the former Russian republics either.

-In all of India, there are only 43 reported cases. That does not make any sense to me.

-The lack of reporting for an infectious disease that is far more infectious than the flu indicates to me that COVID-19 is already in the population and is already spreading.

-Because it is relatively asymptomatic amongst much of the younger populace, it is not being seen/found. And as such, is a potent vector for spreading rapidly in communities.

-Universities around the country (not just my institution, the University of Utah) are prohibiting travel.

All of this means that if this is in fact as bad as it sounds, then we need to take active measures to limit the spread and the impact upon our healthcare infrastructure that is already going to be strained and we need to flatten the curve.

 

This *feels* very different from previous outbreaks, and the lack of coordination from the White House to CDC and .mil folks is astoundingly disturbing. Don’t forget, back in 2018, the Trump administration shut down the CDC foreign disease outbreak monitoring program.  This was the program that helped monitor foreign disease outbreaks and help define what sequences vaccine makers should use.

We have thus been relying upon WHO for information, and they even offered us primers for COVID-19 to make test kits.  But someone in the Trump administration told the CDC not to take them and we’d develop our own. My information says that those US developed primers had problems recognizing the epitope, and now people want to know *why* we rejected WHO primers. Speculation is that they wanted to use IP from the US made primers to capitalize on those markets.

This is infuriating and another example of why we should not be using economic models to address disease outbreaks, a position I’ve been taking on Twitter for some time now.

The WHO certified this outbreak as a pandemic on the 11th.  I have some real concerns, as some, like the former CDC director are advising that in a worst case scenario, up to 1.6 million Americans could be killed by COVID-19.  My concern is that from his numbers, even if this outbreak is just 1% infectious, and just moderately severe, that means 16,350 possible deaths.  That is not the bad part though. More importantly, 16,350 deaths will likely mean ICU beds are needed, and we only *have* 100k beds in the US.  Additionally, optimal occupancy for intensive care units (ICU) in the United States is accepted to be 70-75%. This means that at 1% infection rate, and moderately severe mortality rates, ALL of our ICU bed capacity could be occupied by COVID-19.  If it is worse, and the numbers from Italy’s example suggest it is, this could reach crisis levels quickly.

For all these reasons, I am taking active measures to reduce the exposure of people in my laboratory and help to reduce pressure on our healthcare systems.

 

Bryan William Jones, PhD

 

Müller Cell Metabolic Signatures: Evolutionary Conservation and Disruption in Disease

We have a new manuscript out in Trends in Endocrinology & Metabolism, Müller Cell Metabolic Signatures: Evolutionary Conservation and Disruption in Disease.

Authors: Rebecca L. Pfeiffer @BeccaPfeiffer19, Robert E. Marc @robertmarc60, and Bryan William Jones @BWJones.

This manuscript functions as both a review and presents some exciting new data demonstrating how the glutamate cycle is disrupted during retinal degenerative disease.

Abstract: Müller cells are glia that play important regulatory roles in retinal metabolism. These roles have been evolutionarily conserved across at least 300 million years. Müller cells have a tightly locked metabolic signature in the healthy retina, which rapidly degrades in response to insult and disease. This variation in metabolic signature occurs in a chaotic fashion, involving some central metabolic pathways. The cause of this divergence of Müller cells, from a single class with a unique metabolic signature to numerous separable metabolic classes, is currently unknown and illuminates potential alternative metabolic pathways that may be revealed in disease. Understanding the impacts of this heterogeneity on degenerate retinas and the implications for the metabolic support of surrounding neurons will be critical to long-term integration of retinal therapeutics for the restoration of visual perception following photoreceptor degeneration.

Optic Cup Morphogenesis Requires Neural Crest-Mediated Basement Membrane Assembly

We have a new manuscript out in Development, Optic cup morphogenesis requires neural crest-mediated basement membrane assembly.

Authors: Chase D. Bryan @CDBE30, Macaulie A. Casey, Rebecca L. Pfeiffer @BeccaPfeiffer19, Bryan W. Jones @BWJones, and Kristen M. Kwan @BlockInTheBack

This is a collaborative project out of the Kwan lab that we helped out with some of the ultrastructural work and analysis.

Abstract: Organogenesis requires precise interactions between a developing tissue and its environment. In vertebrates, the developing eye is surrounded by a complex extracellular matrix as well as multiple mesenchymal cell populations. Disruptions to either the matrix or periocular mesenchyme can cause defects in early eye development, yet in many cases, the underlying mechanism is unknown. Here, using multidimensional imaging and computational analyses in zebrafish, we establish that cell movements in the developing optic cup require neural crest. Ultrastructural analysis reveals that basement membrane formation around the developing eye is also dependent on neural crest, but only specifically around the retinal pigment epithelium. Neural crest cells produce the extracellular matrix protein nidogen: impairing nidogen function disrupts eye development, and strikingly, expression of nidogen in the absence of neural crest partially restores optic cup morphogenesis. These results demonstrate that eye formation is regulated in part by extrinsic control of extracellular matrix assembly.

Pathoconnectome Analysis of Müller Cells in Early Retinal Remodeling

We have a new manuscript out in Clinical Neurophysiology, An Update on Retinal Prostheses. PubMedDirect Link PDF here.

Authors: Rebecca L Pfeiffer, James R Anderson, Daniel P Emrich, Jeebika Dahal, Crystal L Sigulinsky, Hope AB Morrison, Jia-Hui Yang, Carl B Watt, Kevin D Rapp, Mineo Kondo, Hiroko Terasaki, Jessica C Garcia, Robert E Marc, and Bryan W Jones.

Abstract: Glia play important roles in neural function, including but not limited to amino acid recycling, ion homeostasis, glucose metabolism, and waste removal. During retinal degeneration and subsequent retinal remodeling, Müller cells (MCs) are the first cells to show metabolic and morphological alterations in response to stress. Metabolic alterations in MCs chaotically progress in retina undergoing photoreceptor degeneration; however, what relationship these alterations have with neuronal stress, synapse maintenance, or glia-glia interactions is currently unknown. The work described here reconstructs a MC from a pathoconnectome of early retinalremodeling retinalpathoconnectome 1 (RPC1) and explores relationships between MC structural and metabolic phenotypes in the context of neighboring neurons and glia. Here we find variations in intensity of osmication inter- and intracellularly, variation in small molecule metabolic content of MCs, as well as morphological alterations of glial endfeet. RPC1 provides a framework to analyze these relationships in early retinal remodeling through ultrastructural reconstructions of both neurons and glia. These reconstructions, informed by quantitative metabolite labeling via computational molecular phenotyping (CMP), allow us to evaluate neural-glial interactions in early retinal degeneration with unprecedented resolution and sensitivity.

 

 

An Update On Retinal Prostheses

We have a new manuscript out in Clinical Neurophysiology, An Update on Retinal Prostheses. PubMedDirect Link PDF here.

Authors: Lauren N. Ayton, Nick Barnes, Gislin Dagnelie, Takashi Fujikado, Georges Goetz, Ralf Hornig, Bryan W. Jones, Mahiul M.K. Muqit, Daniel L. Rathbun, Katarina Stingl, James D. Weiland, Matthew A. Petoe.

Abstract: Retinal prostheses are designed to restore a basic sense of sight to people with profound vision loss. They require a relatively intact posterior visual pathway (optic nerve, lateral geniculate nucleus and visual cor- tex). Retinal implants are options for people with severe stages of retinal degenerative disease such as retinitis pigmentosa and age-related macular degeneration.

There have now been three regulatory-approved retinal prostheses. Over five hundred patients have been implanted globally over the past 15 years. Devices generally provide an improved ability to localize high-contrast objects, navigate, and perform basic orientation tasks. Adverse events have included con- junctival erosion, retinal detachment, loss of light perception, and the need for revision surgery, but are rare. There are also specific device risks, including overstimulation (which could cause damage to the retina) or delamination of implanted components, but these are very unlikely.
Current challenges include how to improve visual acuity, enlarge the field-of-view, and reduce a com- plex visual scene to its most salient components through image processing. This review encompasses the work of over 40 individual research groups who have built devices, developed stimulation strategies, or investigated the basic physiology underpinning retinal prostheses. Current technologies are summarized, along with future challenges that face the field.

 

 

Kevin Rapp At TEM1

This is Kevin Rapp, a technician in the lab running some test captures on our original JEOL transmission electron microscope.  This microscope is likely the highest use electron microscope on planet Earth, having been run effectively non-stop for over a decade.  This microscope, purchased through a generous gift from Martha Ann Dumke Healey made possible our entire connectomics initiatives.

From Jonesblog.

 

Becca Pfeiffer At TEM2

This is Becca Pfeiffer setting up a test capture on our new JEOL electron microscope. We’ve customized this scope, like our previous scope, and it is taking us a little while to track down some variables with a piece of equipment this complex and hammer them down. My thanks to Becca, Jamie and Kevin for working through this together.

From Jonesblog.

Contrast Sensitivity Isocontours Of The Central Visual Field

We have a new manuscript out in Nature Scientific Reports, Contrast Sensitivity Isocontours Of The Central Visual Field.

Authors: Agnes Y. J. Choi, Lisa Nivison-Smith, Jack Phu, Barbara Zangerl, Sieu K. Khuu, Bryan W. Jones, Rebecca L. Pfeiffer, Robert E. Marc & Michael Kalloniatis.

Abstract: Standard automated perimetry (SAP), the most common form of perimetry used in clinical practice, is associated with high test variability, impacting clinical decision making and efficiency. Contrast sensitivity isocontours (CSIs) may reduce test variability in SAP by identifying regions of the visual field with statistically similar patterns of change that can be analysed collectively and allow a point (disease)-to-CSI (normal) comparison in disease assessment as opposed to a point (disease)-to-point (normal) comparison. CSIs in the central visual field however have limited applicability as they have only been described using visual field test patterns with low, 6° spatial sampling. In this study, CSIs were determined within the central 20° visual field using the 10-2 test grid paradigm of the Humphrey Field Analyzer which has a high 2° sampling frequency. The number of CSIs detected in the central 20° visual field was greater than previously reported with low spatial sampling and stimulus size dependent: 6 CSIs for GI, 4 CSIs for GII and GIII, and 3 CSIs for GIV and GV. CSI number and distribution were preserved with age. Use of CSIs to assess visual function in age-related macular degeneration (AMD) found CSI guided analysis detected a significantly greater deviation in sensitivity of AMD eyes from normal compared to a standard clinical pointwise comparison (−1.40 ± 0.15 dB vs −0.96 ± 0.15 dB; p < 0.05). This work suggests detection of CSIs within the central 20° is dependent on sampling strategy and stimulus size and normative distribution limits of CSIs can indicate significant functional deficits in diseases affecting the central visual field such as AMD.