IL-4/C1q activated astrocyte-derived extracellular vesicles promote stroke infarct recovery by recruiting peripheral leukocytes

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Xin S, Zhang L, Phan NV, An M, Shi L, Carmichael ST, Segura T. IL-4/C1q activated astrocyte-derived extracellular vesicles promote stroke infarct recovery by recruiting peripheral leukocytes. Cell Biomaterials. 2026 Jan 1;

01.01.26 Cell Biomaterials IL-4/C1q activated astrocyte-derived extracellular vesicles promote stroke infarct recovery by recruiting peripheral leukocytes

Regeneration of the necrotic infarct core is not currently a goal of clinical stroke therapies. However, restoring this region would be essential for complete and durable functional recovery after ischemic stroke. We engineered an injectable microporous annealed particle scaffold (MAPS) with covalently anchored extracellular vesicles (EVs) derived from primary astrocytes activated by IL-1α/TNF-α/C1q or IL-4/C1q, and delivered them into the infarct cavity of mice five days post-stroke. Only MAPS with IL-4/C1q-EVs induced robust revascularization, with perfused microvessels forming in the infarct center in 9 days, and motor function restored to near-baseline levels by 8 weeks. These regeneration and functional improvement were associated with selective recruitment of reparative neutrophils and macrophages into the infarct. Neutrophil depletion abrogated angiogenesis, demonstrating that neutrophils are required for mediating tissue repair. This immune-vascular crosstalk is associated with IL-4-induced enrichment of EV cargo, including adhesion- and angiogenesis-related miRNAs, such as miR-377-3p and miR-143-5p.

Xin S Zhang L Phan NV An M Shi L Carmichael ST Segura T

Engineering Dynamic MAP Hydrogels to Modulate Cellular Behavior Toward Pro-Repair

From Connor’s lab diary:

I’ve always enjoyed building things – from Legos as a child, to hardwood furniture as my tastes evolved with time. I am fascinated by how we can engineer materials to direct cellular behavior, from assembly to tissue formation, with spatiotemporal control. My work is inspired by biology, because what’s more inspiring than active building blocks that evolved over eons?

All human tissues exhibit stress relaxing behavior, or rather, they gradually reduce internal stress under a fixed deformation. Recently, it’s become clear that stress relaxing biomaterials modulate cellular behavior, such as migration and morphogenesis, without the need for additional factors. Furthermore, we can tune biomaterial stress relaxation by using dynamic chemistries, because dynamic exchange of reversible bonds can result in stress relaxing behavior. Here in the Segura Lab, I am developing microporous annealed particle (MAP) hydrogels using dynamic chemistries and investigating how we can tune these MAP hydrogels to modulate cellular behavior and promote repair.

Reach out if you want to work with me and mention my project #connor #dynamicmatter

Systems in Conversation

From Emma’s lab diary:

As an avid outdoorswoman, I walk through life with John Muir quotes ringing in my mind. Here’s one of my favorites: “When we try to pick out anything by itself, we find it hitched to everything else in the Universe.” In other words: it’s all connected! My work in the Segura Lab considers this in the biomedical setting by thinking about how local and systemic systems are in conversation, and how we can leverage this communication toward better health.

One of the main applications I’m working on is stroke-immunology. The post-stroke immune response is highly complex: there is local neuroinflammation and contradictory, systemic immunosuppression. This leaves patients highly susceptible to post-stroke infections like pneumonia; many patients survive the initial stroke event but later die from infection. Due to the oppositional nature of these immune responses, efforts to address one generally exacerbate the other.

My project seeks to harmonize the conversation between the post-stroke immune responses through two disease-informed strategies: (1) by targeting key, upstream pathways to treat the two responses before they become too divergent. (2) by synergizing the two responses to promote endogenous healing. To accomplish these goals, I’m leveraging MAP hydrogel and aptamer technologies, creating a specific, targeted therapy. Our previous stroke-related work in the Segura Lab has focused on promoting repair locally; I am interested in the knowledge gap and therapeutic potential that arises when we bring systemic systems into the conversation.

If you’re into engineering the dialogue between local and systemic systems, contact us and add in the message that you’re interested in my project! #emma #systemsinconversation

Combining Optimized MAP Scaffolds with Cortical Stimulation for Post-Stroke Repair

From Jeremy’s lab diary:

Current stroke treatments save lives, but nearly two-thirds of survivors live with long-term disabilities that make everyday life a challenge. Many promising new therapies focus on the later phases of recovery to try to reduce this lasting burden. Two in particular caught my attention. Electrical brain stimulation can encourage neurons to form new connections and even promote the growth of blood vessels. Meanwhile, our lab’s MAP scaffolds create a supportive environment for tissue repair in the injured brain. The challenge is that there are countless ways to design these scaffolds, and traditional low-throughput methods likely miss the most effective ones.

That’s why I’m excited about combining these approaches! MAP scaffolds can be injected directly into the damaged region, while cortical stimulation can be applied to the same area. Rather than interfering with each other, the two could actually work hand in hand. My goal is to make MAP formulation optimization feasible and pair them with stimulation, with the hope that together they can achieve what neither could accomplish alone.

If this topic sparks your interest, reach out and mention that you’re interested in my project! #jeremy #sparkingrepair

Quantifying relationships between pore structure and cell behavior

From Yasha’s lab diary:

Model Behavior? Nope – not referring to the Justin Timberlake, Maggie Lawson Disney channel original movie where a model and a nerd switch lives for a bit. (Please sign me up though.)

For my PhD thesis, I want to uncover structure-function relationships in our MAP scaffolds. Cells tend to be sensitive to micron-scale deviations in their environment. Through our custom LOVAMAP software, we can identify pores in 3D images of packed particles. We now know that pore structures are tunable based on the material properties of the particles in the scaffold. With LOVAMAP, we can also characterize individual pores by 23 geometric descriptors.

We’ve seen cells respond differently to different levels of spatial confinement, so quantifying the effects of pore structure on cell behavior is a worthy cause. I’m specifically interested in how pore structure affects cell branching and cell migration. I will use LOVAMAP to collect data about the pores in a scaffold and develop image processing approaches to segment cellular content in microscopy images of the scaffolds. I will also use simulation approaches to simulate transport behaviors in our material. Exploring the pore data and the cell data will lead to indications of what metrics between the two are statistically correlated. Establishing models like this will eventually lead to more data-driven approaches for designing MAP scaffolds.

To uncover this, I will need to employ methods from image processing and machine learning to reconstruct and identify individual particles and cells from microscopy data as the preprocessing step for LOVAMAP input, as well as measure key aspects about the cells located in each pore.

I’m hoping the output of my thesis will be a curve or a simulation that inputs pore properties about a MAP scaffold and outputs a measurable effect it will have on cellular behavior. So, if you want to help me model behavior (or if you have a spare GPU lying around), let me know! #yasha #modelbehavior

Enhancing stroke recovery in aged populations

From Andrea’s lab diary:

While my research career began in the brain studying tumor angiogenesis and the blood-brain barrier, I found myself in reproductive biology and tissue engineering as a graduate student, where I began to appreciate the power and complexity of hormones. They have so many crucial functions outside the endocrine system and their dynamic, pulsatile profiles are fascinating. I found myself wondering how hormones may impact stroke pathogenesis and brain tissue regeneration. It turns out estrogens have some potent pro-regenerative effects that are difficult to leverage because of their chemical properties. Enter – MAP!

I am exploring how MAP scaffolds may serve as a delivery system to cloak the lipophilic properties of estrogens and improve their therapeutic efficacy at the site of ischemic stroke. This also brings up some exciting opportunities to understand how the intersection of sex, hormones, and aging impact healing after stroke.

If you are a student interested in my work on estrogen, MAP, and stroke (do you have hormones on the brain like me???) contact us and add in the message that you’re interested in my project!” #Andrea #hormonesonthebrain

Enhancing stroke recovery in aged populations

From Cara’s lab diary:

Throughout my undergraduate and graduate research (and now my postdoc), I’ve always had a common theme – and that theme is the central nervous system! Following injury (which includes stroke, traumatic brain injury, and spinal cord injury), the central nervous system, or CNS, fails to heal like other tissues in the body, which can result in chronic paralysis or other neurological issues. In aged populations, recovery can be especially difficult, as many of the cells involved in the injury response not only have to deal with problems caused by damage, but also problems caused by getting older.

The Segura Lab is well known for their prior work delivering materials known as microporous annealed particle (MAP) scaffolds to the infarct site following stroke as a therapy. MAP scaffolds have been shown to affect cellular behavior post-stroke – as well as improve recovery! – in young mouse animal models. When I originally joined the laboratory, a graduate student was just beginning to evaluate his MAP scaffolds in aged mouse models of stroke. With my research, I am continuing to explore how MAP may help stroke recovery in aged populations. But, more specifically, I am working on alternative MAP designs (such as the inclusion of extracellular vesicles derived from various types of neural cells) that can even better help aged populations recover following stroke – such as by targeting cellular senescence, one of the many difficulties cells face as they age.

If you’re a student who is interested in doing research at the intersection of neuroscience and biomaterials, please contact us and add in the message that you are interested in my project! #cara #fightingtheclock

Combining nanofiber vaccine and MAP scaffold technology to reduce inflammatory cytokines

From Pablo’s lab diary:

Did you know that putting MAP scaffolds into wounds causes elevated adaptive immune responses, which leads to enhanced regeneration of the skin? I learned about this in grad school, and it made me think – what if we could focus those immune responses to battle the inflammatory cytokines that drive chronic would healing? Chronic wounds that never heal are a big problem for a lot of people, so I’m working on modifying MAP scaffold particles with a nanofiber vaccine platform to see if I can reduce inflammatory cytokines and heal those wounds. My goal is to develop an active immunotherapy that can increase a special type of antibody – ones that are capable of downregulating dysregulated-cytokine activity at the wound site. Accomplishing this will not only avoid delayed healing in patients with underlying chronic conditions, but it will also prevent the recurrence of chronic wounds.

This collaborative project has been a challenging journey for sure, but we are on our way to exciting results! If you are a student interested in working at the intersection between hydrogels and active immunotherapy for enhanced tissue healing, contact us and add in the message that you are interested in my project! #pablo #mapfibers

Engineering Transplanted Organs for Site-Specific, Adaptive Immunomodulation

Straight from Sydney’s dissertation abstract:

Despite advances in surgical techniques and immunosuppressive therapy, long-term survival of transplanted organs and tissues remains limited by immune-mediated injury, chronic rejection, and the systemic toxicities of lifelong immunosuppression. Standard therapies, while effective at reducing early acute rejection, fail to provide spatially confined immune modulation, leading to off-target toxicity and compromised graft longevity. This dissertation addresses these challenges by developing and translating strategies for localized, graft-directed immunomodulation that are modular, organ-agnostic, and mechanistically grounded.

The first component of this work establishes ORGANIC (Orthogonal Reactive Group Addition for Narrowed Immunosuppressant Capture), a chemical engineering platform that transforms donor organs into programmable, reloadable drug-capture interfaces. Donor tissues are functionalized ex vivo with bioorthogonal azide handles that persist in vivo for at least five weeks and retain the capacity to selectively capture cyclooctyne-modified therapeutics following implantation. Serial administration of click-modified probes resulted in dose-dependent signal enhancement, demonstrating the feasibility of repeated, adaptive targeting and enabling localized drug accumulation at sub-therapeutic systemic exposures. Using both murine and porcine models, we optimized perfusion protocols to achieve uniform azide installation across vascularized grafts, confirmed macromolecular capture at both protein and tissue levels, and demonstrated preservation of structural and functional integrity under clinically relevant conditions. Importantly, translation to porcine-scale organs not only established a viable clinical pipeline but also enabled higher-resolution interrogation of tissue modification due to increased anatomic scale and technical control. Collectively, these findings establish ORGANIC as a modular platform that decouples targeting specificity from endogenous receptor expression and enables post-transplant, organ-directed therapeutic control.

Building on these principles, STEALTH (Surface-Tethered Engineered Alloprotective Ligands for Transplant Health) applies a two-step bioorthogonal chemistry strategy to immobilize immunomodulatory ligands, such as PD-L1, directly on graft surfaces. In fully allogeneic rat kidney transplantation models, surface-tethered PD-L1 significantly improved renal function and extended graft survival without systemic immunosuppression. Mechanistically, PD-L1 attenuated immune-mediated injury while reshaping the graft immune microenvironment, promoting the emergence of regulatory T cell–rich niches consistent with regulatory tertiary lymphoid structures. Notably, preserved graft function occurred despite dense lymphocytic infiltration, demonstrating a decoupling of histologic inflammation from functional rejection and underscoring the need for integrated clinical, cellular, and spatial analyses. Multimodal interrogation, including TUNEL staining and multiplexed immunofluorescence (CODEX), revealed reduced cell death and spatially organized immune populations, supporting a protective role for PD-L1 beyond canonical T cell inhibition. Segmental analysis further suggested that PD-L1 confers protection against ischemia-reperfusion injury, indicating a non-canonical cytoprotective role for this co-inhibitory ligand. In addition to regulatory T cells, CD56⁺ cell populations localized to Bowman’s capsule were enriched, consistent with emerging descriptions of renal regenerative niches. Together, these findings demonstrate that localized presentation of inhibitory ligands can reprogram the graft-immune interface and preserve function independent of traditional markers of rejection.

Translation of STEALTH to porcine kidneys focused on clinical feasibility and scalability, including the development of gravity-driven perfusion protocols compatible with standard transplant workflows, optimization of preservation solution exchange to maintain conjugation efficiency, and validation of uniform cortical ligand distribution. Autotransplantation of STEALTH-modified grafts demonstrated immediate reperfusion, recovery of urine output, and absence of off-target toxicity. In parallel, we established a stringent allogeneic porcine transplant model (SLA-mismatched Yucatan-to-Yorkshire) characterized by rapid and reproducible rejection within one week, providing a clinically relevant platform for therapeutic testing. Ongoing studies are leveraging this model to define the effective dosing and durability of STEALTH-mediated protection in large animals. Together, these results demonstrate that STEALTH has the potential for effective, organ-specific immune modulation across scales, from mechanistic rodent models to clinically relevant large-animal systems.

Complementing these translational strategies, we turned to reductionist murine models to interrogate the fundamental principles governing graft-directed immune modulation. Building on the historical role of skin transplantation as a foundational model in transplant immunology, we leveraged fully allogeneic murine skin grafts in C57BL/6 recipients as a tractable platform to study how biomaterials and defined antigens shape the local immune microenvironment and enable therapeutic discovery. To establish a biomaterial framework, we compared conventional wound matrices, including Woun’dress and bulk hydrogels, with microporous annealed particle (MAP) scaffolds, enabling direct evaluation of how material architecture and chemistry influence alloimmune responses. Drawing on prior observations of chirality-dependent immune engagement in wound healing, we engineered MAP hydrogels composed of L- and D-peptide microgels and demonstrated that stereochemistry directs early antigen-presenting cell activation, lymph node trafficking, and downstream T cell polarization in the transplant setting. A racemic formulation (R-MAP) attenuated co-stimulatory signaling and reduced donor-specific antibody formation, achieving durable modulation of humoral alloimmunity beyond systemic CTLA4-Ig therapy and highlighting chirality as a tunable parameter for local immune control.

In parallel, we evaluated a complementary reporter-based system using GFP and luciferase in the NoGlow mouse model to enable longitudinal, non-invasive tracking of graft viability and highlighted the utility of reporter-tolerized mice for studies in transplantation, extending this model from oncologic studies to transplant. While NoGlow mice were engineered to mitigate xenoantigen-driven rejection of these reporters, we also leveraged the inherent immunogenicity of GFP and luciferase in non-tolerant wild-type NoGlow- settings to model antigen-specific immune responses. In this context, GFP and luciferase functioned dually as quantitative readouts of graft survival and as defined xenoantigens, enabling high-resolution profiling of rejection kinetics where NoGlow recipients served as tolerized controls against which to benchmark rejection prevention. This platform was applied to accelerate screening of localized biomaterials and systemic therapeutics as immunosuppressive strategies, including comparative studies with Woun’Dress, bulk hydrogels, and granular materials as well as combination therapy with belatacept and rapamycin, where reporter signal dynamics revealed differential effects on graft viability and function that were not captured by endpoint analysis alone. By integrating real-time imaging with terminal histologic and immunologic assessment, this system enabled rapid, high-throughput comparison of biomaterials and therapeutic regimens, directly informing the design of protective, locally acting interventions. Together, these studies establish murine skin transplantation as a powerful discovery platform and identify biomaterial stereochemistry and real-time, antigen-specific tracking as synergistic tools for engineering spatially confined immunomodulatory strategies.

Collectively, this dissertation integrates ex vivo organ perfusion, bioorthogonal chemistry, biomaterial engineering, longitudinal imaging, and multi-scale transplantation models to systematically develop localized immunomodulatory strategies. The findings demonstrate that engineered chemical and biomaterial interfaces can: (1) transform organs and tissues into programmable therapeutic platforms, (2) preserve graft function and structural integrity, and (3) modulate both innate and adaptive immune responses in vivo. By bridging mechanistic discovery with translational implementation, this work establishes a foundational framework for organ- and tissue-directed immunotherapy and provides a path toward improving transplant outcomes while minimizing systemic immunosuppression.

Read the full dissertation here: TBD

Generate images & GIFs

Part of LOVAMAP’s power is its ability to offer visual insight into all that is void space. Visualization is an important aspect of science communication, and we put a lot of effort into making beautiful images and videos that help the scientific community see what we’re talking about. Below we’re highlighting a single 3-D pore that represents a pocket of empty space surrounded by particles.

Segment void space

Our goal is to isolate all of the distinct pockets of empty space throughout the scaffold – seems like the obvious way to break up the space, right? But doing this accurately is so much more complicated than you might think. LOVAMAP uses the spatial landmarks as a basis for segmenting the void space into these natural open pockets, which we called ‘3-D pores.’ Below you can get a taste for how crazy and diverse pore shapes can be!

Advancing Wound Healing: from Surgical Technology to New and Improved Hydrogel Therapies Engineering Injectable Glycan-Functionalized Biomaterials to Enhance Neuroimmune Regeneration after Ischemic Stroke

Straight from Yunxin’s dissertation abstract:

In the injured brain, shifts in sialylation dysregulate Siglec-mediated neuroinflammation and NCAM-dependent neurogenesis and plasticity, yet most biomaterial strategies rarely utilize glycans as bioactive cues in neural repair¹. Meanwhile, injectable microporous annealed particle (MAP) hydrogels offer a permissive, minimally invasive architecture^2,3 yet are usually combined with labile biologics^2,3 and seldom exploit glycan-specific signaling. This dissertation addresses that combined gap by engineering and testing a polysialic acid (PSA)–functionalized MAP scaffold for post-stroke regeneration. We covalently conjugated PSA to hyaluronic acid (HA) microparticles using end-selective, bioorthogonal chemistry to control ligand orientation and density without altering scaffold mechanics. We implemented rigorous quality controls including sialic acid derivative selection, endotoxin removal, conjugation efficiency, and enzymatic stability.

In vitro, tethered PSA shifted neural progenitor cells (NPCs) from a Sox2-high transitional state toward early neuronal commitment over one week, with increased Tuj1 and decreased Sox2 relative to RGD-only MAP controls. In parallel, primary macrophage phenotyping indicated reprogramming toward a more regulatory phenotype, characterized by reduced iNOS, increased MHCII and CD86, and altered cytokine secretions, with decreased TNF-α and increased IL-10. In a photothrombotic mouse model, PSA-MAP was delivered during the subacute window. At day 7 (early subacute), flow cytometry showed fewer iNOS-positive and more Arg1-positive cells in both macrophages and microglia, indicating early immunomodulation. At day 15 (later subacute), histological profiling indicated a myeloid niche shifted toward resident microglia with a lower inflammatory tone. We also observed early neural repair with increased Sox2 progenitor presence and distribution, enhanced NF200 axonal signal and penetration into the lesion without concomitant astroglial and vascular increase at day 15. Collectively, the findings show that localized, tethered PSA presentation modulates detrimental innate immune activation and establishes conditions that support early neural ingress and axonal entry while maintaining minimal cargo complexity. This work establishes a glycan-centric approach that couples MAP architecture with active carbohydrate cues toward translational neuroimmune regeneration after ischemic stroke.

Read the full dissertation here: TBD

Bioengineering Strategies to Modulate Inflammation, Synaptogenesis, and Extracellular Vesicle Production for Ischemic Stroke Repair

Straight from Nhi’s dissertation abstract:

Ischemic stroke remains a leading cause of adult disability, with acute therapeutic strategies such as intravenous thrombolysis and mechanical thrombectomy largely unchanged since 2015. Beyond these interventions, functional recovery relies primarily on physical rehabilitation, and more than half of stroke survivors aged 65 or older live with long-term mobility impairment. This work develops and characterizes astrocyte-inspired biomaterials and biophysical technologies that engage the endogenous window of neural plasticity during the subacute phase following stroke to promote neural network rewiring and enhance the translational potential of bioactive therapeutics.

The first study investigated the effect of scaffold chemistry on post-stroke tissue response using dibenzocyclooctyne (DBCO)-modified microporous annealed particle (MAP) hydrogels. DBCO functionalization inhibited astrocyte migration but, at low modification levels, significantly enhanced axonal elongation, identifying DBCO as a bioactive cue capable of modulating lipid deposition and regenerative outcomes. The second study employed MAP scaffolds to co-deliver synaptogenic thrombospondin-1 (TSP-1) and angiogenic vascular endothelial growth factor (VEGF) nanoparticles into the infarct cavity. This treatment promoted synaptogenesis without disrupting angiogenesis, though excessive synapse formation constrained axonal extension, leading to modest behavioral recovery. Finally, an acoustofluidic stimulation platform was developed to improve the yield of astrocyte-derived extracellular vesicles (EVs). This device increased EV secretion up to eight-fold across reactive astrocyte states, providing a scalable biomanufacturing tool and a potential diagnostic platform for assessing astrocyte mechanoreactivity. Collectively, these studies advance the development and design of technologies which promote post-ischemic neural circuit reorganization and functional recovery.

Read the full dissertation here: TBD

Investigating the Impact of Biomaterial Chirality on Host-Material Interactions

Straight from Elle’s dissertation abstract:

Wound healing is a complex process that often culminates in scar tissue rather than functional regeneration, representing a major clinical challenge. Biomaterials provide a means to reshape these mechanistic pathways to improve healing outcomes beyond closure rates alone. Microporous annealed particle (MAP) scaffolds are particularly advantageous in the wound healing space due to their modularity and superior tissue integration compared to traditional nanoporous systems. This work characterizes the immune engagement underlying the regenerative capacity of D-chiral MAP (DMAP) scaffolds and directly compares it to an L-chiral (LMAP) scaffold with equivalent mechanical and chemical properties.

We first examined T cell populations implicated in hair follicle neogenesis and T cell-dependent humoral responses. Contrary to our initial hypothesis, these T cell populations were neither recruited nor expanded in the DMAP-treated mice. To capture the broader microenvironmental landscape, we investigated DMAP using a time-course proteomic analysis. DMAP promoted a broad enrichment of immune-related pathways at day 4, followed by enrichment of keratinization and lipid-associated pathways by days 14 and 21. As DMAP-mediated regeneration was previously shown to depend on adaptive immunity, we next assessed the role of infiltrating and resident lymphocytes in the regenerative mechanism. Regenerative repair persisted even when lymphocyte infiltration was restricted, indicating that infiltrating adaptive immune cells are not required. Instead, DMAP activated B cells in the skin draining lymph nodes, driving germinal center formation, the production of biomaterial-specific antibodies, and pro-regenerative serum factors. Remarkably, this circulating milieu was sufficient to confer regenerative healing even in wounds treated with LMAP. Finally, we investigated dendritic cell (DC) involvement in the repair mechanism through both comprehensive wound flow profiling and simplified in vitro assays. DMAP supported the accumulation of DCs that were activated but non-inflammatory. In vitro culture of bone marrow-derived DCs demonstrated direct maturation only in MAP scaffolds with the D-amino acid peptide in a crosslinker format. Taken together, these findings demonstrate that an early immune response, coordinated humoral immunity, and key innate-adaptive linking cells are central to DMAP’s function as a regenerative biomaterial.

Read the full dissertation here: TBD

Regenerating skin by reducing stress… on cells

From Aleja’s lab diary:

After I joined the lab, I read the work of a previous postdoc in the lab – Don Griffin – and learned how MAP scaffolds promote tissue regeneration in skin wound healing. Two questions stuck in my brain: How does MAP work to regenerate skin? And how can I improve our MAP technology? Prof. Segura and I had a lot of fun conversations trying to find the perfect focus for my project, and I remember the first time she pitched this great idea: maybe the unique mechanical properties of our granular scaffolds are the key to MAP’s superior ability to regenerate tissue!

We hypothesized that our microparticles may be dissipating the energy exerted by cells, which in turn influences cell behavior. Prof. Segura suggested studying fibroblasts since these cells are involved in skin wound healing and are very susceptible to mechanical cues. Once fibroblasts receive mechanical (and chemical) cues, they ‘activate’ into another cell type called myofibroblasts, which secrete collagen and other components of the extracellular matrix as well as contract the wound. This process is a feedback loop known as the fibroblast-myofibroblast transition, and it controls the balance between regeneration and scarring. My project is to study this process and investigate whether our material can modulate the communication between fibroblasts and myofibroblasts.

But if you’re interested in working on a skin project that studies my favorite cells, contact us and add in the message that you’re interested in my project! #alejandra #reducingstress

Engineering biomaterials, culturing tolerance

From Holly’s lab diary:

Prior to joining the Segura Lab, my world was in the operating room. As a surgery resident at Duke, I’ve performed many procedures, and as you know, surgery requires cutting through the patient’s healthy tissues to gain access to internal structures. After we close them up, the wound healing process begins and usually a scar forms – but in some unfortunate cases, the wound never properly heals and takes much longer to close. We are trying to understand the molecular signals that lead to healthy wound healing and a return to normal function.

Before I was a surgeon, I spent my PhD investigating the roles of adult stem cells as well as their anti-inflammatory properties, and I later learned that biomaterials could be designed to mimic the pro-healing actions of stem cells. Since I was drawn to transplantation surgery during residency, I became interested in developing a biomaterials-based approach for improving the immune response to soft tissue transplants – and this brought me to the Segura lab. When I first approached Prof. Segura, I was struck by her welcoming attitude toward new applications of her MAP hydrogel technology. Although she was not familiar with the challenges of soft tissue transplantation, I explained what I wanted to do, and she was immediately on board. The plan is to use MAP scaffolds to promote immune tolerance. As you may have already read, MAP scaffolds promote blood and lymphatic vessel formation in healing tissues, both of which are critical processes in reconstructive and transplant surgeries like skin grafting and kidney transplantation, respectively.

Skin grafting (my current topic) is a cornerstone of reconstructive surgery. Over 160,000 skin grafts are performed annually for patients after burns, trauma or vascular disease (Serebrakian, et al. Plast Reconstr Surg Glob Open, 2018). Despite various FDA-approved dressings, up to half of skin grafts are rejected. Repeated skin grafting requires multiple surgeries, which increases risk of morbidity and mortality. Although a variety of cadaveric and synthetic alternatives exist, they are limited by allogeneic responses, dysregulated inflammatory reactions, worse scarring, poor functional outcome, and just overall lower quality of life. My research aims to improve the outcome of soft tissue reconstruction through the development of hydrogel biomaterials that can coordinate the interactions of immune cells to improve tolerance in the wound bed of allotransplantation.

Generate images and GIFs

Coming soon!

Return descriptor data

LOVAMAP does some powerful computing to generate an ever-growing list of descriptors that quantitatively characterize the MAP scaffold. These measurements describe features of the void space between the particles, the pathways throughout the scaffold, integrin-binding hotspots, and more. By quantifying the microarchitecture of our scaffolds, researchers can study how scaffold geometry affects other measurable properties, like infiltrating cell behavior and material mechanics. Ultimately, these types of analyses will help us optimize our material for the best possible therapeutic effect.

3-D pore descriptors

3-D pores (or pockets of open void space) form such weird shapes!! How do you describe something that looks so weird? If someone asked you to come up with different ways to measure a cube, that’s easy – you could measure length, volume, corner angles, etc. 🥱 But for these 3-D pores, it’s more complicated. So we came up with 30 different measurements to describe them, like reporting their volume or the number of surrounding particles that form the space. We also distinguish between 3-D pores that are completely inside of the scaffold vs. those that are touching the edge (like the massive purple one below).

Non-3D pore descriptors

LOVAMAP analyzes more than just 3-D pores. For example, we study paths that course throughout the void space of a scaffold. (You can see a distinct path highlighted in red below.) The measurements taken for a path give rise to some of our non-3D pore descriptors.

Global descriptors

LOVAMAP computes 20 global descriptors for each scaffold. ‘Global’ refers to the fact that a single value is reported for the descriptor, which captures information spanning the entire scaffold. For example, the most common global descriptor reported in the literature for granular biomaterials is Void-Volume Fraction, where the top of the fraction is the volume of scaffold that’s occupied by empty space and the bottom of the fraction is the total volume of the scaffold.

Microporous annealed particles (MAP) scaffolds

The magic of MAP comes from interlinking the HMPs in a granular scaffold so that the particles get stuck in place. There are many, many different techniques for doing this, but the very first MAP scaffold was designed to naturally crosslink in the body. This was great because it meant we could load a syringe with HMPs, inject into the body to form a granular scaffold, then wait for the body’s natural clotting factor to chemically link the particles together. Since then, we’ve also become fond of interlinking using UV light. The benefit of interlinking (or annealing) HMPs is that we create a stable structure that sticks around for longer and makes cells extra happy. We’ve seen MAP scaffolds promote aspects of wound healing in disease models like stroke, myocardial infarction, skin wounds, and spinal cord injury. And once the MAP scaffold has completed its job, it naturally gets degraded by the body.

Extract spatial landmarks

LOVAMAP relies on accurately identifying key spatial landmarks within the void space of MAP scaffolds. These landmarks are subtypes of the medial axis, which are central locations in space, and they serve as a starting point to orient LOVAMAP to the space. Our custom algorithms are highly effective at locating these landmarks because we take advantage of information stored in the graphical representation of granular material. Once the information has been stored, we’re ready to start dissecting and analyzing the MAP scaffold.

Non-invasive therapeutics

We’re motived to develop non-invasive treatment options so that patients can avoid invasive surgical procedures whenever possible. Although surgery is oftentimes the best and only option for treating a condition, the process introduces secondary surgical wounds that both increase the risk for bacterial infections and also require additional energy and resources to heal. Technological advances are tending toward minimizing how much cutting has to be done, but even in the field of biomaterials, surgical procedures are often required to implant therapeutic materials. This is because many products on the market are bulk materials, such as sponges or mesh sheets, that have to be cut to the appropriate shape and physically placed into the wound site.

Our lab has been focused on a different approach – generating materials that are flowable so that they can be injected via needle or catheter to the target location without the need for extensive surgery. Even for sites that are easily accessible, such as skin wounds, our flowable materials offer a benefit because upon injection, they naturally fill the shape of the wound for a seamless interface between the material and surrounding tissue – even if the tissue is moving!

Aside from injectable hydrogel materials, we are also working on intravenous (IV) delivery and transdermal delivery of nanoparticles. IV delivery is a very common route of administration for many therapeutics today, so it makes sense to use this approach when delivering nanosized materials. Transdermal delivery across the skin barrier removes the need for needles and enables home therapy using patches. Of course, the skin has evolved to be a tough barrier to penetrate, so we need to outsmart evolution and find a side door.

Wound healing

We are inspired by our body’s ability to heal. This occurs through a process called endogenous regeneration, which takes advantage of the activation of ‘plasticity’ upon injury. Plasticity is the ability to transform local cells into tissue factories that produce extracellular matrix and other multicellular structures that got destroyed. The goal of plasticity is to produce brand-spanking-new tissue. As you may have guessed, these pathways are usually dormant and only awaken once the body recognizes that an injury has occurred. So this begs the question – if tissues have the ability to fix themselves back to working order, why do some injuries cause permanent damage? This is because there are two competing forces in healing: plasticity and scarring. Scarring is a pro-survival mechanism that evolved to ensure that we survive an injury. Regenerating tissue takes time and resources – time that we may not have if the injury starts inhibiting important bodily functions or introducing deadly bacterial infections. Thus, at the same time that pro-regenerative pathways are activated to promote functional recovery, there are also pro-survival mechanisms activated that favor a quick-fix. Usually, scarring wins, and scar tissue gets deposited that is not completely functional.

Our main goals are to 1) scale-down scarring so that our tissues have a chance to regrow, and 2) deliver key signals that promote tissue regeneration. Luckily we know that our body actively works to repair itself after injury, so we only have to deliver the pro-healing components that are low or lacking in the wound environment. And the good news is that biomaterials can be designed to do just that – modulate the pathways that are responsible for scarring, while at the same time deliver key therapeutics to promote functional tissue deposition.

Heterogeneity

MAP is a collection of tiny particles, but nobody says we have to use the SAME particles throughout our scaffold. In fact, having the option to easily combine different types of particles into one scaffold is a big plus with our material. We can include particles of different shapes, sizes, and stiffnesses…made from different polymers…with different degradation rates…carrying different types of drugs… Really the world is our MAP-oyster! And not only can we precisely choose the proportion of these particle types, but we can also vary where in the scaffold they lie. Do you want everything to be mixed like rainbow Dippin’ Dots? Do you want particle types to be nicely layered like a cake? You choose. This is all to say, the heterogeneity offered by MAP allows for a highly personalized, versatile therapeutic material.

Injectability

In the same way that it would be difficult to fit a sponge into a needle, it’s difficult for a non-granular matrix to be injected. And if something can’t be injected, then getting it to the right place might require some surgery. Luckily MAP is granular, so all of the tiny particles can fit inside a syringe. And – thanks to the concept of particle jamming – after injection, the particles reform into a scaffold that perfectly conforms to the shape of the wound. Once the material has been deposited, the last step is to perform some chemistry magic to interlink the particles so they don’t go anywhere. The injectability of MAP makes it much more convenient for physician’s to use, and patients can avoid undergoing unnecessary surgery – everyone wins!

Outer porosity

Let’s first cover what MAP is not – MAP is not a solid hunk of material, like Jello. Imagine a cell approaching a giant wall of Jello. It would have to eat away at the material in order to get inside and hang out. We want to make our materials inviting to cells so that there’s minimal effort for them to get right in and start healing that wound. MAP is genius in that way because it’s made of a bunch of microscopic particles of Jello packed together, and the space between the particles is also on the micron-scale. Do you know what else is on the micron-scale? Cells. So with MAP, the micron-sized openings (or ‘pores) allow cells to easily crawl into the scaffold without wasting time and energy degrading the material first. This process of cell movement is called ‘cell infiltration,’ and the structure of MAP makes for some effortless cell infiltration. Do you see those brain cells stained in red making their way into the MAP stained in green?

Inner porosity

The picture below really says it all! MAP is designed so that the space between the particles is large enough to fit cells. Back in the day, researchers would create materials that resemble… Jello swiss cheese 🤔 The idea was to create a material that’s filled with interconnected holes (or ‘pores), which would allow cells to move around freely. The structure of MAP is essentially the opposite of that! The holes are now the particles, and the cheese(?) is now empty void space. Cells can still move around freely, and in fact, they seem to prefer the concave surface of the particles. We want to create materials that keep cells happy and push them toward healing wounds, so this is good news.

Hydrogel microparticles (HMPs)

The first step is to design the particles that compose our MAP material. Each particle is actually a network of entangled polymers (like a microscopic hairball), and we typically work with hydrogel polymers that swell in water – hence the term HMPs (also referred to as µgels). The process starts with some good ‘ole polymer chemistry, where we customize our material to fit the therapeutic application. Once the polymers are just right, we form them into the desired shape, then lock the shape into place through a process called polymerization. Most of our MAP experiments use spherical HMPs produced through emulsion or microfluidic techniques, but we’ve been experimenting with other shapes as well. (I know we just dropped some technical terms, so if you’re unfamiliar and want to learn more about HMPs, click here, we won’t tell anyone 🤫)

Read in formatted data

LOVAMAP is a software designed for analyzing granular material. For this reason, LOVAMAP takes in a specific data format that highlights individual particles – basically each voxel (3D pixel) associated with a particular particle should be labeled with a unique identifying number. This ensures that our software is analyzing exactly what you intended. Alternatively, if you’re simulating spherical particles with a known center and radius, go ahead and input the sphere data – our code can handle it.

Ninjabyte Computing

If you’ve ever watched a Marvel movie or played a video game like Witcher 3, you know graphics have come a long way. But you may not know how much serious math and research goes into 3D animation. Technologies in simulation and visual effects are incredibly powerful, yet most scientists don’t have access to them because software engineers in animation don’t often chill with scientists. Ninjabyte Computing was founded to connect the science community with high-performance animation tools used in industry.

The company is headed by Peter Cheng, PhD, who offers the perfect blend of expertise in mathematics, computer science, and animation. Ninjabyte Computing uses professional visual effects software, like SideFX Houdini, to generate physically-accurate simulations of randomly packed particles with varying shapes, sizes, stiffness, and textures. These simulations are fantastic mimics of a range of MAP scaffold types, which makes the data perfect to analyze with LOVAMAP in order to study our scaffolds. Ninjabyte Computing also renders beautiful images and videos of our material to help explain our work. One of their images was featured on the cover of Nature Materials!

While our collaboration focuses on MAP simulations, this only scratches the surface of animation technology. We’re happy to share the love for Ninjabyte, and we recommend reaching out to ninjabytecomputing@gmail.com for any inquiries.

Tweaking the immune system

The goal with this approach is to design materials that communicate with our body’s immune system and urge it to generate a lasting pro-regenerative response in the wound. The immune system is most commonly known for helping us fight disease. When external objects enter our body, specialized cells will recognize them as foreign and trigger a specific immune response depending on the type of invader. Viruses, for example, often trigger a danger! danger! response that causes the immune system to produce antibodies against the virus for long-lasting defense. This means you’re protected from getting sick if you get infected with the same virus again. Materials made in lab, however, don’t always elicit the same immune response, which means our materials can be explicitly engineered to tweak the immune system in a certain direction.

It turns out that in addition to fighting foreign invaders, immune cells are also primary players in healing the body. Inflammation, for example, is an early-stage part of wound healing that relies on the hustle and bustle of immune cells. As you probably know, too much inflammation can be a bad thing that actually delays healing. This is why RICE (Rest, Ice, Compression, Elevation) is used to treat twisted ankles and sprained knees. RICE helps keep inflammation down and is often accompanied by the administration of anti-inflammatory drugs such as ibuprofen, which helps our body heal faster. I’m sure it comes as no surprise that modulating inflammation is one of the key features of our materials. In fact, it has recently been discovered that the same cells involved in creating immunity against viruses could be used to promote healing, so we’re learning how our materials can transition from controlling inflammation to generating long-lasting immune responses.

Do these lipids make my brain look fat?

From Briana’s lab diary:

I’m a chemist, and I love the art of synthesis – creating new polymers from scratch with unique designs. I also have experience making nanoparticles, so Prof. Segura and I discussed using nanoparticles as drug carriers that would target the brain after stroke. Before joining the lab as a postdoc, I worked on developing dendritic polymers that have both a hydrophobic (‘fat-loving’) and a hydrophilic (‘water-loving’) part, so I’m incorporating this chemistry into my current project. By using these amphiphilic polymers to form nanocapsules, we could not only co-deliver both hydrophobic and hydrophilic drugs, but we could also leverage the lipid (fat) content of the nanocapsules to cross the blood-brain barrier. In this way, our material could reach the brain through IV injection into the arterial system as opposed to direct injection into the brain, making it even less invasive.

After talking with Prof. Segura, I also learned that the brain is 50% lipids by weight. Lipids do much more than make us fat – they’re active players in signaling and normal tissue function. I’ll be reading and learning about lipid biology, especially in the context of stroke, so that I can guide my synthesis to include brain-active materials. By using self-assembling lipids, I aim to make nanoparticles that are entirely bioactive.

Angiogenic Granular Biomaterials for Brain Repair after Ischemic Stroke

Straight from Kevin’s dissertation abstract:

There is still a significant need to develop regenerative therapies that improves functional recovery for stroke patients that develop disabilities. Not only does an ideal therapy stimulate tissue reorganization for positive indications of regeneration, it must also produce functional recovery as that has the largest quality of life impact on patients with stroke-related disabilities. As such, we have established a consistent photothrombotic stroke model and behavioral assessment workflow for our studies. Patients, and stroke animal models, undergo stroke-induced enhanced endogenous plasticity and pro-repair mechanisms in the subacute phase after stroke onset, making it an attractive opportunity for therapeutic intervention.

We have designed a biocompatible granular hydrogel, microporous annealed particle (MAP) scaffold to be injected with our angiogenic clustered VEGF heparin nanoparticle (CLUVENA) into the stroke infarct 5 days after photothrombotic stroke. Our combined MAP-CLUVENA was able to attenuate astrogliosis and promote significant angiogenesis with perfused vessels growing throughout the infarct as early as day 15. At day 35, we observed signs of vessel maturation and axonal sprouting that led to functional recovery. Taking the next step in translation by improving clinical relevance, we repeated the study in an aged female mouse model and showed MAP-CLUVENA still maintains therapeutic efficacy in attenuating astrogliosis and enhancing angiogenesis throughout the infarct. Although numerous preclinical studies still need to be done, our engineered MAP-CLUVENA hydrogel is a promising intervention for subacute ischemic stroke clinical translation.

Read the full dissertation here: Angiogenic Granular Biomaterials for Brain Repair after Ischemic Stroke

Glycobiology of stroke

From Juhi’s lab diary:

Did you know that our body uses sugars to regulate its activity? In the brain, sugar molecules are everywhere – they’re attached to biomolecules and covering all of the cells. Sugars, which are called glycans in our world, can help cells communicate with each other and with their environment. And cells will display different glycans depending on their current state. As you may imagine, the glycans that get added and displayed during disease or injury are different to those that get displayed during a healthy state.

I am interested in understanding how glycosylation changes with aging and after a stroke, and I want to understand if part of the success of our previous materials are because of changes in the glycosylation profile. My ultimate goal is to design novel glycan-decorated materials to promote recovery after stroke.

If you’re a student who agrees that glycans > proteins and nucleic acids and you’re interested in their impact on stroke recovery, contact us and add in the message that you’re interested in my project! #juhi #sweetbrain

Trapping the pro-repair environment

From Shangjing’s lab diary:

I’m currently a postdoc in the lab, and my previous PhD project revolved around designing biomaterials (actually MAP gels) for bone tissue engineering. I delivered stem cells from the material to promote bone formation. I joined the Segura lab because I wanted to learn from the creators of MAP scaffolds and challenge myself to design materials for neural applications, which is completely new to me. The project Prof. Segura proposed sounded really interesting, and with some reading, we have a really cool approach.

The fact that brain injury may actually help you learn new things fascinates me. This occurs through a process called plasticity. When we are first born, our brain is very plastic, and we are able to learn many new things. However, as we age, our brains are less plastic, meaning neurons in the brain aren’t able to form as many new pathways and connections. On the one hand, having our brains settle on stable connections helps us become good at things, but unfortunately, it also hinders us from picking up new things easily. It turns out that after injury, our body turns on plasticity as a way to heal the injury.

My project is to leverage this plasticity window with a ‘repair soup’ to promote healing after stroke. In particular, I am working on strategies that would trap locally-secreted plasticity-proteins into our biomaterial in an effort to extend the window of cellular change. The end goal of course is to reduce physical impairment caused by stroke.

Hydrogel-Mediated Gene Delivery from Granular Scaffolds for Applications in Biologics Manufacturing and Regenerative Medicine

Straight from Evan’s dissertation abstract:

Nucleic acid delivery has applications ranging from tissue engineering to biologics development and manufacturing to vaccines and infectious disease. To improve delivery and extend viable expression over time, we turn to biomaterials as a method for sustained nucleic acid release and enhanced cell culture or tissue interaction. Here, we describe how cationic polymer and lipid condensed nucleic acids can be effectively loaded into injectable granular hydrogel scaffolds by stabilizing the condensed nucleic acid into a lyophilized powder, loading the powder into a bulk hydrogel, and then fragmenting the gel into hydrogel microparticles. The resulting microgels contain non-aggregated nucleic acid particles, can be annealed into an injectable microporous scaffold, and can effectively deliver nucleic acids to cells with a sustained rate of expression.

We explore how this technology can improve the production of biologics, like antibodies and viruses, to overcome limitations of current batch processes. Our scaffolds allow for continuous biologics manufacturing, with sustained production upwards of 30 days. We also explore how our platform can improve tissue regeneration in disease models like dermal wounds by delivering nucleic acid drugs, namely DNA, mRNA, and therapeutic viruses. The loaded granular scaffolds allow cells to readily repopulate the missing tissue and drugs be locally released and taken up over time. Overall, our scaffold delivery approach is a customizable platform that can be tuned for many different applications.

Read the full dissertation here: Hydrogel-Mediated Gene Delivery from Granular Scaffolds for Applications in Biologics Manufacturing and Regenerative Medicine

Modulating Macrophage Response with Microporous Annealed Particle Scaffolds

Straight from Yining’s dissertation abstract:

When designing biomaterials for clinical applications, the performance of these platforms hinges on their interaction with the host immune system. A failure in engaging and incorporating the correct immune response would lead to a foreign body response and subsequent rejection of the materials. To improve the biocompatibility of biomaterials and avoid undesired immune reactions, the key immunomodulatory cell type macrophage needs to be engaged and its phenotype modulated properly and in a timely manner. Therefore, the design parameters of biomaterials should be carefully considered in the context of macrophage modulation. Microporous annealed particle scaffolds (MAPS) are a new class of immunomodulatory granular materials generated through the interlinking of microgels. The modular nature of MAPS offers enormous tunability in not only the individual microgel design but also the homogenous or heterogenous microgel assembly into the bulk scaffold. We leveraged the plug-and-play feature of MAPS to study the effect of two design parameters, microgel crosslinking peptide (comprised of L- or D-amino acids) and spatial confinement (achieved through varying microgel size), on macrophage modulation and host responses.

We uncovered that a fine balance between pro-regenerative and pro-inflammatory macrophage phenotypes in MAPS with D-amino acid-based crosslinker was an indicator for regenerative scaffolds in a subcutaneous implantation model. We also discovered that scaffolds comprised of large microgels with pore sizes that can accommodate ~40 µm diameter spheres induced a more balanced pro-regenerative macrophage response and better wound healing outcomes with more mature collagen regeneration and reduced inflammation level. The role of spatial confinement on macrophage response was further explored in vitro, where we demonstrated that size-dependent macrophage response to M1/M2 cytokine stimulations was tied to the change in cell morphology and motility. This work offers valuable insights into the dynamic immune response to synthetic porous scaffolds with a specific focus on macrophages and establishes a foundation for further optimization of immunomodulatory pro-regenerative outcomes for wound healing and biomaterial implants.

Read the full dissertation here: Modulating Macrophage Response with Microporous Annealed Particle Scaffolds

Engineering the microstructure and spatial bioactivity of granular biomaterials to guide vascular patterning

Straight from Alex’s dissertation abstract:

In tissues where the vasculature is either lacking or abnormal, biomaterial interventions can be designed to induce vessel formation and promote tissue repair. The porous architecture of biomaterials plays a key role in influencing cell infiltration and inducing vascularization by enabling the diffusion of nutrients and providing structural avenues for vessel ingrowth. Microporous annealed particle (MAP) scaffolds are a class of biomaterial that inherently possess a tunable, porous architecture. These materials are composed of small hydrogel particles, or microgels, that pack together to produce an interconnected, porous network.

We first demonstrated that the particle fraction in MAP scaffolds serves as a bioactive cue for cell growth. To control this bioactive cue, we developed methods to form MAP scaffolds with user-defined particle fractions to reproducibly assess mechanical properties, macromolecular diffusion, as and cell responses. We then modulated the microstructure of the MAP scaffolds by changing microgel size as well as the spatial bioactivity using heterogeneous microgel populations to promote de novo assembly of endothelial progenitor-like cells into vessel-like structures. Through a combination of in silico and in vitro experimentation, we found that the microstructure (dimension of the void), integrin binding, and growth factor sequestration were all shown to guide vascular morphogenesis. We then demonstrated that the findings produced in a reductionist model of vasculogenesis translated to an in vivo effect on vessel formation in both dermal wounds and glioblastoma tumors.

Read the full dissertation here: Engineering the microstructure and spatial bioactivity of granular biomaterials to guide vascular patterning

Advancing Wound Healing: from Surgical Technology to New and Improved Hydrogel Therapies

Straight from Drew’s dissertation abstract:

Wound healing is a vastly complicated process. While this can be said about many biological functions in the body, wounds present a particularly difficult problem due to their inherent irregularity or uniqueness. Because different wounds behave and heal differently, or not at all, different therapies must be developed to treat them effectively. The research presented here details several approaches to progress not only the entire field of wound healing research, but also focuses on hydrogel technology improvements. Using titanium 3D printing, cap-able splints were constructed to not only ease the surgical process but also enable efficient daily wound access for treatment administration or wound tracking over time without the need to completely undress and redress the wound. The titanium splints did prove effective for daily monitoring but did still require some surgical prowess.

To remove the need for surgical skills, an adhesive wound splint was developed by incorporating ethoxylated polyethyleneimine (EO-PEI) into the traditional polydimethylsiloxane (PDMS) polymer recipe resulting in adhesive PDMS (aPDMS). The aPDMS splints drastically reduced surgery time per animal without compromising wound splinting performance. Traditional bulk hydrogels have been used in wound healing research but have yet to be clinically implemented in a widespread manner due in part to their resistance to cellular infiltration or integration with the host. Using hyaluronidase (HAase) on a hyaluronic acid (HA) based hydrogels to partially degrade the surface of bulk gels yielded a looser nano-scale mesh size that enhanced cellular infiltration into the gel and granted better access to nanoparticle therapy loaded within. Finally, a biologically active viscous salve loaded with heavy chains (HC) of the serum protein Inter-α Inhibitor (IαI) was designed to leverage HC’s ability to mitigate the inflammatory response such that normal wound healing regeneration could ensue.

Read the full dissertation here: Advancing Wound Healing: from Surgical Technology to New and Improved Hydrogel Therapies

Bioengineering Microporous Annealed Particle Scaffolds to Recruit Neural Progenitor Stem Cells and Promote Angiogenesis in the Stroke Core

Straight from Kat’s dissertation abstract:

There remains a significant gap in the need for regenerative therapies for stroke compared to what is currently available. An ideal therapy would be one that stimulates the formation of new tissue with the ability to regain any function previously lost due to stroke. Therefore, methods exploiting the plasticity of the brain and modulating endogenous cellular responses to promote repair in the stroke core after ischemia have become highly attractive. However, this process of neural regeneration is complex and requires a series of controlled biological events, such as recruitment and differentiation of neuron progenitor cells (NPC’s), angiogenesis, and axonogenesis. Biomaterials are now commonly used to research tissue regeneration and cellular mechanisms, both in vitro and in vivo.

We have designed a biocompatible biomaterial from macroporous annealed particles (MAP) hydrogels for injection into the stroke core five days after a photothrombotic stroke. Our hyaluronic acid-based material has been modified to dictate NPC fate in vitro through maintained stemness and the formation of neurospheres or towards Tuj1 positive NPCs, as well as enhance angiogenesis and the recruitment of endogenous NPCs after stroke. Within only 10 days after injection, we have observed the first case of NPC differentiation, enhanced recruitment, and angiogenesis after stroke, and at 30 days after injection we observe significant angiogenesis throughout the entire stroke core.

Read the full dissertation here: Bioengineering Microporous Annealed Particle Scaffolds to Recruit Neural Progenitor Stem Cells and Promote Angiogenesis in the Stroke Core

Stem cell delivery

Tbd

We injected MAP into a skin wound, and you can see cells infiltrating between the particles.

Re-opening the window for cellular change

The most difficult wounds to treat are those that are chronic or well-past the phase of endogenous plasticity – when tissue is still being remodeled. For these wounds, we want to re-activate plasticity so that the body has another opportunity to heal itself properly. As you can imagine, this is harder to achieve than when wounds are still fresh in the acute or sub-acute phase. Our strategy is to first identify the molecules that are present in wounds during endogenous plasticity. We will then introduce materials that induce blood vessel formation as a source for replenishing the molecules that are present during plasticity. The goal is to determine the minimum molecular signals needed to re-open the window for cellular change.

Extending the window for cellular change

We’ve explored this approach the most. During the latter stages of wound healing, the microenvironment is in a state of endogenous plasticity, which means the tissue is still undergoing changes as it heals. The goal here is to place materials into wounds that extend the plastic time period by recruiting progenitor cells and other important factors. We primarily do this by designing materials that promote blood vessel formation. Blood vessels are a key component in tissue repair – they deliver important nutrients, regulate the immune response, and guide cell migration and tissue deposition. By maintaining and controlling the process of vessel formation with our materials, we’re thinking we can extend the window of plasticity and promote tissue regeneration. You can learn more about our approaches to promote vessel formation by reading the articles on Our Publications page that deal with therapeutic angiogenic material (filter by Topic: angiogenesis)!

Another way we’re extending endogenous plasticity is by designing materials that allow for rapid cellular infiltration while maintaining structural support. We do this by forming a microscopic scaffold made of packed hydrogel microparticles that are stuck together, which forms pockets of void space that are large enough for cells to fit. Alright, you guessed it – we’re describing MAP… you might as well read about it here.

Wound healing

Tbd

We injected MAP into a skin wound, and you can see cells infiltrating between the particles.

Granular scaffolds

The word ‘scaffold’ implies a supporting structure or framework, so at this step, we’re creating a framework using the HMPs as building blocks. The term ‘granular’ has a specific definition as well, but we’ll leave you to read this review for the nitty-gritty details. The take-home point is that granular materials transition from a ‘liquid-like’ state to a ‘solid-like’ state when particle jamming occurs – so to create a solid structure from HMPs, we pack them together. Now we have a granular scaffold with empty space in between the particles to support cell ingrowth. But remember, a granular scaffold is just a collection of particles jammed together, so it’s still a dynamic system where everything can slosh around.

Scarless healing

You could call this the Holy Grail of wound healing – everyone dreams of applying a topical cream that miraculously prevents scarring. It’s no wonder scar treatments are a $17.94 billion market [1]. But we’re motivated by more than cosmetic benefits. Many people hear ‘scar’ and think of visible scars that tell an interesting story. But scarring is a physiological process that can occur at any site of injured tissue, including internal organs. Think of scarring like the body filling up the injured site with cement. On the one hand, the cement (or scar tissue) has prevented further damage locally…but on the other hand, the region that contains the cement is no longer normal tissue, and it can’t function properly. Depending on the size and location of the scar, this can lead to compromised organ function and serious health issues.

The medical field is in need of a material that can prevent scarring and recapitulate* the native tissue environment. We are developing biomaterials that are intended to be placed in a wound prior to the formation of a scar. The goal is for our material to modulate the wound environment in a way that promotes normal tissue deposition while reducing scar formation. This means less cement and more healthy, functioning tissue.

[1] Grand View Research: Market Analysis Report. Scar Treatment Market Size, Share & Trends Analysis Report By Scar Type (Atrophic, Hypertrophic & Keloid Scars, Stretch Marks), By Product (Injectables, Topical, Laser), By End Use, And Segment Forecasts, 2019 – 2026.

* Fun fact: Lindsay in our lab was responsible for changing the official Merriam-Webster definition of the word ‘recapitulate’ to include the way scientists have been using it for years:

In vivo work

In vivo studies are performed in a living organism and are an essential and exciting part of the research we do. These types of experiments are invaluable because they not only teach us about biocompatibility, but we also get the full effect of a living tissue. This means we gain access to the complicated signaling pathways that would be impossible to recreate in a test tube, and therefore, we get a complete picture of the healing process. Understanding our in vivo results bring us one step closer to developing a product that can help real patients.

During an in vivo study above, we injected one of our biomaterial scaffolds into a controlled skin wound.

In vitro work

In vitro studies are performed in the lab outside of a living organism – think test tubes and petri dishes. These types of experiments allow us to characterize important properties of our scaffolds, like how much they deform or how well the molecules we’ve attached get released. It also offers us the luxury of studying the effects of our material on specific phenomenon, like cell behavior and vessel growth, in a controlled environment. It’s basically Step 1 in understanding what we’ve created.

Look how our gel squishes during compression testing to measure its stiffness!

Bells and whistles

We hate to be boring, so we’re always exploring new features to incorporate into our materials. Most of our scaffolds include additional bioactive signals that help make our material more effective at promoting healing, like:

peptides / proteins
nucleic acids
small molecules
cells

These additions can be used to attract and influence cells that make their way into the scaffold, or the material can be designed as a drug depot that releases factors at desired time points.

Above you can see DNA nanoparticles (highlighted in green) that have been loaded into a scaffold.

Biomaterial scaffold

A biomaterial scaffold is exactly what it sounds like – a structure made from building blocks that provides support. In this case, the building blocks we use are nanoscopically small because our scaffolds interact with tissue at the microscopic level. The scaffolds act as a supplemental extracellular matrix that provides physical support in compromised tissue, and they also serve as a framework to help nearby cells move around and do their job. You can see below that a typical scaffold looks like a little slab of clear Jello.

The scaffold itself can also behave as a drug and influence biological processes in the tissue surrounding it. This is possible because cells in the tissue are mechanoresponsive, which means that they can sense the structural features around them and respond to any changes. We can engineer the structural features of our scaffold to influence cell behavior, which leads to therapeutic action.

When we make scaffolds out of hydrogel microparticles and link these particles together, we create MAP – but you can read more about that here.

Poorly healing wounds

We all know that tearing your skin creates a wound. And even though skin wounds are a pain (literally), most heal right up – possibly with the help of some stitches. But the bigger and deeper the wound, the harder it is for the body to heal. What’s worse – if a patient has an underlying disease that stunts wound closure, like diabetes, wounds are more susceptible to chronic inflammation, infection, and poor healing. For some perspective, in 2014 there were 8.4 million claims for wound healing related diagnosis codes in Medicare patients alone [1]. Although there is a long history of therapies that have been developed to improve wound healing (check out reference [2] below), there is no universal approach to treat the ones that fail to close up. This means that despite the thousands of products available, many individuals continue to suffer from painful, long-lasting wounds that severely impact their quality of life.

The challenge of fixing a disrupted natural process in the body motivates us to develop a biomaterials-based approach that can break the inflammatory cycle and turn a non-healing environment into a healing one.

[1] Nussbaum SR, et al. An Economic Evaluation of the Impact, Cost, and Medicare Policy Implications of Chronic Nonhealing Wounds. Value in Health. (2018) 27 – 32.

[2] Han, G and Ceilley, R, 2017. Chronic wound healing: a review of current management and treatments. Advances in therapy, 34(3), 599 – 610.

Stroke-induced disability

A stroke is like a heart attack of the brain – it occurs when oxygen can’t get delivered to the organ. But unlike a heart attack, a stroke will leave a patient with substantially increased risk for lifelong disabilities [1]. In fact, stroke is one of the leading causes of serious long-term disability in the United States [2], and in 2018 alone, over 7.8 million individuals suffered from a stroke [3]. Sadly, although thrombolytic drugs are used to minimize damage within the first few hours of stroke onset, there are no therapeutic drugs available to treat damaged brain tissue after stroke. Patients must therefore rely on rehabilitation efforts like physical therapy and video-assisted training to reduce their disability and regain independence.

After a stroke, the brain tissue that was starved of oxygen is irreversibly injured, and the body attempts to repair the injury by forming a scar – but scar tissue is no longer functional. Patient disability is the direct result of non-functional scar tissue at the stroke site. Rehabilitation efforts attempt to recover function by training the brain to rebuild neuronal connections that bypass the stroke site. The brain is an adaptive organ, so figuring out new ways to form connections without the original tissue is challenging, but not always impossible.

Instead of working around scar tissue, we’re motivated to develop a material that can be placed directly into the stroke cavity to promote endogenous tissue regeneration, which is the body’s way of rebuilding functional tissue after injury. Theoretically, the body has all of the tools necessary to replace injured tissue with healthy tissue, and our material will offer physical and chemical cues to help guide this process in the stroke cavity. The end-goal is minimal scar tissue and maximal functional recovery.

[1] Dhamoon MS, et al. (2017) Disability Trajectories Before and After Stroke and Myocardial Infarction The Cardiovascular Health Study. JAMA Neurology. 74(12): 1439-1445.

[2] Centers for Disease Control and Prevention (CDC). (2009) Prevalence and most common causes of disability among adults: United States, 2005. MMWR Morb Mortal Wkly Rep. 58: 421–426.

[3] NCHS, National Health Interview Survey, 2018. Table A-1a: 1-9.

LOcal Void Analysis of MAP scaffolds (LOVAMAP)

Straight from Lindsay’s dissertation abstract:

Our lab designs hydrogel microparticles (HMPs) that are interlinked to form microporous annealed particle (MAP) scaffolds for wound healing applications. The therapeutic effects of MAP are attributed, in part, to the void space between particles that creates inherent micro-porosity through which cells can infiltrate and migrate unhindered. Cell behavior is influenced by local geometry, and our goal is to design scaffolds that influence cells toward pro-healing behaviors. To accomplish this, we need a methodology for quantitatively characterizing the void space of MAP scaffolds in order to study the relationships between internal microarchitecture and therapeutic outcomes.

The work presented here is a visually-rich dissertation that covers our approach for analyzing the void space of packed particles. We use techniques from computational geometry and graph theory to develop a robust methodology for segmenting the void space into natural pockets of open space and outputting a set of descriptors that characterize the space. Our methods are developed using simulated MAP scaffolds covering a range of particle compositions, including mixed particle sizes, stiffnesses, and shapes. Our software, called LOcal Void Analysis of MAP scaffolds (LOVAMAP), has allowed us to study many aspects of void space, including global descriptors like void volume fraction, local ‘pore’ measurements of size and shape, and additional features like ligand availability, paths, isotropy/anisotropy, and available regions for unhindered migration based on size. LOVAMAP is an enabling technology that can be used for analyzing real scaffolds or studying simulated scaffolds to inform material design. It serves as a platform for void space analysis that can easily be built upon to encompass ever-growing innovations in scaffold characterization.

Read the full dissertation here.

Modified building blocks

Our starting materials are hydrogel polymers, such as

hyaluronic acid
polyethylene glycol
poly(propylene sulfide)
heparin
poly(allyl mercaptan)

We chemically engineer these polymers to precisely fit our needs, like adding handles that allow the strands to link to one another and form a gel. Our polymer modifications include end-group modifications, side-chain modifications, and even attachment of biologically relevant ligands that make our materials more biocompatible and useful for therapeutic applications. We also use these polymers to create higher-level building blocks called hydrogel microparticles, which you can read more about on our MAP Scaffolds page.

Ischemic Stroke Project

Ischemic strokes are characterized by a sudden stop of blood flow to the brain due to a clot. The lack of blood flow rapidly kills brain tissue leaving > 40% of patients with a physical disability. There are currently no therapies to decrease stroke related disabilities. Although all current therapeutic treatments for stroke are administered immediately after stroke onset and focus on preventing tissue death, our goal is to develop materials to treat stroke related disability that can be administered days to months after the stroke occurs. Our materials are delivered intracranially, directly into the stroke core, to promote and encourage the brain plasticity needed to form new brain tissue and reduce disability.

Although their name would imply that NPCs become neural cells (e.g., nerves), I am interested in how these cells also play a role in blood vessel formation. There is a synergistic relationship between growing blood vessels in the brain and NPCs. Low blood vessel growth results in low NPC migration, but also NPCs secrete factors that are essential for blood vessel sprouting and maturation. My studies revolve around the use of MAP scaffolds as well as our VEGF delivery technology CLUVENA to promote NPC migration and vascularization. In addition, I am changing the physical and biochemical cues of the scaffold to modulate NPC phenotype and mature vessel formation.

If you’re interested in working on strategies to promote recovery after stroke focusing on NPCs and vascularization, contact us and add in the message that you’re interested in my project! #kat #brainheroes

VEGF – vascular endothelial growth factor
CLUVENA – Clustered VEGF Nanoparticle hydrogel (developed by our lab!)

Sept 2015 – June 2019

Elias Sideris

Clearview Healthcare Partners

Elias obtained his PhD from the Segura Lab in 2019 with a thesis titled “Hydrogel scaffolds and their influence on the brain.” He is now a consultant at Clearview Healthcare Partners based in San Francisco.

Sept 2015 – June 2019

Elias Sideris

Clearview Healthcare Partners

Impact Factor:

02.02.20
Nature Materials

Hydrogel micopraticles for
Biomedical aplications

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