2021-02-26
Dr. Sara Sheibani, an affiliated academic member at Department of Anatomy and Cell Biology, and Dr. Kaustuv Basu, staff scientist at the Facility for Electron Microscopy Research, 捆绑SM社区, are part of an international team of researchers that has developed a new method to better understand how nanomedicines 鈥� emerging diagnostics and therapies that are very small yet very intricate 鈥� interact with patients鈥� biomolecules.
With the emergence of nanotechnology decades ago and its application in nanomedicine, the use of nanoscale materials, such as biocompatible nanoparticles as drug delivery system, diagnosis, and therapeutic tools has contributed significantly to the advancement of biomedical research. However, despite substantial progress, laboratory and clinical research in nanomedicine currently face serious challenges in research and development, clinical trials, and successful commercialization processes.
One of the most promising aspects of nanomedicine has been its capability to offer site-specific targeted treatment. 鈥淭argeted, personalized medicine involving nanomaterials has the potential to revolutionize diagnosis, drug delivery, and therapeutics,鈥� says Dr. Sheibani. 鈥淢ost studies in nanomedicine, however, have overlooked several important factors including the alteration of functionalized nanoparticles (NPs) upon exposure to culture medium and body fluid.鈥�
Leveraging 捆绑SM社区鈥檚 state-of-the-art microscopy capacity
To address some of these challenges, Drs. Sheibani and Basu collaborated with Dr. Morteza Mahmoudi, Assistant Professor in the Department of Radiology and the Precision Health Program at Michigan State University, who is a world leader in designing and developing nanomaterials for diagnostic and therapeutic applications. In a recent joint study, the results for which were published online January 25 in the journal听,听 Drs. Sheibani and Basu and colleagues at 捆绑SM社区 used the most advanced state-of-the-art cryo-microscopy techniques听at the 捆绑SM社区 Facility of Electron Microscopy Research to investigate the morphological feature of proteins and biomolecules coating on the surface of NPs referred to as a corona (not to be confused with the novel coronavirus), the Latin word for crown. This corona contains clues about how nanoparticles interact with a patient鈥檚 biology. Now, the team has shown how to get an unprecedented view of that corona at atomic scale.
鈥淥ne of the main challenges has been our poor understanding of the mechanism(s) of protein鈥損rotein interaction within the corona and the relationship and association of biomolecules with the surface of NPs,鈥� notes Dr. Sheibani. The application of cryo-electron microscopy, cryo-electron tomography, 3D reconstruction and image processing, and image simulation showed the variation in the structure and distribution of the biomolecules in the coronas of hundreds of individual NPs. It also enables the visualization of the individual biomolecules.
Findings may offer promise for future clinical applications
鈥淥ur findings demonstrate that the application of therapeutic NPs is more challenging than predicted in the published literature,鈥� adds Dr. Sheibani. 鈥淏iosystems, including the immune system, respond to NPs at the level of a single NP. The heterogeneous nature of the biomolecular corona (BC), therefore, significantly affects their safety and biological efficacy. The results of our study could lead to a better understanding of the function of the BC and its nonuniform behavior, at a resolution of a single NP, which significantly affects the outcomes of in vitro and in vivo experiments as well as most of the reported clinical trials. It helps the nanomedicine community to define the accuracy and reliability of proteomics and analytical chemistry data of the BC at the surface of NPs; the latter is important for defining the suitability of various types of NPs for clinical applications.鈥�
Image: Fig: Representation of distribution pattern of clusters of biomolecules of the surface of individual NPs. The image is a snapshot generated from the 3D volume of the following movie.
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2021-02-09
Electronic Cryomicroscopy: A New Era for Biology
A clever microscopy technique puts researchers in full view: it allows to observe molecules in action without altering them. This is enough to push the boundaries of science and accelerate the discovery of drugs, among others for COVID-19.
"Wait, I'm going to turn the camera." Joaquin Ortega, with whom I speak on Zoom, shows me his second computer. The quality of the image by interposed screens leaves something to be desired, but the small clusters pointed out by the biochemist clearly stand out against the greyish and grainy background. "You see? These are ribosomes, the cell machines that make proteins, he explains to me with a hint of excitement in his voice. The microscope is taking these photos live. The instrument in question, located on 捆绑SM社区 premises, is partially automated and can be remotely manipulated by the researcher, who is at home. A chance in these times of pandemic.
For the novice eye, these small clumped balls could be dust on the lens as well as cells magnified by any magnifying glass that is somewhat inefficient. Except that electronic cryomicroscopes like this, there are only a handful in the world. And these ribosomes are actually 1,000 times smaller than a cell.
It's simple, this type of device, which uses a beam of electrons to photograph biological samples frozen at -180 degrees Celsius, now offers a resolution of a few tenths of a nanometer. Enough to see individual atoms within a molecule, as demonstrated simultaneously听one German and the other听in the spring of 2020. "We crossed the final barrier of resolution," said Holger Stark of the Max Planck Institute for Biophysical Chemistry in Germany, who led one of the studies. "It opens up a whole new world," commented its British competitor, Radu Aricescu, in the journal听. Because once the grey images are superimposed and cleaned by computer, the molecules reveal themselves with incredible precision, like balls of wool whose every loop, each fiber can be seen in three dimensions.
And that's not all: electronic cryomicroscopy or cryo-ME, which won the Nobel Prize to three chemists in 2017, pushes the boundaries of science by allowing these molecules to be crunched into action. Drugs blocking an enzyme, protein complexes in full assembly, antibodies attaching to their targets or viruses attacking a cell: almost everything is now observable on the spot, like never before. The technique, by adding up hundreds of photographs, captures molecules from all angles and orientations. "You see the chemistry being done," says Joaquin Ortega.
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2020-10-28
Transforming a coronavirus protein into a nanoparticle could be the key to an effective COVID-19 vaccine.
Changing makeup of a specific protein has the potential to neutralize the virus
Researchers from 捆绑SM社区 are part of an international team led by the University of Buffalo that has discovered a technique that could help increase the effectiveness of vaccines against SARS-CoV-2, the virus that causes COVID-19. The group鈥檚 study, titled 鈥淪ARS-CoV-2 RBD Neutralizing Antibody Induction is Enhanced by Particulate Vaccination,鈥� was published online in the journal听Advanced Materials听on October 28 <>.
COVID-19 has caused a disruptive global pandemic, infecting at least 40 million and causing more than 1 million deaths worldwide. Since it began spreading in early 2020, biomedical researchers have been actively pursuing an effective vaccine. Now, researchers suggest one approach that may be effective is designing vaccines that partially mimic the structure of the virus.
鈥淭he most important contribution of this paper is it opens a novel approach to develop a COVID-19 vaccine,鈥� says Prof. Joaquin Ortega, Professor in the Department of Anatomy & Cell Biology at 捆绑SM社区 and a co-author of the study. 鈥淲e found that presenting one of the most important antigens in the SARS-CoV-2 virus, the receptor-binding domain (RBD), in the surface of nanodevices called liposomes induces a much stronger antibody response than when the RBD antigen is administered by itself,鈥� explains Prof. Ortega. He noted that liposomes are used extensively as nanocarriers for drug delivery in cancer. 鈥淭his paper shows that liposomes are also a unique platform for antigen presentation to the immune system and the development of effective vaccines.鈥�
Importantly, the findings from this study create a flexible COVID-19 vaccine development platform. The paper also shows the new approach works efficiently when adsorbing the RBD antigen to the liposome鈥檚 surface. 鈥淗owever, the approach is, in principle, extendable to any other antigens from the Sars-CoV-2 virus,鈥� notes Prof. Mike Strauss, Assistant Professor in the Department of Anatomy & Cell Biology at 捆绑SM社区, and a co-author on the paper. 鈥淲e still do not know which of the viral antigens are causing the most robust immunity. Besides, specific antigens may generate adverse effects, including lung tissue damage, a significant issue encountered in the past during the attempts to develop vaccines against SARS-CoV-1 in the 2000s. Having a flexible platform for vaccine development, such as the one described in our paper, allows us to create 鈥榓-la-carte鈥� vaccines that only incorporate those antigens triggering a beneficial immune response but excluding those responsible for adverse effects.鈥�
Leveraging 捆绑SM社区鈥檚 advanced microscopes for vaccine platform development
捆绑SM社区鈥檚 contribution to this vaccine platform鈥檚 development was in the structural characterization of the nanoparticles used to deliver the antigen to the immune system, using the research infrastructure at the Facility for Electron Microscopy Research (FEMR) at 捆绑SM社区, the largest and most versatile cryo-electron microscope platform in Canada. This facility houses some of the most advanced and fastest electron microscopes in the world, producing extremely high-resolution images that are essential to characterize nanoparticles, such as those used as antigen carriers in these vaccines. Cryo-electron microscopy (cryoEM) images constitute the gold standard technique to ensure the liposomes carrying the antigens in the vaccine have the desired size and structure to induce a robust immune response.
Professors Strauss and Ortega, who are respectively the Technical Director and Scientific Director at the FEMR, loaded approximately three microlitres of the vaccine mixture into the Thermo Scientific Titan Krios microscope at the FEMR. In the span of one hour, the Krios microscope produced over 300 images of the small lipidic vesicles contained on the vaccine. These images verified that the RBD antigen was being adsorbed in the lipidic surface of the liposome, which is the optimal location for antigen presentation to the immune system.
鈥淭he FEMR cryo-EM images showing the structure and location of the RBD antigen in the surface of the lipidic vesicles was essential data for the development of this SARS-CoV-2 vaccine,鈥� says Prof. Ortega. 鈥淭here is no other method that allows for the direct visualization of the antigen adsorbed to the liposomes鈥� surface. Other biophysical methods can produce an indirect measurement suggesting the antigens may have been incorporated. Still, only cryo-electron microscopy is capable of visualizing these antigens on the surface of the liposomes directly.鈥�
While the paper shows that this SARS-CoV-2 vaccine induces a robust immune response in mice animal models, the researchers say that like any other vaccine, it must undergo extensive additional testing and clinical trials before it can be administered to human communities. They also note the evidence presented in this study shows the RBD antigen benefits from being in a particle format, which could help inform future vaccine design that targets this specific antigen.
Authors on the study include Jonathan Lovell, Wei-Chiao Huang, Shiqi Zhou, Xuedan He听听and Moustafa T. Mabrouk, all from the University of Buffalo Department of Biomedical Engineering; Kevin Chiem and Luis Martinez-Sobrido, both from Texas Biomedical Research Institute; Ruth H. Nissly, Ian M. Bird and Suresh V. Kuchipudi, all from the Animal Diagnostic Laboratory, Department of Veterinary and Biomedical Sciences at Pennsylvania State University; Mike Strauss and Joaquin Ortega from the Department of Anatomy and Cell Biology at 捆绑SM社区; Suryaprakash Sambhara from the Immunology and Pathogenesis Branch of the U.S. Centers for Disease Control and Prevention; Elizabeth A. Wohlfert from the Department of Microbiology and Immunology at UB; and Bruce A. Davidson from the Department of Anesthesiology and the Department of Pathology and Anatomical Sciences at UB.
The study was supported by the U.S. National Institutes of Health,听the 捆绑SM社区 MI4 Interdisciplinary Initiative in Infection and Immunity,听and the Facility for Electron Microscopy Research (FEMR) at 捆绑SM社区. The FEMR is supported by the Canadian Foundation for Innovation, Quebec Government and 捆绑SM社区.
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2020-08-12
Nanotubes in the eye that help us see
Researchers find a new structure by which cells in the retina communicate with each other, regulating blood supply to keep vision intact
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A new mechanism of blood redistribution that is essential for the proper functioning of the adult retina has just been discovered in vivo by researchers at the University of Montreal Hospital Research Centre (CRCHUM).
Their study was published in听Nature.
"For the first time, we have identified a communication structure between cells that is required to coordinate blood supply in the living retina," said Dr. Adriana Di Polo, a neuroscience professor at Universit茅 de Montr茅al and holder of a Canada Research Chair in glaucoma and age-related neurodegeneration, who supervised the study.
"We already knew that activated retinal areas receive more blood than non-activated ones," she said, "but until now no one understood how this essential blood delivery was finely regulated."
The study was conducted on mice by two members of Di Polo's lab: Dr. Luis Alarcon-Martinez, a postdoctoral fellow, and Deborah Villafranca-Baughman, a PhD student. Both are the first co-authors of this study.
In living animals, as in humans, the retina uses the oxygen and nutrients contained in the blood to fully function. This vital exchange takes place through capillaries, the thinnest blood vessels in all organs of the body. When the blood supply is dramatically reduced or cut off -- such as in ischemia or stroke -- the retina does not receive the oxygen it needs. In this condition, the cells begin to die and the retina stops working as it should.
Tunnelling between cells
Wrapped around the capillaries are pericytes, cells that have the ability to control the amount of blood passing through a single capillary simply by squeezing and releasing it.
"Using [the FEI Helios Nanolab 660 DualBeam at the Facility for Electron Microscopy Research at 捆绑SM社区]听to visualize vascular changes in living mice, we showed that pericytes project very thin tubes, called inter-pericyte tunnelling nanotubes, to communicate with other pericytes located in distant capillaries," said Alarcon-Martinez. "Through these nanotubes, the pericytes can talk to each other to deliver blood where it is most needed."
Another important feature, added Villafranca-Baughman, is that "the capillaries lose their ability to shuttle blood where it is required when the tunnelling nanotubes are damaged -- after an ischemic stroke, for example. The lack of blood supply that follows has a detrimental effect on neurons and the overall tissue function."
The team's findings suggest that microvascular deficits observed in neurodegenerative diseases like strokes, glaucoma, and Alzheimer's disease might result from the loss of tunnelling nanotubes and impaired blood distribution. Strategies that protect these nanostructures should then be beneficial, but remain to be demonstrated.
This research was supported by the Canadian Institutes of Health Research.
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